Structure and Function of the Limbic System, (Progress in Brain Research, Vol. 27)

PROGRESS I N BRAIN RESEARCH

ADVISORY B O A R D W. Bargmann

H. T. Chang E. De Robertis

J. C. Eccles J. D. French H. HydCn

J. Ariens Kappers S. A. Sarkisov J. P. SchadC

F. 0. Schmitt

Kiel Shanghai Buenos Aires Canberra Los Angeles

Goteborg Amsterdam Moscow Amsterdam Brookline (Mass.)

T. Tokizane

Tokyo

J. Z . Young

London

P R O G R E S S IN B R A I N R E S E A R C H VOLUME 27

STRUCTURE AND FUNCTION OF THE LIMBIC SYSTEM EDITED BY

W. ROSS A D E Y Brain Research Institute, University of California, Los Angeles ( U.S.A.) AND

T. TOKIZANE Institute of Brain Research, University of Tokyo, Tokyo (Japan)

ELSEVIER PUBLISHING COMPANY A M S T E R D A M / LONDON / N E W YORK

1967

tttttttt

335 J A N VAN GALENSTRAAT, P.O. BOX 211, A M S T E R D A M

A M E R I C A N ELSEVIER P U B L I S H I N G COMPANY, INC. 52 VANDERBILT AVENUE, NEW YORK, N.Y. 10017

ELSEVIER P U B L I S H I N G C O M P A N Y L I M I T E D R I P P L E S I D E C O M M E R C I A L ESTATE BARKING, ESSEX

This volume contains the Proceedings of a SYMPOSIUM O N THE STRUCTURE A N D FUNCTION O F THE LIMBIC SYSTEM

organized by the Brain Research Institute, University of Tokyo, and held in connection with the XXIZZrd Znternational Congress of Physiological Sciences at Hakone, Japan in September 1965

LIBRARY O F C O N G R E S S CATALOG C A R D N U M B E R 67-12719

W I T H 266 I L L U S T R A T I O N S A N D 30 TABLES

A L L R I G H T S RESERVED T H I S BOOK O R ANY P A R T T H E R E O F MAY N O T BE R E P R O D U C E D I N A N Y FORM, I N C L U D I N G PHOTOSTATIC O R M I C R O F I L M F O R M , W I T H O U T W R I T T E N P E R M I S S I O N FROM T H E P U B L I S H E R S

,

I,,

,

I

P R I N T E D I N TWE N E T H E R L A N D S

List of Contributors

W. R. ADEY,Departments of Anatomy and Physiology, and Brain Research, University of California, Los Angeles, Calif. (U.S.A.). E. SH. AIRAPETYANTS, The Pavlov Institute of Physiology, USSR Academy of Sciences and The University of Leningrad, Leningrad (U.S.S.R.). P. ANDERSEN, Laboratory of Neurophysiology, Institute of Anatomy, University of Oslo, Oslo (Norway). F. BERGMANN, Department of Pharmacology, Hebrew University, Hadassah Medical School, Jerusalem (Israel). V. M. BUCHER, Department of Physiology, University of Zurich, Zurich (Switzerland). M. CHAIMOVITZ, Department of Pharmacology, Hebrew University, Hadassah Medical School, Jerusalem (Israel). C. D. CLEMENTE, Department of Anatomy and Brain Research Institute, Univexsity of California, Los Angeles, Calif. (U.S.A.). A. COSTIN,Department of Pharmacology, Hebrew University, Hadassah Medical School, Jerusalem (Israel). M. R. COVIAN, School of Medicine, Department of Physiology, Ribeir2o Pr&to,S. P. (Brad). J. M. R. DELGADO, Department of Physiology, Yale University School of Medicine, New Haven, Conn. (U.S.A.). Department of Pharmacology, University of Michigan, Ann Arbor, E. F. DOMINO, Mich. (U.S.A.). A. T. DREN,Department of Pharmacology, University of Michigan, Ann Arbor, Mich. (U.S.A.). M. D. EGGER,Departments of Anatomy and Psychiatry, Yale University,'School of Medicine, New Haven, Conn. (U.S.A.). E. ENDROCZI, Department of Physiology, Medical School, PCcs (Hungary). J. P. FLY", Department of Anatomy and Psychiatry, Yale University School of Medicine, New Haven, Conn. (U.S.A.). S. S. Fox, Department of Psychology, and Mental Health Research Institute, University of Michigan, Ann Arbor, Mich. (U.S.A.). J. A. GERGEN,Department of Physiology, Bowman Gray School of Medicine, Winston-Salem, N.C. (U.S.A.). K. HIROSE,Shionogi Research Laboratory, Shionogi & Co., Ltd., Fukushima-ku, Osaka (Japan). R. D. HUGHDINGLE,Department of Psychology, and Mental Health Research Institute, University of Michigan, Ann Arbor, Mich. (U.S.A.).

R. W. HUNSPERGFJR, Department of Physiology, University of Zurich, Zurich (Switzerland). N. ITOIGAWA, Second Department of Physiology, Osaka University, Medical School, Osaka (Japan). M. KAWAKAMI,Second Department of Physiology, Yokohama University School of Medicine, Yokohama (Japan). R. =DO, Shionogi Research Laboratory, Shionogi & Co., Ltd., Fukushima-ku, Osaka (Japan). K. F. KILLAM, Department of Pharmacology, Stanford University School of Medicine, Palo Alto, Calif. (U.S.A.). E. KINGKILLAM,Department of Pharmacology, Stanford University School of Medicine, Palo Alto, Calif. (U.S.A.). F. KLINGBFJRG, Department of Clinical Neurophysiology, Neurological-Psychiatric Clinic, Karl-Marx University, Leipzig (German Democratic Republic). N. KOBAYASHJ, Department of Physiology, Faculty of Medicine, Kanazawa University, Kanazawa (Japan). J. C. LIEBESKINDE, Department of Physiology, and Mental Health Research Institute, University of Michigan, Ann Arbor, Mich. (U.S.A.). K. LISSAK, Department of Physiology, University Medical School, PCcs (Hungary).

T. WMO,Laboratory of Neurophysiology, Institute of Anatomy, University of Oslo,

Oslo (Norway). A. MATSUSHITA, Shionogi Research Laboratory, Shionogi & Co., Ltd., Fukushimaku, Osaka (Japan). K. MIYAMOTO, Second Department of Physiology, Osaka University Medical School, Osaka (Japan). F. NAKA,Department of Physiology, Faculty of Medicine, Kanazawa University, Kanazawa (Japan). H. NAKAO, Department of Neuropsychiatry, Kyushu University School of Medicine, Fukuoka (Japan). H. NIKI, Department of Psychology, College of General Education, University of Tokyo, Tokyo (Japan). J. H. O B m , Department of Physiology, and Mental Health Research Institute, University of Michigan, Ann Arbor, Mich. (U.S.A.). J. OLDS,Department of Psychology, The University of Michigan, Ann Arbor, Mich. (U.S.A.). T. ONO, Department of Physiology, Faculty of Medicine, Kanazawa University, Kanazawa (Japan). P. L. PARMEGGIANI, Istituto di Fisiologia umana dell'Universit8, Bologna (Italy). P. PASSOUANT, Laboratoire de Pathologie ExpCrimentale, Facultd de Mtdecine, Universitt de Montpellier, Montpellier (France). L. PICKENHAIN,Department of Clinical Neurophysiology, Neurological-Psychiatric Clinic, Karl-Marx University, Leipzig (German Democratic Republic). K. H. PRIBRAM, Stanford University, School of Medicine, Palo Alto, Calif. (U.S.A.).

C. PTERNITIS, Laboratoire de Pathologie Exptrimentale, Facultt de Mtdecine, Universit6 de Montpellier, Montpellier (France). K. SETO,Second Department of Physiology, Yokohama University School of Medicine, Yokohama (Japan). T. S. SOTNICHENKO, The Pavlov Institute of Physiology, USSR Academy of Sciences and the University of Leningrad, Leningrad (U.S.S.R.). M. B. STERMAN, The Sepulveda V. A. Hospital, Sepulveda, Calif. (U.S.A.). E. TERASAWA, Second Department of Physiology, Yokohama University School of Medicine, Yokohama (Japan). Y. YAMAGUCHI, Second Department of Physiology, Osaka University Medical School, Osaka (Japan). K-I. YAMAMOTO, Department of Neuropharmacology, Shionogi Research Laboratory, Osaka (Japan). N. YOSHII,Second Department of Physiology, Osaka University Medical School, Osaka (Japan).

Neuronal Mechanism of Feeding Y U T A K A OOMURA, HIROSHI OOYAMA, TETSURO YAMAMOTO, FUMIHIKO N A K A , N O B U Y A S U KOBAYASHI A N D TAKETOSHI O N 0 Department of Physiology, Faculty of Medicine, Kanazawa University, Kanazawa (Japan)

tt has been generally acknowledged that food intake is controlled by hypothalamic activity, through the observations of a decrease (Hetherington and Ranson, 1940) or increase (Hetherington and Ranson, 1942; Brobeck, 1946) in food intake due to a lesion within the hypothalamus. In 1951, by systematic experiments, Anand and Brobeck clarified the hypothalamic mechanism of feeding. Using rats and cats, they observed that the animals with bilateral lesions made by current passage through an extreme lateral part of the lateral hypothalamic area (LH) at the same rostrocaudal level as the ventromedial hypothalamic nucleus (HVM), showed aphagia (and adipsia) which finally ended in death, if gastric feeding was not applied, due to starvation in spite of the availability of food, and that bilateral lesions of the HVM produced hyperphagia and obesity supposed to be a release phenomenon from an inhibitory mechanism of the HVM. From their results, the LH (at the rostrocaudal plane of the HVM) was termed the feeding center, and the HVM the satiety center. These observations were supported later by Delgado and Anand's (1953) experiment on cats which produced an increase in food intake by electrical stimulation through chronically implanted electrodes in LH. It was demonstrated further that the animals actually ate food, even to the point of satiety, upon stimulation of the LH (Smith, 1956; Miller, 1960). Conversely, by electrical stimulation of the HVM, the cats stopped chewing in the middle of feeding activity and even dropped the food from their mouths (Oomura et al., 1967). Owing to extensive connections between the hypothalamus and various limbic structures, the dual hypothalamic function on feeding is significantly modulated and integrated by those structures (Stevenson, 1964). Many experiments' of lesions of the amygdala (AM) produced hyperphagia and obesity in cats (Koikegami et al., 1955; Green et al., 1957; Fuller et al., 1957; Wood, 1958; Morgane and Kosman, 1960). Fonberg and Delgado (1961) observed, using cats with implanted electrodes, an inhibition of food intake by electrical stimulation of the baso-lateral nucleus of the AM. Oomura et al. (1965, unpublished observation) also confirmed the same result by giving repetitive electrical stimulation to the lateral nucleus of the AM as in the instance of the HVM stimulation. Similar behavioral changes were also induced by septa1 stimulation (Fonberg and Delgado, 1961; Oomura, 1965, unpublished observation). Further, Morgane (1961a) showed that damage to the pallidofugal fiber system of the globus pallidus (GP) in the far lateral hypothalamic area caused consistent References p. 31-33

2

OOMURA

et al.

aphagia and adipsia, though animals lesioned in the pallido-hypothalamic tract immediately dorsal to the fornix columns developed a hyperphagia. Emphasis was, therefore, centered on the possible importance on feeding behavior of the two directed fiber trajectories to the LH and HVM respectively. The relationship between the LH and the mesencephalic tegmentum is emphasized neuroanatomically (Nauta, 1958) and functionally (Morgane, 1964). Hyperphagia was produced by lesion of the dorsolateral tegmentum (Sprague et al., 1961) and of periaqueductal gray matter beneath the superior colliculus (Skultety and Gary, 1962), but no effect was obtained in cats by electrical stimulation on the dorsomedial tegmentum (griseum centrale) during food intake (Oomura, 1965, unpublished observation). In this way, our explanations for food intake may become more precise but also more complex. Analytical studies by single unit recording within the feeding and satiety centers are still relatively few (Sawa et al., 1959; Barraclough and Cross, 1963; Tsubokawa and Sutin, 1963; Oomura et al., 1964; Anand et al., 1964). Oomura et al. (1964) observed the following reciprocal relations between the HVM and LH, when the simultaneous recording of the spontaneous unitary discharges (SUDs) were carried out from them. SUDs in one center were decreased in number by repetitive electrical stimulation of the other center where the SUDs were increased. Moreover, using a statistical treatment of the time series of SUDs, we proved a significant negative cross-correlation between both centers. Striking control and modifications on electrical activities of the HVM and LH by the influence of the limbic system such as the AM, septum, GP and mesencephalic tegmentum were also verified. From these experiments, the neuronal control mechanism of feeding will become clearer. METHODS

Forty-five cats, 2.54.0 kg in weight, were used under artificial respiration and ether anesthesia. To decrease bronchial secretions, atropin was sometimes administered intraperitoneally in doses of 25 mg/kg body weight. In the later experiments on the effect of stimulation of the septa1 nucleus and of the dorsomedial mesencephalic tegmentum, animals were immobilized with gallamine triethiodide (flaxedil) and all wound margins and pressure points were infiltrated with 1% procaine (reinfiltration every 3 h). Concentric bipolar electrodes (tip diameters about 0.1 mm) for stimulation were implanted unilaterally through small openings in the cranium into the following regions according to Jasper and Ajmone-Marsan’s stereotaxic atlas (1954) : HVM (A 11.5,L 1.5, H 4 . 5 ) , L H ( A 1l.O,L3,H-3.5)lateralandmedialnucleusoftheAM (A 11.0, L 10.5, H -6.0, and A 11.0, L 8.0, H -6.0, respectively), lateral and medial parts of the GP (A 14.0, L 7.5, H -1.0, andA 14.0,L6.0,H-2.0,respectively),septal nucelus (SEP) (A 16.0, L 1.0, H 3.0), and the dorsomedial part of the mesencephalic tegmentum (A 2.0, L 2.0, H --0.5), and the electrodes were fixed there with dental cement. For extracellular recordings of the action potentials of neurons of HVM and LH, recording microelectrodes of glass pipettes filled with 6 M NaCl (about 0.1 p, tip diameter; DC resistance 40-60 MQ) or tungsten microelectrodes (about 1 p, tip

N E U R O N A L M E C H A N I S M OF F E E D I N G

3

diameter) coated with vinyl lacquer, were inserted simultaneously into the HVM (A 11.5, L 1.3, H -5.5) and LH (A 11.5, L 3.0, H -3.5) using motor-driven micromanipulators. In some experiments, an additional recording electrode was introduced into the lateral nucleus of the AM (A 11.5, L 10.5, H -6.0). The intervals of time between successive unitary discharges in each sample taken on films were measured under an enlarger and punched in paper tapes, which were then fed for statistical calculation into a general purpose electroniccomputer, NEAC 2230. When the microelectrodes had been inserted into the desired positions, and stable neurons firing with SUDs were detected, lack of neuronal responses to visual, auditory, and other skin stimuli was first verified by standard procedure. Secondly, changes in the activity of the HVM or LH on stimulation of other parts of the brain were recorded. For electrical stimulation, square pulse currents of 0.1 msec duration, 1-50 A amplitude lasting 3-5 sec were applied. c/sec, and 10-6 to At the end of each experiment, the brain, with the recording and stimulating electrodes in position, was perfused by neutral formol-saline solution (10 %). Thus the tips of the electrodes were fixed at the site of the last recorded unit. Electrode sites were verified histologically. Each neuron used as data for this paper was located within the HVM, LH or other desired region. RESULTS

( A ) Relationship between the HVM and LH activity

( 1 ) Spontanegus unitary discharges (SUDs) from the HVM and LH and the depth OJ anesthesia Depth of anesthesia. Simultaneous recording of SUDs from the HVM and LH longer than 40 sec was so difficult, though the recording electrodes were placed at the desired postions, that only 13 of the experiments could be treated statistically. Our predictable rough identification of HVM neurons during the experiment was derived from the observation of an increase and decrease in SUDs frequency upon repetitive electrical stimulation of 50 c/sec for a few seconds applied to the AM and LH respectively. SUDs were, in general, much influenced by the depth of ether anesthesia (Oomura et al., 1964). When the cat was in a half arousal stage (we called this lignt anesthesia), showing responses or moving the ears spontaneously to various sensory stimuli, the frequency (number of impulses/sec) of SUDs was 8-20 c/sec in HVM, while it was 2-6 clsec in LH. Whereas, under the deeply anesthetized condition, showing no response to stimuli and with their muscle tonus relaxed, it decreased to 0-3 c/sec in HVM and increased to 7-20 in LH. It has been believed that though the neuronal activity of the hypothalamus is affected inhibitorily by pentobarbital (Brooks, 1959; Stuart et al., 1964) and chloralose (Stuart et al., 1964), it is not, or only slightly, affected by ether except at the neocortex. In these experiments, however, SUD in the hypothalamus was undoubtedly affected by ether. Fig. 1 shows 16 instances of the mean frequencies of the simultaneous SUDs recordings from the HVM (Y, ordinate) and LH (X, abscissa) References p. 31-33

4

OOMURA

-

et al.

15L

> v

fI

-H

510-

C

f

5-

L

Fig. 1. Mean frequency (number of impulses/sec) of the spontaneous unitary discharges (SUDs) recorded simultaneouslyfrom the HVM and LH under deep as well as light anesthesia. Summarized from 16 records. Ordinate, mean frequency in HVM or);abscissa, that in LH 0. The straight line drawn by Y = - 1.1 X 18 with a correlationcoefficient r,, = -0.78. In mean frequency,increase in the HVM and decrease in the LH indicates light anesthesia (Oomura er d.,1967).

+

under various anesthetic stages, where the electrode tips were proved histologically to be in the right places. When the HVM neurons increased in frequency, those in LH decreased, and vice versa, showing the relation expressed by the equation, Y = -1.1 X 18, with a correlation coefficient, rxy = -0.78 (P< 0.01). Throughout the whole time course of change in depth of anesthesia produced by altering the amount of ether inhalation, continuous simultaneous SUDs recording from both centers was extremely difficult ,so that only three experiments were successful. At the intqmediate stage from deep to light anesthesia, the frequency was increased in HVM and decreased in LH, then became nearly the same. In another case, the frequenciesof HVM and LH under light anesthesia, 10.1 & 1.1 (N = 44)and 4.4 f 2.8 (N = 44)respectively, became 10.0 f 2.2 (N = 6 2 ) and 9.5 3.4l (N = 60) respectively during the intermediate stage. Discharge pattern. First, the discharge patterns in HVM and LH were determined from the histograms of frequency and interspike intervals of SUDs recorded simultaneously from both. Sampling periods for which these histograms were based were taken from the sequential ordering of neuronal activity, which was verified statistically as the stationary process by the F-test mentioned in the next section. The close accord of the observed and theoretical distributions was estimated by the Chi-square test (P < 0.05). Under deep anesthesia, the frequency distribution agreed with the results which Oomura et al. (1964) have already given, e.g. as illustrated in Nos. 1, 2, 5 and 12 of Table I, whenever the mean frequency of the HVM was as low as 2-3 c/sec, differences between mean values of frequency and standard deviations were small, and the pattern corresponded with the Poisson distribution, while the mean frequency of the LH was as high as 10-20 c/sec, with large differences between mean values and standard deviations, and it corresponded with the Gaussian distribution. In regard to the pattern of the histogram of interspike intervals, as also shown in Nos. 1, 2, 5 and 12 of Table I and Fig. 2. the distribution of HVM, mean intervals 25&900 msec fit an

+

TABLE I

a ! %

2

E

e :: i bJ

DIS T RIB UT IO N PATTERN, T H E M E A N FREQUENCY ( N U MBER OF IMP U LS ES /S EC ) A N D MEA N I N T E R S P I K E I N T E R V A L O F S P O N T A N E O U S UNITARY D I S C H A R G E S (SUDS) RECORDED SIMULTANEOUSLY FROM H V M A N D L H (Oomura el al., 1967)

From the histogram of number of impulses/sec (with mean & standard deviation), the Poisson or Gaussian distribution is determined. From the histogram of the interspike intervals,the exponential (Exp.) or r (order 2, T2,)distribution is determined. Total number in parentheses. For determination of the pattern, X2-test was performed at P < 0.05, e.g. ~5~ > 4.6,5 degrees of freedom. Two and three units from the HVM were recorded simultaneously by the same electrode in Unit Nos. 1 and 5 respectively. In No. 1, 3.2 f 1.5 shows mean frequency measured without differentiation of the two spikes.

LH

HVM Unit No.

1

2 5

Number of impulseslsec Mean

Pattern

Mean

3.2 f 1.5 (45) 2.0 f 1.4 (71) 1.2 f 0.4 1(66) 2.5 & 1.6

Poisson X52 > 4.6 Poisson X2 > 4.8 Poisson Xz2 > 3.2 Poisson Xa2 > 4.2 Poisson Xs2 > 4.6 Poisson x 4 2 > 5.3

250 f 140 (199) 434 & 383 (118) 912 & 648 (52) 330 & 250 (30) 350 f 250 (137) 282 f 2 3 5 (303) 375 i-300 (166)

;:1

(77) 2.5 f 1.4 3.3 f 1.8

-

7 8

9 12

-

Interspike interval (msec)

15.8 f 2.1 (36) 15.1 f 3.1

(44)

14.7 i 2.2 (50) 2.2 & 1.3 (46)

-

not Poisson Gauss

3.3 Poisson X32 > 5.1 Xl2>

65 5 47.5 (310) 70 rt 15 (630)

Pattern

Exp. X72

> 9.6

Exp. X2 > 4.1 Exp. X32 > 3.8 Exp. x32> 1.1 Exp. X32 > 7.0 Exp. Xe2 > 4.0 Exp. X72 > 12.3

XS2

> 9.2

not Fa

Number of impulseslsec Mean

19.1 f 3.2 (45)

Pattern

Gauss x 1 2

> 0.1

Interspike interval (msec) Mean

47 f 1 (825)

Pattern

z

m

C

w

0

z

k

r2

> 11

x72

r m 3 ct X

k

10.8 & 2.3 (77) 11.5 & 1.3 (92)

Gauss x22

> 1.1

Gauss Xi2 > 2.4

110 63.7 (177)

172 X? > 13

5 m 3

0 .rt

w

B

m

ElZ 0

6.3 & 2.7 (36) 2.1 f 2.8 (44)

Poisson x 4 2> 1 Poisson x 2 2>3

19.8 f 3.0 (46)

Gauss

406 f 220 (40) 343 70 (95)

Exp. x 5 2

> 10.1

Exp.

X32 >

7.1

Xa2 > 3.2 v,

6

OOMURA

et al.

exponential curve, whereas that of LH fits a curve of a drawn from the following equation (1): n = N/ 2 -

"Xi

+1

I' distribution of order 2 ( r ~ )

a2xe-ax dx

Xi

where a = =;x, mean interval; N, total number. X

Under light anesthesia, as already mentioned, the SUD frequency increased in the HVM to 10-15 c/sec and decreased in the LH to 3-7 c/sec. Consequently, completely reversed patterns of SUDs could be obtained. As illustrated in Nos. 7, 8 and 9 of Table I, the HVM frequency was no longer the Poisson distribution but a Gaussian one, the Poisson distribution being in LH. Accordingly, the distribution pattern of the interspike interval in the HVM was a TZwith mean intervals of 60-70 msec, as shown in Fig. 2, whereas in the LH it was exponential with mean intervals of 120-550 msec. From these results, it is proved that when the SUD frequency decreased greatly either in the HVM or LH, the neuronal discharges arose from a perfect Poisson process, whereas after an increase in activity level they appeared more or less in a regular manner. The results also indicate an intimate reciprocal relationship between the HVM and LH activities, with a highly significant inverse relation of SUDs with a coefficient of - 0 . 7 8 . When recorded units in one center discharged according to a pattern of distribution, the other units in the same region or center would be expected to discharge in the same pattern, too. This postulate was verified by simultaneous recording of SUDs of two units in one center. As with the two examples shown in Nos. 1 and 5 of Table I, two series of SUDs having different amplitudes (three units in No. 5) were recorded simultaneously by the same microelectrode in HVM. The distribution patterns of frequency and of interspike interval were Poisson and exponential distributions respectively in both units. Their mean values, however, were different, which means that they were not working in perfect synchronism. Thus it is likely that various units in the same center, at least in the vicinity, act approximately uniformly. (2) Correlationfunction between the activity of the HVM and LH Simultaneous recording of SUDs for more than 30 sec in the HVM and LH, was treated for an estimation of the auto- and cross-correlation function. Before these statistical treatments, the stationary state of the SUDs had to be checked. Neither the mean nor the degree of variability of the activity (discharge frequency) under study was expected to change significantlythroughout the sampling perigd. For the stationary state test, an F-test was employed, the analysis of variance described by Werner and Mountcastle (1963), and the criterion of stationary state was that of no significance level at P < 0.05. (i) Correlation calculated from the frequency at relatively long z. The auto- and cross-correlation functions were evaluated from the record in which the stationary state was already verified by the following equation (2) :

NEURONAL MECHANISM OF FEEDING

7

7

\,

Deep anesthesia

HVM (40 rec) N=58

534 f 5N.O msec

Fig. 2. Histograms of interspike interval of SUDS in the HVM (left) and the L H (right), recorded simultaneously. Under deep anesthesia, mean interval and standard deviation (2 r t b ) in the HVM is 534 f 574.0 msec (total number N = 58). The distribvtion pattern fits the theoretical exponential curve drawn from the equation, n (N

= 441), and the distribution pattern

equation, n

=N

,.Xi+l

J Xi

a2x e-&" dx, (a

=N

4eX

X

(continuous curve). In LH, 86 f 44.7 msec

fits the theoretical r-function of order 2 ( r z ) , drawn from the

=

*

5). Under light anesthesia, the mean interval, 65 & 47.5 X

msec (N = 310) in the HVM, and the distribution pattern fits the r a type, while in the LH it was 406 f 222.0 mSec (N = 42) and fits the exponential curve. Continuous lines are theoretical curves. n

(yi + k -

i) (xi - x)

[XY(tk), cross -correlation coefficient from x to y series at the time lag. of ket. yi (y! + k), number of SUD in ith {(i 4- k)th} time bin ti (ti + k) of y series; Xi (Xi + k), References p. 31-33

00

TABLE I1 C R O S S - C O R R E ~ A T I O N SOF

SUDS BETWEEN HVM

AND

LH (Oomura et al., 1967)

Calculatedfrom equation(2). T = 1 sec except where specified as T = 0.5 sec. Total number in parentheses. Cxlxain No.1 : corss-correlations calculated from two SUDS(XI and xa) recorded simultaneously from the HVM (No. 1 in Table I). X = HVM; Y = LH. Unit no.

1

Anesthesia

Location of electrodes

D=P

Correct

-0.46 (42) t = 0.5

D=P

Correct

-0.44

D=P

Correct

D=P

Correct

D=P

Correct

D=P

Correct

-0.32 (36)

Light

Correct

-0.27 (49)

Light

Correct

0

I

2

3

4

5 see

-0.36

5YX1

(64)

-0.36

CXZY

(62)

- 0.32

5YXZ

(65) 2

CYX

3

5YX

(34) t = 0.5 4

-0.55

5YX

(17)

5

5X1Y 5X2Y 5XaY

6

SXY

No correlation (92) 0.21 (92)

-0.21

0.16 (185)

-0.18

t = 0.5 5YX

7

tXY

8

tXY

(90) (1 82) 0.19 (184)

-0.12 (182)

e0 *

b

P

% rn

$w

‘p

Y

9

CYX

10

CXY

-0.31 (41) t = 0.5

0.42 (37)

-0.32

Light

Correct

Light

Correct

(43)

11

5XY

0.28 (64)

0.26 (63)

Intermediate Correct

0.34 (61)

No correlation (46)

12

Deep

LH, increases in frequency by AM stimulation

Deep

LH, correct; HVM, 2 mm dorsal from HVM

0.25 (63) 0.29 (63)

Deep

Correct

0.23 (89)

Light

Correct

Deep

Correct

D ~ P

Correct

i ,

bJ

13

5XY 5YX

14

CYiYa

15

h y a

0.31 (89)

0.24 (84)

0.24 (84)

0.12 0.18 (260) (259) t = 0.5 0.38 (55)

t = 0.5

0.15 0.12 (254) (253)

10

OOMURA ef

al.

the same as above but for the x series; % (i), mean number of SUDs per unit time z calculated for the whole period; n, total number of ti. The criterion of a statistical significance level of the cross-correlation followed the central limit theorem (Freund, 1960), and the value of the correlation function over 1.96/dn is significant(the whole period used for the computation was nt). Table I1 summarizes the data obtained. Only significant values of the cross-correlation function during spontaneous activity between the HVM and LH are included. Negative cross-correlations at 1 to 3 sec were obtained in Nos. 1 , 2 , 3 , 4 , 5 and 6, except No. 5x1. For example, in No. 2, when the LH neuron discharged at more than average frequency for 1 t (0.5 sec), the HVM neuron discharged less than its average, sixfold z (3 sec) afterwards. In other words, low activity of the HVM neuron followed high activity of the LH neuron with a delay of 3 sec. In Nos. 1 and 5, two (XI and XZ)and three (XI, x2 and x3) unitary discharges with differept amplitudes were recorded from the same electrode in two HVMs respectively. In the former, the cross-correlations between one unit of HVM, XI and that of LH, and between the other unit of HVM, x2 and that of LH were also significantly negative at 2, and 4 and 1 sec respectively. In the latter, a significant correlation did not appear between one unit of HVM and LH, but significant correlations were between two other units of HVM and the same LH unit. In No. 6 (also in No. 5-&), positive cross-correlations preceded negative ones, i.e. during and after 1 sec of LH discharging more than the average, the HVM unit also discharged more than the average and then less than the average two sec later. Significant negative cross-correlations between the spontaneous activities in HVM and LH existed under light anesthesia as shown in Nos. 7, 8, 9 and 10 of Table 11. On the other hand, only positive cross-correlations were obtained in Nos. 11 and 13. As to the correlation in No. 1 1 , the same units as those of No. 10;were under an intermediate stage from a light anesthesia to a deep one due to an increase in ether inhalation. In No. 12, a cross-correlation function was insignificant, though the LH electrode was in the designed place in LH, probably due to an abnormal response of the LH neuron. That is, the SUDs were not decreased in number by repetitive stimulation applied to the AM, but augmented instead. From the evidence that the activity of the LH neurons was lowered by AM stimulation in more than 80 % of the recorded units, a typical example is given in Fig. 7, the LH neuron of:No. 12 may be supposed to be a special one not affected by the HVM. In No. 13, the HVM electrodewaslocated 2 mm dorsal from the upper margin of the HVM, while the LH electrode was within the intended place, and only positive cross-correlationwas obtained. The physiological function of the region of the former site in the hypothalamus is still unknown. As described before, and clear from Nos. 1 and 5 of Table I, in the discharge patterns of two units recorded simultaneously from one HVM, the distributions of the frequency were of the Poisson type, and of the interspike interval were exponential, showing that their activities were all Poisson processes though each value was different. The cross-correlation function between the activity of the two units, XI and XZ,cleaily indicated significant positive values (lower part in Table IT), while significant negative correlations appeared between XI and LH and between X2 and LH. Nos. 14 and 15 of Table I1 show similar positive correlations between two units acting spontaneously in

11

N E U R O N A L M E C H A N I S M OF F E E D I N G

LH. Consequently,one may suppose that in the same center, or at least in neurons in the very vicinity, the patterns of neuronal activitiestend to be the same, though asynchronous, and between the two centers, HVM and LH, they are reciprocal. As to the autocorrelation function calculated at t = 0.3 to 1 sec, no periodicity of this function appeared at any time, as it did on the SUD in the thalamic neurons (Werner and Mountcastle, 1963). (ii) Auto- and cross-correlation at short unit time. Serial dependencies of the discharge occurring over a time range shorter than the unit time lag, z of 0.3 to 1 sec, escaped detection by the above method. Therefore, the correlations at short unit time, z of 10 msec, were calculated. A c t o c o r r e I a t iD n N=90,

Deep a n c s l h e s l a

1=900

L 1 8 h t anesthesia

L H

0.10005 -

L H

iarf(il

Hvu

I l f ( ' )

015.010-

ODS-

-005-010-015-

-015-

d

Fig. 3. Autocorrelation (5 (t)) calculated at short unit time (t = 10 rnsec). Upper: under deep anesthesia (No. 2 in Table I and 11; N = 907, i.e. total period, Nt,taken for the calculation is 9.07 sec), (t) of the HVM unit is almost zero except at t = 0 (discharge pattern of the HVM is a Poisson distribution. In the LH, ((t) is negative until about 50 msec except at t = 0. The values of [(t) closely fit the 1 theoretical curve (dotted line) drawn from the equation [(t) = - kAAt (A =.: ,x-, mean discharge X

interval). Lower: under light anesthesia (No. 8 in Table I and II, N = 900), [(t) of the LH unit is almost zero (dischafge pattern shows a Poisson distribution). [(t) of the HVM unit is negative until about 40 msec (discharge pattern, r2 type), also fits the theoretical curve (Oomura el al., 1967). ReJerences p. 31-33

12

OOMURA

et al.

First, Nos. 2 and 8 of Table I1 as typical examples of deep and light anesthesia respectively, were analyzed. The autocorrelation functions of the HVM unit of No. 2 and LH unit of No. 8 were almost zero up to 100 msec, as shown in Fig. 3. Since their discharge patterns indicated the Poisson process, this result was natural. About the LH unit of No. 2 and HVM unit of No. 8, whose discharge patterns were more or less regular with interval distributions of I'z type, the autocorrelation functions were negative up to about 50 msec, except at t = 0, then approached zero, as shown in Fig. 3. When the distribution pattern of discharge intervals corresponded to the r2 type, the autocorrelation function was obtained theoretically, which is given in the equation (3) : CX (t) = -1e-41t (3) 1 il = -..-;x, mean SUD interval X

The significant level of the autocorrelation function is not yet determined, but as shown in Fig. 3, the results and the theoretical curves fit very well, which implies physiologically that once a neuronal unit discharged, a lowered state of excitability lasts for the following 50 msec due to some mechanism. Since the mean intervals of such SUDs were always more than 40 msec, this lowered excitabilityis not attributable to the post-tetanic depression (Eccles, 1964) or post-tetanic hyperpolarization (Eccles, 1953) but probably due to neuronal refractoriness (Lloyd, 1951), accompanied by positive after-potential or recurrent inhibition. The meanings of equation (3) will be discussed in more detail later. As to the autocorrelation function at the short t,no periodicity was found in the present experiment as it was in the thalamic neuron every 50 to 70 msec (Poggio and Viernstein, 1964). (iii) Correlation calculatedfrom discharge intervals. To calculate the auto- and crosscorrelation functions from the discharge intervals, the method described by Gerstein and Kiang (1960) was employed. When the total period of the record for analysis, T, and the total number of SUDs in HVM and LH, X and Y contained in T, is extremely big, a conclusion could be derived from plxu(t),an expression in their method. But in our experiments, the amount of T, X and Y were usually so small (T, 20 sec; 40 impulses for X or Y at minimum) that tpXy(t)had to be normalized and tested for its significance level. Therefore, ex= (t)was evaluated by the following equation (4):

xi, yi, impulse number in HVM and LH respectively during A i t . At AT X--, Y--, mean impulse number in HVM and LH respectively.

T T The statistically significant level of the cross-correlation was also tested by the central limit theorem (Freund, 1961).

P

TABLE I11 C O M P A R I S O N B E T W E E NT H E CROSS-CORRELATION COEFFICIENT (X, HVM ;Y, LH) C A L C U L A T E D FROM EQUATION EQUATION (4) (DIGITAL) (Oomura et al., 1967) t = 333

msec; N, 57. Case of No. 2 in Table I.

1.66

2

2.33

0.14 -0.07

0.21

0.09

0.07 -0.09

-0.29* 4 . 1 3

0.07 4 . 1 9

0.02 -0.07

0.17 -0.12

-0.29* 4 . 2 0 -0.03

(2)

( A N A L O GUE) A N D

Q

Time lag

0.33

Analogue -0.02 Digital

*

-0.09

Significant

0.66 -0.07

0.02

1

1.33

2.66

3

3.33

3.66

4

0.07 -0.25*

4.33

4.66

5 sec

0.19 -0.25*

0.12

-0.22* -0.03

-0.27*

0.19

m

0

e

m m

L

w

14

OOMURA

et al.

Table 111 shows the cross-correlation of No. 2 in Table calculated by this method at t = 333 msec. For comparison, the cross-correlation calculated by equation (2) is shown. The significant value of the cross-correlation appearing at 3, 4 and 4.66 sec by the latter, are also all significant in the result calculated by the equation (4). ( 3 ) Change in SUD provoked by electrical stimulation After stable spontaneous firing neurons had been detected, responses to repetitive electrical stimulations in HVM or LH were investigated. The results were coincident with those obtained by Oomura et al. \1964). When LH was stimulated, the SUDs in 30 units out of 44 of the HVM decreased or ceased, in 5 units increased and in the remaining 9 units did not change. With increased stimulus intensity, this decrease lasted not only during the stimulation but also for several seconds following it, and then the SUD returned to its original level. One typical example (50 c/sec, 3 V for 3 sec) is shown in Fig. 4. Conversely, HVM repetitive stimulation inhibited SUDs in 15 LH units out of 24 (Fig. 4). The remaining 3 LH units tended to be facilitated slightly by 15t

.

*

H.V.M

10

10

30

SIC

Fig. 4. Left two records:inhibition of SUD in the HVM (left) and LH (right) by repetitive stimulation (50 c/sec, 0.1 msec duration, 3 V, for several seconds) upon the LH and HVM respectively. Top to bottom and left to right. Stimulation artifacts are seen in the middle of each left column. Horizontal bar: 1 sec (one sweep). Right two curves, inhibitoryeffect in the HVM (top) and the LH (bottom) as seen in the records. Ordinates, impulses per sec; abscissae, time (sec). Arrows indicate stimulation periods. Right and left units are different (Oomura et al., 1967).

stronger stimulation, and 6 units did not respond at all to stimulation of any strength. The inhibitory effect did not always become clear with the lowering of stimulus frequency. It became apparent, however, when the cross-correlation function between stimulus and SUDs was evaluated by the method mentioned on p. 12. Stimulation of the LH even at 1 c/sec still affected the HVM activity. For example, as shown in Fig. 5, the significantly negative cross-correlation (t = 25 msec) which lasted more than 75 msec, was followed by significantly positive correlations at 125 and 275 msec. This negative value may indicate a long-lasting inhibition caused by hyperpolarization of

NEURONAL MECHANISM OF FEEDING

15

Cross-correlation 1 = 25nrec

N = 1380

L H IWSSlirn - H V M

0.15

SUO

2 -lH&largeqikc)

Jk ...................

?set

ft

........................

- ao

-0.05

1

Mean Freqirec Ilk0 2 6 t I x 6 5rO 3(S t )

1

Mean Freqirec

18f05(St)~37rOd(St)

Fig. 5. Upper: inhibition of HVM activity by LH stimulation. Inset records show a series of HVM SUDs, and 1 c/sec stimulation (0.1 rnsec, 3 V indicated as an artifact by a dot) was applied to the LH for 34.5 sec. To make clear the inhibitory effect of stimulation on the HVM SUD, cross-correlation C(t) between stimulation pulses and SUDSin the HVM are shown. Method described on p. 12. t = 25 msec, N = 1380. Significant negative cross-correlation (statistically significant level indicated by dotted lines) lasts more than 75 msec, then significant positive one follows at 125 and 275 msec, which may indicate a post-inhibitory excitation. Lower: facilitation of LH activity by the same LH stimulation. Two SUDS in the LH with different amplitudes (large, LH1 and small, LHz) recorded by one electrode, increase in discharge number by 10 c/sec LH stimulation (0.1 msec, 5 V, for 59 sec). In LH1, the SUD of 1.1 0.2 (mean frequency standard error) before stimulation increased to 6.5 f 0.3 during stimulation. In LH2,1.8 f 0.5 increased to 3.7 f 0.4 .T(t) between stimulation pulse and LHI unitary discharges indicate a significant positive value at 10 msec after each stimulation, which may be the latency from stimulation to response. This is followed by a significant negative value at 40 to 50 msec. This negativity may correspond to the post-excitatory inhibition. In LH2, a significant negative correlation lasting for 30 msec appears, then becomes positive at 40 msec and again becomes negative at 40-50 msec (Oomura et al., 1967).

*

the membrane followed by a post-inhibitory excitation or rebound. In contrast to the above, SUDs in LH increased up to about twice their original frequency upon LH repetitive stimulation. This facilitation lasted for a few seconds. If a mechanism of spreading depression had been responsible for the HVM inhibition by the LH stimulation, a decrease or even a cessation of SUDs shoud have appeared at first in the LH References p . 31-33

16

OOMURA

et al.

neuron which was in the vicinity of applied electrical stimulation, before giving any inhibition in the HVM neuron which was in a remote place. If, furthermore, the HVM inhibition had been caused secondarily by the increased activity in LH, it should have appeared with more delay than the latency of facilitated activity in LH which was more than 15 msec (Fig. 9). Thus these inhibitions in HVM, considering the long-lasting inhibition more than 75 msec together, imply some inhibitory synaptic mechanism. By much stronger stimulation of LH up to 10 V or so, an increase in its SUD frequency was followed by a decrease and then a complete disappearance. This could be explained by an electrotonic spread of the stimulation to the HVM, which facilitated the HVM activity,and the latter in its turn inhibited the LH. In 33 units of the SUDSof the LH, discharge frequencies increased notably by the LH stimulation in 19 experiments, decreased in 6, and in the remainder was not altered. Although SUDs in the LH were modulated by LH stimulation, statistical analysis gave more precise knowledge. The cross-correlation functions at 10 msec t between stimulation and SUDSare shown in Fig. 5 (lower). Two spontaneous unitary discharges with large and small amplitudes were recorded in one recording electrode during stimulation at 10 c/sec and 5 V applied through the stimulatingelectrode 0.5 mm apart from the recording electrode. The frequency of SUDs of the large unit, whose original value was 1.1 f 0.2 c/sec, increased up to 6.5 f0.3, while that of the small one increased from 1.8 f 0.5 to 3.7 f0.4, on stimulation. A significant positive cross-correlation appeared on the large unit from 10 to 20 msec after the stimulation, then the correlation became negative at 40 to 50 msec. On the small unit, a significant negative correlation lasting for 30 msec appeared at first, then became positive at 40 msec and again became negative at 50 msec. In other experiments, the latency of driven unitary discharges in the LH by a single volley applied to the same LH was also analyzed statistically: the curve of post-stimulus latency and probability of response showed an extremely large latency of approximately 15 msec, and this latency was not shortened by stimulation of multi volleys. Therefore, the significant positive cross-correlations at 10 to 20 msec in the large units may indicate the latency of the response; then the following negative correlations show the post-excitatory inhibition (or post-spike hyperpolarization). In the small unit, the significant positive correlation at 40 msec followed by the negative one from 50 msec, can be explained in the same way, though an elucidation of the first negative correlation is not easy and needs further study. (B) Relationship among the limbic system, HVM and L H ( I ) Functional interaction between the A M and hypothalamus To investigate the functional relationship between the HVM or LH and the AM, simultaneous recordings of SUDs from the LH and lateral nucleus of the AM (lat. principal nucleus) were first carried out with 8 successful results under flaxedil immobilization. In three instances in which clearly theoretical discharge patterns in both AM and LH were yielded, a significant negative cross-correlation between both SUDs of AM and LH resulted, but in the other 5 in which definite distribution patterns were not determined, significant cross-correlations alone did not result, but

17

N E U R O N A L M E C H A N I S M O F FEEDING

sometimes positive and negative correlations appeared mixed. Moreover, in the three experiments, the SUD frequencies were alike in both AM and LH, 10-15 c/sec. As shown in the middle and lower parts of Fig. 6, when the number of SUDs in the AM was low (about 10 c/sec) and high in the LH (about 20 c/sec) the frequencies and interspike intervals were Poisson and exponential distributions in the AM, and Gaussian and rz distributions in the LH respectively, exactly the same as with the HVM and LH in the deep ether anesthesia. A significant negative cross-correlation (t,500 msec) from the LH to AM at 3 sec was also found (upper, Fig. 6). By stimulation of the lateral nucleus of the AM, in general, an increase in SUD frequency resulted in the HVM and a decrease in the LH. One example of simultaneous recordings of SUDs in Cross-corielat ion AH-

LH

LY

P( I )

Interval histogram

n

-1H

T = 500 msec

LH

80

N=311

I =4 9 f 2 6 9 n s e c

i= got29.lmsec

0

80

40

P

E -

200

0

400 0

200 msec

100

Frequency hist ogram

An N= 37

n

i=9B+4.0

2

10

I

=36

'Ol 12 Impmmb. I sec

= 20.4 f 2 . 5

24

36

Fig. 6. Middle aad lower: discharge patterns of SUDS recorded simultaneously from the lateral nucleus of the AM and LH. Histogram of the interspike interval of the AM fits the theoretical exponential distribution (continuous line) with the mean interval of 90 f 29.1 msec (% f a), while that of LH to rs distribution with 49 f 26.6 msec. Histogram of the frequency of the AM fits a Poisson distribution with mean frequency of 9.8 f 4.0 c/sec, while that of the L H fits a Gaussian one with 20.4 f 2.5. Upper: cross-correlation of SUDs between the AM and LH calculated from the lower record, t = 500 msec; total period, 13 sec. Significant negative correlation at 3 sec, indicating a hardness of unitary discharging in the AM more than the average after 3 sec when LH unit discharged more than the average for 500 msec. References p. 31-33

18

OOMURA

et ul.

the HVM and LH is shown in Fig. 7A. In A, to make the effect of the AM stimulation clearer, the moving average of the SUD series, mean impulse numbers per 5 sec of the HVM or LH SUD were successively taken at every 1 sec interval. Responding to 10 c/sec stimulation at 10 V for 15 sec, the frequency of SUDs in the HVM increased up to more than twice its normal frequency in the fist few seconds and stayed at this level during the stimulation. It decreased along with the cessation of stimulation. In the LH, the frequency decreased to zero and even continued in this state for about 15 sec after the cessation of stimulation. To obtain the precise time course of the facilitation and inhibition on the HVM and LH activities due to the AM stimulation, the cross-correlation functions between stimulation and SUDs were calculated on a simultaneous recording of the HVM and LH units different from those in A. In Fig. 7B, by stimulation with 10 c/sec at 10 V for about 20 sec, significant positive correlations (z, 10 msec) at 40 and 130 msec were evident in the HVM, whereas a significant negative one (z, 25 msec) at 175 msec was followed by significant positive ones at 200 and 300 msec in the LH. In one example, the upper part of Fig. 7B, the latency of units in the HVM responding to more than 3 stimuli of the lateral nucleus of the AM at 100 c/sec was approximately 40 msec; on increasing stimulus number at 100 c/sec, the latency was not shortened but a multiple response appeared.A complete inhibition of the SUDs in the LH by the same stimulation lasted more than 100 msec as shown in the lower inset. Taking account of these values, the first positive cross-correlation at 40 msec in the HVM is attributable to the latency of a response from a pulse of the stimulation, and the second one at 130 msec also accounted for the latency, since the second pulse of the stimulation fell 100 msec later. In regard to the LH, the first negative cross-correlation at 175msec would be derived from the inhibitory mechanism, and the second positive correlation at 200 msee would be due to the post-inhibitory excitation, but hardly explicable on that at 300 msec. Furthermore, close functional interactions, as well as other evidence, were found between either the AM and HVM or the LH. First, the summation of synaptic potentials with about 14 msec latency could be recorded in the HVM by repetitive stimulation of the lateral nucleus of the AM, as

A

E f f e c t 0 1 A L s t i m . o n HVM I L H

set

NEURONAL MECHANISM OF FEEDING

-005

- 0.1 0

19

-

1

Fig. 7. (A) Increase in SUD frequency in the HVM (upper) and decrease in the LH (lower) by repetitive stimulation of the lateral nucleus of the AM (0.1 msec, 1-0 c/sec, 10 V, for 15 sec). Ordinates: the moving average of SUDs, i.e. mean impulse numbers/5 sec were successively taken at every 1-sec interval. (B) Upper records of insets in the right show responding unitary discharges in the HVM by volley stimulations of the lateral nucleus of the AM at 100 c/sec (0.1 msec, 10 V). The latency of units responding by more than 3 stimuli is about 40 msec (by increasing stimulus number, the latency measured from the onset of stimulation is not shortened but a multiple response appears). Upper left curve shows cross-correlation, [(t), between stimulation pulse (10 c/sec, 10 V, for 19.8 sec) upon the AM, Z, and unitary discharges in the HVM, 5 = 10 msec. Note significant positive correlations at 40 and 130 msec. These 40 and 130 msec are attributable to the latency of responses by lO-c/sec stimulation. Lower records of insets in the right show inhibitions of LH SUDS lasting more than 100 msec, by volley stimulation of AM (at 100 c/sec, 0.1 msec, 10 V). Lower left curve shows t(t) between 2 (10 c/sec, 10 V, for 19.5 sec) and SUDs in the LH, T = 25 msec. A significant negative correlation at 175 msec is followed by a positive one at 200 and 300 msec. Simultanems recording from the HVM and LH.

shown in Fig. 8, though the neuronanatomical connection between this nucleus and the HVM have not been confirmed, probably either through an unknown direct synaptic pathway or indirectly through the medial principal nucleus. Second, when conditioning single stimuli of the AM with a fixed intensity were followed by test stimuli of the LH with various intensities, the probability of LH unitary response was calculated. One example of this kind of experiment is shown in Fig. 9. At two times, 10 and 40 msec, the probability of LH response was lowered. PresumabIy the first lowering was due to a direct inhibition from the AM to the LH and the second to an inReferences p . 3 1 3 3

20

OOMURA

et al.

-

Fig. 8. Summating responses in the HVM by repetitive stimulation of the lateral nucleus of the AM, with a latency of about 14 msec. These responses are probably synaptic potentials produced in the HVM neuron. Vertical bar, 5 mV; horizontal one, 100 q e c .

direct pathway from the HVM whose activity was driven with a latency less than 40 msec by the conditioning stimulus. Twenty-four out of 40 HVM units were facilitated by the repetitive AM stimulation, though 5 other units were inhibited slightly; histological identification showed that 3 units were actually located in regions adjacent to the HVM, probably related to regions in the LH. Eleven units did not respond to any stimulation. In regard to LH units, 32 out of 55 units were inhibited by AM

I

t 1Y c o d .

1

I

30

1

I

60nstc

Interval O f t B t l Shock Fig. 9.Effect of conditioning stimulation of the lateral nucleus of the AM on LH-LH response. Right record in the inset shows a preceding AM conditioning stimulation (fixed intensity of 10 V, 0.1 msec) (first pulseat theleft)andaunitaryresponseoftheLH(thirdspikefrom theleft) by single test stimulus (0.1 msec) of LH (middle pulse, stimulus artifact). Left curve: probability of LH response (ordinate) against intervals of the conditioning and test stimulus. The probability is lowered at 10 and 40 msec. See text.

N E U R O N A L M E C H A N I S M OF F E E D I N G

21

stimulation, 11 units were facilitated to some extent and 6 units never responded. The remaining 6 units increased in frequency first, then decreased. (2) The efect of septa1 stimulation on the SUDs in the HVM and LH As already clearly demonstrated by Tsubokawa and Sutin (1963), and in the present experiments too, unitary dischargesin the HVM evoked by single or volley stimulation at 100 clsec of the central part of SEP under flaxedil immobilization appeared with latencies extending from 10 to 40 msec. Though the latencies were not definitely shortened by the increase in number of stimuli at 100 c/sec, the mean impulse numbers for 100 msec from the cessation of volley stimulation were proportionally increased by an increasing stimulus number at 100 c/sec with a positive significant correlation coefficient (in one instance, 0.86). Some neurons were fired by both AM and SEP stimulations. In contrast, with these effects of brief stimulation, on repetitive stimulation, say 50 c/sec for 10 sec, only three out of 11 HVM units increased in frequency and two units decreased after cessation of stimulation, though they were not altered during stimulation. No obvious change was seen in the remaining units. LH units were markedly inhibited by SEP stimulation. As shown in Fig. 10, in the upper records (A, B, C), a considerably long period of cessation of the LH SUDs

A

C

o . n l l Cross-corrrlation

-0.8

_____ ____-__ - -

1 ::lI S t P 4* l!ai US stiu. I I=Ill1

Fig. 10. Effect of SEP stimulation upon L H SUDs. Upper left (A, B, C ) : SUDS i0 the L H disappear for about 200 msec (B) and 350 msec (C), after single (A), 3 (B) and 4 (C) stimuli at 100 c/sec (0.1 msec, 7 V). After the inhibition, increases in number of SUDS result. As shown upper right, an increase in number of stimulations at 100 c/sec (7V, abscissa, X ) leads to a decrease in S U D numbers for 50 msec after stimulation (ordinate, Y,mean impulse number/50 msec), and this relationship is expressed by Y = - 0.242 (X - 2.5) 0.625, with a significant negative correlation coefficient of -0.825. Lower: cross-correlation between stimulus pulse of SEP stimulation (3 stimuli at 100 c/sec, 7 V, for 15 sec) and S U D s in the LH, t = 10 msec. Sigdicant negative correlation lasting more than 120 msec was followed by a significant positive one at 210-240 msec.

+

References p . 31-33

22

OOMURA

et al.

followed the stimulations of SEP, even after only a single stimulation, but was more evident after .volley stimulation, e.g. with three stimuli at 100 c/sec there was an inhibition lasting approximately 200 msec and with 4 stimuli more than 350 msec. In the right inset, the relationship between the inhibition and the volley stimuli are shown ; the mean impulse number of LH SUDs for 50 msec after stimulation was approximately inversely proportional to the stimulus number at 100 c/sec with a significant negative correlation coefficient, -0.83. To obtain a precise picture of the temporal pattern, the cross-correlation function (r, 10 msec) between SEP stimulation (three stimuli at 100 c/sec, 7 V for 15 sec) and LH SUDs was also calculated (lower, in Fig. 10). A significant negative cross-correlation lasting more than 120 msec was followed by a signscant positive one at 210-240 msec. The negative correlation is caused by the inhibition mentioned above. The latter positive one is due to post-inhibitory excitation which is proved in record c in Fig. 10. An increase in discharge frequency preceded by the inhibition is clearly seen at about 250 msec after the volley stimulation. Eleven out of 16 LHunits behaved asmentioned above on repetitive SEP stimulation, one unit increased in frequency and 4 were not altered. (3) The effect of lateral and medial parts of the GP on HVM and LH activities Repetitive stimuli of 10-50 c/sec for several seconds were applied to the lateral part of the GP. These effects on both the HVM and LH were found to be similar to the effects of stimulation of the lateral nucleus of the AM. As shown by the movement of the average curves (average frequency/sec>of the simultaneous recordings of SUDs in both the HVM and LH in Fig. 11, the SUDs of the HVM unit were increased in frequency not only during stimulation (10 c/sec, 4 V, for 7 sec), but also thereafter up to 2-3 times the original rate. This state lasted about 10 sec and then gradually returned to the original level. Six out of the 16 HVM units were affected similarly. The latency of the responding unit to single or multiple volleys at 100 c/sec stimulation was 30 to 60 msec as shown in the right upper records of Fig. 1 1. In regard to the LH units, an inhibition of activity was produced by stimulation, i.e. as shown by the moving average curve, the SUD decreased to almost zero in frequency during the stimulation and gradually went up beyond the original level. This increase was believed to be due to the post-inhibitory excitation. As shown in the upper and middle records in the right lower part of Fig. 11, the noticeable inhibitions lasting 50-100 msec produced by single volleys were followed by an increase in number of unitary discharges. As becomes clear in the records, the larger the stimulus intensity, the longer the inhibition. The lowest record shows a considerably longer period of inhibition, more than 250 msec on injury discharges from a single volley. Ten of the 17 LH units behaved almost identically. Three units increased in frequency, and 4 units were not affected by the stimulation. The effects of the medial part of the GP were in the opposite direction from those of the lateral one. In three HVM units out of 5, SUDS were decreased in frequency and gradually returned to the original level when the repetitive stimulation (50 c/sec for 5 sec, 10 V) of the medial part was switched on and off. Two units increased slight-

23

N E U R O N A L M E C H A N I S M OF F E E D I N G

HVM SUO n

L - 6 P 4 HVM L-BP

Train

s r i m . la% 4 v

1 rtlavll I t I W W

lmnr

Fig. 11. Effect of GP stimulation on SUDs in the HVM and LH. Right upper records show unitary discharges in the HVM responding to 2 stimuli of the lateral part of the G P at 100 c/sec with latencies of 30 to 60 msec. Stimulus intensity is different in all records, but around 7 V. Right, lower records: upper twoshow post-inhibitoryexcitationin theLHneuron by singlestimulations of the lateral part of the GP. The inhibitions last 50-100 msec. The lowest record shows a long lasting inhibition of injury discharges of a LH neuron more than 250 msec from a single stimulation of the lateral part of the GP. Left: upper and middle, HVM and LH SUDs increase and decrease in frequency respectively after repetitive stimulations of the lateral part of the GP (10 c/sec, 0.1 msec, 4 V, for 5 sec). Simultaneous recording from the HVM and LH. Lower, LH SUDSincrease in frequency during repetitive stimulation of the medial part of the GP (100 c/sec, 17 V, for 15 sec).

ly in frequency. On the LH SUDs, three of 11 units increased in frequency during the stimulation as shown in the lowest curve of the moving average in Fig. 11. One unit was inhibited, but 7 units were not affected by the repetitive stimulation. The latency of LH units responding to a single stimulation of the medial part of GP was 10-20 msec. ( 4 ) The effects of the mesencephalic tegmentum on HVM and LH activities Stimulation at 5-15 V, 1&50/sec of the dorsomedial part of the periaqueductal gray substance had different effects upon both HVM and LH units under flaxedil immobilization. In two out of the 10 HVM units, the moving average curve of HVM SUDS as shown in Fig. 12, decreased in frequency not only during stimulation for 3 sec but also for some time after it, then gradually returned to the original level in nearly 10 sec. But the frequency of 3 HVM units was increased and of 5 units was not altered by the stimulation. It is therefore difficult to reach a conclusion from the results. Fifteen out of the 33 LH units were considerably increased in frequency up to about five fold on stimulation of the periaqueductal gray substance for 3 sec. This effect usually lasted more than 10-15 sec after cessation of stimulation (Fig. References p . 31-33

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12), except in one instance in which the increase in frequency caused by 10 clsec stimulation went back once to the original level on cessation of stimulation but was seen again 14 sec later and continued for about 5 sec. Seven units were inhibited and the other 11 units remained unaffected by the stimulation. Unitary discharges of the LH units responding to the single volley stimulation of the dorsomedial mesencephalic tegmentum were usually 10-15 msec in latency, but in some units, as shown in the lower records in Fig. 12, repetitive firings appeared which increased in frequency due to increased stimulus intensity though the latency was extremely long.

-L I

%-

'150-

100 50

-

LH

SUO

50

I

1

I

.

I

gT

Ch 1 5 V

I

I

Fig. 12. Effect of stimulation of dorsomedial part of the dorsomedial mesencephalic tegmentum CDMT) on SUDs in the HVM (upper) and LH (middle). SUDS in the HVM decreasein frequency for about 10 sec after the stimulation (50 c/sec, 13 V), and increase in the LH for about 10 sec (50 clsec, 7.5 V). Lower records: repetitive M n g in LH neuron by DMT stimulation. On increasing the intensity of volley stimulation (9 stimuli at 50 c/sec), the number of unitary response increases.

DISCUSSION

( I ) Reciprocity of SUDs in H V M and LH. The present experiments have established the intimate reciprocal relations between the activities of the HVM and LH. Examples are as follows. (a) By applying a repetitive stimulation upon the HVM for 2-3 sec, SUDs of the LH disappeared for a considerably long period (3-15 sec, Fig. 4) and exactly the reverse relation holds true, even for single stimuli (Fig. 5). (b) Under light

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25

anesthesia in which animals showed responses to various kinds of sensory stimulation, SUDs in the HVM were high in frequency and low in the LH, both having characteristically different discharge patterns, and under deep anesthesia the relation between the frequencies was completely reversed as were the discharge patterns. In an unanesthetized and unrestrained cat, when the EEG of the HVM showed an arousal pattern of fast waves and that of the LH slow waves with high amplitude, a very slight anesthesia also produced a completely reversed pattern in EEG (Oomura et al., 1967). (c) From statistical treatments, between the SUDs in the HVM and LH there appeared a significant negative cross-correlation, i.e. when either the HVM or LH discharged more than the average in frequency for a unit time, say 250 msec, several units of time later, the other discharged less than the average. The influences of changes in blood composition (Anand et al., 1964; Iki, 1964; Oomura et al., 1964) and of visceral impulses (Anand 1961) on the HVM and LH activities were excluded from this discussion; (a> HVM and LH units were markedly affected in reverse direction by stimulating the various limbic structures. Behaviorally, as mentioned in the introduction, food intake was elicited by LH stimulation and stopped by HVM stimulation. Mechanisms and physiological functions of these reciprocities are of interest. The mutual inhibition in the eye of Limulud is well known. The frequency of an optic nerve discharge caused by illumination of one ommatidium of the Limulus eye, was decreased by an additional illumination of the neighboring ommatidium (Hartline and Ratliff, 1957). This is called the lateral inhibition which is mediated through some lateral connections in the optic nerve plexus between sensory units (Hartline et al., 1956). A big difference is that the inhibition in the hypothalamus is between neurons in the HVM and those in the LH, but not between two neighboring neurons in the same centers, whereas the lateral inhibition is between sensory units of the same kind. In our experiments, the same kind of discharge patterns and positive cross-correlations were usually confirmed between two neurons in the same center (Tables I and 11).Functionally, however, it is reasonable since the physiological functions of the HVM and LH were supposed to be of another kind; as though in two antagonistic muscles, a hyperexcitation of one center causing the hypoexcitation of the other antagonistic center, and vice versa, is in no way contradictory. The lateral inhibition works to emphasize the contrast between bright and dark areas in the visual field at their borders. The two physiological functions, the motivations of food intake and satiety, may be also emphasized in contrast, through the reciprocity of activities in the LH and HVM. Here, the contrast is not spatial but temporal. In this respect, a detailed spatial differentiation in the same center would not be required as in the eye, so mutual facilitations instead of inhibition exist between two neurons in the same center, LH or HVM. Even though it be teleological, the functions of the reciprocity do not seem difficult to understand; but to know the exact pathway through which the inhibitions are delivered between the two centers, we have much more to study, and at this stage conjectures may be inevitable. Though the HVM and LH are 2-3 mm apart from each other, SzentAgothai et al. (1962, Fig. 22) clearly demonstrated by the Golgi and Cox method that the dendrites of the HVM and LH neurons orient into mediolateral directions more than 2 mm on the References p. 31-33

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et ul.

same frontal plane, to establish contacts with neurons in each others' nuclei as well as with those in the same nucleus. They showed further by even very small lesion in the HVM, numerous degenerated fibers of extremely fine caliber in the LH, showing lots of contacts of many collaterals of the HVM with the LH neurons (Szenthgothai et al., 1962, p. 67). These connections may be considered as pathways for the reciprocal inhibition. As the neurons in the same nucleus seem to have a similar kind of connection between each other, and their activities are facilitatory rather than inhibitory, the above conjecture seems at first rather unreasonable, because it suggests that activity of one neuron, say in HVM, exerts a facilitatory effect on another nucleus, without any internuncial neuron. A few of our recorded SUDS,not described in this paper, leave some suspicion of such interneurons, though the examples were too few. As for the difficulty of one neuron exerting opposite effects on different neurons without any internuncial neuron, observations on molluscan neurons may furnish some explanation. Upon ACh application in ganglia of several species of molluscus, a type of neuron called a D cell depolarizes, and another called an H cell hyperpolarizes (Tauc and Gerschenfeld, 1962; Strumwasser, 1962; Kerkut and Cottrell, 1963 ; Oomura et al., 1965). Moreover, it was found that the impulses discharged from a single neuron actually exert direct synaptic excitatory actions on D cells, and inhibitory actions on H cells, probably both mediated by ACh. The differentiation of excitation or inhibition in this event is provided by the specific properties of the ACh receptor sites. Oomura et ul. (1965) demonstrated that these receptor sites or postsynaptic membranes are not different in their reaction to the ACh, but the difference in cell metabolism maintains differentlevels of concentration of internal chloride in D and H cells, which lead to the differentiation of depolarization and hyperpolarization, but induced by chloride permeability increase by ACh. In view of this information, the difficulty mentioned earlier may not be so grave a problem. The difference in the chemical affinities between neurons in the LH and those on the HVM is conceivable, from physiological functions attributed to them, and from the results of behavioral observation upon local applications of ACh, adrenalin, etc. to these centers (Grossman, 1962; Wagner and de Groot, 1963).

The slow time course of reactions to electrical stimuli is another problem. The latency of LH unitary discharge responding to the LH single shock was almost 15 msec (Figs. 5 and 9). As the latencies did not decrease much when the strength of stimulation was increased, supposing the latency to be solely the conduction time from the site of stimulation to the recording electrode, around 0.5 and 1 mm maximum, the conduction velocity comes out to be 3-7 cm/sec. There is still the problem as to whether active conduction takes place in the dendrite (Rall, 1962). Though Chang (1951) gave a value of 1-2 m/sec for the conduction velocity of the dendrite, Cragg and Hamlyn (1955) and Andersen (1960) obtained the value of 30-50 cm/sec and 18-60 cm/sec respectively on the apical dendrite of rabbit hippocampus (CAI), and Hild and Tasaki (1962) about 10 cm/sec (38") on the dendrite of the tissue culture neuron of kittens and rats. Our values on the conduction velocity in the LH are smaller than this last and the feast value. The reactions of neurons in these centers seem to be rather more sluggish than those in the mammalian spinal cord or cortex on which most

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quantitative analyses were carried out, so the slow velocity may not be inconceivable, or it may imply that each neuron even in the LH is connected functionally by interneurons as Eccles (1951) proposed for the cortex neurons of mammals. While there was little evidence of the interneuronal activity, almost all inhibitions or excitations of neurons in both centers seem to last fifty milliseconds or more in units of 10. The supposition of a few fast-working interneurons between the HVM and LH alone, unless their reactions are slow, will not be enough to explain such a slow onset of inhibition as seen in the negative cross-correlation which was of the order of seconds (Table 1). As already shown by Sawa et al. (1959), Tsubokawa and Sutin (1963), Oomura et al. (1964), and in the present study, the HVM and LH were in an intimate relationship, i.e. also reciprocal with the limbic system, the AM, lateral and medial parts of the GP, SEP and mesencephalic tegmentum, which may be another factor contributing to the slow time course of reciprocal correlation between HVM and LH. (2) The eflect of anesthesia on the HVM andLH. The anesthesia brought about considerable effects upon the activities of both HVM and LH. Under light anesthesia, the HVM neurons continued their activity well. They could keep discharging at the relatively high frequency of 10-20 c/sec and with a more or less regular pattern. This is common with the repetitive firing in the sensory nerve provoked by a long sustained depolarization elicited at the sensory terminals (Hagiwara, 1954). The high frequency in HVM neurons may also be due to the membrane depolarization. Depolarization at a certain level of the membrane potential greatly helped to produce hyperpolarization following spike (Hagiwara and Tasaki, 1958, Fig. lo). This after-hyperpolarization sometimes lasted more than fifty tens of msec (Kolmodin and Skoglund, 1958). In HVM neurons, with a Gaussian distribution of SUD frequency and TIof the interval pattern, the autocorrelation function was negative up to about 50 msec (Fig. 3), and this negativity may mean a lowered excitability (or relative refractory period) provoked by the after-hyperpolarization (Lloyd, 1951 ; Goldberg ef al., 1964) which could be especially marked when the membrane is depolarized. As another possibility, a recurrent inhibition of the Renshaw type (Eccles et al., 1954) may be considered. But so far we have obtained only few and indefinitive observations suggesting the existence of the recurrent interneuron. In the central nervous system, depolarization is usually maintained by facilitatory synaptic bombardments (Kolmodin and Skoglund, 1958 ; Hunt and Kuno, 1959). Similarly, depolarization in HVM neurons may be maintained first by arrivals of facilitatory synaptic bombardments upon them from the AM (Figs. 7A, B and 8), the lateral part of the GP (Fig. 1 l), the SEP, etc. together with those from the already known positive feedback circuit between the AM and HVM (Oomura et al., 1965, unpublished observation) or with other unknown circuits, or by a decrease in inhibitory synaptic bombardments from the LH (Figs. 1 and 4), or by both. On the contrary, in the LH neurons, the SUD frequency was low and its discharge patterns were attributable to complete Poisson processes. The lowered activity may be due to membrane hyperpolarization. When the membrane potential was high, the after-hyperpolarization following the spike was less marked, or rather after-negativity followed a spike (Hagiwara and Tasaki, 1958, Fig. 10). This corresponds to the lack of References p . 31-33

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initial negativity in the temporal pattern of the autocorrelation of the LH SUDs (Fig. 3). Hagiwara (1954) also demonstrated that the serial autocorrelation of neighboring intervals on discharge records from a toad muscle spindle in which the interspike intervals showed an exponential pattern, was almost zero except at t = 0. Membrane hyperpolarizationmay be due to the increased inhibitory input or decreased excitatory input, OT both. We already know that HVM activity which was inhibitory to the LH, was high and regular under light anesthesia (Fig. 2). The limbic structures were considered as inhibitory areas to the LH, too (Figs. 7-1 1). The decrease in excitatory input to the LH from the dorsomedial mesencephalic tegmentum (Fig. 12) should also be considered. The neuronal activity of the latter probably receives inhibitory influences from the cortex as well as the medullar reticular formation; this is known to occur under light anesthesia or in the half arousal state (French et al., 1955; Magnes et al., 1961 ; Dell et al., 1961). In unrestrained conditions, the SUDs in the cat visual cortex (Hubel, 1959) and in the mesencephalic tegmentum (Huttenlocher, 1961) tended to become random, together with lowered frequency in the arousal state, and increased during sleep. Because of the increase in IPSP and decrease in EPSP, the LH neuron might keep the membrane potential hyperpolarized. Under deep anesthesia, not only the distribution patterns of the SUDs (Table I), but also the autocorrelation function (Fig. 3), were completely reversed in the HVM and LH. Further, the low frequency and randomness of SUDs in HVM neurons were mainly due to an increase in inhibitory input from the LH (Figs. 4 and 5). Low voltage fast waves in the cat cortical EEG under ether anesthesia (Adrian and Matthews, 1934) were descrided as activation sleep by Jouvet (1963). In this sleep, in the mesencephalic tegmentum, its EEG was a low voltage fast wave (Jouvet, 1963), and its SUD frequency was high (Huttenlocher, 1961). Consequently, the neuronal activity in this region should be high under the deep ether anesthesia of the present experiments, and the LH neurons might be much enhanced in their activity through the mesencephalic tegmentum (Fig. 12), while the HVM neurons, or at least some parts of them, might be inhibited (Fig. 12, Tsubokawa and Sutin, 1963). The decrease in the activity of the HVM neurons may also tend to decrease the AM activity. (3) Neuron activity in the HVM or LH. Two contiguous units in the HVM had not only the same discharge patterns but also a positive cross-correlation to each other, though the mean discharge frequencies or mean spike intervals were a little different. They had both negativecross-correlations to SUD simultaneously recorded in the LH. The same relations seem to hold in the units in the nuclei of both hemispheres: the same patterns on activities among units in either HVMs or L H s in both hemispheres, but negative correlations between units in an HVM and the contralateral LH. These observations show that the hypothalamic nuclei are considerably primitive, acting in the same way in the same nucleus. Different and complicated situations are known in the higher centers. For example, in the motor cortex neurons, Creutzfeldt and Jung (1961) found that two units recorded simultaneously by the same electrode sometimes showed reciprocal activity. Asanuma and Okada (1962) discovered that a pyramidal neuron in the motor area responded to stimulation applied exactly to the corresponding point in the opposite

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hemisphere, but was inhibited when the stimulating electrode was moved 1 mm off the point. Recording activities of 3 to 4 units in the lateral geniculate body, by the insertion of a triple or quadruple electrode, whose tips were aligned along a straight line and separated about 150 p from each other, Verzeano and Negishi (1960) recognized circulating neuronal activities, i.e. bursts of impulses were recorded by a component of the electrodes, then by another, repeatedly. But the fourth component of the electrodes often recorded a different or rather inhibiting activity to the other three components. ( 4 ) Relationship between the limbic structure and the hypothalamus. The present experiments on the effects of stimulation of the limbic structure upon the activities of the HVM and LH agreed fairly well with the behavioral experiments carried out by many investigators. The fact that the SUDs in the HVM were increased and those in the LH were simultaneously considerably decreased by AM or SEP stimulation confirmed the findings related to the feeding mechanism in the AM. Koikegami et al., (1955), Green el al., (1957), and Morgane and Kosman (1960) produced hyperphagia in cats by bilateral amygdalectomy, and Fonberg and Delgado (1961) an inhibition of the food intake of hungry cats following stimulation in the AM or SEP. As regards stimulation of the lateral part of the GP, the present results mainly showed the same effects on the HVM and LH activities as with the AM stimulation. This strongly supports the results obtained by Morgane (1961a) that lesions placed on the pallido-hypothalamic pathways from the lateral part of the globus pallidus to the HVM caused considerable hyperphagia. On the other hand, the stimulation of the medial part of the GP resulted in an opposite effect on the SUDs of the HVM and LH. Morgane (1961b) also indicated that rats lesioned in this region showed an aphagia, and postdated that crossing fiber systems, particularly pallidofugal fibers to the LH were the critical elements in the organization of the feeding centers. The significance of the medial mesencephalic tegmentum upon the neuronal feeding mechanism emphasiized by Nauta (1958) and Morgane (1964) was also confirmed by our experiments. However, our results were carried out solely on the region of the periaqueductal gray substance. Further unit investigations are required in order more fully to discuss such interactions.

SUMMARY

To establish neuronal regulation of feeding, the electrical activities of the feeding and satiety centers in the hypothalamus, as well as those of the limbic system and of the mesencephalic tegmentum, were investigated. The cross functional relations among those activities were clarified. Glass pipette electrodes were used to record spontaneous unitary discharges (SUD) simultaneously from the cat’s hypothalamic lateral area (LH), the feeding center, ventromedial nucleus (HVM), satiety center, and lateral principal nucleus of the amygdala (AM), under ether anesthesia or flaxedil-immobilization (routine procaine infiltration into pressure points were carried out). For stimulating, concentric bipolar electrodes were inserted into the regions mentioned above and into the lateral or References p.!31-33

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et al.

medial principal nucleus of the AM, septum (SEP), medial or lateral part of the globus pallidus (GP) and dorsomedial part of the mesencephalic tegmentum. (1) In a half arousal state under light ether anesthesia, in simultaneous recording from the HVM and LH, the HVM-SUD frequency was 10 to 20 impulses/sec on the average, yielding a Gaussian distribution, and the average interspike interval was 50 msec, the interval histogram being a T-distribution of order two (rz), showing more or less regularity. At the LH, the SUD frequency was low, a few impulses/sec, showing a Poisson distribution. The interspike interval was 200-300 msec on the average, the distribution pattern being exponential, revealing a complete randomness or a perfect Poisson process. When the anesthesia became deep, the HVM-SUD frequency was gradually decreased, while that in LH increased, and a completely reversed result in SUD distribution was obtained in the deep ether anesthetic state. (2) When two SUDs with different amplitudes were recorded simultaneously in one center, both showed quite the same discharge patterns, although they were not synchronized. This may indicate that the neurons in one center act in somewhat the same way. (3) After confirming the stationary state of the time series of SUD by variance analyses, we calculated the auto- and cross-correlations: (a) At a comparatively long t (300-1000 msec), there was a significant negative cross-correlation, revealing a marked reciprocal relation between the activities of the HVM and LH, i.e. when one center discharged more than the average for 1 t, several t’s afterwards, the other discharged less than the average. (b) At relatively short t (10 msec,) the autocorrelation function of the SUD with a Poisson distribution was zero except at t = 0, whereas that with the Fa distribution was negative for some tens of msec. Both these autocorrelation functions agreed with the mathematical analysis. The negative value of the correlation was presumed to show a lowered excitability of the neuron due to the after-spike hyperpolarization. (c) Significant positive correlations were obtained between the two SUDs in one center. This means, again, the same type of neuronal activity in the same center. (4) Under flaxedil immobilization, in simultaneous recordings from the LH and the lateral nucleus of the AM, the frequency of SUD was in general high in the LH, yielding a Gaussian distribution, and low in the AM, yielding a Poisson distribution. The cross-correlation function of the SUD between the LH and AM was calculated, resulting in significant negative correlations. These suggest a marked reciprocal relation of the neuronal activities between LH and AM. ( 5 ) Not only the SUDs, but also the electrical stimulation experiments, confirmed the reciprocal relationships between the HVM and LH. The number of the SUDs in the LH was markedly decreased for a considerable time by single or repetitive electrical stimulation of the AM but the reverse was true for the HVM. (6) Regarding the mutual facilitatory circuit, there were intimate functional associations between the HVM and AM, and between the LH and the dorso-medial part of the mesencephalic tegmentum (DMT). By stimulations of the PGS, a considerable increase in frequency of the SUD in the LH was also recorded, and, sometimes, a decrease in the activity of the HVM.

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(7) The lateral part of the G P behaved on the HVM and LH approximately in the same direction as did the AM, whereas the medial part of it behaved as did the DMT. (8) The driven unitary discharges were recorded in the HVM by single or volley stimulation of the SEP. (9) The hypothalamic feeding mechanism, therefore, is well maintained by the reciprocal activity between the HVM and LH as well as by a close linkage of the negative and positive feedback system made by the relationship among the hypothalamus, limbic system and dorsomedial mesencephalon. ACKNOWLEDGMENTS

The author is greatly indebted to Dr. S . Kano, Professor of Mathematics, Faculty of Science, Kagoshima Universtiy, Kagoshima, for the statistical treatments. This work was supported in part by aids from the Ministry of Education, the Rockefeller Foundation (Re GA MNS 6194), the Japan Waksman Foundation (1962), and the U.S. Army Research and Development Group (Far East) (DA-92-557-FEC-37352).

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tures in the lower brain stem. The Nature of Sleep, G. E. W. Wolstenholme and M. OConnor, Editors, London, Churchill, p. 57. MILLER,N. E., (1960); Motivational effects of brain stimulation and drugs. Fed. Proc., 19, 846. MORGANE, P. J., (1961a); Alternations in feeding and drinking behavior of rats with lesions in globi pallidi. Amer. J. Physiol., 201, 420-428. MORGANE, P. J., (1961b); Electrophysiological studies of feeding and satiety centers in the rat. Amer. J. Physiol., 201, 838-844. MORGANE, P. J., (1964); Limbic-hypothalamic-midbrain interaction in thirst and thirst motivated behavior. Thirst, M. J. Wayner, Editor, Oxford, Pergamon Press., p. 429. MORGANE, P. J., AND KOSMAN, A. J., (1960); Relationship of middle hypothalamus to amygdalar hyperphagia. Amer. J. Physiol., 198, 1315-1318. NAUTA,W. J. H., (1958); Hippocampal projections and related neural pathways to the mid-brain in the cat. Brain. 81,319-340. OOMURA, Y., KIMURA, K., OOYAMA, H., MAENO,T., IKI,M., AND KUNIYOSHI, M., (1964); Reciprocal activities of the ventromedial and lateral hypothalamic areas of cats. Science, 143, 484485. OOMURA, Y., OOYAMA, H., AND SAWADA, M., (1965); Ionic basis of the effect of ACh on Onchidium D- and H-neurons. XXIZI Int. Congr. Physiol. Sci., No. 913. OOMURA, Y., OOYAMA, H., YAMAMOTO, T., AND NAKA,F., (1967); Reciprocal relationship of the lateral and ventromedial hypothalamus in the regulation of food intake. Physiol. Behuv., 2, in press. POGGIO,G. F., AND VIERNSTEIN, L. J., (1964); Time series analysis of impulse sequences of thalamic somatic sensory neurons. J. Neurophysiol., 27, 517-545. RALL,W., (1962); Electrophysiology of a dendritic neuron model. Biophys. J., 2, 145-167. SAWA,M., MARUYAMA, N., HANAI,T., AND KAJI, S., (1959); Regulatory influence of amygdaloid nuclei upon the unitary activity in ventromedial nucleus of hypothalamus. Fol.psychiut. neurol.jap., 13, 235-256. SKULTELY, F. M., AND GARY,T. M., (1962); Experimental hyperphagia in cats following destructive midbrain lesions. Neurology, 12, 394401. SMITH,0. A., (1956); Stimulation of lateral and meidal hypothalamus and food intake in the rat. Anat. Rec., 124, 363-364. SPRAGUE, J. M., CHAMBERS,W. W., AND STELLAR, E., (1961); Attentive, affective, and adaptive behavior in the cat. Science, 133, 165-173. STEVENSON, J. A. F., (1964); The hypothalamus in the regulation of energy and water balance. Physiologist, 7, 305-318. STRUMWASSER, F., (1962); Post-synaptic inhibition and excitation produced by different branches of a single neuron and the common transmitter involved. XXIZ Internat. Congr. Physiol. Sci., Vol. 2 No. 801. STUART, D. G., PORTER, R. W., ADEY,W. R., AND KAMIKAWA, Y.,(1964); Hypothalamic unit activity. I. Visceral and somatic influences. Electromceph. clin. Neurophysiol., 16,237-247. SZENTAGOTHAI, J., FLERKO, B., MESS,B., AND H A L ~ Z B.,, (1962); Hypothalamic Control of the Anterior Pituitary, Budapest, Akadtmiai Kiadb. TAUC, L., AND GEPSCHENFELD, H. M., (1962); A cholinergic mechanism of inhibitorysynaptictransmission in a molluscan nervous system. J. Neurophysiol., 25, 236-262. TSUBOKAWA, T., AND S m ,J., (1963); Mesenczphalic influence upon the hypothalamic ventromedial nucleus. Electroenceph. clin. Neurophysiol., 15, 804-810. VERZEANO, M., AND NEGISHI,K., (1960); Neuronal activity in cortical and thalamic networks. A study with multiple microelectrodes. J. gen. Physiol., 43, (6) Suppl. 177-195. WAGNER, J. W., AND DE GROOT, J., (1963); Changes in feeding behavior after intracerebral injections in the rat. Amer. J. Physiol., 201,483-487. WERNER, G.,AND MOUNTCASTLE, V. B., (1963); The variability of central neural activity in a sensory system, and its implications for the central reflexion of sensory events. J. Neurophysiol., 26, 958-977. WOOD,D. C., (1958); Behavioral changes following discrete lesions of temporal lobe structures. Neurology, 8,215-226.

34

Limbic and other Forebrain Mechanisms in Sleep Induction and Behavioral Inhibition * CARMINE D. CLEMENTE AND MAURICE B. STERMAN Department of Anatomy and Brain Research Institute, University of California, Los Angeles, and the Sepulvedu V.A. Hospital, Sepulveda, Calif. (U.S.A.)

INTRODUCTION

Cortical EEG synchronizationis of interest to the behavioral physiologist because it is characteristic of the spontaneously occurring cortical spindle pattern associated with sleep and other states of central nervous system suppression. Perhaps one of the first contributions which implicated the brain stem’s influence ovei the state of the EEG, was the classical observation of Bremer (1935) who showed that transaction of the neuraxis at the high cervical level resulted in an EEG pattern of wakefulness, while a more rostra1 transection at the midbrain level, resulted in an EEG pattern similar to sleep. The functional importance, however, of the intrinsic brain stem mechanisms involved in the maintenance of an alert, active EEG was not appreciated until Magoun with his associates Moruzzi (1949) and Lindsley and Bowden (1949) reported their well known stimulation and lesioning experiments in the late 1940’s. Evidenced further by the work of French and Magoun in the primate (1952), this research led to the formation of the concept of an ascending reticular activating system in the core of the brain stem which extends rostrally into the thalamus. Other experiments performed earlier by Dempsey and Morison (1942a, b; 1943) indicated that low frequency stimulation of the intralaminar thalamic muclei was capable of driving the electrical activity of the cerebral cortex in the opposite direction, producing instead, the recurrent EEG spindle burst pattern termed ‘recruitment’. These findings stressed, in our view, the potential for a differential influence from various areas in the central nervous system upon ongoing electrocortical activity. Hess (1954, 1957) contributed significantly to this concept by demonstrating the induction of behavioral sleep with low frequency electrical stimulation of nuclei in the medial thalamic mass. In his distinguished studies on the functions of the hypothalamus Hess differentiated two areas subserving, generally speaking, antagonistic functions. The sympathetic nature of the response, and the generalized behavioral activation resulting from stimulation of the caudal hypothalamus, led him to call this area the

*

Aided by a grant from the U. S. Public Health Service (MH-10083).

MECHANISMS I N SLEEP I N D U C T I O N A N D B E H A V I O R A L I N H I B I T I O N

35

‘ergotropic zone’. More anteriorally located hypothalamic zones produced generalized somatomotor suppression and parasympathetic responses to stimulation, and to these areas he ascribed the term ‘trophotropic zone.’ He included the medial thalamic areas, capable of inducing sleep in this latter functional system. In an evaluation of other brain stimulation studies, our attention was drawn to the preoptic-basal forebrain area, in which there appeared to exist potent suppressor mechanisms affecting a variety of peripheral functions. We felt that other mechanisms may exist there which would be reflected in the EEG and perhaps also in the animal’s behavior. If our thinking in these matters was consistent with the many correlating observations of others, the effects of stimulation here should tend to synchronize the electrical activity of the cerebral cortex and perhaps also produce behavioral suppression.

METHODS A N D R E S U L T S

I. Acute stimulation and recording experiments The first series of electrophysiological experiments were performed in the brains of acutely prepared adult cats. General anesthetic drugs often depress cortical electrical activity thereby masking the cortical influences of stimulation in subcortical areas. Because of this a neuromusclular blocking agent, Flaxedil, was used in conjunction with local anesthesia at incision sites in an effort to achieve recordings against the background of an activated and responsive cerebral cortex. Operations were carried out under general ether anesthesia and electroencephalographic recording was achieved by the use of phonograph needle electrodes placed into the calvarium over the cerebral cortex. Upon an EEG background characterized by low voltage fast activity, the effects of stimulating various basal forebrain sites with concentric stereotaxically placed bipolar electrodes were observed. It was found that stimulation in certain preoptic and basal forebrain areas produced an immediate, sustained and diffuse cortical synchronization. This response was observed upon stimulation at low voltages (2-2.5 V at 0.75 msec duration) and was found most effective at lower frequencies (5-7 per sec). Examples of this cortical synchronization can be seen in Fig. 1. The subcortical points, which upon stimulation resulted in an onset of cortical synchronization, were mapped anatomically and the results from 25 exploration experiments are shown collectively in 4 cross-sectional diagrams of the cat brain in Fig. 2. At A15.5 and A16 a distinct concentration of positive stimulation sites was observed. This focus extended caudally into the preoptic region of the hypothalamus, but at more posterior hypothalamic sites, electrical stimulation did not result in the onset of cortical synchronization. Other experiments were carried out to assess the interaction between stimulation of both the brain stem reticular activating system and the basal forebrain-preoptic synchronizing sites. Figs. 3 and 4 show the results of such an interaction experiment. In the upper set of traces in Fig. 3 is illustrated the well known cortical arousal phenomenon resulting from high frequency stimulation in the reticular activating system. The second series of traces in Fig. 3 shows, however, that if in the course of this arousal, References p . 47

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the basal forebrain zone was simultaneously stimulated at low frequency, a cortical synchronization was induced, superimposed upon the original activated EEG. The reverse interaction effectwas also achieved and can be seen in the first series of traces of Fig. 4. If basal forebrain stimulation preceded stimulation of the reticular formation the induced cortical synchronization was then replaced by arousal. The second series of traces in Fig. 4. shows the sensitivity of both the arousal response and the induced cortical synchronization to the intravenous administration of 15 mg/kg of barbiturate anesthesia, Nembutal. This non-anesthetizing dose of barbiturate anesthesia clearly established the ineffectiveness of basal forebrain stimulation following barbiturate administration, and perhaps indicates why this rather pronounced effect has not been observed before. 10

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Fig. 2. This figure shows 4 frontal diagrams of the cat brain taken from the Jasper-Ajmone Marsan atlas. The black circles indicate points where stimulation resulted in cortical synchronization;the negative points are designated by minus signs. (From Sterman and Clemente, 1962a). References p. 47

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ZI. Chronic behavioral experiments In conjunction with acute experiments, other adult cats and adult monkeys were prepared for long-termed chronic study. Bipolar strut electrodes were placed into the regions which were found capable of inducing synchronous cortical activity upon electrical stimulation in acute preparations. The animals were permitted to recover and the effects of stimulation in these unrestrained animals were observed. EEG activity was recorded through leads taken from a number of small screws placed into the calvarium at the time of operation. Bilateral stimulation at basal forebrain sites in these chronically prepared animals at levels of 2-3 V, 0.75 msec and at frequencies of 5-7 per sec resulted in the appearance of drowsiness and sleep within 1-2 min from the commencement of stimulation. This behavioral transition was accompanied by a correlated shift in the frequency and amplitude characteristics of the electroencephalogram. The low voltage fast activity of the activated EEG, within a few seconds from the onset of stimulation, gave way to the spindling patterns observable during the initial stages of sleep (Fig. 5). When the animals appeared to be behaviorally asleep, they could be aroused by tapping on the cage and then returned to sleep once again by basal forebrain stimulation. In other animals, electrodes were also placed into the reticular activating system and these cats could successivelybe aroused by stimulating the reticular formation at high frequency and put back to sleep by stimulating the basal forebrain region at low frequencies. The sequence of events initiated by the onset of electrical stimulation in the basal forebrain usually consisted of a cessation of ongoing behavior, a retreat on the part of the cat to some corner of the cage, the assuming of the reclining position, and the closing of the eyelids. These behavioral correlates of sleep were accompanied by the EEG spindling patterns characteristic of sleep. The effective stimulation parameters and the time required for the onset of sleep seem to be dependent on several factors. Generally, stimulation at 1-3 V with 0.5-0.75 msec duration was effective for frequencies varying from 5-250 impulses per sec. In some animals the onset of sleep was seen to occur with latencies as short as 5-10 sec after the start of stimulation, whereas others required as long as 2-3 min between the onset of stimulation and the occurrence of drowsiness or sleep. The average time required was about 30 sec. i t was found that factors such as the time of day, time of feeding and the numbers of times that the animal had been stimulated previously proved to be important determinants with respect to the ease with which these effects could be induced. Fig. 6 shows an animal in which sleep was induced with bilateral basal forebrain stimulation at higher frequencies (I 50 c/s, 1 V, 0.75 msec). The transition in this animal from alert behavioral posture seen on the left to the quiescent sleep posture seen on the right occurred approximately 20 sec after the onset of stimulation and was accompanied by a rapid transition from an EEG of wakefulness to an EEG of sleep. III. Conditioning experiments

in another series of adult male cats electrodes were chronically implanted into the brain and recording stainless steel screw electrodesplaced into the skull bilaterally over

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MECHANISMS I N SLEEP INDUCTION AND BEHAVIORAL INHIBITION

41

the frontal, parietal, and occipital lobes of the cerebral cortex. After a postoperative period of two weeks, these animals were allowed to become familiar with an observation cage and the effective basal forebrain stimulation values determined,with respect to the induction of synchronized EEG activity and behavioral sleep. Conditioning experiments were carried out in these animals as follows: A tone of a given frequency was presented for 10 sec in advance of basal forebrain stimulation and was continued throughout an employed 20-sec period of brain stimulation. The tone and stimulation overlapped for a period of 20 sec and terminated simultaneously. An intertrial interval of 30 sec lapsed between the termination of one trial and the subsequent presentation of the next tone. Trials were repeated every minute throughout the conditioning session. Initially, the presentation of a 2000 c/s tone evoked either a desynchronization of the EEG in the non-alerted cat or little change in the flat low

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voltage pattern of the alerted animal. After a few pairings of the tone with bilateral stimulation in the basal forebrain, a conditioned EEG and behavioral response became apparent. This is illustrated in Fig. 7, where the presentation of the 2000 c/s tone on trial 38 evoked a shift in the predominantly desynchronized EEG to one of low frequency high amplitude synchronized activity. At the same time, the alerted animal suddenly showed sleep preparatory behavior at the onset of the tone. After the establishment of a conditioned EEG synchronization to the onset of a 2000 c/s tone, the References p. 47

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CLEMENTE A N D STERMAN

frequency of the tone was changed to 4000 c/s between trials 45 and 46. Upon hearing the 4OOO c/s tone for the first time, the animal's EEG response generalized to the previous tone of 2000 c/s. On trial 46, instead of following the 4000 c/s tone by low frequency stimulation in the basal forebrain, the animal was stimulated by high frequency low voltage pulses in the miedal thalamic nuclei. These stimulation parameters characteristically induced an alert EEG. In our conditioning experiment the initial response to

Fig. 6. Behavioral response to high frequency electrical stimulation of the basal forebrain area in the cat (150 c/sec, 1 Volt, 0.75 msec). In this alert but unaroused animal (left) the induction of sleep was accomplishedin approximately 20 sec and was accompaniedby an equallyrapid EEG shift to a pattern of slow wave activity. (From Sterman and Clemente, 1962b.)

this alerting stimulus was a transitory synchronization which was shortly followed by the expected activation of the EEG. Further pairings of the 2000 c/s tone with basal forebrain stimulation and the 4000 c/s tone with high frequency stimulation in the medial thalamic nuclei resulted in the establishment of classical conditioned responses. These experiments indicated that stimulation in the basal forebrain could be paired with a previously neutral conditioning sensory stimulus in such a manner that the presentation of the tone alone was able to establish the conditioned synchronized EEG response. We should like to point out that the basal forebrain area is the only non-specific central nervous system site which has been found capable of producing a conditioned synchronization in the ongoing EEG activity. This classical Pavlovian conditioning procedure lends further evidence to the strong functional relationship existent between the basal forebrain area and the cerebral cortex. IV. Pathway studies

An additional series of acute cat preparations was studied in an effort to determine electrophysiologically the connections of the basal forebrain area with other parts of

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CLEMENTE A N D STERMAN

the central nervous system. Adult cats were surgically prepared for artificial respiration under ether anesthesia and placed in a stereotaxic instrument. Various areas of the cerebral cortex were exposed, and cortical and subcortical electrodes were located for evoked potential studies. Wound margins and pressure points were infiltrated with xylocaine and a neuromuscular blocking agent administered. Following recovery from the ether anesthesia stereotaxicplacement of electrodes in the basal forebrain area was verified by electrical stimulation through the induction of EEG slow wave activity as the criterion for correct localization, Single biphasic shocks were then delivered to this area, while the electrical activity of various cortical and subcortical sites was explored systematically on a multiple beam oscilloscope. Evoked potentials were observed in a discrete region of the ventrolateral frontal cortex, in the temporal lobe, amygdala, hippocampus, septum, midline thalamus and mesencephalic reticular formation (Fig. 8). The general region of the anterior sylvian gyrus and the more rostra1 temporal lobe cortex were the only cortical areas which showed a direct electrical evoked potential to basal forebrain stimulation. The electrical activity recorded in the hippocampus consisted of a short and a long latency bimodal response. Subsequent lesion studies indicated that the short latency component was mediated by a basal forebrain-septalhippocampal pathway, whereas the long latency component required the integrity of temporal-ammonic tracts. Responses were also evoked in the basal forebrain region by stimulation of the orbital gyrus in the cerebral cortex and in the hippocampus by stimulation of this same cortical area. These findings suggest the involvement of the orbital gyrus and lateral orbital regions of the cerebral cortex and a ‘limbic loop’ including septum, amygdala, hippocampus and temporal cortex, in a descending inhibitory pathway through the basal forebrain ultimately expressing its influence upon regulatory nuclei in the thalamus, mesencephalon, and pontine brain stem (Fig. 8). Induced synchronization observed in midline thalamic nuclei (Clementeand Sterman, 1963)and more generally throughout the cerebral cortex upon stimulation of the basal forebrain region as well as the orbital gyrus, may, therefore, reflect an integrated inhibitory influence upon brain stem structures.

DISCUSSION

It is our interpretation that the forebrain areas stimulated in these experiments, resulting in the production of widespread cortical synchronization in restrained animals and arrest reactions and sleep in behaving animals constitutes a functional brain system which expresses its behavioral suppressor influence over a wide variety of somatic and visceral functions. Furthermore, it is felt that the level of cortical activation or synchronization may be regulated by dually active systems : the reticular activating system of Moruzzi and Magoun (1949) and this forebrain corticalsynchronizingsystem. We wish to propose that these two systems act reciprocally, perhaps through relays in the limbic circuit and in the diffuselyprojecting thalamo-cortical system, of which the latter has been shown capable of activating the EEG with high frequency stimulation

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other workers. The problems raised will be discussed more fully in the paper in preparation. Skultety (1963), working with caged cats, produced rage reactions from the two anterior thirds, and attempts to escape from caudal parts of the central gray matter of the midbrain, but not from the adjacent tegmental fields, a territory from which Hunsperger (1956, Fig. 2) produced flight reactions in the freely-moving animal. The location of the active area for threat within the amygdala established by Fernandez de Molina and Hunsperger (1959) agrees well with the findings of Naquet (1953), MacLean and Delgado (1953), Magnus and Lammers (1956), Shealy and Peele (‘1957) and Wood (1958), but does not tally with the findings of Kaada et al. (1954) and Ursin and Kaada (1960). According to these authors, the zone concerned with fear occupies lateral portions of the amygdala, and the zone concerned with anger occupies medial and ventral portions of this structure. Fernandez de Molina and Hunsperger, however, obtained turning of the eyes and head, or repeated jerking of eyes and head to the side opposite stimulation from the lateral amygdaloid nucleus (Fig. lo), a reaction recalling the ‘attention response’ evoked by Ursin and Kaada (1960) from this nucleus and more medially lying structures by applying weak stimulation. For the anxiety reactions obtained by Sano (1958) and Fangel and Kaada (1960) and others from cortical structures, see Hunsperger (1963). Hilton and Zbroiyna (1963) have recently emphasised that defence reactions according to these authors threat, wildly running movements and jumping up the walls of the cage in which the cats were enclosed - can be traced from the basal nucleus of the amygdala into the preoptic area and rostral hypothalamus by way of the anterior amygdaloid area and a narrow connection dorsal to the optic tract. The band connecting the amygdala and preoptic/hypothalamic region, according to these authors, seems to correspond to the ventral amygdalofugal pathway described by Nauta (1961). Multiple electrolytic lesions placed in this band abolished the responses from the amygdala. It is not clear from their illustration (Fig. 5) whether or not the stria terminalis system was involved. Neither threat nor flight was obtained by Fernandez de Molina and Hunsperger (1959) by stimulation of the area lying beneath the pallidum between the anterior amygdaloid region and the preoptic area and rostral hypothalamus, an area comprising the diffuse fibres described by Johnston (1923), Fox (1943), and Nauta (1961). This area has recently been re-explored by Hunsperger using bipolar stimulation. The effects yielded were compared with former results obtained with monopolar stimulation. All threat and flight responses obtained from the forebrain and hypothalamus with stimuli up to 2.5 V at frequency 8 or up to 1.5 V at frequency 17 (in other words, stimuli not exceeding three times the lowest threshold for these responses) were considered positive, and were plotted in diagrams based on the Hess atlas. The extent of the areas in the forebrain and hypothalamus yielding threat and flight is illustrated in Fig. 11 in 5 frontal sections. Negative points with regard to these reactions are also shown. This figure should be compared with Fig. 1 of Hilton and ZbroyZna (1963) and Figs. 5 and 6 of Ursin and Kaada (1960). The stipplled areas, sections 326,352, indicate the region through which the diffuse fibres (d.f.) run. It will be observed that the responsive points for threat and flight are grouped in the References p.

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Fig. 11. Schematic illustration of areas in the amygdala, stria terminalis bed and hypothalamus yielding threat, flight or mixed reactions. Shows region below the pallidurn yielding negative findings with regard to affective reactions. 5 frontal sections (atlas of Hess, S 326-430). Effects obtained at intensities up to 2.5 V a t 8 per sec and up to 1.5 V a t 17 per sec. 0 = threat with growling and/or hissing; 0 = flight; C ) = hissing followed by flight; 0 = negative points with regard to affective reactions.

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121

Fig. 12. Effects obtained by bipolar stimulation of region belov, the pallidum and location of electrode serving as cathode. (a, left) ipsilateral twitching of the eyelid and rno’xients of the tongue as if to eject a foreign body (anode in region rostra1 to cathode); (a, right) ips.kteral twitching of the upper lip and repeated jerking of eyes and head to side opposite stimulation (anode in region caudal to cathode). (b) the electrode serving as cathode lies beneath the pallidum at the level of Fox’s association bundle b (see arrow).

References p. 125-127

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amygdala (sections 378,404,430). By way of the stria terminalis, they can be traced to the bed of the stria at the level of the anterior commissure, shown in section 326, and from this level, along the hypothalamic component of the stria (section 352) into the preoptic and hypothalamic fields (sections 378,404,430). The question of direction of conduction in this path remains open (Fernandez de Molina, and Hunsperger, 1962; Hilton and Zbroiyna, 1963). Again it will be observed that negative points with regard to threat and flight lie scattered above the optic tract (sections 430, 404) and in the region beneath the pallidum through which the diffuse fibres pass (sections 378, 352, 326). The reactions obtained from these ‘negative points’ (a total of 18) include pupillary dilatation, sniffing, salivation, tongue movements, facial motor effects, and repeated jerking of eyes and head to the side opposite stimulation, the latter an effect, as already mentioned, recalling the ‘attention response’ of Kaada. Mewing was obtained only once. Fig. 12a shows two examples of responses obtained by bipolar stimulation of the region of the diffuse fibres. The track of the electrode that served as cathode is shown in Fig. 12b and lies beneath the pallidum among diffuse fibres, including Fox’s b bundle. Fig. 12aleft, shows the effects obtained when the electrode that served as anode was placed in the innominate substance of Reichert -twitching of eyelids and movements of the tongue as if to eject a foreign body. Fig. 12a right, shows the effects produced when the electrode that served as anode was placed more caudally in the region between the optic tract and the entopeduncular nucleus - twitching of the upper lip and repeated jerking of eyes and head to the side opposite stimulation. All these observations lend no support to the contention that the responsive fields for threat and fiight (defence reaction according to Hilton and Zbroiyna) in the amygdala and the hypothalamus are connected by way of a direct ventral route that passes in the region between the optic tract and pallidum. The role played by the diffuse system -a subject to which Koikegami and Yoshida (1953), Ursin and Kaada (1960), Hilton and ZbroZyna (1963), Karli and Vergnes (1964), have drawn attention requires further investigation. Substratumfor mewing

The field for mewing obtained during passage of current also extends from the brainstem to forebrain. At the level of the midbrain, it occupies ventral portions of the central gray matter adjacent to the more dorsally lying region for threat pattern. At Fig. 13. Schematic illustrationof points in the forebrain and diencephalonyielding mewing. 8 frontal sections (atlas of Hess S 287-469). Lightly shaded areas: hippocampus/fornix system and region of transitionlateral preoptic area/hypothalamus, dotted area: gray matter of the hypothalamus and preoptic area. 0 = mewing evoked duringpassageofcurrent; 0 = mewing occurring after cessation of stimulation ; //// = responsive field for threat response in the amygdala, stria terminalis system and hypothalamus ; \\\\ = responsive field for flight in the hypothalamus and preoptic area; 0 = negative points with regard to affective reactions.

AFFECTIVE B E H A V I O U R

References p . 125-127

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diencephalicand forebrain levels (Fig. 13, S a-h) active points in the preoptic region and anterior hypothalamus border on and intermingle with the active area for flight and threat, and are further traceable into the septum (including precommissural fornix), fornix (S a, b), and fimbria/hippocampus (S f-h), the latter findings supporting results obtained by Magnus and Lammers (1956), MacLean (1955, 1957b) and Parmeggiani (1960). Another group of active points is situated in anterior and dorso-medial parts of the thalamus, and appears to follow the track of the stria medullaris to the habenula. Eleven points were stimulated in this latter structure and the tractus Meynert (S g,h ), but mewing was only obtained from one. It has been shown by Wallenberg (1902), Burgi and Bucher (1960, 1963), that the stria medullaris, besides ipsilateral fibres, conveys a bundle that crosses in the habenular commissure, runs forward on the contralateral side and distributes (arrows, section c, Fig. 13) to the region of transition lateral preoptic area/hypothalamus, an area from which marked mewing responses were consistently obtained as the sole effect (S c, d). The question arises whether the effects obtained from the stria medullaris region are due to activation of the focal field on the side opposite stimulation. This focal field (transition lateral preoptic area/hypothalamus -the lightly-shaded area in Fig. 13, b-d) also receives fibres from the hippocampus/fornix system and septum according to Valenstein and Nauta (1959). We suggest that mewing obtained from these latter structures may be secondarily reinforced by activation of this focal field. The active points for mewing obtained after cessation of stimulation are widely dispersed in structures of the diencephalon and forebrain and include the hippocampus /fornix system, anterior nuclei of the thalamus, cingulum, and finally, the anterior two thirds of the amygdala (S d, e). These active points either lie in regions adjacent to those yielding mewing during passage of current, or in structures which are anatomically directly or indirectly connected with the focal field for mewing described previously. Substratum for pattern suggestive of reaction to pain

It is difficultto assign a specific substratum to responses suggesting reaction to pain, as such responses were never obtained as threshold answers from the midbrain, hypothalamus or forebrain. The most convincing reaction expressing pain (illustrated in Fig. Sa) was evoked from ventral parts of the periaqueductal gray matter, a region which according to Walker (1942) and Morin et al. (1951) receives spinoreticular fibres that accompany the spinotectal and spinothalamic tracts. It may be possible that this ventral region constitutes a relay station for afferent nociceptive fibres, or an ontophylogenetically ancient system subserving pain reactions. These aspects, and the somewhat different findings of Spiegel et ~ l(1954) . in the cat, have been discussed in the review of Hunsperger (1963, pp. 52, 53). SUMMARY

(1) The data presented show that the affective patterns elicited by electrical stimulation of the forebrain and brain stem in unanaesthetized freely-moving cats are threat

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and flight, mewing of various types, and an effect suggestive of reaction to pain. (2) The threat and flight reactions and mewing are obtained ast hreshold answers. All three can be assigned to specific areas; the reaction suggesting pain perception, however, is only obtained with strong stimulation from points lying within the responsive fields for mewing or threat. (3) The affective patterns obtained from the midbrain, hypothalamus and forebrain, although grossly similar, vary according to the level stimulated. (4) Seismographic recording of tonic muscular activity and of heart beats supplies further information on the somato-motor and autonomic changes associated with threat and flight. (5) The threat and flight responses elicited from the hypothalamus show adaptation to changes in environmental conditions. REFERENCES ABRAHAMS, V. C., HILTON,S. M., AND ZBRO~YNA, A., (1960); Active muscle vasodilatation produced by stimulation of the brain stem: Its significance in the defence reaction. J. Physiol. (Lond.),154, 491-5 13. ANDY,0.J., AND AKERT,K., (1955); Seizure patterns induced by electrical stimulation of hippocampal formation in the cat. J. Neuropath. exp. Neurol., 14, 198-213. BURGI,S., UND BUCHER, V. M.. (1960); Markhaltige Faserverbindungen im Hirnstamm der Katze. Monographien aus dem Gesamtgebiet der Neurologie und Psychiatrie. Fasc. 87, Berlin, Springer. BURGI,S., AND BUCHER,V. M., (1963); Stria terminalis and related structures. Progress in Brain Research Vol. 3. The Rhinencephalon and Related Structures, W. Bargmann and J. P. Schade, Editors, Amsterdam, Elsevier pp. 163-169. J., (1955); Hippocampe et Epilepsie: A propos d'une Sirie d'Expiriences sur le Cobayeet CADHILLAC, le Chat et de I'Exploration Electrique de la Corne d'dmmon chez 1'Homme. Montpellier, Paul Dehan. W. B., (1939); The Wisdon of the Body. New York, Norton. CANNON, CANNON, W. B., (1953); Bodily Changes in Pain, Hunger, Fear and Rage. 2nd ed. Boston, Branford. CORTI,U. A., GASSMANN F., UND WEBER, M., (1955); Unruhebestimmung bei Menschen und Tieren. Verh. Schweiz. Naturforsch. Ges., Pruntrut, 164-167. R. W., UND WYSS,0. A. M., (1961); Registrierung der vertikalen KraftCORTI, U. A., HUNSPERGER, komponente der endogenen Korpererschiitterung (Haltetonus). Reaktionstisch fir Tierversuche. Pfliigers. Arch, ges. Physiol., 274, 95. FANGEL, CH., AND KAADA, B. R., (1960); Behavior 'attention' and fear induced by cortical stimulation in the cat. Electroenceph. clin. Neurophysiol., 12, 575-588. FERNANDEZ DE MOLINA, A., AND HUNSPERGER, R. W., (1959); Central representation of affective reactions in forebrain and brainstem : Electrical stimulation of amygdala, stria terminalis, and adjacent structures. J. Physiol. (Lond.), 145, 251-269. FERNANDEZ DE MOLINA, A., AND HUNSPERGER, R. W., (1962); Organization of the subcortical system governing defence and flight reaction in the cat. J. Physiol. (Lond.), 160, 200-213. Fox, C. A., (1943): The stria terminalis, longitudinal association bundle and precommissural fornix fibers in the cat. J. comp. Neurol., 79, 277-295. HESS,W. R., (1932); Beitriige zur Physiologie des Hirnstamms. I . Die Methodik der lokalisierten Reizung und Ausschaltung subkortikaler Hirnabschnitte. Leipzig, Thieme HESS,W. R., (1949); Das Zwischenhirn. Basel, Schwabe. HESS,W. R., (1957); The Functional Organization of the Diencephalon.New York, Grune and Stratton. HESS,W. R., UND BRUGGER,M., (1943); Das subkortikale Zentrum der affektiven Abwehrreaktion. Helv. physiol. pharmacol. Acta, 1, 33-52. V., (1945/46); Zur Physiologievon Hypothalamus, Area HESS,W. R., BRUGGER,M., UND BUCHER, praeoptica und Septum, sowie angrenzender Balken- und Stirnhirnbereiche. Mschr. Psychiat. Neurol., 3, 17-59. A. W., (1963); Amygdaloid region for defence reactions and its HILTON,S. M., AND ZBROZYNA, efferent pathway to the brainstem. J. Physiol. (Lond.), 165, 160-173.

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HUNSPERGER, R. W., (1956); Affektreaktionen auf elektrische Reizung im Hirnstamm der Katze. Helv. physiol. pharmaeol. Acta, 14, 70-92. HUNSPERGER, R. W., (1963); Comportements affectifs provoques par la stimulation electrique du tronc drkbral et du cerveau anterieur. J. Physiol. (Paris), 55, 45-97. HUNSPERGER, R. W., (1965); Mektreaktionen auf Mittelfrequenzreizung (5000 Hz) im Hypothalamus der Katze. Helv. physiol. pharmacol. Acta, 23, C25-C28. HUNSPERGER, R. W., UND DIETIKER,M., (1962); Die Methodik der Registrierung der endogenen Korpererschiitterung (Haltetonus) an der Katze. Isolierung von Storschwingungen. Helv. physiol. pharmacol. Acta, 20, C7-C9. HUNSPERGEX, R. W., UND G w ~ Z D Z ,B., (1964); Lokalisation zentralnervoser Strukturen fur Miauen auf Grund von Hirnreizversuchen. Verh. Schweiz. Naturforsch. Ges. Zurich, 228-238. HUNTER,J., (1950); Further observations on subcortically induced epileptic attacks. Electroenceph. clin. Neurophysiol., 2, 193-201. HUNTER,J., AND JASPER, H. H., (1949); Effects of thalamic stimulation in unanaesthetized animals. The arrest reaction and petit mal-like seizures, activation patterns and generalized convulsions. Electroenceph. elin. Neurophysiol., 1, 305-324. JOHNSTON, J. B., (1923); Further contributions to the study of the evolution of the forebrain. J. comp. Neurol., 35, 337481. KAADA,B. R., JANSEN, J. JR., AND ANDERSEN, P., (1953); Stimulation of the hippocampus and medial cortical areas in unanesthetized cats. Neurology, 3, 844-857. KAADA,B. R., ANDERSEN, P., AND JANSEN,J. JR., (1954); Stimulation of the amygdaloid nuclear complex in unanesthetized cats. Neurology, 4,48-64. KARLI,P., ET VERGNES,M., (1964); Nouvelles donnks sur les bases neurophysiologiques du comportement d’agression intersgcifique rat-souris. J. Physiol. (Paris), 56, 384. KARPLUS, J. P., UND KREIDL,A., (1909); Gehirn und Sympathicus. I. Zwischenhirnbasis und Halssympathicus. Ppigers Arch. ges. Physiol., 129, 138-144. KARPLUS,J. P., UND KRUDL, A., (1910); Gehirn und Sympathicus. 11. Ein Sympathicuszentrum im Zwischenhm. Pfliigers Arch. ges. Physiol., 125,401416. KARPLUS, J. P., UND KREIDL,A., (1928);Gehirn und Sympathicus. VIII. (1) Zur zentralen Regulierung der Irisbewegungen; (2) Bemerkungen zur Schmenemphdlichkeit der vegetativen Hypothalamuszentren. Pj7iigers Arch. ges. Physiol., 219, 613-618. KOIKEGAMI,H., AND YOSHLDA,K., (1953); Pupillary dilatation induced by stimulation of the amygdaloid nuclei. Folia psychiat. neurol. jap., 7, 109-126. LISSAK,K., GRASTYAN, E., CSANAKY, A., UKESI, F., AND VEREBY, GY., (1957); A study of hippocampal function in the waking and sleeping animal with chronically implanted electrodes. Acta physiol. pharmacol. neerl., 6, 415459. MACLEAN, P. D., (1955a); The limbic system (‘visceral brain’) and emotional behavior. Arch. Neurol. Psychiat. (Chic.), 73, 13CL134. MA CLEAN,^. D.,(l957b); Chemical and electrical stimulationof hippocampus in unrestrainedanimals. I. Methods and electroencephalographic findings. Arch. Neurol. Psychiat. (Chic.), 78, 113-127. MACLEAN, P. D., (1957); Chemical and electrical stimulation of hippocampus in unrestrained animals. II. Behavioral findings. Arch. Neurol. Psychiat. (Chic.), 78, 128-142. MACLEAN, P. D., AND DELGADO, J. M. R., (1953); Electrical and chemical stimulation of frontotemporal portions of limbic system in the awaking animal. Electroenceph. clin. Neurophysiol., 5, 91-100. MAGNUS, O., AND LAMMERS, H. J., (1956); The amygdaloid nuclear complex. Part I: Electrical stimulation of the amygdala and periamygdaloid cortex in the waking cat. Fofia psychiat. neerl., 55, 555-581. M o m , F., SCHWARTZ, H. G., AND O’LEARY,J. L., (1951); Experimental study of the spino-thalamic and related tracts. Acta Psychiat. (Kbh.), 26, 371-396. NAKAO, H., (1958); Emotional behavior produced by hypothalamic stimulation. Amer. J . Physiol., 194,411418. NAQUET,R., (1953); Sur les fonctions du rhinenckphale d’aprb les resultats de la stimulation chez le chat. Thesis, Marseille. NAUTA,W. J. H., (1961); Fibre degeneration following lesions of the amygdaloid complex in the monkey. J. Anat., 95, 515-531. PARME~IANI, P. L., (1960); Reizeffekte aus Hippocampus und Corpus mammillare der Katze. Helv. physiol. pharmaeol. Acta, 18, 523-536. SANO,T., (1958); Motor and other responses elicited by electrical stimulation of the cat’s temporal

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lobe. Folia psychiat. neurol. jap., 12, 152-176. SHEALY, c.W., AND PEELE,T. L., (1957); Studies on amygdaloid nucleus of cat. J. Neurophysiol., 20, 125-139. SKULTETY, F. M., (1963); Sti nulation of periaqueductal gray and hypothalamus. Arch. Neurol. Psych iat. (Chic.). 8, 608-620. SPIEGEL, A. E., KLETZKIN, M., AND SZEKELY, E. G., (1954); Pain reactions upon stimulation of the tectum mesencephali. J. Neuropath. exp. Neurol., 13, 212-220. URSDT, H., AND KAADA, B. R., (1960); Functional localization within the amygdaloid complex in the cat. Electroenceph. clin. Neurophysiol., 12, 1-20. E. S., AND NAUTA,W. J. H., (1959); A comparison of the distribution of the fornix VALENSTEIN, system in the rat, guinea-pig, cat and monkey. J. comp. Neurol., 113, 337-363. A. E., (1942); The somatotopical localization of the spinothalamic and secondary trigeminal WALKER, tracts in the mesencephalon. Arch. Neurol. Psychiaf. (Chic.), 48, 884-889. A,, (1902); Das basale Riechbundel des Kaninchens. Anat. Anz., 20, 175-1 87. WALLENBERG, WOOD,CH. D., (1958); Behavior changes following discrete lesions of temporal lobe structures. Neurology, 8, 215-220. WYSS, 0. A. M., (1950); Beitrage zur elektrophysiologischen Methodik. 11. Ein vereinfachtes Reizgerat fiir unabhangige Veranderung von Frequenz und Dauer der Impulse. Helv. physiol.pharmaco1. Acta, 8, 18-24. WYSS,0.A. M., (1963); Die Reizwirkung mittelfrequenter Wechselstrome. Helv. physiol. pharmacol. Acta, 21, 173-188. WYSS,0.A. M., (1965); Beitrage zur elektrophysiologischen Methodik. V. Ein Reizgerat zur konventionellen Impulsreizung. Helv. physiol. pharmacol. Acra, 23, 26-30. YASUKOCHI, O., (1960); Emotional responses elicited by electrical stimulation of the hypothalamus in the cat. Folia Psychiat. Neurol. h p . , 14, 2-267.

128

Facilitation and Inhibition in Centrally Induced Switch-off Behavior in Cats HIROYUKI NAKAO Department of Neuropsychiatry, Kyushu University School of Medicine, Fukuoka (Japan)

(1) Switch-off behavior ( S O B ) An experimental box designed for escape learning, as shown in Fig. 1, has been used throughout the present studies. This box has two small windows in a wall. In front of

Fig. 1 . Apparatus used for the switch-off behavior. The cat switches off the stimulation in response to central stimulation(0)yieldmg a flight response, or to sensory stimulation(0).A buzzer is interrupted by the animal trained with the buzzer as the warning signal of a grid shock.

one window is a plate, pushing of which breaks the stimulation circuit. The animal placed in the box can be trained to push the plate to turn off the stimulation delivered to himself. This type of response will be designated as the switch-off behavior (SOB) in the present experiment.

( 2 ) Switch-off behavior induced by sensory stimulation The animal placed in the experimental box learns to push the plate to terminate a grid shock to the feet or a warning buzzer of the grid shock (Nakao, 1958). Experiments have been carried out to see whether brain stimulation can be substituted for stimulation of sensory receptors in the SOB. Delgado et al. (1954) have shown that

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pain-like responses accompanied by a strong aversive drive can be elicited by stimulation in the neighborhood of the medial lemniscus or the spinothalamic tract. In recent experiments (Aoki and Nakao, 1965), it was found that medial lemniscal stimulation at pontine levels evoked the SOB (Fig. 2C).

( 3 ) Hypothalamic switch-off behavior Hess (1949) demonstrated in cats that flight reactions develop during hypothalamic stimulation. This result suggests that the SOB may occur when a flight response is produced by hypothalamic stimulation. The animal with a hypothalamic electrode that could give rise to a flight response, was placed in the experimental box, and the hypothalamus was stimulated monopolarly. The animal pushed the plate while trying to get out of an inadequate opening of a window, switching off the hypothalamic stimulation. After such an accidental pushing had been repeated several times, the animal learned to remove the hypothalamic stimulation by pushing the plate. Studies revealed that the response time, i.e. the time from the onset of stimulation to switching off, decreased to a certain level after training trials had been repeated a few hundred times. The response time was constant with constant intensity, and increased as the intensity decreased. The term hypothalamic SOB will be applied to the SOB induced by hypothalamic stimulation. The response was considered to be positive if an animal maintained a highly stable value of the response time during several hundred trials. If a response time increased or remained unstable, the response was discarded in the present study. Mapping studies showed that the sites yielding the SOB fall medially in the hypothalamus except for the ventromedial region. A low threshold area was found in the anterior hypothalamic nucleus (Fig. 2A). ( 4 ) Mesencephalic switch-off behavior Studies carried out by stimulation of the brains of freely moving cats indicated that emotional responses were obtained not only from the hypothalamus but also from the central gray matter of the midbrain (Spiegel et al., 1954; Delgado, 1955; Hunsperger, 1956; Skultety, 1963). In these reports there was a diversity of opinion concerning the nature of the response elicited on electrical stimulation of the central gray matter. The purpose of the present study was to test whether the flight response from the midbrain can be used to motivate the SOB. At first, more than 400 points in the border between the hypothalamus and the midbrain, and in the dorsal half of the midbrain, were stimulated in order to find the points yielding the SOB. It was found from these pilot experiments that a few points within the central gray matter at the middle and caudal portions could elicit the SOB. The next experiments were concentrated on the central gray matter of the midbrain to determine more precisely the anatomical structures involved in the SOB. For this purpose more than 300 points were explored within this area. The points evoking the SOB were found within the points inducing forward locomotion. The localization of the electrodes is presented in Fig. 2B. References p . 143

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A

B

Fig. 2. Representative sections of the cat’s brain to show the points stimulated to induce the switchoff behavior in the hypothalamus (A), the midbrain (B), and the pons (C). Numbers above sections indicate the anterior and posterior stereotaxic plane. Abbreviations: F,fornix; Mb, mammillary body; ML, medial lemniscus; NO, nucleus of oculomotor nerve; NT, nucleus of trochlear nerve; NvT, trochlear nerve; OT, optic tract; Vm, nucleus of ventromedialis.

It has been shown that pain-like responses can be evoked by stimulation of the central gray matter (Spiegel et al., 1954; Delgado, 1955). In the SOB experiments cats with electrodes yielding pain-like responses, as shown by high-pitched screeching,were not used. (5) Effect of stimulation of basal forebrain areas an hypothalamic switch-off behavior

In these experiments, the influence of basal forebrain stimulation on the response time of the hypothalamic SOB was studied. Double stimulation is the overlap of a proper stimulus for the hypothalamic SOB and an additional stimulus to another

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Fig. 3. Theeffect ofstimulationofasecondareaon thehypothalamic switch-off behavior (s). Symbols: A,complete inhibition, i.e. no performance; A,marked inhibition; A, slight inhibition; x, no effect; 0, indefinite effect; 0 , marked facilitation; 0, slight facilitation. Abbreviations: fx, fornix; vm, nucleus of ventromedialis.

0L-

1 2 3 4 s

---1 2 3 4 s

1 2 3 4 s

1 2 3 4 5

1 2 3 4 5 trial

Fig. 4. The effects of additional stimulation on the response time of the switch-off behavior. The location of the electrodes and the intensities of additional stimulation: A, lateral hypothalamus (15.5), 0.5 V; B, lateral hypothalamus (13.0), 1 V; C, diagonal band (17.0), 1 V; D, anterior hypothalamic area (13.5), 1 V; E, anterior hypothalamic area (13.5), 0.5 V, 60 c/s A. C., each. Proper stimulation of the switch-off behavior was applied in anteromedial portion of the hypothalamus. The intensities of proper stimulation: A, 0.5 V; B, 0.5 V; C, 1 V; D, 1 V; E, 0.8 V, 60 c/s,A. C., each. Crosses show no performance of the response. References p . I43

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point. The additional stimulus was started 1 sec before the onset of the proper stimulus, and was adjusted to subthreshold strength when applied to medial hypothalamic points from which the SOB could be elicited. At other sites the stimulus intensity was adjusted to threshold intensity, when it evoked overt responses. The animal was able to turn off both stimuli by pushing a plate in the experimental box. Response time was measured to the proper stimulus. An experimenterinterrupted the stimulation if the animal failed to switch off the stimulation during 10 sec, and the trial was qualified as no performance. The results are illustrated in Fig. 3. Stimulation of the medial hypothalamic area with the exception of the ventromedial nucleus had a facilitatory effect on the hypothalamic SOB (Fig. 4D, E). Stimulation of the ventromedial nucleus caused a reaction accompanied by snarling and other full rage manifestations and, when applied as an additional stimulus, resulted in a complete inhibition of the hypothalamic SOB. It has been reported that the lateral hypothalamus and the ventromedial hypothalamus mediate opposing effects on feeding and self-stimulation (Anand, 1961; Hoebel and Teitelbaum, 1962; Olds, 1962). Stimulation of the lateral hypothalamus, however, inhibited the hypothalamic SOB (Fig. 4A, B) as ventromedial stimulation did. Thus, both areas seemed to have the same effect in regard to the SOB, but the mechanism of the inhibition was different. The SOB was disturbed by exploratory behavior during stimulation of the lateral hypothalamus, while the SOB was inhibited by cessation of locomotion during ventromedial stimulation. Among lateral hypothalamic points which were expected to have an inhibitory effect, some points induced a facilitatory effect on the hypothalamic SOB. This may be due to initiation of a forward movement elicited by lateral hypothalamic stimulation. A zone situated at the base of the brain, just rostra1 to the optic chiasma, which has been termed the basal forebrain synchronizing area (Sterman and Clemente, 1962), showed an inhibitory effect on the hypothalamic SOB (Fig. 4C). This effect was induced by unilateral stimulation of the area which caused sniffing and exploring. Although it is hard to determine whether the change in response time in double stimulation is attributed to modification of the reinforcing property or of some performance capacity, it seems very likely that changes in the intensity or the characteristic of the reinforcing play a main role. ( 6 ) Hypothalamic switch-oJ behavior and limbic after-discharges

Stimulation of the areas described above showed stimulus-bound effects on the hypothalamic SOB. The situation is different in limbic structures, because their stimulation readily causes after-discharges wherever stimulated, and when the discharges are induced it has a tendency to spread throughout limbic structures and even the whole brain (Goodfellow and Niemer, 1961; Walker and Udvarhelyl, 1965). The only exception is the anterior cingulate gyrus whose after-discharges seldom propagate to other areas (Nakao, 1963). Observation of animals during such after-discharges has shown that there is little or no evidence of convulsive phenomena in the body’s musculature despite the intense

7

1

m

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discharges. It has, however, been reported that there is an apparent loss of an animal’s ability to perform an avoidance task to respond to sensory stimulation during hippocampal after-discharges (Delgado and Sevillano, 1961). It is therefore of interest to assess the effects of after-discharges from some limbic foci on the centrally-induced SOB. Bipolar stimulation and recording were made in the following studies. (a) Hippocampal after-discharges The first of this series of experiments dealt with the effects of dorsal hippocampal after-discharges on the hypothalamic SOB (Nakao, 1962). Symptoms of hippocampal after-discharges have been described in detail (Andy and Akert, 1955; Delgado and Sevillano, 1961). Slight changes in appearance of the animal were produced until the after-discharges propagated to the ipsilateral amygdala. During the after-discharges, which were localized in the dorsal hippocampus, there was no appreciable alteration of the hypothalamic SOB. When there was propagation of the after-discharges to the amygdala, a partial loss of the performance ability occurred (Fig. 5E, F, G). As the discharges developed in the amygdala, ipsilateral facial twitchings were observed, often with salivation. Sometimes the after-discharges in the amygdala suddenly changed into the typical amygdala 4-6 c/sec spiking called the reactive after-discharge (Delgado and Sevillano, 1961), coinciding with the start of masticatory movements. A complete loss of the performance of the hypothalamic SOB was observed when the reactive after-discharges appeared in the amygdala (Fig. 51, J). Our observations of the appearance of these animals indicated that the responses were impaired to some extent in the twitching phase and completely inhibited in the masticatory phase of hippocampal after-discharges. These results appear to indicate that the active participation of the amygdala inhibits the hypothalamic SOB during hippocampal after-discharges, because facial motor effects have a clear correlation with the activity of the amygdala.

(b) Amygdaloid after-discharges

Amygdaloid stimulation commonly produces immediate results, namely ipsilateral facial movements and often masticatory movements accompanying slow spiking afterdischarges. The animal, contrary to expectation, maintained normal performance during the slow after-discharges in the amygdala which corresponded to the reactive after-discharges of the amygdala observed during hippocampal after-discharges (Fig. 6B, C, D, E). In other words, twitching and mastication, which are correlated with inhibition of the hypothalamic SOB during hippocampal after-discharges, are not accompanied by inhibition of the hypothalamic SOB during amygdaloid afterdischarges. When fast activity was superimposed on the propagated slow after-discharges in the ipsilateral dorsal hippocampus, an increased response time was often observed, and the severity of the deterioration increased as the fast discharges developed in the hippocampus. After the propagated hippocampal after-discharges became fast discharges, the animal lost its performance ability (Nakao and Yoshida, 1963) as illus-

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a I-1LGv . -a

1202-6-13-65

Fig. 8. The effects of hypothalamic after-discharges on the hypothalamic switch-off behavior. The abbreviations are the same as in Fig. 5.

trated in Fig. 6F. The present result reveals that the functional deficits of after-discharges do not always depend on the slow discharges of the amygdala.

Cingulate after-discharges Anterior cingulate stimulation induces short after-discharges which are localized in the cingulate gyms and which may propagate slightly to other areas. Posterior cingulate stimulation induces after-discharges which propagate to other structures. Localized after-discharges did not disturb the hypothalamic SOB performance (Fig. 7,2B), but the after-discharges with some propagation in the dorsal hippocampus caused an increase in response time. The response was abolished when marked after-discharges appeared in the dorsal hippocampus accompanied by some propagated after-discharges in the amygdala (Fig. 7, IB). It was sometimes observed that after cessation of cingulate after-discharges, propagated after-discharges in the dorsal hippocampus (c)

References p. 143

lreC

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continued to develop, or spontaneous after-discharges started anew in the dorsal hippocampus as if the dorsal hippocampus were stimulated, and the after-discharges developed and propagated. The effect of these after-discharges on the hypothalamic SOB was similar to the effect of hippocampal after-discharges induced by electrical stimulation of the dorsal hippocampus (Nakao, 1964). ( d ) Hypothalamic after-discharges The site within hypothalamus which upon stimulation elicits after-discharges is the fornix (Nakao, 1963).As the after-discharges propagate to the hippocampus and then to the amygdala, no localized after-discharges in the hypothalamus are obtained. The behavioral responses during the after-discharges, and the inhibitory effects on the hypothalamic SOB were similar to those observed in hippocampal after-discharges. The results of this experiment are shown in Fig. 8. The animal regularly switched off the hypothalamic stimulation during hypothalamic after-discharges (Fig. 8, 1); it therefore was tempting to suggest that hypothalamic after-discharges do not affect the hypothalamic SOB. Hypothalamic after-discharges, however, disrupted the response when the after-discharges developed fully in the amygdala (Fig. 8, 2G).

( 7 ) Mesencephalic switch-of behavior and limbic after-discharges (a) Hippocampal after-discharges

Fig. 9 shows one instance in which a complete inhibition of the mesencephalic SOB occurred during hippocampal after-discharges (Fig. 9E, F).This impairment is attributed to full propagation of the after-discharges into the amygdala, especially to the appearance of slow discharges in the amygdala. Similar results have been obtained in a study of the hypothalamic SOB, as described above. ( b ) Amygdaloid after-discharges A complete performance of the mesencephalic SOB was observed during propagated after-discharges in the hippocampus which were synchronized with the after-discharges in the amygdala and were evoked by amygdaloid stimulation. (Fig. 10, IB, C). The appearance of fast discharges in the propagated after-discharges in the hippocampus was an indication of complete loss of the performance ability (Fig. 10, lD, 2B, C). The findings observed here were also found in the hypothalamic SOB. ( 8 ) Comparison of inhibitory effects of limbic after-discharges on hypothalamic and

mesencephalic switch-off behavior

This study has been extended to investigate whether or not a distinction can be made between the inhibitory effects of limbic after-discharges upon hypothalamic and mesencephalic SOB. Animals were trained to push a plate either on hypothalamic stimulation or onmesencephalic stimulation. These stimulations were delivered alternately before, during, and after limbic after-discharges. References p. 143

140

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Fig. 11. Comparison of the hypothalamic switch-off behavior and the mesencephalicswitch-off behavior during hippocampal after-discharges.The conventions are the same as in Figs. 5 and 9.

The inhibitory effects on each SOB can appear in various combinations. Inhibition intensity on each SOB could not be compared unless one response still remained after abolition of the other. In general, the inhibition increased as time passed during afterdischarges. If the mesencephalic SOB suffers from limbic after-discharges much more than the hypothalamic SOB, the latter will be performed after abolition of the mesencephalic SOB. The converse is true if the mesencephalic SOB remains after the hypothalamic SOB is abolished. Sixteen animals were prepared to compare these two kinds of SOB under limbic after-discharges. Results obtained from 12 of them are available for comparison. With only one exception, the animals performed the hypothalamic SOB after the mesencephalic SOB was abolished during hippocampal (Fig. 11G) or amygdaloid after-discharges (Fig. 12G, I). The results indicate that the hypothalamic SOB resists hippocampal or amygdaloid after-discharges more than the mesencephalic SOB. References p.!143

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Fig. 12. Comparison of the hypothalamic switch-off behavior and the mesencephalic switch-off behavior during amygdaloid after-discharges. The conventions are the same as in Figs. 5 and 9

(9) Summary

This report is concerned with a learned response in which the cat pushes a plate to terminate the stimulation applied. This response is termed the switch-off behavior (SOB). Studies have been carried out on the neural mechanism that affects the SOB. The findings were as follows: (1) The hypothalamically induced SOB (hypothalamic SOB) was facilitated by stimulation of the medial hypothalamus except the ventromedial nucleus, and inhibited by stimulation of the lateral hypothalamus, the ventromedial nucleus, or the anterior portion of the basal forebrain areas. (2) Limbic after-discharges initiated from the hippocampus, the amygdala, the cingulate gyrus, or the hypothalamus inhibited the hypothalamic SOB to various extents.

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(3) Similar results were obtained with the mesencephalic SOB during hippocampal or amygdaloid after-discharges. (4) The inhibitory effects of the hippocampal or amygdaloid after-discharges on the mesencephalic SOB were more severe than those found on the hypothalamic SOB. REFERENCES ANAND,B. K., (1961); Nervous regulation of food intake. Physiol. Rev., 41, 677-708. ANDY,0. J., AND AKERT,K., (1955);Seizurepatternsinduced by electrical stimulation of hippocampal formation in the cat. J. Neuropath. exp. Neurol., 14, 198-213. AOn, I., AND NAKAO,H., (1967); To be published. DELGADO, J. M. R., (1955); Cerebral structures involved in transmission and elaboration of noxious stimulation J. Neurophysiol., 18, 261-275. DELGADO, J. M. R., ROBERTS,W. W., AND MILLER,N. E., (1954); Learning motivated by electrical stimulation of the brain. Amer. J. Physiol., 179, 587-593. J. M. R., AND SEWLLANO, M., (1961); Evolution of repeated hippocampal seizures in the DELGADO, cat. Electroenceph. clin. Neurophysiol., 13, 722-733. GOODFELLOW, E. F., AND NIEMER, W. T., (1961); The spreadof after-dischargefromstimulationofthe rhinencephalon in the cat. Electroenceph. clin. Neurophysiol., 13, 710-721. HESS,W. R., (1949); Das Zwischenhirn : Syndrome, Lokalisationen, Funktionen, Basel, Schwabe. HOEBEL, B. G., AND TEITELBAUM, P., (1962); Hypothalamic control of feeding and self-stimulation. Science, 135, 375-377. HUNSPERGER, R. W., (1956); Affektreaktionen auf elektrische Reizung im Hirnstamm der Katze. Helv. physiol. Pharnaacol. Acta, 14,70-92. NAKAO,H., (1958); Emotional behavior produced by hypothalamic stimulation. Amer. J. Physiol., 194,411-418. NAKAO,H., (1958); Study of a learned behavior motivated by hypothalamic stimulation in cats. Sei-shin-kei-shi, 60, 1396-1401 (In Japanese). NAKAO,H., (1962); The spread of hippocampal after-discharges and the performance of switch-off behavior motivated by hypothalamic stimulation in cats. Fol. psychiat. neurol. yap., 16, 168-180. NAKAO,H., (1963); Learned behavior motivated by hypothalamic stimulation and brain stimulation and brain lesion in cats. No-shinkei., 15, 1117-1129 (In Japanese). NAKAO,H., (1964); The spread of cingulate after-discharges and the performance of switch-off behavior motivated by hypothalamic stimulation in cats. Fol. psychiat. neurol. jap., 18, 153-160. NAKAO,H., AND YOSHIDA, M., (1963); Effect of amygdaloid after discharges on switch-off behavior motivated by hypothalamic stimulation in cats. Fol. psychiat. neurol. Jap., 17, 221-229. OLDS,J., (1962); Hypothalamic substrates of reward. Physiol. Rev., 42, 554-604. SKULTETY, F. M., (1963); Stimulation of periaqueductal gray and hypothalamus. Arch. Neurol., 8,608-620. SPIEGEL,E. A., KLETZKIN, M., AND SZEKELY, E. G., (1954); Pain reactions upon stimulation of the tectum mesencephali. J. Neuropath. exp. Neurol., 13, 212-220. C. D., (1962); Forebrain inhibitory mechanisms: sleep patterns inSTERMAN, M. B., AND CLEMENTE, duced by basal forebrain stimulation in the behaving cat. Exp. Neurol., 6, 103-117. WALKER, A. E., AND UDVARHELYL, G. B., (1965); Dissemination of acute focal seizures in the monkey. II. From subcortical foci. Arch. Neurol., 12, 357-380.

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The Limbic System and Behavioral Reinforcement JAMES OLDS Department of Psychology, The University of Michigan, Ann Arbor, Mich. (U.S.A.)

What neurohs of the brain are involved in voluntary actiohs and how are they involved? Evarts (1964) discovered patterns of activity of pyramidal neurons characteristic of sleep, paradoxical sleep, waking, and activity. Can observations of this type be extended to permit an understanding of differences in pattern in these and other neural systems which occur while a neuron is involved in an ongoing voluntary act? Prelirhinary experiments reported here suggest that we will be able to make the appropriate observations. Because the neurons involved in any particular pattern of voluntary behavior might be hard to find in the course of normal microelectrode explorations, a method was adopted to circumvent the difficulty. To assure that a neuron under study would be involved in the final voluntary pattern, a neural discharge pattern itself was chosen as the ‘behavior’ to be reinforced in a conditioning experiment. Under these circumstances, if neurons at a recording site could participate in a voluntary pattern, then some behavior would be ‘shaped’ by these procedures whose performance involved an increment in the pace of a neuron under study. The questions that could then be asked concerned (1) the differences in firing pattern as between ‘voluntary’ and ‘involuntary’ bursts of discharges of the neuron, (2) concomitant variation in neighboring neurons, (3) changes in degree of voluntary control attainable contingent on variation in peripheral stimulation, (4) the relative difficulty involved in bringing neurons of different anatomical structures under voluntary control, (5) the behavior patterns associated with the voluntary control of neural patterns in particular brain areas. METHOD

In these experiments, patterns of neural activity were recorded from the brain during periods of ‘quiet waking’ by means of implanted microelectrodes. Patterns which occurred rarely at the outset were ‘conditioned’ to occur more frequently by methods of operant conditioning. Afterwards, these conditioned brain responses were brought under control of ‘discriminative’ stimuli so that periods of operant neural behavior could be alternated with control periods at the will of the experimenter. This permitted observation of the neural and behavioral concomitants of these neural responses when they occurred as components of voluntary behaviors. The neural concomitants were recorded simultaneously from other implanted microelectrodes, some nearby in the

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same structure, other farther away in the brain ; behavioral concomitants were recorded cinematographically. The method also permitted observation of pattern differences between the activity of neurons in the ‘random’ state and activity of the same neurons in the ‘operant’ state. The methods used have been described in more detail in a previous paper (Olds, 1965).

( A ) Preparation (1) Nichrome wires of 67 p diameter with formvar insulation of about 10 p thickness were used as microelectrodes. The wires were insulated at the factory (Driver-Harris Company, Harrison, N. J. ( U S A . ) or Johnson-Matthey Co., Ltd., 73-83 Hatton Garden, London, E C1, England) and cut with scissors. The cut cross section of the tip was the uninsulated portion of the probe. (2) Microelectrodes were implanted in ventral midbrain, aimed at the pons, and in dorsal forebrain, aimed at the hippocampus, and in other parts of the brain for special tests. Stereotactic guidance was used to lower each probe to its target area, and electrophysiological guidance was then used to bring it into a region of recordable neural activity. Recurrent negative spikes of constant amplitude (200 to 500 p V ) with signal to noise ratio of more than 2 to 1, and with duration of 0.2 to 0.7 msec were taken to be single unit responses if they were dependent on movement of the microelectrodes in a 10 to 100 p range. If one or several unit responses in the target area were recordable for a 15-min period, the probe was fixed to the skull by means of acrylic. A screw which had been placed in the skull previously, and a Z bend previously placed in the microelectrode wire helped to hold the acrylic to the skull and the wire to the acrylic. Four to nine microelectrodes were placed in each rat. These plus a larger, uninsulated ground electrode which was also planted in the brain were attached to a 10 contact plug which was also fixed to the skull with acrylic. (3) A test for stable recording was made 1 to 2 weeks after surgery. A 12 inch, 10lead cord of microdot cable was fixed a t its lower end to the 10 contact plug. At the upper end it was fixed to a 10 wire swivel commutator which was mounted on a counterbalanced arm. High-impedance, solid-state preamplifiers were also mounted on the counterbalanced arm. The animal was placed in a cylindrical plastic cage which was 10 to 15 inches in diameter and 11 inches high; the counterbalanced arm was mounted above. The animal was relatively free to move around. All probes were tested to find whether unit responses were still recordable. If unit responses were observed in recordings from a given probe a week after surgery they were usually recordable indefinitely. ( B ) Training This was begun about 2 weeks after surgery. (1) Pretraining: a subject was trained continuously for about 4 days in a movement detecting, feeding cage to remain relatively motionless in order to obtain food, and to go rapidly to eat when a pellet dispenser discharged, making a characteristic noise. (2) Setting of unit discriminators: the animal was then transferred again to the plastic recording cage with cable, commutator, and counterbalanced arm. One of the References p. I64

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unit responses or a set of indiscriminable unit responses recorded from a single probe was chosen for conditioning. The original choice was made by visual inspection of many ‘stopped’ traces on the cathode ray tube of a storage oscilloscope. A simple pattern recognition device was then set to respond whenever one of the chosen action potentials occurred. The device could be set to respond to the action potentials of one neuron if the response was large and well differentiated. More often, however, it was set to respond to several of the unit responses recorded from the same probe. These were then grouped together for the purposes of the ensuing experiment. The criteria for recognition were (1) minimum amplitude, (2) maximum amplitude, (3) minimum ‘fall’ time, and (4) maximum fall time. These criteria were specified by setting 4 potentiometers. The settings were adjusted repeatedly until almost all identified events were clearly within the chosen unit response class by visual inspection and almost all rejected events were clearly outside the chosen class. Thereafter the response of the pattern recognition device was taken as the definition of an occurrence of a unit response. When compared with human judgments of photographed wave forms, the pattern recognition device ‘correctly’ identified about 9 of 10 unit responses and misidentified about 1 of 100 interfering patterns, particularly those emanating from the muscles of the head. Because the unit responses were typically firing at rates in the 10/ sec range and head muscle artifacts during jaw movements were at a rate of 1000/sec, the 1 out of each 100 which were counted as units constituted a substantial source of interference. For this reason, the whole experiment was conducted during those periods when there was no detectable movement or muscle activity. Movement of the animal and muscle activity from jaws and head were detected directly by 3 detectors, 2 mechanical and 1 electrical. Electrical pulses from any one of these caused the analysers and the recorders to be blocked during the movement and for a period of 2 sec thereafter. Thus all recordings were made during artifact-free periods. Long-run records were made from the neurons by counting each pulse which met the amplitude and time-constant criteria of the primary pattern recognizer. An electronic counter was used to count all the spikes of one class; it was set to readout, reset and start again each time it reached a predetermined number. Each such readout made a spike on a moving paper record. The predetermined number was chosen on the basis of convenience between 2 and 127 with a view to obtaining a record with the maximum of detail compatible with a clear separation of spikes. The paper moved at 2 cm/min. As mentioned previously, both movement of the recording paper and counting of spikes were stopped automatically by an electronic gate whenever detectable movement of the animal occurred or when interfering muscle activity reached a criteria1 level. (3) Setting of the ‘burst’ discriminator: a secondary pattern recognition system was set to respond electrically whenever, during an artifact free period, a particularly high frequency of the chosen unit responses occurred. The frequency chosen was so high that it occurred by chance only once every 5 to 15 min during the first several hours of recording (see Fig. 1). The electrical response of this secondary pattern recognizer was used to trigger an event marking pen which recorded its occurrence. During con-

Fig. 1. Three series of 4 sec tracings; highest spikes = 500 pV. The series on theleft and that in themiddle were taken at random during an extinction period; they illustrate to some degree the unconditioned rate of the unit responses recorded from an experimental probe (in this case it was probe No. 6 medially placed in the hippocampus of rat 1588). The series on the right was taken by a method which photographed only bursts which metthecriterionofthe secondary pattern discriminator; in this case the criterion was 20 responses in a 4 sec period. The photographed burst was occurring ‘purposefully’during an acquisition period. Similar bursts sometimes occurred by ‘accident’ during extinction periods (see Fig. 10) but they tended to have a somewhat different pattern. The dots above the unit responses represent the output of the primary pattern discriminator; they indicate which spikes were counted as members of this particular unit group.

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ditioning periods, this response also triggered the food magazine or the application of a reinforcement stimulus. ( 4 ) Reinforcement training: during the first day, the burst discriminator was continuously coupled to the pellet dispenser and each of the high frequency patterns which occurred during an artifact-free period caused a 45 mg pellet to be presented to the animal. The high frequency burst discharged a pellet only if it started at least 2 sec after the last movement or artifact and providing there was a 100-msec period of artifact-free recording after the completion of the burst. This latter requirement was enforced by a delay of reinforcement device which permitted cancellation of reinforcement if artifact occurred during the delay interval. (5) ‘Discrimination’ training: during the second and all following days of conditioning, 2-min periods of acquisition were alternated with 8-min periods of extinction. The onset of each acquisition period was signaled by a stimulus change which was ordinarily sustained for the whole 2-min acquisition period. In these cases, the beginning of each extinction period was signaled by a reversal of the stimulus change. In special tests, only the onset of the acquisition period was signaled; the only indication of the onset of extinction in these cases was the failure of reinforcement. During acquisition periods the secondary pattern discriminator was coupled to the feeder or to a brain stimulator so that each high frequency burst was reinforced either by a pellet or by a $ sec train of brain stimulation (30 pA, 60 c/s, sine wave) in the hypothalamus which had been shown in pretests to have positively reinforcing value. Training was considered to be complete when the animal showed clear and regular discrimination of the acquisition signal by a stable augmentation in rate of the chosen unit pattern during each acquisition period. ( C ) Recordings

In addition to recordings from the unit responses chosen for conditioning, one to three control unit responses were simultaneously recorded and discriminated by means of other discriminators utilizing sometimes other brain probes, sometimes the same brain probe. These units were also counted electronically and marked in convenient multiples on separate channels of the moving record. ( D ) Histological analysis After experiments were completed, brains were cut in 50 p sections. Every other section was saved and mounted; from these, two series were prepared, one stained with Weil stain and the other with cresyl-violet. This permitted precise determination of the electrode track and estimation of the locus of the recording tip. An unusually high proportion of probes fell in or near their target. This was probably due to the clearly recognizable response patterns which often characterized different brain regions when the large fine wire microelectrodes were used. The characteristic responses served to assist the experimenter in homing on target structures during the neurophysiologically guided phase of the implantation procedures.

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RESULTS

( A ) Histological findings The hippocampal probes were found to be in the ridge of pyramidal neurons in the hippocampal gyrus (see Fig. 2). The pontine probes were just posterior and ventral to the interpeduncular nucleus. Some were placed medially in the pontine gray, others were placed 1 to 3 mm away from the midline.

Fig. 2. Photomicrograph of Weil-stained section showing the track of a microelectrodein the hippocampus. The probe penetrated through the alveus and was lodged among the pyramidal cells of the Ammons horn. This was the experimental probe No. 5 which was conditioned in animal 1404. It was implanted at 2.5 mm lateral from the midline.

( B ) The recordings ( I ) The spike patterns: in the recordings there were repeating single and multiplespike patterns (see Fig. 3). Action potentials of constant amplitude and large signal to noise ratio were common. Often several of these were observed in recordings from a single probe. A multi-spike pattern which was particularly frequent when probes were in the hippocampus had the form of a decrescendo of 3 to 7 spikes each one about 213 of its predecessor in amplitude; the interspike interval within such a group was relatively brief and constant, giving the impression that despite the amplitude differences, the series of spikes represented a repetitive discharge from a single element. If single or multi-spike patterns were observed in recordings made a week after implantation they usually recurred indefinitely over the 2-week to 3-month period of an experiment. In response to the question of whether these long duration recordings were made from References p . 164

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Fig. 3. Typical neuronpatterns recordedfrom one probe in the hippocampus(probeNo. 6 in rat 1588)

the self-same neurons, the answer is that in some cases it appeared that identical responses from the same elements were recorded for the whole period. In these cases, not only was the amplitude relatively stable, but also the relative sizes of several spikes, or the signal to noise ratio, or the pattern of a decrescendo burst would remain relatively constant over the whole period. Other times gradual changes occurred suggesting a modification of tissues or movement or erosion of the probe; even in these cases it often appeared possible to follow a given unitary response for several weeks through the course of such a change with the conviction that despite the change in amplitude or template this was still the ‘same’ response. (2) The long run variability; The unit discriminator was usually set to respond to any member of a group of units rather than to a single unit. However, there were often two non-overlapping groups of units counted separately from the same probe in order to study correlations among proximal groups of neurons. The paper speed of 2 cm/min (i.e. 3 sec/mm) set limits on the analysis reported here. When the analysis system was set to read out at rates appreciably higher than 1 per sec, the detailed record disappeared. Thus gross but not fine changes in rate were observed. The highest sustained rates of the chosen response or response class determined the actual settings of the counting devices. When several units were counted together, rates were often very high and a correspondingly high counter setting was used; when a single unit was counted rates were often very low and the counter was set to read out at some relatively low number. Whatever the setting, the counter upon reaching this number would

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make a mark on the moving record, and then reset and start counting again within a 0.5-msec interval. The most striking aspect of the records so formed was that all discriminators at one time or another exhibited rate changes that had the appearance of waves with 10 to 20 min periodicity; for several unit groups in the same animal, these waves were to some degree synchronized. The waves had the form of a gradual crescendo of readout rate up to a maximum followed by a relatively abrupt slowing. Because the 10 to 20 min periodicity was similar to that recently reported for rat sleep-cycles by Roldan ‘et al. (1963) it is appealing to suppose that these were waves of sleep and waking, and that the waking occurred at the point of abrupt slowing. Slow wave records were not made however and it is therefore not possible to offer direct evidence in behalf of this supposition. Besides these 10 to 20 min cycles, there were longer run changes which are so far unexplained. When the average response rates for a given discriminator for different 10-min periods were compared it was not unusual for these to vary by a factor of 2 or more during a period of several hours; sometimes there was even a 5-fold variation (Figs. 4 and 5). Several discriminators responding to different groups of neurons recorded from the same probe or from other probes in the same structure often gave evidence of a positive correlation among these long run changes of rate. Correlations appeared to be higher within structures than between structures and to be positively related to anatomical proximity of the two groups of neurons. Negative correlations of these gross changes were not observed frequently ;this was particularly true prior to conditioning. Because two negatively related neurons could occur during alternating small time intervals it was not impossible for such neurons to appear positively correlated when rates were averaged over periods of several minutes. It is only these longer run averages which are presented in the present report and this may account for a predominance of positive correlations. Later, analyses in more detail may reveal a larger number of negative relations. ( 3 ) Classical Pavlovian conditioning ? During operant conditioning experiments, the long run variability of neuron responses (analysed grossly by averaging over several minute periods) was regularly brought under some control so that response rates for the series of 2 min acquisition periods were often systematically different from response rates during the interspersed series of 8 min extinction periods. During the early phases of discrimination training, these controlled changes of rate followed an unexpected course that seemed better interpreted in terms of Pavlovian conditioning than in terms of operant conditioning. This was particularly true in the case of certain hippocampal unit responses. In these cases, before the ‘conditioned’ unit response pattern showed any evidence of discrimination, one of the other ‘control’ patterns would present clear evidence that a discrimination already existed (see Fig. 6). The animal, however, still made the ‘operant’ unit response on a chance basis and by the same token the animal still earned reinforcements on a chance basis. One might say that the animal already knew what to expect but did not yet know what to do. In some cases, during these early phases of conditioning the animal actually made fewer of the operant responses References p. 164

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Fig. 4. Long run variability in 4 different hippocampal neuron groups recorded simultaneously. The period of 6 h represents artifact-free time taken from a longer period of real time (about 12 to 20 h in this case). In all figures and in the text the time baseline is got by using a clock which ran continuously during quiet periods and stopped completely during and 2 sec after periods of detectable movement or artifact. Rates were averaged for 10-min periods: the top point in each case represents the rate for the most active period of an hour, the corresponding low point represents the rate for the quietest period of the same hour. It should be noted that the different curves are plotted on different ordinates and the origin is not zero in any of the cases; low points and high points of the 6-h curves are noted by numbers indicating responses per minute simply to aid inspection. All probes were in hippocampal gyrus (HPC) with laterality (L) at 1,2.5 and 3.5 mm from the midline as indicated. The two probes at 3.5 mm were a twisted pair of microelectrodes cut to the same depth.

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during acquisition than during extinction periods. If this happened repeatedly when the animal was hungry and if the difference was stable and significant, it was assumed that the unit responses had an a priori negative relation to some aspect of the acquisition period, being depressed by the visual signal or by the expectancy or immediacy of food or by similar causes unknown. Even in such cases, there was often a full reversal during the later phases of training so that a stable difference in favor of the acquisition period occurred. Because these ‘classically conditioned’ changes in rate of the experimental neurons or other neurons were observed mainly with the hippocampal neurons and not in experiments on pontine neurons it is possible to suppose that they derived from some unconditioned or previously established relation between hippocampal neuron activity and the hunger drive and food rewards which were used to shape operant behavior in these experiments. ( 4 ) Operant conditioning: when this was successful, the high frequency response of the chosen unit group occurred far more frequently during the acquisition periods than during the corresponding extinction periods. While this could occur without any overall shift in response rate (by alternation of very high frequency intervals with silent intervals) it was ordinarily associated with a clear increment in the rate of the chosen unit responses during the acquisition periods. The criteria1 high frequency response often underwent an almost all or nothing change as between acquisition and extinction. That is, there would be no sustained frequencies high enough to meet criterion during extinction periods, even though these would occur often during acquisition. At other times these high frequency responses would occur in acquisition and extinction periods but would occur with considerably greater frequency during acquisition. In either case it was common for the background unit response rate to undergo a sizeable increment in rate during acquisition usually amounting to at least a doubling (see Figs. 7, 8 and 9 and Table I). Sometimes rates went up even 5 or 10 fold during acquisition periods and very rarely the units would be almost silent during extinction periods. ( 5 ) Covariation: in the cases of successful conditioning, it was the rule for other unit groups to show clear covariation or clear inverse variation whenever the acquisiReferences p . 164

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tion or extinction changes occurred. While the degree and sign of the correlation changed depending on the anatomical proximity of neuron groups and depending on other unknown factors, it was a rare occurrence for a relative absence of correlation to obtain between two groups of neural responses, particularly insofar as the changes caused by conditioning were concerned.

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In the case of pontine probes, all the other units recorded from the pons showed clearcut positive or negative changes as between acquisition and extinction as soon as one group was conditioned to the point where it exhibited a clear discriminative response. This seemed to indicate that in conditioning one pontine neural group, the procedures actually conditioned all of them. Most pontine units were faster in acquisition periods and slower in extinction, thereby following the changes of the conditioned units. In two cases, however, a medial lateral distinction occurred; a lateral unit group was conditioned and other lateral units followed its accelerations and decelerations; medial neurons in these cases, however, exhibited inverse variation. In another experiment, a control unit group in anterior lateral hypothalamus was uncorrelated with a conditioned group in the pons; in this case there was neither positive nor negative correlation. The picture with hippocampal neurons was somewhat different. In these cases also the conditioned change in response rate of the chosen units had widespread ramifications. More units showed positive correlation with the conditioned neuron group than negative. However, independence of two neuron groups within the hippocampus appeared possible (see Fig. 9), and negative correlation between the experimental units and closely neighboring units occurred even when more distant neurons were positively correlated. One interesting observation in a hippocampal experiment was an apparent change in correlation pattern which accompanied conditioning. The unit groups involved had seemed positively correlated when 10-min intervals were used as a basis for comparison prior to conditioning (see Fig. 4 :6 and 7). Later, one was chosen as the experimental group and its rate was greatly augmented during 2 min acquisition periods after training was complete. At this point, the other group was a control set and its rate was greatly depressed in'acquisition (see Fig. 7 : 6 and 7). (6) Interspike intervals: Whenlthe high criteria1 rates of the experimental unit (which constituted the operant response) occurred during extinction periods after

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conditioning was complete, they seemed to have a different pattern from that exhibited during the more 'purposeful' responding of acquisition. Extinction responses seemed to be constituted by one or several very fast bursts of the kind which Evarts found to characterize pyramidal neurons during sleep. There were brief and regular interspike intervals within these bursts and relatively long silent periods between them (see Fig. 10). Acquisition responses on the other hand seemed to be more often com-

Fig. 10. Photographed extinction bursts. These should be compared with correlated acquisitionbursts (right hand group in Fig. 1).

posed of interspike intervals which were not so regular or so brief; and the longer silent periods which intervened between the bursts in extinction did not appear so often in the course of acquisition responses. In the case of one hippocampal neuron group interspike interval histograms were plotted from photographic records of successful high frequency bursts. The histograms did not show any significant difference between acquisition and extinction bursts even though the photographs had seemed to indicate that such a difference existed (see Fig. 11). When a similar set of histograms was plotted for a conditioned pons unit, the expected difference did appear (see Fig. 12). Extinction bursts had more very short and very long intervals; acquisition bursts had more of in-between duration. ( 7 ) Special tests: Tests for the validity of the operant conditioning were made by (a) changing the discriminative C.S. to find whether these were responses to the acquisition signal, (b) changing the reward to test for an apriori relation of the unit response to the References p . 164

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J. O L D S

expectancy of food, (c), radically changing the visual surroundings, and (d) radically changing the drive level to test for a dependency of the conditioned response on aspects of the internal environment. (a) Discrimination evidenced in the behavior of the experimental neurons was not 351

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greatly impaired by modification of the acquisition signal. Even when this was changed from a light sustained for the whole acquisition period to a bell at the onset only, operant behavior continued to show a marked change at the beginning and shortly after the end of the acquisition period (see Figs. 13 and 14). This appeared to indicate that the pattern involved was not an unconditioned response to the original signal. Moreover, when the unit behavior was extinguished quickly without any extinction signal, it appeared from the record of responses that a definite operant behavior pattern was tried several times and then stopped altogether. This suggested that the operant response might be quite a definite act from the animal's point of view even though it required no overt behavior. Not too much should be made of the absence of

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Fig. 13. Photographed tracings showing rate of control (CONT) units and experimental (EXP) units in the pons. Control units made a mark on the top channel of each tracing when a predetermined number was reached. Experimental units caused a step to occur in the second tracing when a predetermined number was reached, thereby converting the response rate into the slope of a line. Criteria], high frequency patterns (BURSTS) of the secondary pattern discriminator made marks on the lower channel of each tracing. Different control probes in the pons were numbered 1 and 5 ; one control probe in anterior hypothalamus was numbered 7.The experimental probe in the pons was numbered 3. The ‘L‘ number for a given pons probe indicates its distance from the midline. Two minute acquisition periods were alternated with 8 min extinction periods; the recorder and the timer ran only during periods of artifact-free time. Therefore, the actual acquisition and extinction periods were of longer and unknown duration.

an extinction signal, however, as the animal still had the cessation of pellet discharges and pellet-dispenser noises to go by. Even though these occurred after the responses, they still could influence much of the behavior because what was after one response was before the next. (b) When animals were shifted from food reinforcement to brain stimulus reward, this often resulted in a relapse to undifferentiated unit behavior. Then during the course of a single day, conditioning would recur. During such a period of reconditioning with a hippocampal unit probe an interesting series of changes took place which might be considered a foreshortened version of the changes that occurred during the original course of conditioning (see Fig. 15). For the conditioned unit, rates not only rose during successive acquisition periods but also declined during successive extinction periods. For a control unit, rates first stayed at initial levels for both acquisition and extinction periods, then a brief discrimination appeared, the control units following the experimental ones, becoming accelerated during acquisition. Finally, the response rate of the control units fell sharply and there was again no discrimination evidenced in their behavior but now they were responding at a rate considerably below their initial level. Referencrs p . 164

160

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Fig. 14. Photographed tracings showing rates of control and experimental neuron groups in the hippocampus. In ‘bell only’ tests there was an auditory rather than a visual discriminative stimulus, and it was not sustained throughout the acquisition period. In ‘brain reward’ tests, an electrical stimulus in anterior lateral hypothalamus was used for reward rather than a food pellet. In thecurare test, the animal was paralysed and respirated. During the acquisition period there were very few hippocampal responses at the beginning. Therefore, the animal was rewarded first for ‘2 bursts’; i.e. two discharges in a & sec period; then for ‘4 bursts’, and so on up to ‘8 bursts’. This produced a considerable augmentation of the hippocampal discharge rate; in the ensuing extinction period it dropped back toward the prereinforcement level. It did not, however, become augmented during the second acquisition period, and shortly thereafter the animal died. Apparently, the anterior hypothalamic stimulation combined with the curarization t o cause death.

(c) Major modification of the visual stimulus environment had a debilitating influence on the unit behavior. In these tests the cage doors were opened and photoflood lights were used to illuminate the whole area for cinematographic recordings. Direct observation of the animal before and during these photo-flood tests made it clear that the animal was ‘trying’ to get pellets in roughly the same way after the major change in visual environment. Nevertheless, these efforts were not nearly so effective. The animal made fewer operant responses during acquisition tests; and the unit behavior showed less evidence of discrimination between acquisition and extinction. These tests lasted only for short periods and therefore it was not clear whether the changes would have been gradually erased after continued training in the brightened environment. In spite of the poor performance there was still some evidence of discrimination in the brightly lighted environment. Moreover, some of the poor performance may

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have been due to a general reduction in hunger produced by the warming effect of the flood lights. (d) A surprising but in retrospect predictable result was the very great influence of the drive state even when the animal was responding for a brain reward whose reinforcing value in other tests appeared to be unaffected by such drive changes. Such tests were made in only 2 cases and in both cases unit response probes were in the hippocampus. Discrimination evidenced in the behavior of the experimental unit broke down and was not restored until the animal was returned to the high drive state of the original training. It seems possible that this was due to an intrinsic relation between the visceral drives and hippocampal responses.

( C ) The correlated behaviors ( 1 ) Cinematographic recording: Although neither the unit analyses nor the operant behavior could take place during periods of detectable movement or detectable muscle artifacts, there were nevertheless subtle and undetectable movements which accompanied or preceded many operant responses. Cinematographic recordings showed a slow head movement to left or right as a more or less constant predecessor or concomitant of the ‘purposeful’ pontine unit behavior which occurred during acquisition periods. Even if the animal appeared to be standing quite still during the 2-sec period immediately preceding a reinforced burst of units, the head movement was nevertheless apparent in the recordings which immediately preceded the quiet period. Other times a movement which was barely discernible to the eye and which escaped completely the mechanical detectors appeared as the overt component of the pontine operant response*. When hippocampal units were conditioned, there were sometimes clearcut sniffing movements of the tip of the nose combined with whisker movements that similarly accompanied the operant responses. However, with hippocampal responses there were

*

Stimulation of a pontine probe in later tests often seemed to provoke almost the same movement.

References p . 164

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times when the animal emitted apparently purposeful operant unit bursts without any observable overt responding at all. (2) Curarizedpreparation: One animal with conditioned neurons in the hippocampus and positive reinforcement probe in anterior lateral hypothalamus was tested for operant behavior after paralysis induced by i.p. injection of 50 mg/kg of flaxedil. The animal had previously been tested in a high drive condition with hypothalamic stimulation as reward. Clear discrimination behavior had been maintained over a 6-h period immediately preceding the flaxedil induced paralysis. The rat was respirated through the nasal openings by means of a nose grip respirator of a type designed and made in our laboratory and which has been used successfully in many of our experiments. The animal was hyperventilated utilizing air mixed with 5 % carbon dioxide. The animal with respiratory device attached was replaced in the plastic recording cage after having been out of it for about 10 min during curarization procedures. Recording and conditioning procedures had been underway prior to the 10-min interruption and they were reinstituted as soon as the animal was replaced in the recording cage. The rate of the experimental unit responses had fallen considerably, and even though the visual acquisition signal was turned on the animal showed no signs of producing the 16 resp sec burst required to earn a brain stimulus reinforcement. In order to shape the operant behavior, therefore, the burst requirement was set back first from 16 to 2. Then after the first reinforcement it was set forward from 2 to 4; then 4 to 6 and then 6 to 8. During this period the experimental unit responses increased their rate about 60 fold from 32 per min up to 32 per sec. This rate was maintained for 2 min and then acquisition was terminated. During the ensuing extinction period the rate fell again to its low, post curare level; 8 min later a second acquisition test was imposed. No shaping methods were used this time and the response rate did not rise above chance levels. Five min later the animal died, death apparently being caused by interaction of the parasympathetic influences of anterior hypothalamic stimulation and the effects of curare. Non-self-stimulating rats were tested repeatedly before and after the test rat. These withstood and recovered well from the curarization procedure. Animals are now being prepared for further curare tests. The reinforcing electrodes in these new cases are in posterior hypothalamus, where, it is hoped, reinforcing stimulation will not kill the curarized animal. The curarization test was considered a limited success because (1) the reinforcement procedure caused a large increase in rate of the hippocampal response in the paralysed animal at first and (2) when the increment failed to appear a second time the animal was already in a debilitated condition preceding death. However, it would require more evidence than is yet at hand to permit a conclusion that hippocampal neurons can be voluntarily controlled without recourse to overt behavior.

+

SUMMARY A N D CONCLUSIONS

1. Responses composed of patterns derived from pontine and hippocampal units were conditioned to occur more frequently by rewarding the animal after each occurrence of the response.

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2. These patterns were readily conditioned even though detectable movement and muscle artifacts were interdicted before, during and after the response. 3. By rewarding the animals only for those responses made in the presence of a particular stimulus, these unit patterns were brought under the control of this discriminative stimulus. 4. When unit groups in the pons became conditioned all other units in the pons gave evidence of the conditioning. Most other units followed the conditioned response being accelerated during acquisition and decelerated in extinction. But there were two cases where lateral units were conditioned and medial units gave evidence of inverse variation. 5. No similar correlation was observed between pontine units and units in anterior lateral hypothalamus. 6. When unit groups in the hippocampus were conditioned many other units in the hippocampus varied directly and a few inversely with the operant-induced changes in the conditioned units. However, there were cases of independent hippocampus units which did not follow or invert a conditioned change in the experimental unit. 7. In one case, a hippocampal unit which seemed to vary directly with the experimental unit when hour to hour changes were considered prior to conditioning later showed clear evidence of inverse variation when the acquisition and extinction periods were compared after conditioning was complete. 8. Sustained t sec high frequency responses constituted the operant response. When they occurred ‘accidentally’ during extinction they appeared to have more stereotyped bursts and longer intervening silent intervals than when they occurred ‘purposefully’ during acquisition. 9. When pontine units were conditioned, the operant response seemed to include a movement of eyes or head which escaped the movement detectors. 10. When hippocampal units were conditioned the operant response sometimes included movement of nose or whiskers but sometimes there was no obvious peripheral component of the behavior. 11. Hippocampal neurons responded in controlled fashion in midconditioning prior to the emergence of the discriminated operant response. This suggested an a priori connection between the hippocam pal responses and the feeding schedule cycles produced by the alternated acquisition and extinction. 12. Suggesting a similar connection of some hippocampal neurons to hunger drive was the fact that a neural response shaped under high drive conditions with food reward could be transferred to brain reward only if the high drive of training were maintained. 13. Preliminary evidence gave grounds for the hope that it would eventually be possible to observe operant responses from hippocampal neurons when the animal was paralysed with a curarizing agent.

References I. 164

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ACKNOWLEDGMENTS

This work was supported by grants from The National Science Foundation, The National Aeronautics and Space Administration, and the U.S. Public Health Service to Dr. J. Olds. The author is grateful to Fred F. Coury (BSEE) for designing analytic circuits, to Mr. William E. Wetzel for designing brain probes and cages, to Mr. Giulio Baldrighi for surgery and neurophysiological work at the time of surgery and to Mr. Floyd F. Foess for assistance with all aspects of the experimental work. REFERE N C E S EVARTS,E. V., (1964); Temporal patterns of discharge of pyramidal tract neurons during sleep and waking in the monkey. J. Neurophysiol., 27, 152-171. OLDS,J., (1965); Operant conditioning of single unit responses. Proc. XXIIZ Znt. Congr. physiol. Sci., Tokyo, Sept. ROLDAN, E., WEISS,T., AND FIFKOVA, E., (1963); Excitability changes during the sleep cycle of the rat. Electroenceph. clin. Neurophysiol., 15, 775-785.

165

Further Studies on the Effects of Amygdaloid Stimulation and Ablation on Hypothalamically Elicited Attack Behavior in Cats M. D A V I D EGGER A N D JOHN P. FLY" Departments of Anatomy and Psychiatry, Yale University School of Medicine, New Haven, Conn. (U.S.A.)

Evidence on the role of the amygdala in the integration of aggressive behavior is contradictory and confusing. However, most investigators agree that an important function of the amygdala is a modulatory one, modulating the activity of the hypothalamus in particular. If we assume that the amygdala has anatomically distinct regions that suppress or facilitate the hypothalamus, it might be possible to explain many apparently contradictory data. Earlier we found that the amygdala does indeed contain anatomically distinct regions capable of suppressing or facilitating an attack response elicited by stimulation in the hypothalamus (Fig. 1;Egger and Flynn, 1963). However, because we used bipolar needle electrodes with tip separations of 2 mm or more, the anatomical localizations were not precise. To determine more sharply the anatomical loci of suppression or facilitation of the attack response, in 9 cats we stimulated in the amygdala with monopolar electrodes. Experiment I

Methods The details of our stimulation and recording methods have been described (Egger and Flynn, 1963; Wasman and Flynn, 1962). Because the attack response we studied was directed at a rat, we selected our experimental subjects from among 'non-ratters'. Most tame cats do not attack rats. During an experimental session, a cat and an anesthetized rat were placed together in an observation cage. The electrodes, implanted in the cats at least 7 days before testing, were connected to a relay so that they could be switched selectively from connection with an EEG recorder to connection with a stimulator. EEG recordings were taken from the amygdala before and after each stimulation, to determine if electrical after-discharges had occurred. Data recorded during trials on which after-discharges did occur were not included in analyses of the effects of amygdaloid stimulation. Electrical stimulation consisted of biphasic, rectangular-shaped pulses. The electrodes were stainless steel needles, insulated except for 0.6 mm at the tip. Stimulation was unilateral. References p . 180-182

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Fig. 1.Sitesof the tips of amygdaloidelectrodesprojected on diagramsof frontal sections (based on the atlas of the cat brain in Bur& et al., 1962). Filled circles depict electrodesthrough which stimulation produced a suppression of hypothalamically elicited attack behavior; open circles depict electrodes through which stimulation produced a facilitation of hypothalamically elicited attack behavior; and crosses depict electrodesthrough which stimulation produced no consistent effects. The lettered lines connecting the poles of bipolar electrodes identify the electrodes of individual cats (after Egger and Flynn, 1963). AAA = area amygdalaris anterior; AB = nucleus basalis amygdalae; AC = nucleus centralis amygdalae; ACO = nucleus corticalis amygdalae; AHL = area hypothalamica lateralis; AL = nucleus lateralis amygdalae; AME = nucleus amygdalae medialis; APL = area praeopticalaterak; CA = commiissura anterior; CH = chiasma opticum; CI = capsula interna; EN = nucleus entopeduncularis; GP = globus pallidus; P = putamen; SO = nucleus supra0pticus;TO = tractus opticus.

In order to stimulate in two regions of the brain simultaneously (actually, in very rapid succession), we interlocked the outputs of two electrically isolated stimulators in such a way that the two regions of the brain were never stimulated at the same instant, so no inadvertent stimulation of regions lying between the electrode sites could occur. stimulation in the amygdala was monopolar; that in the hypothalamus was bipolar.

STIMULATION A N D A B L A T I O N I N THE A M Y G D A L A

167

The stimulation frequency was 62.5 pulses per sec, with the onset of hypothalamic stimulation typically preceding that of the amygdala by 6 msec. We measured the latencies of initial movement and of attack following the onset of electrical stimulation with stop watches. The attack latency was defined as the interval from the onset of brain stimulation until the cat touched the rat with tooth or claw. Stimulation trials were separated by at least 4 min. Two levels of hypothalamic stimulation were used; the higher level was obtained by increasing the duration of the stimulus pulses. Typical values of hypothalamic stimulation were 0.40 mA, biphasic, peakto-peak, with 1.0 msec per pulse at the low level of stimulation and 1.5 msec per pulse at the high level. Amygdaloid stimulation intensities were chosen at levels below the threshold for electrical after-discharges. We reasoned that a more meaningful anatomical localization of effects might be obtained if we could avoid after-discharges. Typically, the amygdala was stimulated at 0.15-0.20 mA, biphasic, peak-to-peak, 3.0 msec per pulse. Rarely did stimulation of the amygdala alone at these levels elicit observable behavioral reactions. Experimental sessions were separated by at least 46 h. The effects of simultaneous stimulation in the amygdala and the hypothalamus were evaluated systematically for a selected amygdaloid electrode in two successive sessions. During each session, ‘single’, i.e. stimulation in the hypothalamus alone, and ‘dual’, i.e. stimulation in the amygdala and hypothalamus together, trials occurred in ABBA order. The effects of dual stimulation at both low and high levels of hypothalamic stimulation were tested. Each test session consisted of 12 pairs of single and dual stimulation trials. In addition, at least twice during each session, the amygdala alone was stimulated. During trials in which no attack occurred, stimulation was continued for 20 sec; otherwise, stimulation was terminated when an attack occurred. Those trials during which no attack occurred were included in the data analyses as if an attack had occurred at 20 sec. All data on attack and initial movement latencies were transformed to their reciprocals. We shall refer to these reciprocals as ‘speeds’. Because the speed distributions were less skewed than the latency distributions, where possible, statistical analyses were performed on the speeds. In the remainder of this paper, we shall refer to the cats from our earlier study, stimulated with bipolar electrodes in the amygdala, as members of the ‘bipolar’ group ; and to the cats from the present study, stimulated with monopolar electrodes in the amygdala, as members of the ‘monopolar’ group. Results Amygdaloid stimulation in 5 of the 9 monopolar cats produced a statistically significant suppression of attack, i.e. an increase in the time from the beginning of stimulation to the occurrence of an attack as a result of adding stimulation in the amygdala to that in the hypothalamus (Table I ; Fig. 2). In two of the 9 cats, facilitation of attack occurred, i.e. a decrease in the time to attack when both the amygdala and the hypothalamus were stimulated. References p . 180-182

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TABLE I EFFECTS OF ELECTRICAL STIMULATION I N THE AMYGDALA ON LATENCIES OF HYPOTHALAMICALLY ELICITED ATTACK ~

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STIMULATION A N D ABLATION I N THE AMYGDALA

I69

The localization of effects in the amygdala for the monopolar cats agreed with those of the bipolar, and, as hoped, enabled us to define their anatomical loci more precisely : in general, suppression was elicited most consistently in the magnocellular portion of the basal nucleus of the amygdala, and in the anterior and medial portions of the lateral nucleus of the amygdala (Fig. 3). Facilitation was elicited in the dorsolateral portion of the posterior part of the lateral nucleus. One exceptional cat, A, showed facilitation of attack even though, from the location of its electrode in the amygdala, we would have expected suppression. The electrode of cat D is not depicted in Fig. 3. Stimulation through this electrode, located at the interface of the hippocampus and the optic tract, produced no consistent effects on hypothalamically elicited attack. Similarly located bipolar electrode placements also failed to produce consistent effects. At the end of the two test sessions, we stimulated the amygdala by itself at increasing current levels until after-discharges occurred following not more than 20 sec of stimulation. In only one cat, C, did stimulation below the threshold for after-discharges elicit a behavioral response. During stimulation of the amygdala, this cat assumed a defense-like crouch and hissed. At current levels that evoked after-discharges, growling, hissing, and a characteristic amygdaloid-seizure facial twitch also occurred. Stimulation through this electrode, when paired with hypothalamic stimulation, produced a marked suppression of hypothalamically elicited attack.

Discussion Electrical stimulation in the amygdala suppressed or facilitated hypothalamically elicited attack. Opposing effects were elicited from different parts of the amygdala. These effects were not due to the elicitation of behavioral responses by amygdaloid stimulation, because in only one cat did amygdaloid stimulation alone elicit a behavioral response at the current levels used in dual stimulation. The suppression effect was not due to arrest of all motor activity, because at the levels of amygdaloid stimulation used, the cats were responsive and displayed normal behavior patterns (Egger and Flynn, 1963 ;Fonberg and Delgado, 196 1). Furthermore, amygdaloid stimulation was considered to have been effective in suppressing hypothalamically elicited attack behavior only if attack latencies were increased, but latencies of initial movement were unaffected. Finally, the dual stimulation effects were not associated with after-discharges in the amygdala, because data from trials during which electrical after-discharges occurred were excluded. We would like to argue that the effects of amygdaloid stimulation we have observed are due to modulation by the amygdala of neural activity in the hypothalamus. Neuroanatomists have described at least 3 pathways from the amygdala to the hypothalamus in the cat (Fox, 1940,1943; Hall, 1963; Szentiigothai et al., 1962; Valverde, 1963). One pulse per sec electrical stimulation through the electrodes in the amygdala in our cats evoked electrical responses at the hypothalamic electrodes used to elicit attack (Egger and Flynn, 1963). Furthermore, many units in the hypothalamus are influenced by electrical stimulation in the amygdala (Sawa et al., 1959; Stuart et al., 1964; Tsubokawa and Sutin, 1963; Wendt and Adey, 1960). The firing rates of some References p. 180482

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SUPPRESSION

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FACILITATION

Fig. 3. Triangles depict electrodes through which stimulation produced a suppression of hypothalamically elicited attack behavior and open circles depict electrodesthrough which stimulation produced a facilitation of hypothalamically elicited attack behavior. Letters in squares identify the electrodes of individual cats. Broken limes connect the poles of bipolar electrodesof cats, M, N, P, and V from Fig. 1. This figure depicts the locations of monopolar and bipolar electrodesof all cats meeting the following criteria: the effect of amygdaloid stimulation on attack speed, as well as on the interval between initial movement and attack, had to have a statistical signiticance exceeding the 0.01 probability level in the first 24 pairs of trials or less. AHA = area hypothalamica anterior; A1 = nucleus intercalatus amygdalae; APM = area praeoptica medialis; CD = nucleus caudatus; C PYR = cortex pyriformis;F = fornix;HVM = nucleus ventromedialis hypothalami; NCAST = nucleus commissurae anterioris et striae terminalis; NHD =

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units in the hypothalamus can be slowed by stimulation in one part of the amygdala, and speeded up by stimulation in another part of the amygdala, in analogy with the contrasting behavioral effects of amygdaloid stimulation reported here (Egger, 1965). None of these observations, however, tells us anything related directly to attack responses elicited by hypothalamic stimulation. We reasoned that more direct evidence of neural effects of the amygdala on hypothalamic mechanisms concerned with attack behavior might be provided by examining the effects of small lesions placed at the tips of electrodes in the amygdala for which we had determined the stimulation effects. Experiment II

Methods Fifteen cats, 7 monopolar and 8 bipolar, received bilateral amygdaloid lesions. Cats in the bipolar group received anodal lesions at the tips of both poles of the bipolar electrodes in the amygdala. Six cats, 2 monopolar and 4 bipolar, served as shamlesion controls. Anodal electrolytic lesions were made by passing 3.CL5.0 mA direct current for up to 20 sec. A lead clipped to the nape of the cat’s neck served as cathode. Between making a lesion and testing its effect, 27 to 36 days elapsed for the monopolar, and from 2 to 36 days elapsed for the bipolar cats. We assessed the effects of the lesions by comparing attack speeds on corresponding trials of experimental sessions before and after lesioning. Identical intensities of hypothalamic stimulation were used in pre- and post-lesion test sessions. Results Five cats attacked significantly faster after receiving amygdaloid lesions : monopolar cats A and F, and bipolar cats M, N, and 0 (Table 11). The mean pre- and post-lesion attack latencies of cat A are summarized in Fig. 4.Similar data of Cat 0 are summarized in Fig. 5. All 5 cats that attacked faster following lesioning, and 5 of the 10 cats that did not attack faster following lesioning were first tested at least 21 days after lesioning. None of the 6 sham-lesion cats attacked faster during the ‘post-lesion’ test. Four of the 5 cats that attacked faster following lesioning had shown statistically significant suppression of attack during amygdaloid stimulation. The 5th cat, A had its electrodes in an amygdaloid region associated with suppression of attack, namely, the magnocellular portion of the basal nucleus of the amygdala, but it had shown facilitation of attack during amygdaloid stimulation. All 5 cats that attacked faster following lesioning had bilateral lesions that included a common locus : the border region of the magnocellular portion of the basal nucleus of the amygdala and the dorsomedial portion of the lateral nucleus of the amygdala nucleus hypothalamicus dorsalis; NHP = nucleus hypothalamicus posterior; NT OLF LAT = nucleus tractus olfactoriilateralis;NTS = nucleus triangularis septi; PV = nucleus paraventricularis; RE = nucleus reuniens; SCH = nucleus suprachiasmaticus; TMT = tractus mammillo-thalamicus ; V = ventriculuslateralis ;VM = nucleus ventralis medialis;VPL = nucleus ventralis posterolateralis ; ZI = zona incerta. For key to additional abbreviations, see Fig. 1. References p . 180-182

I 72

M. D. EGGER A N D J. P. F L Y N N

Fig. 4. Mean latencies of attack during hypothalamic stimulation for cat A before and after amygdaloid lesions. The total height of each pair of bars represents mean attack latencies for low and high levels of hypothalamic stimulation. The shaded portions at the bottom of each bar show latencies of initial movement. The first post-lesion test (session 2) was given 34 days following lesioning; the last (session 5), 178 days or approximately 6 months following lesioning.

(Fig. 6). These lesions also encroached upon the ventral and medial borders of the central nucleus. None of the 10 cats that did not attack faster following lesioning had lesions that included this region (Fig. 7). Lesions of 3 cats are not depicted in Fig. 7: that of cat D, that had no bilaterally symmetric component to its lesion; and those of cats I and T, that attacked more slowly following lesioning. Cat I had a unilateral amygdalectomy prior to electrode implantation. Post-mortem examination of its brain revealed signs of hydrocephalus and apparent shifts in electrode postitions. The decrease in attack speed following SEC

la

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Fig. 5. Mean latencies of attack during hypothalamicstimulationfor cat 0before and after amygdaloid lesions. The total height of each pair of bars represents mean attack latencies for low and high levels of hypothalamic stimulation. The shaded portions at the bottom of each bar show latencies of initial movement.Thefirstpost-lesiontest(session 3) was given 36 days, the second (session 4) 38 days following lesioning.

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173

lesioning in cat T is unexplained. Sham-lesion cats W, X, and Y also attacked more slowly in ‘postlesion’ tests. Both of the monopolar cats that had shown facilitation of attack during dual stimulation were lesioned. One, cat A, attacked faster following lesioning. The other, cat G, tended to attack more slowly following lesioning, but this tendency was not statistically reliable (P < 0.20). Cats with amygdaloid lesions were tested in the following behavioral situations : they were handled, placed near barking dogs, and placed together in their home cages with mice. No effects of our small amygdaloid lesions were observed in any of these situations. The destruction of a circumscribed region of the amygdala, in which stimulation generally produced suppression of attack, was followed by statistically significant increases in attack speed at fixed intensities of hypothalamic stimulation. All 5 cats that had lesions in this region attacked faster; not one of the 10 cats with lesions in other parts of the amygdala, or of the 6 cats without lesions, attacked faster. The probability of the consistency in location of the effective lesions occurring by chance among the 15 lesioned cats is less than 0.001. Discussion Although in his review, Gloor (1960) concluded that stimulation in the amygdala failed ‘to reveal any consistency in localization of the various amygdaloid stimulation responses’, others have subsequently published data supporting localization of function in the amygdala (e.g. Fonberg, 1967; Koikegami, 1963, 1964; Kreindler and Steriade, 1963, 1964; Ursin and Kaada, 1960). In our earlier experiment using bipolar stimulation in the amygdala, and in the present experiment in which we used monopolar stimulation in the amygdala, electrical stimulation in different regions of the amygdala produced opposing effects on hypothalamically elicited attack behavior. Some anatomical evidence suggests that these regions in the amygdala may be connected to the hypothalamus via different pathways (Fox, 1943; Ban and Omukai, 1959; Hall, 1963). Yoshida (1963) has also found that stimulation in the amygdala can affect responses elicited by hypothalamic stimulation in cats. Table I11 lists selected effects from studies of cats in which electrical stimulation elicited different effects from different regions in the amygdala. While differing in details, these studies of effects of stimulation are remarkably consonant with a functional division within the amygdala, with the dorsomedial amygdala in one functional category, and the lateral and ventral amygdala in another. This bipartite functional division of the amygdala should be considered only a first approximation of a more complicated organization. The dorsomedial division appears related to some sympathetic functions, arousal, and defense ;the lateral and ventral division to a heterogeneous grouping which Koikegami (1964) referred to as parasympathetic and extrapyramidal motor functions. Koikegami (1963) contrasted a proposed functional division of the amygdala with the traditional anatomical division of the amygdala into corticomedial and basolateral nuclear groups (Fig. 8). A similar functional division of References p. 180-182

174

M. D. EGGER A N D J. P. PLY"

Fig. 6. Bilateral amygdaloidlesions of monopolar cats A and F, and of bipolar cats M, N, and 0.A l l these lesions were followed by faster speeds of attack for fixed levels of hypothalamic stimulation. All lesions involved the border region between the magnocellular basal nucleusand the lateral nucleus.

the amygdala is consistent with many of the data in Table 111, as well as with those reported in this paper (Fig. 9). Table I11 does not include studies or effects that differ with the rest of the Table (e.g. Kaada, et al. 1954; the localizations of effects of stimulation on gastric motility, Shealy and Peele, 1957) or in which agreement was uncertain (e.g. Koikegami et al.,

S T I M U L A T I O N A N D A B L A T I O N I N THE A M Y G D A L A

175

Fig. 7. Bilateral amygdaloid lesions of monopolarcats G, E, and H, and of bipolar cats, P, Q, R, and S. None of these lesions was followed by a change in speeds of attack for fixed levels of hypothalamic stimulation.

1957; Kreindler and Steriade, 1963; Wood et al., 1958). However, the major portion of published data provides strong support for functional localization within the amygdala of the cat. Our experiments showed that hypothalamically elicited attack responses can be suppressed by stimulation in a region in the amygdala in which others have elicited References p . 1 8 6 1 8 2

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M. D. EGGER A N D J. P. FLY"

f Fig. 8. Schematic diagrams showing two divisions of amygdaloid nuclei. The traditional anatomica division of the amygdala into corticomedial and basolateral divisions on the left is contrasted with a proposed functional division on the right. From Koikegami, 1963.

defense-like responses. However, we used intensities of amygdaloid stimulation below threshold for elicitation of behavior, hence lower intensities than those used by investigators studying defense reactions elicited by stimulation in the amygdala. In addition, attack directed at a rat is a different response from the defense reactions elicited by others during and following amygdaloid stimulation. Defense reactions are often associated with attack at a threatening object or person. But the attack response we used in our studies was directed preferentially at a rat (Egger, 1962; Levison and Flynn, 1965). Defense reactions are characterized by sympathetic activation. Our attack responses often occurred with minimal signs of sympathetic activation. It should not be surprisingthat regions related to defense might act to suppress attack at a non-threatening object such as a rat. Following bilateral amygdalectomy in cats, some investigators have observed increased docility (Schreiner and Kling, 1953;Brady et al., 1954; Shealy and Peele, 1957; Kling and Hutt, 1958), some have observed an increase in aggressive behavior and rage reactions (Spiegel et al., 1940; Bard and Mountcastle, 1948), and others have failed to detect any changes in affective behavior (Summers and Kaelber, 1962). The data on the effects of partial lesions of the amygdala in cats are still incomplete and less consistent than those on the effects of stimulation (Green et al., 1957; Horvath, 1963; Kling et al., 1960; Macchi et al., 1963; Morgane and Kosman, 1959; Nakao, 1960; Ursin, 1965a, 1965b; Wood, 1958). Small bilateral lesions in the amygdala in our cats produced consistent and reproducible effects on the latencies of hypothalamically elicited attack. Nakao (1960) found no effects of amygdaloid lesions on thresholds for eliciting a variety of behaviors during hypothalamic stimulation, not, however, including attack behavior. In 4 of Nakao's cats, lesioning of the amygdala was followed by faster speeds of performing a learned escape response to turn off hypothalamic stimulation. These 4 cats all had bilateral lesions involving the region that, when injured in our cats, was followed by faster attack speeds (Fig. 10). The cat in Nakao's study in which the lesions produced

177

STIMULATION A N D ABLATION IN THE AMYGDALA

T A B L E I1 E F F E C T S OF L E S I O N S I N THE A M Y G D A L A O N L A T E N C I E S OF H Y P O T H A L A M I C A L L Y ELICITED

ATTACK

Mean attack latencies (sec) Prelesion

cafi stimulation efects2

Bilateral lesions4 including ABM-ALDM

Bilateral lesions4 not including ABM-ALDM

M B OB A M N B F M P B E M Q B

H M D M R B S B

G M T B I M

No lesions

BM C M

VB YB WB XB

S***

S* F*** S***

S**

s*** S* S*

F*** S*

s*** s*** s*** F** F**

Low level of

hypothalamic stimulation3

High level of hypothalamic stimulation3

Statistical significance of post-lesion increase ( I ) or of post-lesion decrease ( D ) in attack speed

B

A

B

A

13.9 10.2 8.3 10.2 3.9

7.0 5.4 5.9 5.3 3.5

3.6 5.2 4.9 5.5 2.8

2.7 3.7 3.6 3.0 2.1

I*** I*** I*** I** I**

13.3 9.3 9.3 5.6 6.7 14.8 11.4 10.4 9.3 7.8

11.5 8.7 7.8 5.7 6.4 16.3 8.2 10.8 19.9

0

8.2 3.6 6.4 2.8 3.2 4.3 7.1 4.6 5.3 6.9

6.8 3.7 8.4 2.8 4.0 5.1 6.7 5.3 12.2

D**

5.9 7.0 8.5 8.6 4.5 15.1

7.0 6.3 6.1 17.1 6.3 20.0

4.1 4.0 4.5 6.6 3.0 4.7

4.0

0

3.4 5.5 10.6 5.5 12.4

D* D*** D***

‘B’refers to bipolar and ‘M’ refers to monopolar pre-lesion electrical stimulation in the amygdala. ‘S’ refers to suppression and ‘F’ refers to facilitation of hypothalamically elicited attack during simultaneous stimulation in the amygdala. 3 ‘B’ refers to attack latencies before lesioning; ‘A’ refers to attack latencies after lesioning. 4 ‘ABM-ALDM’ refers to the border region between the magnocellular portion of the basal nucleus of the amygdala and the dorsomedial portion of the lateral nucleus of the amygdala. 5 No attack responses could be elicited. See text. * P < 0.05;** P < 0.01; *** P < 0.001. 1 2

the smallest effect on escape latencies, cat 21 1, had the least involvement of this region. Fernandez de Molina and Hunsperger (1962) found no effect in 3 cats of bilateral lesions in the amygdala on latencies of hissing and fiight elicited by hypothalamic stimulation. But lesions in our cats followed by faster attack speeds had no effects on latencies of initial movements. In conclusion, much evidence supports localization of function in the amygdala in cats. Although denial on the basis of direct evidence (Kling et al., 1960) cautions us against unqualified acceptance of this attractive hypothesis, apparently contradictory reports on the effects of amygdalectomy may after all be explained on the basis of differences in site and extent of amygdaloid damage. References p. 180-182

178

M. D. EGGER A N D J. P. FLY”

Fig. 9. A functional division of the amygdala of the cat consonant with many published data on the effects of electrical stimulation in the amygdala. SUMMARY

Electrical stimulation in the magnocellular portion of the basal amygdaloid nucleus, and in the anterior and medial portions of the lateral amygdaloid nucleus generally suppressed hypothalamically elicited attack behavior in cats. Stimulation in the dorsolateral portion of the posterior part of the lateral nucleus facilitated the attack behavior.

STIMULATION A N D ABLATION I N THE AMYGDALA

-

179

..

3

2

I

0

Fig. 10. In these 4 cats, the amygdaloid lesions depicted were followed by faster speeds of escape from hypothalamic stimulation. From Nakao, 1960.

Lesions in the same region of the amygdala in which stimulation generally suppressed attack produced a facilitation of the hypothalamically elicited attack response. This facilitation was observed in all 5 cats with bilateral amygdaloid lesions involving the border region between the magnocellular basal nucleus and the lateral nucleus. None of the 6 sham-lesion cats, and not one of the 10 cats whose amygdaloid lesions failed to include this circumscribed region showed the facilitation. ACKNOWLEDGEMENTS

Most of the data reported here were collected while the senior author held a United States Public Health Service postdoctoral traineeship in the Department of Psychiatry, Biological Sciences Training Program, Yale University School of Medicine. This research was supported by grants from the National Science Foundation and the United States Public Health Service. We thank Mildred Groves, Zaven Khachaturian, and Dirk van Loon for technical assistance, and Louis G. Audette for drawing the figures. References p . 180-182

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M. D. EGGER AND J. P. FLYNN

TABLE III SELECTED STUDIES OF EFFECTS OF ELECTRICAL STIMULATION IN THE AMYGDALA OF THE CAT

Experimenters

Egger and Flyp, 1963

Sites of stimulation tending to be in dorsomedial amygdala

Sites of stimulation tending to be in lateral and ventral amygdala

Suppression of hypothalamically Facilitation of hypothalamically elicited attack behavior elicited attack behavior Growling, hissing Sniffing, retching

Femandez de Molina and Hunsperger, 1959 Decreased blood pressure Gastaut, 1952; Morin et ul., Increased blood pressure 1952 Snitting, searching Hilton and Zbroiyna, 1963; Defense reaction, including prowlintz and extension of claws Zbroiyna, 1963 Inkbitionof knee jerk and of Facilitation of knee jerk and of Kaada, 1951 cortically induced movements cortically induced movements Koikegami and Fuse, 1952 Increased amplitude of respira- Decreased amplitude of respiration tion Inhibition of gastrointestinal Rise in body temperature Koikegami et al., 1952 motility Kreindler and Steriade, 1964 Acceleration-desynchronization Synchronizationof neocortical electrical activity in the form of neocortical electrical activity of spindles and slow waves Respiratory inhibition M a c h and Delgado, 1953 Respiratory acceleration Mastication Magnus and Lamers, 1956 Growling Effects other than arrest of Arrest of eating and mousing, Norris, Jr., 1963 activity or arousal arousal Cowering, snitling, licking Undirected rage Shealy and Peele, 1957 Decreased corticosteroid levels Increased corticosteroid levels Slusher and Hyde, 1961 in adrenal vein in adrenal vein Cowering, flight, searching Growling and hissing Ursin and Kaada, 1960 Respiratory acceleration, Respiratory inhibition, searching Wood, 1958 growling and biting Sneezing, seeking Yoshida, 1963 bge Facilitation of firing in short Inhibition of firing in short Zbroiyna, 1963 ciliary nerve ciliary nerve, with dilation of sympathectomized pupil

REFERENCES BAN,T., AND O m , F., (1959); Experimental studies on the fiber connections of the amygdaloid nuclei in the rabbit. J. comp. Neurol., 113,245-279. BARD, P., AND MOUNTCASTLE, V. B., (1948); Some forebrain mcchanisms involved in expression of rage with special reference to suppression of angry behavior. Res. Publ. Ass. nerv. rnent. Dis., 27, 362404. BRADY, J. V., SCHREINER, L., GELLER, I., AND KLING,A., (1954); Subcorticalmechanisms in emotional behavior: the effect of rhinencephalic injury upon the acquisition and retention of a conditioned avoidance response in cats. J. comp. physiol. Psychol., 47, 179-186. BmS, J., PETR,~~, M., AND ZACHAR, J., (1962); Electrophysiological Methods in Biological Research. New York, Academic Press. EGGER, M. D., (1962); Some effects of amygdaloid stimulation and ablation on hypothalamically elicited attack behavior in cats. Diss., Doctor of Philosophy. Graduate School, Yale University, New Haven, Conn. EGG=, M.D., (1965); Effects of amygdaloid stimulation on hypothalamic units. AbstractsofPapers, XXZZZ Znt. Congr. Physiol. Sci., Tokyo, p. 429.

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EGGER, M. D., AND FLYNN,J. P., (1963); Effects of electrical stimulation of the amygdala on hypothalamically elicited attack behavior in cats. J. Neurophysiol., 26, 705-720. FERNANDEZ DE MOLINA, A., AND HUNSPERGER, R. W., (1959); Central representation of affective reactions in forebrain and brain stem: electrical stimulation of amygdala, stria terminalis, and adjacent structures. J. Physiol. (Lond.), 145, 251-265. FERNANDEZ DE MOLINA, A., AND HUNSPERGER, R. W., (1962); Organization of the subcortical system governing defense and flight reactions in the cat. J. Physiol. (Lond.), 160, 200-213. FONBERG, E., (1967); The role of amygdaloid nucleus in animal behaviour. Progr. Brain Res., Vol. 22 (in pres). FONFIERG, E., AND DELGADO, J. M. R., (1961); Avoidance and alimentary reactions during amygdala stimulation. J. Neuroph.vsiol., 24, 651-664. Fox, C. A., (1940); Certain basal telencephalic centers in the cat. J. comp. Neurol., 72, 1-62. Fox,C. A., (1943); The stria terminalis, longitudinal association bundle and precommissural fomix fibers in the cat. f.comp. Neurol., 79, 277-295. GASTAUT, H., (1952); Corrdlations entre le systeme nerveux vdgdtatifet le systbme de la vie de relation dans le rhinenckphale. J. Physiol. (Paris), 44,431470. GLOOR, P., (1960); Amygdala (Chapt. 48). Hancibook of Physiology, Neurophysiology. J. Field, H. W. Magoun and V. E. Hill, Editors. 11, pp. 1395-1420. GREEN,J. D., CLEMENTE, C. D., AND DEGROOT, J., (1957); Rhinencephalic lesions and behavior in cats. An analysis of the Kliiver-Bucy syndrome with particular reference to normal and abnormal sexual behavior. J. comp. Neurol., 108, 505-545. HALL,E. A., (1963); Efferent connections of the basal and lateral nuclei of the amygdala in the cat. Amer. J. Anat., 113, 139-151. HILTON,S. M., AND ZBROZYNA, A. W., (1963); Amygdaloid region for defense reactions and its efferent pathway to the brain stem. J. Physiol. (Lond.), 165, 16CL173. HORVATH, F. E., (1963); Effects of basolateral arnygdalectomy on three types of avoidance behavior in cats. J. conip. physiol. Psychol., 56, 380-389. KAADA,B. R., (1951); Somato-motor, autonomic and electrocorticographic responses to electrical stimulation of ‘rhinencephalic’ and other structures in primates, cat and dog. Acta physiol. scand., SUPPI.83,24, 1-285. KAADA,B. R., A~DERSEN, P., AND JANSEN, JR., J., (1954); Stimulation of the amygdaloid nuclear complex in unanesthetized cats. Neurology, 4, 48-64. KLING,A., AND HUTT,P. J., (1958); Effect of hypothalamic lesions on the amygdala syndrome in the cat. Arch. Neurol. Psychiat. (Chic.), 79, 511-517. KLING,A., ORBACH, J., SCHWARTZ, N. B., AND TOWNE, J. C., (1960); Injury to the limbic system and associated structures in cats. Arch. gen. Psychiot., 3, 391-420. KOIKEGAMI, H., (1963); Amygdala and other related limbic structures; experimental studies on the anatomy and function. I. Anatomical researches with some neurophysiological observations. Acta med. biol. (Niigata), 10, 161-277. KOIKEGAMI, H., (1964); Amygdala and other related limbic structures; experimextal studies on the anatomy and function. 11. Functional experiments. Acta med. biol. (Niigata), 12, 73-266. KOIKEGAMI, H., DODO,T., MOCHIDA, Y., AND TAKAHASHI, H., (1957); Stimulation experiments on the amygdaloid nuclear complex and related structures. Effects upon the renal volume, urinary secretion, movements of the urinary bladder, blood pressure and respiratory movements. Folia psychiat. neurol. jap., 11, 157-206. KOIKEGAMI, H., AND FUSE, S., (1952); Studies on the functions and fiber connections of the amygdaloid nuclei and periamygdaloid cortex. Experiments on the respiratory movements (2). Folia psjchiat. neurol. jap., 6, 96103. KOIKEGAMI, H., KUSHIRO, H., AND KIMOTO, A., (1952); Studies on the functions and fiber connections of the amygdaloid nuclei and periamygdaloid cortex. Experiments on gastrointestinal motility and body temperature in cat. Folia psyehiut. neurol. jup., 6, 76-93. KREMDLER, A., AND STERIADE, M., (1963); Functional differentiation within the amygdaloid complex inferred from peculiarities of epileptic afterdischarges. Electroenceph. clin. Neurophysiol., 15, 811-826. KREINDLER, A., AND STERIADE, M., (1964); EEG patterns of arousal and sleep induced by stimulating various amygdaloid levels in the cat. Arch. ital. Biol.. 102, 576586. LEMSON, P. K., AND FLY”, J. P., (1965); The objects attacked by cats during stirnulation of the hypothalamus. Anim. Behav., 13,217-220. MACCHI, G., CARRERAS, M., AZZALI,G., DALLA ROSA,V., AND LECHI,A., (1963); Rinencefalo e com-

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portamento istintivo: Effetti provocati da lesioni operate sul lob0 piriforme. (Studio sperimentale nel gatto). Boll. Soc. ital. Biol. sper., 39, 37-41. MACLEAN, P. D., AND DELGADO J. M. R., (1953);Electricalandchemical stimulation offrontotemporal portion of limbic system in the waking animal. Electroenceph. clin. Neurophysiol., 5, 91-100. MAGNUS, O., AND LAMME=, H. J., (1956); The amygdaloid-nuclear complex. Folia psychiat. neerl., 59, 555-582. MORGANE,P. J., AND KOSMAN, A. J., (1959); A rhinencephalic feeding center in the cat. Amer. J . Physiol., 197, 158-162. M o m , G.,NAQUET,R., AND BADJER,M., (1952); Stimulation electrique de la region amygdalienneet pression artbrielle chez le Chat. J. Physiol. (Paris), 44, 303-305. NAKAO,H., (1960); Hypothalamic emotional reactivity after amygdaloid lesions in cats. Folia psychiat. neurol.jap., 14,357-366. NORRIS, JR., F. H., (1963); Arrest of activity by temporal lobe stimulation. Neurology, 13, 895-898. SAWA,M., MARUYAMA, N., HANAI, T., AND UI, S., (1959); Regulatory influence of amygdaloid nuclei upon the unitary activity in ventromedial nucleus of hypothalamus. Folia psychiat. neurol. jap., 13, 235-256. SCHREINER, L.,AND KLING,A., (1953); Behavioral changes following rhinencephalic injury in cat. J. Neurophysiol., 16, 643-659. SHEALY,C. N., AND PEELE, T. L., (1957); Studies on amygdaloid nucleus of cat. J . Neurophysiol., 20, 125-1 39. SLUSHER, M. A., AND HYDE,J. E., (1961); Effect of limbic stimulation on release of corticosteroids into the adrenal venous effluent of the cat. Endocrinology, 69, 1080-1084. SPIEGEL, E. A., MILLER, H. R., AND OPPENHEIMER, M. J., (1940); Forebrain and rage reactions. J. Neurophysiol., 3, 538-548. STUART,D. G., PORTER, R. W., AND ADEY,W. R., (1964); Hypothalamic unit activity. 11. Central and peripheral influences. Electroenceph. d i n . Neurophysiol., 16, 248-258. Sumwxs, T. B., AND KAELBER, W. W., (1962); Amygdalectomy: effects in cats and a survey of its present status. Amer. J. Physiol., 203, 1117-1119. SZENTAGOTHAI, J., FLERK~, B., Mess, B., AND HALASZ,B., (1962); Hypothlumic Control of the Anterior Pituitary. Budapest, Akadkmiai Kiad6. TSUBOKAWA, T., AND S m , J., (1963); Mesencephalicinfluence upon the hypothalamic ventromedial nucleus. Electroenceph. clin. Neurophysiol., 15, 804-810. URSIN,H., (1965a); The effect of amygdaloid lesions on flight and defense behavior in cats. Exp. Neurol., 11, 61-79. URSIN,H., (1965b); Effect of amygdaloid lesions on avoidance behavior and visual discrimination in cats. Exp. Neurol., 11,298-317. URSIN,H., AND KAADA, B. R., (1960); Functional localization within the amygdaloid complex in the cat. Electroenceph. din. Neurophysiol., 12, 1-20. VALVERDE, F., (1963); Amygdaloid projection field. The Rhinencephalon and Related Structures, Vol. 3, Progress in Brain Research. W. Bargmann and J. P. Schadk, Editors. Amsterdam, Elsevier, pp. 20-30. WASMAN, M., AM) FLY", J. P., (1962); Directed attack elicited from hypothalamus. Arch. Neurol., 6, 220-227. WENDT,R. H., AND ADEY,W. R., (1960); A study of evoked unit activity in the hypothalamus. Anat. Rec., 136, 301 (abstract). WOOD,C. D., (1958); Behavioral changes following discrete lesions of temporal lobe structures. Neurology, 8, 215220. WOOD,C. D., Scxo'rm~ms,B., FROST,L. L., AND BALDWIN,M., (1958); Localization within the amygdaloid complex of anesthetized animals. Neurology, 8,477480. YOSHIDA,M., (1963); Effects of amygdaloid stimulation on emotional responses produced by hypothalamic stimulation in cats. Psychiat. neurol. jap., 65, 863-879 (English summary, pp. 71-72). ZBROZYNA, A. W., (1963); The anatomical basis of the patterns of autonomic and behavioural response effected via the amygdala. The Rhinencephalon and Related Strucrttres, Vol. 3, Progress in Bruin Research. W. Bargmann and J. P. Schadk, Editors. Amsterdam Elsevier, pp. 5 M 8 .

183

Influence of Labyrinthine Stimulation on Hippocampal Activity A. COSTIN, F. B E R G M A N N

AND

M. CHAIMOVITZ

Department of Pharmacology, Hebrew University, Hadassah Medical School, Jerusalem (Israel)

We have shown previously that labyrinthine stimulation has a direction-dependent influence on optic nystagmus. E.g. when both labyrinthine and optic nystagmus are directed towards the same side, they enhance each other, while antagonistic stimulations produce mutual depression (Bergmann and Costin, 1966). Similarly, sideposition - a vestibular stimulus that does not evoke nystagmus - enhances ipsiversive optic nystagmus and inhibits contraversive eye movements (Costin et d.,1966). These phenomena can be satisfactorily explained by the non-symmetrical distribution of the fibers in the photic and vestibular pathways. Th&observations mentioned suggest the possibility that labyrinthine stimulation may also exert an asymmetric influence on other responses induced by unilateral stimulation of a cerebral structure. The present study shows that under certain conditions labyrinthine nystagmus to left or right has a different influence on the afterdischarge following unilateral electrical stimulation of the dorsal hippocampus. METHODS

Adult rabbits of either sex, weighing between 2 and 3.5 kg, were used. All surgical procedures were carried out under ether. At the end of the operation the wounds were infiltrated with 2 % xylocain, and the animal was allowed to recover from general anesthesia, before the experiment was started. Three pairs of bipolar, concentric electrodeswere placed in the dorsal hippocampus, at positions bC-bD (Monnier and Gangloff, 1961). One pair served for stimulation and the other two for recording of the left and right hippocampal potentials on a Schwarzer electroencephalograph. At the end of the experiment the animal was killed, and the position of the electrodes was checked by histological examination. For angular acceleration and deceleration, the rabbit was placed on a Toennies rotating chair. After a predetermined angular velocity had been attained, the rotation was maintained at constant speed for 60 sec before deceleration began. In order to avoid interference by photic stimuli, both eyes were kept covered with small black hoods throughout these experiments. References p . 188

184

A. COSTIN

et al.

a

b

C

t

I

1

5

10

15

20

25

200pv

30

35 sec

Fig. 1. Influence of perrotatory nystagmus on the discharges of the resting hippocampus. Male rabbit, 2 kg. Counterclockwise acceleration at 10"/sec2(between arrows) evokes 14 eye beats to left (see nystagmogram in c). After 10 sec, when an angular velocity of lOO"/sechad been reached, the rotating chair was maintained at this speed. After-nystagmus 9 beats during 12 sec. Note that concomitantly with the nystagmus, a 0-rhythm appears in both records, (a) from the right, and (b) from the left dorsal hippocampus.

a

b

C

I

t 1

I

I

I

I

I

200pv I

I

5 10 15 20 25 30 35 sec Fig. 2. Influence of sudden arrest of rotation on the potentials of the resting hippocampus. The same rabbit as used for Fig. 1. After the animal had been rotated counterclockwise for 1 min at a constant speed of lOO"/sec,it was suddenly arrested (at arrow) causing 26 eye beats to the right during 11 sec (c). The nystagmus caused fast, synchronized potentials in both hippocampi, outlasting the eye movements for about 20 sec (a and b).

L A B Y R I N T H I N E S T I M U L A T I O N OF H I P P O C A M P U S

185

Fig. 3. Asymmetric effect of angular accelerationon hippocampal after-discharge (HAD). Male rabbit, 2.6 kg. Stimulation of left dorsal hippocampus at bC, for 5 sec (arrows); 40 c/sec, pulse duration 2 msec, current strength 0.15 mA. Recording electrodes in left hippocampus at bD. (a) Control: hippocampal stimulation, without rotation, produces an HAD of 30 sec duration; (b) simultaneous counterclockwiseacceleration at 10"/sec2,fcr 10 sec; reduction of HAD to 3 sec; (c) simultaneous clockwise acceleration at 10"/sec2,for 10 sec; prolongation of HAD to 60 sec. RESULTS

I . Effect of labyrinthine stimulation on the discharges of the resting hippocampus In the awake, non-stimulated animal, angular acceleration evoked a hippocampal 8rhythm, expressing itself in synchronization and in an increase in amplitude. In Fig. 1, counterclockwise acceleration at lO"/secZ produced during 10 sec 14 eye beats to the left; the changes in the hippocampogram appeared simultaneously with the first eye movement. When after 10 sec a velocity of 100"/sec was maintained, perrotatory nystagmus persisted for another 7 sec, but with decreasing strength. At the same time, the hippocampal potentials decayed and returned to their resting pattern. Analogous observations were made during deceleration. The duration of the hippocampal aftereffect clearly depends on the strength of the vestibular stimulus. Thus in Fig. 2, after sudden arrest of the rotating chair the synchronization outlasted the postrotatory nystagmus considerably. The changes in the left and right hippocampus parallel each other, and either side may be used to measure the effect of clockwise or counterclockwise rotation. II. Effect of labyrinthine stimulation on the after-discharge, following hippocampal excitation In Fig. 3a, electrical stimulation of the left hippocampus for 5 sec produced an afterdischarge for 30 sec. Counterclockwise acceleration at 1O"/secz reduced this period to 3 sec (Fig. 3b), while clockwise acceleration prolonged it to 60 sec (Fig. 3c). The directional effect of perrotatory nystagmus could be demonstrated only with difficulty in References p . I88

1616

A. COSTIN

et ai.

experiments in which the hippocampal after-discharge (HAD) was short (see Table IA), but if the latter lasted for more than 30 sec, the inhibitory effect of labyrinthine nistagmus became manifest. On the other hand, the enhancement was sometimesweak or absent, even when the HAD extended over a considerable period. The data in the table establish the rule that vestibular nystagmus to the left inhibits the HAD evoked from the left hippocampus. It was observed previously (Gangloff and Monnier, 1957; Costin et al., 1963) that the HAD can be considerably prolonged by application of chlorpromazine. This drug TABLE I EFFECT OF LABYRINTHINE STIMULATION O N HIPPOCAMPAL AFTER-DISCHARGE

(HAD)

Duration of HAD (see) Side of hippocampal stimulation

( A ) Untreated animals : Left Right Right Left Left Left Left Left Right Right Right

Control

Clockwise acceleration

Counterclockwise acceleration

13 28 25

11 28 18

15 24 23

21 29

30 40 50 115

75 35 70 130 60 13 14 5

7 16 15 20 5 55 50 110

12 80 20 100

6 13 22 140

12 90 22 40

70 117

(B) Before and after treatment with 2 mg/kg chlorpromazine i.v. : Right (before) Right (after) Left (before) Left (after)

TABLE 11 LOSS OF ASYMMETRIC INHIBITORY INFLUENCE O N

HAD

W I T H INCREASING ANGULAR

ACCELERATION

Female rabbit, 3 kg; stimulation of left dorsal hippocampus at bC for 5 sec;40 c/sec, 2 msec, 1.5 mA. Angular accelerations lasted for 10 sec. Direction of rotation

Control Counterclockwise clockwise Counterclockwise Clockwise

Angular acceleration

Duration of HAD (see) 130 20 130 4 7

187

L A B Y R I N T H I N E S T I M U L A T I O N OF H I P P O C A M P U S

TABLE I11 LONG-LASTING INHIBITORY EFFECT OF LABYRINTHINE STIMULATON ON HIPPOCAMPAL AFTER-DISCHARGE

(HAD)

Male rabbit, 2.8 kg; stimulation of left dorsal hippocampus for 5 sec at 40 c/sec, 2 msec, 0.3 mA. Counterclockwiseacceleration at 20"/sec2,for 10 sec. ~

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65

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permitted the demonstration of the asymmetric effect of vestibular nystagmus on the HAD even in animals that before treatment were not affected by angular acceleration (Table IB). The asymmetric effects of Fig. 3 can be demonstrated only when the intensity of labyrinthine stimulation is not too high. Angular accelerations above 20"/sec2 usually produced inhibition of the HAD, irrespective of the direction of rotation (Table 11). Although after a short while the hippocampal potentials reassume their resting appearance, the after-effect of labyrinthine stimulation has not passed. If hippocampal stimulation is repeated without rotation, the inhibitory effect remains manifest for about 30 min (see Table 111). On the other hand, the enhancement of the HAD, represented in Fig. 3c, vanishes shortly after the end of the rotation. DISCUSSION

At first glance, no relation is apparent between vestibular nystagmus and HAD. However, Spiegel(l932) has shown that labyrinthine stimulation facilitates cortical seizures. The present findings on hippocampal discharges thus enlarge the observations of this author. The experiments described here establish a pronounced asymmetric effect on the HAD, following labyrinthine stimulation under certain conditions. The duration of after-discharges, evoked by stimulation of the dorsal hippocampus, is reduced by ipsiversive nystagmus. On the other hand, the effect of contraversive nystagmus movements is not as clear-cut, since the HAD sometimes remains unchanged while in others it is markedly prolonged (see Fig. 3c and Table I). It is most conspicuous that a more intense vestibular stimulus, such as increased angular acceleration,may produce inhibition even when hippocampal excitation is combined with contraversive nystagmus (see Table 11). These observations, together with the identical effect of clockwise and counterclockwise rotation on the discharges of the resting hippocampus (Figs. 1 and 2), suggest that each vestibular pathway has connections to both hippocampi, forming both References p . 188

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et al.

excitatory and inhibitory synapses. However, the quantitative distribution of the two types of synapses to either side appears to be unequal. In addition, the effect of vestibular impulses, arriving at the hippocampus, also depends on the state of activity of the latter structure. Electrical excitation of the hippocampus produces a well-organized pattern of discharges. The stimulated region probably serves as pacemaker for the hippocampal neurons on both sides (Gutman et al., 1963). This means that the activity spreading from the pacemaker suppresses all independent, local discharges. It appears possible that this mechanism also paves the way for the inhibitory effect of labyrinthine nystagmus on the HAD. REFERENCES BERGMANN, F., AND COSTIN,A., (1965); Studies on the physiological relationship between retina and labyrinth. Israel J. Med. Sci., 1, 13661372. COSTIN, A., BERGMANN, F., AND CHAIMOvITz, M., (1966); Influence of side position of the head on central and flash nystagmus in the rabbit. Acta oto-laryng., 61, 323-331. Cam, A., GUTMAN,J., AND BERGMANN, F., (1963); Relationship between caudate nucleus and dorsal hippocampus in the rabbit. Electroenceph. elin. Neurophysiol., 15, 997-1005. GANGLOFF, H., AND MONNIER, M., (1957); Topic action of reserpine, serotonin and chlorpromazine on the unanesthetized rabbit's brain. Helv. physiol. pharmacol. Ada, 15,83-104. GUTMAN, J., a m ,A., AND BERGMANN, F., (1963); Constancy of hippocampal afterdischargeunder various conditions of stimulation. Electroenceph. clin. Neurophysiol., 15, 989-996. MONNIER, M., AND GANGLOFF,H., (1961); Rabbit Brain Research, Vol. 1. Elsevier, Amsterdam. SPIEGEL, E. A., (1932); Rindenemgung (Auslosung epileptiformer Anfalle) durch Labyrinthreizung. Versuch einer Lokalization der corticalen Labyrinthzentren. Z. ges. Neurol. Psychiat., 138,178-196.

189

Studies on the Neurovegetative and Behavioral Functions of the Brain Septa1 Area MIGUEL R. COVIAN School of Medicine, Department ofphysiology, Aibeirrio Prgto, S.P. (Brazil)

The septal area is a small structure of the limbic system almost completely surrounded by cerebrospinal fluid, rostra1 and anterior to the anterior commissure, ventral to the splenium of the corpus callosum, related to autonomic, reproductive, emotional and other behavioral responses. We have studied in anesthetized cats, rats and rabbits, and in unanesthetized cats, the effects of electrical septal stimulation upon blood pressure and respiration (Part I). In anesthetized rabbits a conditioned blood pressure response was obtained (Part 11). In rats the role of the septal area on drinking behavior for water and NaC1, using the standard ‘two bottle’ self-selection procedure, was investigated (Part 111). Part I was done in collaboration with J. Antunes-Rodrigues, E. M. Krieger, J. J. O’Flaherty and C. Tim0 Iaria; Part I1 with M. C. Lico; and Part I11 with C. G. Gentil and A. Negro Vilar. (I) EFEECTS OF S T I M U L A T I O N OF T H E S E P T A L A R E A U P O N B L O O D P R E S S U R E

A N D RESPIRATION

The experimental data available on the central nervous control of cardiovascular activity, as well other bodily activities, show that this control is exerted at different levels of the neuroaxis. Since the pioneer work of Karplus and Kreidl (1909,1910) the hypothalamus has been considered one of the most important structures controlling neurovegetative functions. Changes in blood pressure and respiration following the electrical stimulation of several areas of the cerebral cortex in animals and man have also been reported; Delgado (1960) has published a good review on this subject. During recent decades, as a result of investigations carried out in several laboratories, increasing emphasis is being placed on the role played by the limbic system in the regulation of visceral functions and behavior. The first part of our presentation will deal with the blood pressure and respiratory changes elicited in anesthetized and unanesthetized animals by electrical septal stimulation. Anesthetized animals. Cats and rats were anesthetized with a-chloralose, 70 mg/kg i.v. Rabbits received urethane, 1.25 mg/kg given in half doses i.p. and i.v. respectively. Conventional stereotactictechniques were used to introduce bipolar electrodesinto the References p . 215-217

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septal area according to the co-ordinates of Jasper and Ajmone-Marsan (1954) for cat, of De Groot (1959) for rat, and of Sawycr et al. (1954) for rabbit. An AEL stimulator with an isolation unit was used to deliver unidirectional rectangular pulses of 50 c/sec, 10 msec of pulse duration and an intensity of 3-5 V. Systemic arterial pressure was measured with a Statham P23-AA transducer connected to a catheter placed in a femoral artery. Respiration was recorded through a pneumograph attached around the chest. Changes in blood pressure and respiration were recorded synchronously on a polygraph. A tracheal cannula was inserted into each animal, and artificial respiration was used when required. Body temperature was maintained by radiant heat and measured with a telethermometer with a probe introduced in the colon. At the end of each experiment the head was perfused with 4% formaldehyde, and the brain submitted to routine histological procedures to determine the position of the stimulating electrodes.

Results The effects produced by septal stimulation on blood pressure and respiration in the cat have been reported elsewhere (Covian et al., 1963);they will be brieffly summarized and some new unpublished results will be described in more detail. In 21 of 25 intact cats a fall in blood pressure was elicited that outlasted the electrical stimulation, applied during 30 sec, by about 3-5 min. When a pure depressor effect was elicited it began gradually during the period of stimulation, and was accentuated after the end of the stimulus; however, when respiratory changes were present the latency of the response was shorter. Occasionally both changes were concomitant but either effect could appear independently of the other. The fall in blood ptessure was sometimes preceded by a slight pressor reaction, and in some cats the depressor effect was only observed after the end of stimulation. Fig. 1 shows the depressor reactions obtained in one cat, without any significant change in either the heart rate or the frequency or depth of respiration. The blood pressure measurements made before, during, and after the stimulation were 162/122, 138/98, and 95/64 mm Hg respectively; the fall in pressure lasted for 5 min after stimulation was stopped : in the last period there was a diminution in pulse pressure due to the greater fall of the systolic pressure. Fig. 2

Fig. 1. Depressor reaction in the chloralosed cat. Blood pressure (lower record) values before, during and after electricalstimulation (solid line) of the septal area: 162/122,138/98 and 95/64 mm Hg. The fall lasted for 5 min after cessation of stimulation. No change in respiration (upper record) or heart rate.

NEUROVEGETATIVE A N D BEHAVIORAL FUNCTIONS OF THE SEPTAL

AREA^^^

Fig. 2. Depressor reaction in the chloralosed cat. Blood pressure (upper record) values before, during and after electrical stimulation (solid line) of septal area: 142/98,117/72and 75/30 mm Hg. The fall outlasted by 4 min the stimulation. Respiration (lower record) showed an expiratory apnea of 12 sec during stimulation. Heart rate dropped 12 beats/min.

Fig. 3. Depressor reaction in the chloralosed rat. Mean blood pressure (lower record) fell 45 mm Hg during and after electricalstimulation of the septal area. The fall lasted for 3 min 50 sec after cessation of stimulation.No signifcant change in respiration (upper record) occurred.

Fig. 4. Diagram showing the electrode location in rat of Fig. 3. References p. 215-21 7

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Fig. 5. Respiratory change (upper record) in chloralosed rat without any change in blood pressure (lower record) during electrical stimulation of the septal area.

Fig. 6. Diagram showing the electrode location in rat of Fig. 5.

depicts the result elicited in another cat in which the alterations in blood pressure were accompanied by mod5cations in respiration and heart rate. The arterial pressure values for the three periods mentioned above were 142198,117172 and 75/30mm Hg; the fall in pressure outlasted the stimulus by 4 min. There was an expiratory apnea during the first 12 sec of stimulation; afterwards normal frequency and amplitude of respiration were regained slowly. Bradycardia occurred during the period of stimulation and persisted during the poststimulatory pressure drop. The heart rate was 180, 160 and 168/min for the three periods mentioned above. In rats the stimulation of the septal area also elicited a blood pressure fall as is shown in Fig. 3. Immediately after the onset of the stimulation there was a small rise

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193

Fig. 7. Blood pressure (lower record) and respiration (upper record) in the chloralosed cat before, during, and after electrical stimulation (solid line) of the septa1 area. A, intact animal; B, after bilateral cervical vagotomy ;C, after atropine injection (1 mg/kg i.v.).

in blood pressure followed by a fall which persisted for 3'50" after the end of stimulation. The mean blood pressure values before and during the stimulation were 115 and 70 mm Hg respectively. The stereotaxic settings were F, 7.5; L, 1.0; H, +3. The electrode position is shown in Fig. 4. In some rats only respiratory changes were observed (Fig. 5). During stimulation an increased amplitude of respiratory movements was elicited. The electrode was placed at F, 7.5; L, 1.0; H, 1.0. In another horizontal

+

References p. 215-217

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M. R. C O V I A N

f

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plane (+2) the respiratory changes were accompanied by a fall in blood pressure. The electrode position is shown in Fig. 6. Midcervical bilateral vagotomy did not abolish the blood pressure fall in cats and rats. In Fig. 7 this is shown for the cat. The effect of septal stimulation was not blocked by atropine in either animal, as shown in Fig. 8 for the rat. To rule out any possible interference of muscular contraction on the depressor reaction, flaxedil was injected in some cats: after flaxedil the blood piessure fall still remained, as can be observed in Fig. 9. The possibility of explaining the effects observed by a substance released by septal stimulation was tested by the crossed circulation technique, and by plasma injection from the stimulated animal into an intact rat. These procedures failed to prove the possibility suggested (Covian et al., 1964). Bradycardia was a concomitant feature very often observed in cats and rats under

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195

Fig. 9. A, blood pressure (lower record) fail following septal stimulation in the chloralosed cat. B,

after flaxedil administration.

chloralose. In Fig. 10 this result is shown for the cat: in the upper record the values of the systolic and diastolic blood pressure are measured every 20 sec, and in the lower record the beats per minute are plotted. It is observed that during stimulation there was a diminution of 15 beats/min. When the stimulus ceased the diminution was accentuated, the rate falling to 40 beats/min. The pre-stimulatory rate was regained 5 min after the withdrawal of the stimulus. Eserine (100 ,ug/kg i.v.) enhanced the fall of the heart beat and its duration. Nevertheless in some cats the bradycardia still remained after vagal sections. This observation suggests the possibility of an inhibition of the cardiocelerator fibers playing a role in the bradycardia due to septal stimulation. Baroreceptor reflex. The interplay between septal stimulation and baroreceptor reflex was studied in the cat. The procedure was as follows: the baroreceptor reflex was obtained by bilateral carotid occlusion, and tested at different intervals during the hypotension elicited by septal stimulation. Fig. 11 shows at A: the control baroreceptor reflex, at B, during the last 5 sec of septal stimulation, at C , 30 sec, at D, 1 min, at E, 1.5 min, at F, 2 min and at G , 3 min after the stimulus withdrawal. In this sequence the diminution of the baroreceptor reflex during hypotension due t J septal stimulation is considerable. References p . 215-21 7

196

M. R. COV IA N

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Fig. 10. Effect of septal stimulation on blood pressure (A) and heart rate (B) in the chloralosed cat.

In (A), upper circles, systolic blood pressure; lower circles, diastolic blood pressure (both measured every 20 sec, as well as heart rate). Solid line, stimulation time 3 min.

Fig. 11. Interaction between septal stimulation and baroreceptor reflex in the chloralosed cat. A, central baroreceptor reflex elicited by bilateral carotid occlusion; B, during the last 5 sec of septal stimulation; C,30 sec after stimulus withdrawal; D, E, F, G, 1 min, 1.5 min, 2 min, and 3 min, respectively after stimulus withdrawal.

Fig. 12. Increased blood pressure (lower record) during stimulation of the septal area without change in respiration (upper record) in the chloraiosed cat. The values were 104p8, 142/90, and 106/60 before, during and after stimulation.

N E U R O V E G E T A T I V E A N D B E H A V I O R A L FUNCTIONS OF THE S E P T A L A R E A

197

Fig. 13. Microphotographsto illustrateelectrode locations in cats that showed depressor responses to septal stimulation. A, anterior septal area; B, anterior septal area, electrode more medially placed; C, median septal area; D, posterior septal area.

Hypertensive efects. In a few cats and rats a pressor reaction was also elicited. Contrary to the depressor effect, rises in blood pressure occurred very quickly after the onset of stimulation, and they never outlasted the period of stimulation. Fig. 12 is the record obtained in one cat in which the systolic blood pressure increased by 38 mm Hg and the diastolic by 32 mm Hg during stimulation. Both in cats and rats the hypertensive effect was inhibited by dibenamine. Respiratory effects. In both species changes in the amplitude and/or frequency of respiratory movements as well as apnea, either in inspiration or expiration, were obtained during stimulation. They were concomitant sometimes with blood pressure changes, but either effect could appear independently of the other. The location oj’sites stimulated. The histological examination of the septal area of all cats and rats showed that the effects were independent of electrode position, for depressor effects were elicited from the anterior, median and posterior zones, as well as from the medial and lateral parts of the septum. Nevertheless, some points were more ‘respiratory’ than ‘depressor’, and in some animals the moving of the electrode 1 mm down or laterally changed the type of response. In one rat and in one cat a fall in blood pressure was changed to a rise when the electrode was more medially placed. Because of these findings a mapping study of the septal area in the cat is under way. Fig. 13 illustrates the electrode placement in the brains of some cats. Rabbits. In rabbits anesthetized with urethane, stimulation of the septal area also resulted in a fall of blood pressure. This depressor reaction was sometimes very persistent as is shown in Fig. 14. Between stimulation numbers 6 and 46 about 6 h elapsed; References p . 215-21 7

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Fig. 15. Effect of stimulationtime upon blood pressure fall due to septal stimulation in the chloralosed cat. G , 23 min; H, 28.5 min. Upper circles, systolic blood pressure; lower circles, diastolic blood pressure (both measured every 20 sec). Voltage, frequency and pulse length were constant.

note the constancy of the response obtained with no change in the position of the electrode or the parameters of stimulation. It is relevant that in cats we have shown (Covian and Timo-Iaria, 1966), that the depressor reaction is maintained during a stimulation time prolonged to 28.5 min as illustrated in Fig. 15. In rabbits also, as in cats and rats, a pressor reaction was obtained once in a while, as shown in Fig. 16. When the electrode was moved 1 mm the depressor reaction was changed to a pressor one. Unanesthetized cuts. In cats with bipolar electrodes implanted in the septal area, and a permanent polyethylene cannula in the abdominal aorta, the results of electrical septal stimulation were studied. As happened in anesthetized cats, a drop in blood pressure was observed together with bradycardia (Fig. 17). After withdrawal of the References p. 215-21 7

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M. R. COVIAN

Fig. 16. Effects of stimulation of the septal area upon blood pressure (lowerrecord) in the rabbit under urethane anesthesia. A, pressor reaction; B, depressor reaction. Both stimulations were made within a short interval. Parameters of stimulation were the same.

Fig. 17. Blood pressure fall in the mesthetized cat due to septal stimulation. Bradycardia was also elicited. Stimulation time, 30 sec.

N E U R O V E G E T A T I V E A N D B E H A V I O R A L F U N C T I O N S OF THE S E P T A L

AREA201

Fig. 18. Depressor reaction in the manesthetized cat. A, stimulation time, 2 sec;B, 4 sec.

stimulus the fall as well as the bradycardia persisted and both regained the prestimulatory values after 3 min had elapsed. Brief stimulation times (2 4,s and 16 sec) also elicited a fall in blood pressure as shown in Figs. 18 and 19. Bradycardia, without significant changes in blood pressure, was elicited with a lower stimulus intensity, and it was abolished by atropine (Fig. 10). Comments

The effects of electrical stimulation of the septal area upon blood pressure and respiration in the curarized cat (Torii, 1961), in the chloralosed cat (Covian et al., 1964; Manning et al., 1963) and in chloralosed and conscious dogs (Gorten et al., 1964) have recently been reported. The septal area is one of the anatomical components of the limbic system, the phylogenetically oldest structure of the cerebral hemispheres and a common denominator in the brain of all mammals, as was pointed out by Broca. MacLean (1949,1958,1959) has made it clear that man shares with other mammals the physiological properties of this system. Changes in blood pressure and respiration evoked by septal stimulation have been reported before by several investigators, but the depressor reactions were not subReferences p . 215-21 7

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M. R. COVIAN

Fig. 19. Depressor reaction in the unanesthetized cat. C, stimulation time 8 sec; D, stimulation time 16 sec.

Fig. 20. A, bradycardia elicited by septal stimulationin the unanesthetized cat. B, bradycardia abolished by atropine (1 mg/kg i.v.).

N E U R O V E G E T A T I V E A N D B E H A V I O R A L F U N C T I O N S OF T H E S E P T A L A R E A

203

jected to any detailed analysis (Hess, 1958; Kabat et al., 1935; Kabat, 1936; Kurotsu et al., 1958; Ranson et al., 1935). MacLean et al., (1960) elicited penile erection (a localized circulatory effect) by electrical stimulation of the septal area in the unanesthetized squirrel monkey. The fact that in our expsriments the same reaction has been obtained in three different species of animals pleads for its physiological importance in the regulation of the neurovegetative functions studied. The most striking feature observed in the experiments described was the consistent, marked and long-lasting fall in blood pressure [this last peculiarity was absent in the rabbit] elicited during stimulation but always accentuated or clearly seen after the stimulus was withdrawn. This reaction did not parallel the changes in respiration or the bradycardia which also occurred frequently. The depressor effect could be the result of either an inhibition of vasoconstrictor fibers (directly and/or indirectly through the vasomotor center) or of a facilitation or stimulation of vasodilator fibers or both. Vagotomy and atropine did not interfere with the fall in blood pressure. Further, Manning et ~ l(1963) . have shown, in the chloralosed cat, that the dexease in contractile force and heart rate induced by septal stimulation was abolished by ruling out the cardioaccelerator sympathetic fibers (stellate ganglionectomy, blocking agents), but the fall in blood pressure was unaffected. This observation points to the peripheral origin of the depressor reaction, that is, a decrease in sympathetic tone to the blood vessels. The assumption that septal impulses could affect the discharge of the medullary vasomotor center is based on the observation that stimulation of a restricted hypothalamic zone, just behind and below the anterior commissure about 2 mm laterally to the midline, resulted in drastic inhibition of sympathetic vasoconstrictor tone. It was suggested by Folkow et al. (1959) that this area constitutes a hypothalamic relay station for cortical inhibitory pathways to lower structures, mainly affecting the discharge of the medullary vasomotor center. The possibility of septal impulses acting through this area is supported by the known connections of the septum with the hypothalamus either directly by way of the medial forebrain bundle or indirectly through the amygdala and fornix. It is also important to recall the connection between the septum and the mesencephalic reticular formation (Nauta, 1956). The interplay between septal stimulation and the baroreceptor reflex studied in the cat also favors the idea of an action of the septal area on the vasomotor center. The blockage of the baroreceptor reflex by the hypotension due to septal stimulation cannot be ascribed to the blood pressure fall, because such a reflex is extremely resistant even when the hypotension is very marked. The suggestion is that impulses originating in the septal area at least partially inhibit the bulbar vasomotor center, thus blocking the action of the reflex mechanisms of the baroreceptor nerves. This idea was already advanced (Candia et al., 1962) to explain the blood pressure fall observed during sleep. A peripheral blockage by a humoral factor can be discarded because we were unsuccessful in demonstrating the release of a humoral factor after septal stimulation (Covian et al., 1964). One argument in favor of some possible participation of vasodilator fibers is the fact that the sympathetic vasodilator tract in its intracerebral course passes through the septal region (Elliasson et d., 1952; Uvnas, 1960). References p . 215-217

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Bradycardia following septal stimulation has been a feature which very often, but not always, accompanied the fall in blood pressure. It was inhibited by atropine but in some cases it persisted after the drug injection or vagotomy thus suggesting an inhibition of efferents which produce cardiac acceleration. The long persistence of the response observed in rabbit, as well as its maintenance during a prolonged stimulation time in the cat, means that the septal area is one of those structures of the brain resistant to fatigability. Delgado (1959) has shown that continuous stimulation reveals fatigability in some areas of the brain while others are very resistant. One can argue that those brain regions resistant to fatigue integrate the physiological mechanisms which are constantly at work, such as those which regulate blood pressure. The septal area could be one of the structures that form part of these mechanisms. The hypertensive reactions observed in some animals before the hypotension (or as a pure effect) suggest the facilitation of some sympathetic efferents. Blockage by dibenamine demonstrated their vasoconstrictional origin. There would be two antagonistic systems intermingled in the septal area and acting on blood pressure: one induces hypertension and the other provokes hypotension. During electrical stimulation both systems would be excited but predominantly the hypotension circuits, shifting the dynamic balance to the side of hypotension. By the end of stimulation in our experiments, the sudden withdrawal of the less effective hypertensor system would unchain the hypotensor one, thus allowing its full effect, and explaining the accentuation of the blood pressure fall after stimulus withdrawal that we have observed. An interesting feature of the hypotension is its prolonged duration, often lasting 3-5 min after the cessation of the stimulus. This seems to be a peculiarity of the effects produced by septal stimulation, for MacLean and Ploog (1960) reported that in unanesthetized squirrel monkeys throbbing erections may wax and wane for periods up to 5 min following septal stimulation. On the other hand Folkow et al. (1959) observed the slow restitution of the blood pressure after the interruption of the stimulation, requiring 3-5 min or more. Kabat and co-workers (1935) observed that the blood pressure fall did not return to its former level until some time after cessation of stimulation in the neighborhood of the anterior commissure. This peculiarity is of uncertain explanation, but we must remember that a fall in blood pressure after septal stimulation in the cat, and simultaneously the appearance of fast waves in the hippocampus, have been reported (Torii and Kawamura, 1960); also hippocampal afterdischarge following electrical stimulation of the septal area has been described (Torii, 1961). It is possible that the depressor reaction reported here is mediated through the hippocampus, and its maintenance after the stimulus withdrawal could accounted for the hippocampal after-discharge due to reverberating circuits. A decrease in the heart rate and irregularity have been reported in kittens during sleep (Jouvet et al., 196l), as well as constant and marked pressure fall in cats during the fast cortical activity phase of sleep (Candia et al., 1962). Increased sleeping time under barbiturate after destruction of the septal area in the rat has also been observed (Harvey et al., 1964; Heller et al., 1960). On the other hand lesions ofthe septal area interfere with the rhombencephalic phase of sleep in cats (Jouvet, 1962). It has been

NEUROVEGETATIVE A N D B E H A V I O R A L F U N C T I O N S OF THE S E P T A L A R E A 2 0 5

reported that destruction of the septal area produces a heightened emotional behavior in the rat (Brady and Nauta, 1953, 1955; Tracy and Harrison, 1956>,sham rage in the cat (Spiegel et al., 1940) and increased sympathetic activity in the cat (Bond et al., 1957). Electrical and chemical stimulation of the septal area in the cat resulted in enhancement of pleasure reactions (Trembly, 1956). According to these reports, and those of self-stimulation in rats (Olds and Milner, 1954) and man (Heath, 1963), the septal region could be regarded as a ‘quieting system’. However, some authors have not found behavioral changes after septal lesions (Brady and Nauta, 1953; Harrison and Lyon, 1957). If we take into account that placidity and sleep are associated with falls in blood pressure and diminished heart rates, it can be suggested that there is a relationship of our findings with these states. (11) C O N D I T I O N I N G OF THE B L O O D P R E S S U R E F A L L D U E TO S E P T A L STIMULAT I O N I N THE A N E S T H E T I Z E D R A B B I T

The regularity and magnitude of the evoked arterial hypotension obtained by septal stimulation led us to test its conditionability. Thus, the unconditioned response elicited by stimulation of the septal area in rabbits under urethane anesthesia was used as a reinforcer of a sensorial (acoustic) stimulus. Each pairing consisted of a click train of lO/sec repetition rate applied during 35 sec, and an electrical stimulation of the septal area applied during the last 15 sec of the conditioned stimulus. The aim of this uncommon approach, such as studying an adaptive reaction during attention blockage, was to elucidate to what extent a high level of vigilance is necessary to obtain a neurovegetative conditioning when a low integrating structure such as the limbic system is activated. The results showed that approximately 10-25 associations were sufficient to obtain the initial conditioned hypotension to the acoustic stimulus. Fig. 21 illustrates the characteristics of a conditioned response ; the acquired response was progressively enhanced in successive reinforcements reaching a magnitude similar to the unconditioned one. The ECG recorded on the acoustic area of the brain shows, in a few experiments, desynchronization due to the acoustic stimulus only during the conditioning. Septa1 stimulation also caused desynchronization as was often observed. It is well to make clear that in some rabbits the unconditioned stimulus provoked a hypersynchronization. Fig. 22 depicts the results obtained in another rabbit in which the ECG did not change, either during conditioning or during septal stimulation. The conditioned hypotension presented all the peculiarities of conditioned responses, namely, external and internal inhibition, generalization, differentiation (Fig. 23) and extinction (Fig. 24). Comments

The right level of anesthesia for obtaining the conditioned response was one of the hard problems we faced. It seems that there is an optimal degree in which the conditioning is elicited, this being an intermediate stage between a deep and a light anesthesia. References p. 215-217

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Fig. 21. Conditioning of the blood pressure fall due to septal stimulation in the rabbit with urethane anesthesia. A, before conditioning septal stimulation elicited the blood pressure fall and ECG desynchronization. After some associations (B) the blood pressure fall was conditioned. The conditioned stimulus desynchronized the ECG and determined a fall which paralleled that obtained by the unconditioned stimulus.

V 3 X V 7 V L d 3 S 3H1 do S N O I 1 3 N n J 'IVXOIAVH3CI a N V ElAIlVL3DElAOXfl3N

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Fig. 23. Conditioning in the rabbit under urethane anesthesia. Differentiation. A, the conditioning has been established to a click train of lO/sec repetition rate; B, there was no response to a stimulus of 31 sec repetition rate; C, the conditioning is again shown to a click train of lo/=.

NEUROVEGETATIVE A N D BEHAVIORAL F U N C T I O N S OF THE SEPTAL A R E A 2 0 9

Fig. 24. Conditioning in the rabbit with urethane anesthesia. Extinction. D, the conditioning being established, the blood pressure fall was elicited by the conditioned stimulus without reinforcement. E, after some applications of the conditionedstimulus alone, extinction appeared. F, after some associations the conditioned response reappeared. References p. 215-21 7

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M. R. C O V I A N

In this regard it is well to quote Galambos and Morgan (1960): ‘... relatively small amounts of anesthetics unfailingly reduce animals and men to a state where no learning whatever is possible. One might naively expect that between the stages of complete anesthesia and none at all a level would be reached where the learning process was, say, only half impaired. Such a stage has, however, never been defined’. Emphasis has been placed on the role played by subcortical structures in the process of conditioning, namely, the thalamus, hypothalamus and mesencephalic reticular formation (Cardo, 1961). It seems that the septal area is another subcortical structure of importance in conditioning. Since Pavlov’s work it is known but not always remembered that the conditional reflex extends to the autonomic system, and that this autonomic conditioning is usually out of consciousness. Horsley Gantt (1964) has clearly pointed out: ‘Traditionally both the medical profession and psychology refuse to admit the existence of conditioning below the conscious level, in spite of the fact that Pavlov began his conditional reflex research with the gastrointestinal system three score years ago’. Our finding is another example of conditioning obtained out of consciousness which raises a number of questions such as the activity and role of the reticular formation in this conditioning; the possibility that lower levels of integration, e.g. limbic structures, can accomplish all the steps of the process and also that the rabbit because of neural peculiarities is a specially suitable animal for this unconscious conditioning. These and other questions involving neurophysiological and even psychological implications claim clarification. (111) A L T E R A T I O N S I N S O D I U M C H L O R I D E A N D W A T E R I N T A K E AFTER SEPTAL LESIONS I N T H E R A T

Considering that an organism acts as an integrated unity, and that appetite behavior requires the interaction of different parts of the central nervous system, therefore not depending on a circumscribed region of the brain, we planned a study to evaluate which parts of the CNS are engaged in the control of NaCl and water intake. Accordingly, systematic studies were undertaken in our laboratory to determine the changes on the free ingestion of 1.5 % NaCl and tap water after localized lesions in the brain of the rat. The normal variations in the intake of both fluids, and those introduced by the technical procedures preceding the local destruction of the hypothalamus, as well as the specific alterations in sodium chloride and water intake, have already been reported (Antunes-Rodrigues and Covian, 1963, 1965; Covian and Antunes-Rodrigues, 1963). Owing to the known connections of the septal area with the hypothalamus, and because of the physiological characteristics of this structure, the role it plays in the regulation of the ingestion of both fluids was investigated. This third section of our communication will deal with our preliminary results. Detailed reports of other aspects of these studies are in preparation. In general, the procedures applied have been reported elsewhere (Antunes-Rodrigues and Covian, 1963; Covian and AntunesRodrigues, 1963). The self-selection method was used, rats of both sexes and weighing

N E U R O V E G E T A T I V E A N D B E H A V I O R A L F U N C T I O N S OF T H E S E P T A L

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about 200 g being kept in individual cages. Each had a food cup filled with a dry mixed diet containing 0.1 12 mEquiv/g Na, and two graduated drinking bottles, filled with 1.5 % NaCl and tap water, respectively. Daily readings were made of the intakes as well as of some environment factors which could interfere with appetite behavior, such as temperature, relative humidity and atmospheric pressure of the rat house. After a control period of about 4 weeks bilateral septal lesions were made by electrolysis under ether anesthesia using the co-ordinates of de Groot’s atlas (1959). A stainless steel electrode of 0.36 mm thick and insulated to the tip was mounted in a KriegJohnson stereotaxic instrument, and a current of 1.5 mA was applied for 15 sec. A control group of sham-operated rats was also studied. After septal lesions were made. observations were maintained as previously to study any alterations in the ingestions. Routine histological procedures were used to localize the site of the lesions. In those rats that showed a change in fluid intake this consisted in an increase in NaCl and a decrease in water consumption. In 10 rats with this alteration the average intake of NaCl solution during the control period of 1 month was of 8.10 ml/day; after the septal lesions the value for the same period was of 31.60 ml/day, and for the two months of postoperative observation it was of 23.1 ml/day. For water the values observed were 20.1, 6.65 and 9.96 ml/day respectively. The total ingestion of fluids was 28.16 ml/day before the operation, and 38.2 and 33.05 ml/day for the aforementioned postoperative periods. Food ingestion did not show any significative change. Fig. 25 illustrates the differences observed in one rat: after the bilateral lesion was made at co-ordinates F, 8.0; L, 0.5; H, +0.5 there was a sharp increase in NaCl (solid line) and a drop in water (broken line) intake which remained for the 76 days of postoperative observation. The total amount of fluid consumption showed a value of 25.3 ml/day during the 30 days preceding the operation and of 35.48 ml/day during the 30 days following it; considering the whole postoperative period this last value was 31.09 ml/day. For NaCl and for the same periods the intake was of 4.1, 33 and 26.25

I5



peut persister durant 1 B 2 min au cours de la reanimation (Fig. 6). Elle est parfois surchargee de courts fuseaux interessant simultantment l'hippocampe, la formation reticulaire et le vermis du cervelet. L'activation hippocampique a des expressions differentes durant l'hypoxie et la reanimation.

H I P P O C A M P E ET HYPOXIES O X Y P R I V E S REPETEES CHEZ LE C H A T

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En cours d‘hypoxie profonde des pointes isoltes ou rythmiques, des fuseaux avec ondes rapides (12 ii 15 c/sec) ou ralenties (6 a 7 c/sec) sont identifites dbs les premiers jours aprbs la naissance. Les rythmes hippocampiques peuvent Ctre les derniers A disparaitre avant la ptriode de tract nul. En cours d’hypoxie modtrte de longues ptriodes d’activitt synchroniste de 6 A 7 c/sec surviennent la fin de la premike semaine. En cours de reanimation l’activation hippocampique est plus precise. Elle se traduit soit par des pointes rythmiques, rtptttes durant plusieurs minutes, soit par des fuseaux rapides (25 30 c/sec) limit& a la come d‘Ammon. Durant ces modifications tlectriques le chaton presente une agitation dtsordonnte associCe a des miaulements (Fig. 7). Ces rtactions de l’archto-cortex i% l’hypoxie s’opposent par leur prkcision, B celles du nto-cortex traduites uniquement par des pointes isoltes. Cette difftrence, en faveur d’un niveau de maturation hippocampique plus avanct, est appuyte par ailleurs par l’existence dbs la naissance d’une activitt rtgulikre hippocampique de 8 a 12 c/sec et rtactive A certaines stimulations sensorielles (olfacto-trigtminte) (Cadilhac et Passouant-Fontaine, 1962). La pbriode du 15dme au 3Odme jour La pCriode comprise entre le 15bme et 30kmejour est caracttriste par l’organisation progressive des ptriodes d’activation et de depression telles qu’elles existent chez l’animal adulte et par l’apparition de dtcharges tonico-cloniques en cours de rtanimation. L’activation corticale, traduite par des rythmes rapides est indiqute dbs le 15bme jour, la synchronisation hippocampique est precise au 30bme jour. Lors de la ptriode d’inhibition, des ondes ralenties corticales surviennent dbs le 15bmejour et sont comparables B celles de l’adulte, a la fin du premier mois (Fig. 8). En ptriode de rtanimation des dtcharges organistes tonico-clonique apparaissent A partir du 15bmejour. Ces dtcharges de longue durte (3 4 min) inttressent la formation rtticulaire et peuvent se propager au cortex (Fig. 9). Elles sont accompagntes sur le plan comportemental, d’une agitation de l’animal, associte a des miaulements plaintifs. Ces dtcharges sont frtquentes et peuvent se rtptter durant la reanimation. Elles surviennent, aprbs la premibre hypoxie et non, comme chez l’animal adulte, aprks plusieurs hypoxies. Elles se reproduisent, dans une proportion de plus de 50%’ au cours des hypoxies successives. Des ltsions histologiques inttressent les neurones des divers champs de la come d’Ammon, mais il n’a pas Ctt observt de nCcrose sectorielle. DISCUSSION ET CONCLUSIONS

La rtpttition d‘hypoxies oxyprives, chez le chat libre de ses mouvements, prtcise les reactions de l’archto-cortex zi ce type d’hypoxie, indiqutes par ailleurs par les ttudes anatomiques classiques. Les rtactions tlectriques de la come d’Ammon varient avec le degrt de l’hypoxie, sa rtpttition et en fonction de l’lge de l’animal. References p. 474475

472

P. PASSOUANT, C. P T E R N I T I S

Fig. 8. Reactions il l’hypoxie profonde d‘un chaton de 15 jours.I, Pkriode d‘activation; 11, Pkriode de dkpression corticale. III, Pkriode de track plat.

Fig. 9. Decharge spontank hippocampo-dticulaireet corticale aprh 3 min 40 sec de rhnimation, chez un chaton de 15 jo-.

Chez I’animal adulte Trois types de risultats miritent &&re isolis : les rtactions ilectriques constantes, les dtcharges Cpisodiqueset la rtsistance de l’activit6 Clectrique de la corne d’Ammon a l’hypoxie. (a) Les riactions constantes correspondent A la synchronisation hippocampique de la piriode d’activation de l’hypoxie et aux pointes hippocampiques qui surviennent lors de la pCriode de dtpression corticale. Ces 2 types de modifications de l’activitt Blectrique de l’hippocampe, dicrites, l’une en piriode d‘alerte, l’autre au cours du

HIPPOCAMPE ET HYPOXIES OXYPRIVES REPETEES CHEZ LE C H A T

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sommeil lent (Rimbaud et al., 1955), objectivent au niveau de l’archto-cortex les 2 ptriodes successivesd’activation et de dtpression provoqutes par I’hypoxie (Dell et al., 1961). La frtquence et la prtcision des pointes hippocampiques peuvent Ctre facilities par l’activation rtticulaire qui persiste durant la ptriode de dtpression corticale. (b) Les dtcharges tpisodiques ne surviennent qu’aprbs plusieurs hypoxies et correspondent aux fuseaux hippocampiques et aux dtcharges hippocampo-rtticulaires. (1) Les fuseaux, de la pCriode terminale de l’hypoxie profonde et du dCbut de la rtanimation, sont localisis l’hippocampe et sans propagation. Ce type d’activation parait exprimer une modification des conditions locales de l’hippocampe en liaison avec l’appauvrissement extrgme et rtpttt du milieu en 0 2 . I1 ne parait pas dtpendre de I’activation rtticulaire qui, dans nos conditions d’exptriences, peut avoir disparu lors de la production de ces fuseaux. Des dtcharges analogues enregistrtes au niveau du cortex ecto-sylvien et du cervelet, la ptriode terminale de l’hypoxie profonde et au dtbut de la rtanimation paraissent traduire aussi, un processus d’activation IocalisCe. La plus grande frtquence des fuseaux hippocampiques est en faveur d‘une atteinte sClective de cette structure. Ce type d’activation localiste parait Ctre la traduction tlectrique d’une souffrance precise des neurones dont la constquence pourrait Ctre un abaissement du potentiel de membrane. Un tel mtcanisme a CtC retenu pour l’activation des neurones spinaux au cours d’hypoxie(Kolmodin et Skoglund, 1959), et pour l’activation qui accompagne la ptriode de rtanimation (Baumgartner et al., 1961). (2) Les dtcharges hippocampo-rkticulaires surviennent en milieu plus riche en 0 2 (hypoxie ,modtrte) lors de la rtanimation et en tant que stquelles post-hypoxiques. Ces dtcharges ont probablement un mtcanisme different selon les conditions de leur production. L‘absence de ltsions hippocampiques, chez les animaux soumis B des hypoxies modtrtes est en faveur d‘une perturbation mttabolique rtversible qui prtctde les ICsions anatomiques irrtversibles observtes aprts des hypoxies profondes rtptttes. La frtquence des dtcharges organistes hippocampo-rtticulaires, comparativement aux dtcharges corticales et gtntralistes, souligne B nouveau la reactivitt particulitre de l’hippocampe a I’hypoxie oxyprive. Une telle sensibilitt va de pair avec la rtactivite exquise de l’hippocampe A la stimulation tlectrique, mtcanique ou chimique et qui est facilitte par l’organisation architectonique de cette structure (Passouant et Cadilhac, 1962). (3) L’activitC Clectrique de l’archto-cortex est plus rtsistante a I’hypoxie que celle du nCo-cortex. Cette constatation en faveur d‘un moindre besoin en 0 2 des structures phylogCnCtiquement anciennes, est appuyCe par certains rCsultats sur la consommation en 0 2 de diverses structures ctribrales: cette consommation &ant plus ClevCe pour le cortex que pour la corne d’Ammon (Quastel et Quastel, 1961). Chez le chaton Deux rtsultats mtritent d’&treisolts : (1) la sensibilitt exquise de l’hippocampe a I’hypoxie, (2) I’expression difftrente de la rtaction hippocampique au cours du premier mois. (1) La sensibilitC de l’hippocampe aux hypoxies oxyprives rtpCttes est plus grande References p . 474475

414

P . P A S S O U A N T , C. P T E R N I T I S

que chez l'animal adulte. Les rCactions hippocampiques (pointes rythmiques, fuseaux) sont plus frtquentes. Les dkcharges tonico-cloniques, apparues d5s le 15bme jour, surviennent des la premiere hypoxie et sont retrouvkes dans une proportion de plus de 50 % au cours des hypoxies suivantes. Ces rksultats en prtcisant la rtactivitt de l'hippocampe ii l'hypoxie dks la naissance, confirment la facilitt convulsivante du jeune animal. (2) L'organisation progressive des rkactions hippocampiques B l'hypoxie au cours du premier mois, confirme les indications dbjh obtenues sur l'tvolution de la maturation de l'hippocampe (Cadilhac et Passouant-Fontaine, 1962; Passouant et al., 1965). C'est ainsi que les dtcharges hippocampiques, provoqutes par l'hypoxie ii partir du 1%me jour, prkisent l'organisation h ce moment des relations hippocampo-rtticulaires. Ces ddcharges hippocampiques sont les premitres ii apparaitre, les dkcharges gtnCralisCes ne survenant qu'apres le premier mois. D'aprbs ces premiers rbultats, indiquant les variations de l'activation de l'hippocampe par l'hypoxie selon l'fige, l'ttude de l'hypoxie pourrait Ctre retenue chez le jeune animal comme un test valable dans l'ttude de la maturation ctrtbrale. BIBLIOGRAPHIE

BAUMGARTNER, G., CREUTZFELD, O., AND JUNG, R., (1961); Microphysiology of cortical neuron- in acute anoxia and in retinal ischemia. Cerebral Anoxia and the Electroencephalogram. Springfield, Thomas, pp. 5-34. BREMER, F., ET THOMAS, J., (1936); Action de l'anoxie et de l'hypercapnie sur l'activite klectrique du cortex drkbral. C.R. SOC.Biol. (Paris), 123, 1256-1260. CADILHAC, J., ET PASSOUANT-FONTAINE, TH., (1962); Dkcharges kpileptiques et activitk klectrique de veille et de sommeil dans l'hippocampe au cours de l'onto&&e. Physiologie de I'Hippocampe. Paris, Centre National de la Recherche Scientifique, pp. 429442. C~GGESHALL, R. E. AND MACLEAN,P. D., (1958); Hippocampal lesions followhg administration of 3-acetylpyridine. Proc. SOC.Exp. Biol. N.Y., 98, 687-689. CREUTZFELD, O., KASAMABU,A., UND VAZ-FERREIRA, A., (1957); Aktivitatsanderungen einzelner corticaler Neurone im akuten Sauerstoffmangel und ihre Fkziehungen zum EEG tn?i Katzen. Pfliigers Arch. ges. Physiol., 263,647-667. DELL, P., HUGEF, A., ET BONVALLET, M., (1961); Effects of hypoxia on the reticular and cortical diffusesystems. Cerebral Anoxia and the Electroencephalogram. Springfield, Thomas, pp. 46-58. EULER,C. VON, (1962); On the signifcance of the high zinc content in the hippocampal formation. Physiologie de I'Hippocampe, Paris, Centre National de la Recherche Scientifque, pp. 135-145. GELLHORN, E., AND HEYMANS, C., (1948); Differentialaction of anoxia, asphyxia and carbon dioxide on normal and convulsive potentials. J. Neurophysiol., 11, 261-274. HUGELIN, A., BONVALLET, M., ET DELL,P., (1959); Activation reticulaire et corticale d'origine chemoceptive au c o w de l'hypoxie. Electroenceph. clin. Neurophysiol., 11, 326340. KOLMODIN, G. M., AND SKOGLUND,C. R., (1959); Influence of asphyxia on membrane potential level and action potentials of spinal mot0 and intemeurones. Actaphysiol. scand., 45, 1-18. LAMMERS, H. J., ET GASTAUT, H., (1962); Relations cyto-architectoniqueset enzymo-architectoniques dam l'hippocampe. Physiologie de I'H*pocampe. Paris, Centre National de la Recherche Scientifique, pp. 1-21. MACLARDY, T., (1960); Neuro syncitial aspects of the hippocampal mossy fibre system. Confin. neurol. (Basel), 20, 1-17. MACLARDY,T., (1962); Pathological zinc rich synapse. Nature (Land.), 195, 1315-1316. NILGES, R. E., (1944); Arteries of the mammalian comu Ammonis. J. comp. Neurol., 80, 117-190. PASSOUANT, P., ET CADILHAC, J., (1962); Place de l'hippocampe dans l'organisation fonctiomelle du cerveau. J. Psychol. (Paris), No. 4.

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PASSOUANT, P., CADILHAC, J., ET PASSOUANT-FONTAINE, TH., (1965); Hippocampe et maturation &rebrale. Actualitis Neurophysiologiques, 6 eme drie. Paris, Masson, (sous presse). PASSOUANT, P., ET PTERNITIS, C., (1965); Activation de l'hippocampe par des hypoxies oxyprives repetks. Acta. physiol. Acad. sci. hung., 26, pp. 123-130. PTERNITIS, C., (1964); Reactions de I'hippocampe a des hypoxies oxyprives repktks. Etude dectrophysiologique et anatomique. Th2se Mid., Montpellier, 106 pp. PTERNITIS, C., ET PASSOUANT, P., (1961); Modifications EEG et de comportement au cours de l'hypoxie repet& et de la reanimation chez le chat libre de ses mouvements. C.R. Soc. Biol. (Paris), 155, 2380-2397.

PTERNITIS, C., PASSOUANT, P., ET CADILHAC, J., (1963); Etat de ma1 post-anoxique, place de l'hippocampe dans l'entretien de l'etat de mal. C.R. SOC.Biol. (Paris), 157, 347-349. QUASTEL, J. H., AND QUASTEL, D. M., (1961); The Chemistry of Brain Metabolism in Healthand Diseases. Springfield, Thomas, 170 pp. RIMBAUD, L., PASSOUANT, P., ET CADILHAC, J., (1955); Participation de l'hippocampe a la regulation des Ctats de veille et de sommeil. Rev. Neurol., 93, 303-308. SCHOLZ, W., (1952); Les nkcroses parenchymateuses electives par hypoxkmie et oligkmie et leur expression topistique. Atti I" Congr. Intern. Istopathol. Sist. Nerv., 1, 321-346. SPIELMEYER, W., (1962); Histopathologie des Nervensystems, Berlin, Springer. SUGAR,O., AND GERARD, R. W., (1938); Anoxia and brain potentials. J. Neurophysiol., 1, 558-572. STAFFORD, A., ASD WEATHERALL, J. A. C., (1960); The survival of newborn rat in nitrogen. J. Physiol. (Lond.), 153,457412. WINDLE,W. F., BECKER, R. F., AND WEILL,(1944); A., Alteration in brain structure after asphyxiation at birth. An experimental study in the guinea-pig. J. Neuropath. exp. Neurol., 3, 224-238.

476

Author Index * Abrahams, V. C., 111 Adam, D., 295 Adamovitch, N. A., 295 Adey, W. R., 3, 85, 171, 218, 228-245, 254, 274, 393,425,429,430,455,457 Adrian, H. O., 27,28 Adrianov, 0.S., 340 Auapetyants, E. Sh., 293-304 Ajmone Marsan, C., 2, 190, 339 Akert, K.,6 6 3 , 1 1 4 , 134,413 Albe-Fessard, D., 271,272 Alcocer-Cuaron, C., 246 Anand, B. K., 1,2,25, 132 Andersen, P., 114, 119, 174, 320,400412, 452,457 Andy, 0. J., 114, 134 Antunes Rodrigues, J., 190,201, 203,210,214 Aoki, I., 129 Archibald, D., 91, 94 Arduini, A., 218,232,290,413-415,429, 430,433,445 Asanuma, H., 28 Asanuma, H., 318 Aserinsky, E., 420 Azzali, G., 176 Babkin, B. P., 300 Badier, M., 180 Bagshaw, M. H., 325,329 Baldwin, M., 175 Ballim, H. M., 360 Balvin, R. S., 305 Ban, T., 85, 173, 203 Bando, T., 384 Barbizet, J., 229 Bard, P., 176 Barker, S. A., 242 Barraclough, C.A., 2, 69 Barrnett, R. J., 241 Baumgartner, G., 462,464,473 Bayyuk, S. H. J., 242 Beck, E. C.,246 Becker, R. F., 469 Bell, F. R., 235 Bergmann, F., 183-188 Beteleva, T. G., 413 Biscoe, T. J., 359 Blackstad, T. W., 401,403

Bliss, C. I., 365 Bond, D. D., 205 Bonnet, V., 257, 271, 338, 423 Bonvallet, M., 23, 46, 462, 464, 473 Borgest, A. N., 295 Boswell, R. S., 433 Bowden, 3. W., 34 Boyajy, L. D., 360 Boyarsky, L. L., 361 Bradley, P. B., 338, 359 Brady, J. V., 176,205,295,433 Bremer, F., 34, 257,271, 290, 338,423,424, 466 Bremner, F., 229 Brinley, F. J., 404,411 Broadwick, M., 333 Brobeck, 5. R., 1 Brodal, A., 430 Brodwick, M., 313 Brookhart, 5. M., 430 Brooks, C. McC., 76 Brooks, D. C., 3 Brooks, V. B., 318 Brown, R. M., 422 Brown, T. S., 228,229,231, 233, 234 Brown, W. C., 360 Briicke, F., 413 Brugge, J. F., 413, 433 Briigger, M., 103, 117, 118 Bruland, H., 378 Brust-Carmona, H., 246 Brutkowski, S., 316 Bucher, V. M., 103-127 Buchwald, N. A., 378 Bunn, J. P., 71 BureS, J., 166 Biirgi, S., 124 Buser, P., 254, 274 Butter, C. M., 316 Candia, O., 203, 204 Cadhillac, J., 114,413,462, 471,473, 474 Cajal, S. Ram6n y, 401 Cannon, W.B., 111 Cardo, B., 210 Carlsson, A., 398 Carreras, M., 176 Caspers, H., 430

* Italics indicate the pages on which the paper of the author in these proceedings is printed.

AUTHOR I N D E X

Cazard, G., 254, 274, 429 Cenacchi, V., 422,430 Chaillet, F., 356 Chairnovitz, M., 183-188 Chambers, W. W., 2 Chamorro, A., 69 Chang, H. T., 26 Charbon, G. A., 203 Chernigovskii, V. N., 295 Chhina, G. S., 2, 25 Chow, K. L., 251 Clark, C. V. H., 307 Clark, W. G., 382-384 Clemente, C. D., 1, 29, 34-47, 176 Coggeshall, R. E., 462 Corazza, R., 413423,429, 430 Cordeau, J. P., 311,425 Corti, U. A,, 111 Costin, A., 183-188 Cotten, M. de V., 203 Cottrell, G. A., 26 Courvoisier, S., 384 Coury, 5. N., 213 Covian, M. R., 189-217 Cragg, B. G., 26 Cramer, M., 243 Creuzfeldt, O., 28, 242, 462,464, 473 Critchlow, V., 72 Crosby, E. C., 62, 85 Cross, B. A., 2 Crossland, J., 340 Csanaky, A , 114 Curtis, D. R., 359 Daigneault, E. A., 360 Dalla Rosa, V., 176 D’Amour, F. E., 365 Dashiell, J. F., 312 De Groot, J., 26, 176, 190,211 Deisenhammer, E., 413 Delgado, J. M. R., 1, 29,4848, 119, 129, 130, 134, 170, 180, 189,204,445 Dernpsey, E. W., 34 Dell, P., 28, 46, 291, 424, 462, 464, 473 Delov, V. E., 295 Dernber, W. N., 313, 316,333 Dement, W., 383,420 Demetrescu, Maria, 290, 424 Demetrescu, M., 290, 423, 424 Denisenko, P. P., 359 Dennis, B., 235 Denniston, R. H., 445 Desmedt, J. E., 359,424 Dews, P. B., 365 Dewson, 111, J. H., 322 Diamond, F. D., 305 Diamond, S., 305 Didio, J., 237 Dietiker, M., 111

477

Dodo, T., 174 Domino, E. F., 337-364 Donhoffer, H., 218, 219, 222, 223,235, 274, 413,425,445 Doty, R. W., 246 Douglas, R. J., 313, 329, 333, 458 D r a c h m a , D. A., 229 Dren, A. T., 337-364 Dua, R. H..445 Dua, S., 2, 25 Ducrot, R., 384 Durnont, S., 290, 424 Dunlop, C. W., 218, 229,235,429 Eccles, 5. C., 12, 27, 338, 404, 452 Eccles, Sir J., 320 Eckhaus, E., 246 Egger, M. D., 63, 165-182 Egyhazi, E., 241 Elkes, J., 359 Elliasson, S., 203 Elul, R., 243, 290, 429 Elwers, M., 72 Endrkzi, E., 246-253 Epstein, 5. A., 296, 300 Evarts, E. V., 144 Everett, J. W., 70, 71, 76, 190 Fangel, C. H., 119, 251,258,273 Farago, L., 445 Fatt, P., 27 Favale, E., 203, 204, 425 Fee, A. R., 76 Feldberg, W., 337, 340 Feldman, S., 457 Fernandez de Molina, A., 65,103, 114, 117-119, 122, 177, 180 Fernandez-Guardiola, A., 62 Fessard, A,, 254 Fifkovk, E., 151 Fisher, A. E., 213 Flataker, L., 365 Flerkb, B., 25,26, 69, 170 Flym, 5. P., 63,165-182 Folkow, B., 203,204 Fonberg, E., 1, 29, 170, 173, 316 Fox, C. A., 119, 170, 173 FOX, S. S., 254-280, 320 Frazier, D. T., 361 French, 5. D., 28, 34 Frost, L. L., 175 Fujita, M., 242, 243, 413, 429 Fuller, J. L., 1 Fulton, J. F., 295 Fuse, S., 1, 29, 180 Fuster, J. M., 242 Galambos, R., 210 Gangloff, H., 183, 186, 413

478

AUTHOR INDEX

Gantt, W., Horsley, 210 Gardner, K. W., 329 Gary, T. M.,2 Gasanov, G. G., 296 Gassmann, F., 111 Gastaut, H., 180, 246,462 Gauthier, C., 423 Gaza, N. K., 295 Geller, I., 176 Gellhorn, E., 63,64,360,413,466 Gerard, R. W., 254, 270,466 Gergen, J. A., 442-461 Gerschenfield, H. M.,26 Gerstein, G. L., 12,452 Giussiani, A., 203, 204 Gloor, P., 63, 64, 85, 173, 384, 452 Gogolslk, G., 218, 356 Gogolof, G., 413 Goldberg, J. M.,27 Goldring, S., 429 Goldstein, A. C., 76 Goldstein, M. H., Jr., 422 Goodfellow, E. F., 132 Goodman, L. S., 366 Gorski, R. A., 69 Gorten, R. A., 201 Grastyh, E., 114,218-223,229,235,274, 413,425,445 Green, J. D., 1,29, 63, 70, 85, 176, 190, 218,232,254,255,272,290,413415, 429,433,442,445 Groot, J., 1, 29 Grossman, S., 213 Grossman, S. P., 26 Gulyaeva, L. N., 295 Gutman, J., 186, 188 Gwbtdt, B., 103, 113 Gygax, P. A., 451 Haberland, K., 445 Haffner, F., 365 Hagiwara, S., 27,28 Halirsz, B., 25,26, 170 Hall, E. A., 170 Hamlyn, M. H., 26,401 Hammel, H. T., 63 Hammerstein, J., 91 Hanai, T., 2,27, 171 Hanking, B. H., 71 Hardy, J. D., 63 Harmony, T., 62 Harris, G. W., 69 Harrison, J. M.,205, 433 Hartline, H. K., 25, 318 Harvey, J. A.. 204,213 Hasegawa, I., 290 Hayward, J. N., 91, 94 Heath, R. G., 205,235 Heller, A., 204

Henatsch, H.- D.. 384 Hendrix, C. E., 218,228,229, 393 Hemhdez-Pebn, R., 28,246 Herzet, J. P., 395 Hess, J., Jr., 46 Hess, W. R., 34,46, 62, 103, 105, 117, 118, 129,203,435 Hetherington, A. W., 1 Heuser, G., 378 Heymans, C., 466 Hiebel, W., 26 Had, G., 46 Hilliard, J., 91,94 Hilton, S. M.,111, 119, 122, 180 Himwich, H. E., 338 Hirose, K., 365-387 Hisaw, F. L., 81 Hodes, R., 62 Hoebel, B. G., 132 Hohlweg, W., 69 Holines, J. E., 223, 425 Holmqvist, B., 402 Hori, Y., 290 Horvath, F. E., 176 Hosli, L., 46 Hugelin, A., 28,462,464,473 Hugh Dingle, R D., 254-280 Humphrey, T., 85 Hunter, J., 114 Hullay, J., 445 Hunsperger, R. W., 65, 103-127, 129,177, 180 Hunt, C. C., 27 Hunt, H. F., 204,213 Hutt, P.J., 176 Huttenlocher, P. R., 28 Hyde, J. E., 96, 180 Hydh, H., 241 Iki, M.,2-4,12,25,27 Ilyutechenok, R. I., 359 Imamura, G., 295, 356 Ingvar, D. H., 384 Iosif, G., 290,424 Isaacson, R. L., 309,313, 316, 333,458 Ito, M., 290 Itoigawa, N., 281-292 Iwamura, Y., 377,380,382, 383 Iwata, K., 254, 272, 413 Jackson, D. C., 63 Sansen, J., 258,273 Jansen, J., Jr., 114, 119, 174 Jarrard, L. O., 307, 309,458 Jasper, H. H., 2, 114, 190,229,239,241, 338, 357 Johansson, B., 203,204 John, E. R., 388,397 Johnston, J. B., 119 Jouvet, D., 204

AUTHOR I N D E X

Jouvet, M., 28, 204, 383, 425 Julou, L., 384 Jung, R., 28,229,413,462,464,472 Junkmann, K., 69 Kaada, B. R., 62, 114, 119, 122, 173, 174, 180,251, 295, 296, 300, 316, 378 Kabat, H., 203, 204 Kado, R. T., 235, 237, 239,241,445 Kaelber, W. W., 176 Kaitor, F., 445 Kaji, S., 2, 27, 171 Kamikawa, K., 228 Kamikawa, Y., 3 Kanai, T., 357 Kandel, E. R., 404,411,452 Karli, P., 122 Karmos, G., 219, 220,425 Karplus, 5. P., 116, 189 Kasamatsu, A., 464 Kato, S., 295 Kasamatsu, A., 464 Kato, S., 295 Kawakami, M., 69-102,413 Kawamura, H., 204,295, 356, 377, 382, 383, 413 Kellhyi, L., 219, 425 Keilicutt, M. H., 316 Kennard, M., 295 Kerkut, G. A., 26 Kiang, N. Y.-S., 422 Kiang, N. Y.-S., 12, 452 Kido, R., 365-387 Killam, K. F., 388-399 Kimble, D. P., 316, 325, 333,458 Kimura, D., 316,458 Kimura, K., 2 4 , 12, 25,27 King, F. A., 433 King Killam, E., 388-399 Kite, W. C., 300 Kjaerheim, A., 401 Kleitman, N., 420 Kletzkin, M., 124, 129, 130 Kling, A., 176, 177 Klingberg, F., 218-227 Knapp, D. A., 359 Kobayashi, N., 1-33 Koella, W. P., 271, 274 Koelle, G., 337 Koepke, J. E., 327 Kogi, K., 295 Koikegami, H., 1,29,48, 62, 63, 71, 85, 122, 173, 174, 180 Koizumi, K., 359 Koketsu, K., 27 Kolmodin, G. M., 27 Kolmodin, G. M., 473 Kooi, K. A., 246 Korhyi, L., 245, 247

Kornmuller, A. E., 229,413 Kosman, A. J., 1,29, 176 Kotljar, B. I., 218, 223 Kreidl, A., 116, 189 Kreindler, A., 173, 175, 180 Kremer, W. F., 295 Krnjevie, K., 357, 361 Kruger, L., 318 Krupp, P., 46 Kubota, K., 380, 384 Kuehn, A., 382,384 Kuniyashi, M., 2-4, 12, 25, 27 Kuno, M., 27 Kurotsu, T., 203 Kushiro, H., 180 Kveim, O., 316 LaGrutta, G., 359, 424 Lammers, H. J., 119, 124, 180,462 Lauer, E. W., 85 Lechi, A., 176 Lena, C., 433,434 Levison, P. K., 176 Levy, C. K., 271 Lewis, P. R., 357, 359 Liberson, W. T.,413 Libeskind, J. C., 254-280 Lim, R. K. S., 340 Lilly, J. C., 63 Lindgren, P., 203 Lindsley, D. B., 34, 322, 235 Lisk, R D., 69 Lisshk, K., 114, 218, 219, 222, 223, 235, 246253,413,425 Liu, C. N., 340 Livingston, R.B., 28,245, 274,451 Lloyd, D. P. C., 12, 27 Lobanova, L. V., 293 Loeb, C., 368,425 Loeser, J. D., 271 L0m0, T., 400-412 Long, J. P., 361 Longo, V. G., 223,338 Lbpez-Mendoza, E., 246 Lorente, de N6, R., 401, 442 Losonczy, H. V.,220 Layning, Y.,404,452 Lux, H. D., 242 Lyon, M., 205,433 Macchi, G., 176 MacGillivray, B., 241 Machne, X.,254,413,429 Macht, M. B., 76 MacIntosh, F. C., 357, 361 MacLardy, T., 462 MacLean, R. D., 65, 114, 119, 124, 180, 201-204,295, 318,433,435,422-446, 459,462

479

480

AUT H O R I N D E X

Madarasz, T., 218,219, 222, 223,235,274 413,425 Maeno, S., 290 Maeno, T., 2 4 , 12, 25,27 Magni, F.,360 Magnus, J., 26, 28, 170 Magnus, O., 119, 124, 180 Magoun, H. W., 34,44,62,203,204,246,356 Mahut, A., 311 Mahut, M., 425 Mancia, M., 430 Manfredi, M., 425 Manning, J. W., 203 Mantegazzhi, P., 338, 357 Manzoni, T., 425,428432,457 Marshall, W. H., 254, 270 Martin, J., 425 Maruyama, N., 2,27, 171 Mason, J. W., 96, 97 Matsushita, A., 365-387 Matsumoto, J., 290 Matthews, B. H. C., 28 Maxwell, D. S., 413,429, 452 Mayer, Ch., 413 McCleary, R. A., 316, 318 McIlwain, J. T., 228 McIver, A. H., 433 McLardy, T., 226 McQueeney, J. A., 458 Megawa, A., 203 Mernpel, E., 316 Mering, T. A., 340 Merrick, A. J., 340 Mess, B., 25,26, 170 Metz, B., 361 Michael, R. P., 69 Michelson, M. J., 359 Miller, G. A., 318 Miller, H. R., 176, 205 Miller, N. E., 1, 128 Milner, B., 318 Milner, P., 205 Mir, D., 54 Mishkin, M., 228, 316 Mitchell, J. F.,357 Mitsuyasu, K., 384 Miyamoto, K., 281-292 Mochida, Y.,174 Moffitt, R. I., 340 Moiseeva, N. A., 296 Mollica, A., 359 Monnier, M.,46, 183, 186, 413 Monroe, R. R., 235 Moore, R. Y.,204,458 Morgan, C. T., 209 Morgane, P. J., 1, 2, 29, 176 Morin, F.,124, 254, 255, 429 Morin, G., 180 Morison, R. S., 34

Morrell, F., 246 Moruzzi, G., 28, 34,44,46,246, 356,430 Mountcastle, V. B., 6, 11, 176, 318 Musyaschikova, S. S., 295 Nacimknto, A., 242 Naka, F., 1-33 Nakamura, Y.,382,413 Nakao, H., 118,128-143, 176, 177, 360 Naquet, R., 62, 64,119, 180 Nauta, W. J. H., 2, 29, 85, 119, 203,205, 246, 295,433,451 Negishi, K., 29 Nicholson, A. N., 359 Nielson, H. M., 433 Niimi, Y.,380 Nilges, R. E., 462 Niemer, W. T., 132,339 Niki, H., 305-317, 383, 458 Nobel, K., 322 Norris, Jr., F. H., 180 Novikova, L. A., 413

bberg, B., 293,204 OBrien, J. H.,254-280 ODoherty, D. S., 48

O'Flaherty, J. J., 194, 201, 203 Okuda, O., 28 Okuma, T., 378 Olds, J., 132,144-164, 205 O'Leary, J. L., 124,430 Ommaya, A. K., 229 Ornukai, F., 173 Ono, T., I-33 Oomura, Y.,1-33 Ooyama, H., 1-33 Oppenheimer, M. J., 176, 205 Orbach, J., 176, 177

Palesthi, M., 425 Papez, J. W., 295 Parkes, A. S.,76 Parma, M., 423 Parmeggiani, P. L., 124, 4 1 3 4 1 Passouant, P., 413, 462475 Passouant-Fontaine, Th., 413, 471, 474 Paton, D. W. M., 356 Paul-David, J., 360 Peele, T. L., 71, 119, 174, 176, 180 Penfield, W. G., 318 Pepeu, G., 357 Petrh, M., 166 Petsche, H., 218,290, 356,413,429 Phillis, J. W., 357 Pickenhain, L., 218-227 Pillat, B., 413 Pisano, M., 425 Ploog, D. W., 65,203,204,445 Poggio, G. F.,12

AUTHOR INDEX

Pompeiano, O., 28,46, 380,413, 429 Porter, R. W., 3, 171, 228, 229, 231, 233, 234 Pressman, G. L., 329 Preston, J. B., 382 Pribram, K., 1,228, 252, 296, 300, 318-336 Pternitis, C., 462475 Quastel, D. M., 473 Quastel, J. H., 473 Rabini, C., 419, 425 RadulovaEki, M., 229,230,235,236,242,425 Rall, W., 26 Ramey, E. R., 48 Randall, L. D., 382 RandiC, M., 359 Ranson, S. W., 1, 203, 204 Rasmussen, E. W., 316 Ratliff, F., 25 Redding, F., 254, 274 Reite, M. L., 445 Rhodes, I. M., 445 Rice, B. F., 91 Riehl, R.-L., 360 Rimbaud, L., 413, 473 Rinaldi, F., 338 Roberts, S., 71 Roberts, W. W., 128, 313, 333 Robinson, B. W., 49,203 Robinson, F., 413 Roldan, E., 151 Rosadini, G., 425 Rossi, G. F., 360,425 Rossi, R. F., 203, 204 Rosvold, H. E., 1, 49, 316 Roth, L. J., 204 Rumbaugh, D. M., 458 Rushmer, R. F., 201 Sacco, G., 425 Sadowski, B., 223 Sagawa, Y., 384 Sailer, S., 219, 226 Sakai, A., 203 Sakai, Y.,384 Salmoiraghi, G. C.. 359 Salvatorelli, G., 419, 426 Sano, T., 119 Sato, T., 242, 243, 413, 429 Sevillano, M., 134 Saul, C. J., 254, 270 Savard, K., 91 Sawa, M., 2, 27, 171 Sawada, M., 1, 26,27 Sawyer, C . H., 69, 70, 76, 91, 94, 190 Schallek, W., 382, 384 Schindler, W. J., 237, 241, 413, 429, 452 Schlag, J. D., 356 Scholz, W., 462

48 1

Schottelius, B., 175 Scholz, W., 462 Schottelius, B., 175 Schreiner, L., 176 Schueler, F. W., 361 Schwartz, N. B., 176, 177 Schwarz, H. G., 124 Schwarzbaum, J. S., 316, 329 Scott, P. P., 69 Sears, T. A., 320 Segundo, J. P., 254, 274,457 Seiden, L. S., 398 Sekiguchi, N., 71 Seto, K., 69-102,71 Sharma, K. W., 2,25 She&', C. N., 71, 119, 174, 176, 180 Sheer, D. E., 48 Shimazu, H., 380 Shirnokochi, M., 290, 291 Showers, M. J. C., 62 Shute, C. C. D., 357, 359 Sidman, M., 367 Siegel, S., 305 Siegfried, J., 254, 274 Silver, A,, 357, 359 Silvestrini, B., 338 Singh, B., 2,25 Skoglund, C. R., 27 Skoglund, C. R., 473 Skultely, F. M., 2 Skultety, F. M., 119 Slater, L., 49 Slusher, M. A., 96, 180 Smith, D. L., 365 Smith, F. D., 27 Smith, 0.A., 1 Smith, 0.A., Jr., 201 Smith, W. K., 295,296 Snider, R. S., 254,257,258,270-272, 339,413 Sokolov, E. N., 226, 235, 324 Sotnichenko, T. S., 293-304 Sovetov, A. N., 295 Spencer, W. A., 404,411,425 Sperti, L., 452 Spiegel, A. E., 12!, 129, 130 Spiegel, E. A., 176, 187, 205 Spielmeyer, W., 462 Spinelli, D. N., 320 Sprague, J. M., 2 Stacey, M., 242 Stafford, A., 469 Stamm, J. S., 252 Stark, L., 62 Stefanis, C., 359 Steg, G., 384 Steiner, F. A., 359 Stellar, E., 2 Steriade, M., 173, 175, 180, 423, 424 Sterman, M. B., 3 4 4 7

482

AUTHOR INDEX

Stevenson, J. A. F., 1 Stolwijk, J., 63 Stoupel, N., 423,424 Stowell, A., 254,257,258,270,271 Straughan, D. W., 359 Stremme, S. B., 63 Strumwasser, F.,26 Stuart, D. G., 3, 171 Stumpf, Ch., 218, 219, 226,290, 356,413 429,452 Sugar, O., 466 Summers, T. B., 176 Sunderland, S., 429 Sutin, J., 2, 21, 27, 28, 171 Swett, J. E., 46 Swett, J. E., 380 Swinyard, E. A., 366 Szabo, I., 219 Szabo, Th., 271,272 Szekely, E. G., 124, 129, 130 Szentitgothai, J., 25, 26, 170 Szerb, J. C., 357, 361 Takahashi, H., 174 Tasaki, I., 26, 27 Tauc, L., 26 Teitelbaum, H., 311 Teitelbaum, P., 132 Terasawa, E.,69-102 Thomas, J., 466 Thompson, J. B.,316, 324 Timo-Iaria, C., 199 Tokizane, T., 295, 356, 377, 378, 380, 383, 384,413 Tolmasskaya, E. S., 295 Torii, S., 201, 204, 295, 360,413, 416 Tower, S. S.,251 T o m e , J. C., 176, 177 Tracy, W. H., 205 Trembly, B., 205 Trendelenburg, U., 360 Tsubokawa, T., 2,21,27,28, 171 Tsuchinashi, S., 76-79 Udalova, G. P., 296 Uemura, T., 76 Udvarhelyl, G. B., 132 Umezu, M., 71 Unna, K. R., 360 Ursin, H., 119, 122, 173, 176, 180 Ushikoshi, I., 71 Usui, K., 71 Uvniis, B., 203 Valatx, J. L., 204 Valdman, A. V., 359 Valenstein, E. S., 124 Valverde, F., 170 Van Reeth, P. Ch., 423,424

Van Zwieten, P. A., 413 Vasilevskaya, N. E.,296 Vaz-Ferreira, A., 464 Vera, C. L., 452 Vereby, Gy., 114 Vereczekey, L., 220,425 Vergnes, M., 122 Verly, W. G., 91 Veneano, M., 29 Viernstein, L. J., 12 Villarreal, J. E., 359 Vinagradova, 0. S., 235 Von Bekesy, G., 318 Voorhoeve, P. E., 402 Voronin, L. G., 218,223 Votaw, C. L., 274 Wagner, H. G., 25 Wagner, H. O., 318 Wagner, J. W., 26 Walker, A. E., 124, 132 Wallenberg, A., 124 Walsh, J., 425 Walter, D. O., 228, 232, 234, 235, 237, 241, 242, 393 Ward, A., 295 Warden, C. J., 367 Warner, H., 49 Wasman, M., 165 Watanabe, H., 1,29 Watanabe, T., 1,29 Weatherall, J. A. C., 469 Weber, M., 111 Wendt, R. H., 171 Weill, A., 169 Weiss, T.,151, 239 Werner, H. G., 25 White, L. E., 301, 360 Whiteside, J. A., 272 Wickelgren, W. O., 309, 316,458 Wikler, A., 360, 382 Windle, W. F., 469 Winter, C. A., 365 Wolstencroft, J. H., 359 Wood, D. C.,1 Wood, D. C., 119, 175, 176, 180 Wyers, E. J., 378 Wyrwicka, W., 43 Wyss, 0. A. M., 103, 111,435 Yamada, T., 71 Yamaguchi, Y., 281-292 Yamamoto, K.-I.,337-36#, 365-387 Yamamoto, T., 1-33 Yamanaka, K., 76,79 Yasukochi, O., 118 Yokoyama, T., 1,29 Yoshida, K., 69-102, 122 Yoshida, M., 134, 173, 180

AUTHOR INDEX

Yoshioka, M., 382 Yoshii, H., 290 Zachar, J., 166 Zanchetti, A., 423

1

Zanocco, G., 413,424,433,435 Zaraiskaia, S. M., Zbroiyna, A., 111, 119, 122, 180 Zubkova, N. A., 294

483

484

Subject Index Acetylcholinelevel, and cholinergic agents, 354-357 and hemicholinium, 354-356 Affective behavior, and brain stem, electrical stimulation, 103-125 and forebrain, electrical stimulation, 103-125 mewing, and brain level, 114, 115, 122-124 zones, 122-124 pain reactions, 115, 116, 124 Alimentary conditioned reflex, and amygdala, 248-251 and hippocampus, 248-251 and reticular formation, 248-251 Amygdala, ablation, and aggressive behavior, 165-180 affective behavior, zones, 119-122 and cortex, evoked responses, 269-272 EEG activity, and ovulation, 81-83, 98, 99 electrical stimulation, and aggression, latency, 168-172 and conditioned reflex, 248-251 estrogen administration, and ARC potential, 79, 80 function, and ovulation, 71-74 and hypothalamus, functional interaction, 16-21, 29 lesions, and attack behavior, 171-175, 177-179 and hyperphagia, 1 , 2 and orienting, regulation, 325-329 progesterone administration, and ARC potential, 79, 80 Amygdala, radio stimulation, responses, 60, 61, 64 stimulation, and affective behavior, 119, 120 and aggressive behavior, 165-180 and attack behavior, 171-177 and estrogen formation, 88-93, 99 and ovulation, 87-91 and pain pattern, 116 and progesterone formation, 88-93,99

Anesthesia, level, and blood pressure, conditioned response, 205-210 Anticholinergic drugs, and behavior, 393-395. Aphagia, and hypothalamus, lesions, 1, 2 Arousal, and hippocampus, theta-rhythm, 415419 Attack behavior, suppression, and amygdala, lesions, 171-175, 177-179 Attention, cerebral processes, and hippocampus, 242,243 and hippocampus, 228-243 Atropine, and behavior, 393-395 Auditory responses, and hippocampus, theta-rhythm, 422427 Avoidance, conditionedreflex, and limbic system, 247,248 Awake-sleep cycle, and cholinergic agonists-antagonists,341-35 1 and cholinergic antagonists, 340, 341 Baroceptor reflex, and septa1 area, stimulation, 195, 196, 203 Behavior, affective -, and brain stem, stimulation, 103-125 and forebrain, stimulation, 103-125 aggressive -, and amygdala, ablation, 165-180 and amygdala, stimulation, 165-180 and drugs, 365-385 scoring sheet, 367 and sinomenhe, 365-385 conditioned reflex, and limbic system, 247,248 and SHR, 221-225 discriminative -, and EEG, patterns, 235,237 and hippocampus, EEG activity, 235-239 and impedance responses, 238-242 efferent control, and limbic system, 318-335 free -, and limbic system, 48-66 and hippocampus,

SUBJECT I N D E X

EEG activity, 229-231 electrical stimulation, 435-437 frequency potentiation, 400, 401, 410 hypoxia, 466, 467 theta-rhythm, suppression, 433-435 inhibition, and sleep induction, 34-47 mechanisms, coordination, 64 and hippocampus, slow-wave activity, 218227 and limbic system, 295 microphysiology, 48-50 orientating reaction, and SHR, 219-222 orienting -, and EEG patterns, 235, 237 and hallucinogenic agents, 235 and psychomimetic drugs, 234-236 reinforcement, and limbic system, 144-163 classical conditioning, 151-153 correlated behavior, 161-163 covariation, 153, 154, 156 histological findings, 149 interspike intervals, 156-158 operant conditioning, 153-155 special tests, 157-161 spike patterns, 149, 150 variability, 15&152 responses, and brain structure, 4 0 W 1 2 and hippocampus, theta-rhythm, 445 sequential -, and limbic system, stimulation, 65 triggering area, specificity, 64, 65 sensory stimulation, 128, 129 and SHR, 218-220 switch-off -, and amygdala, after-discharges, 134, 135, 139, 140, 142 and cingulate, after-discharges, 137, 139 facilitation, 128-142 and hippocampus, after-discharge, 133, 134 139, 141 and hypothalamus, after-discharge, 138, 139 and limbic after-discharge, 132-139 stimulation, 129-132 inhibition, 128-142 mesencephalon, and limbic after-discharge, 139-141 stimulation, 129-132 trigger mechanisms, and hypothalamus, 64 Blood pressure, conditioned response, level of anesthesia, 205-210 and subcortical structures, 210

485

fall, conditioning, and septal area, stimulation, 205-210 and septal area, stimulation, 189-205 Blood sugar, and limbic system, 296 Brain, septal area, and behavioral functions, 189-215 lesion, and sodium chloride intake, 21&215 and water intake, 210-215 and neurovegetative function, 189-215 stimulation, and baroceptor reflex, 195, 196,203 and blood pressure, 189-205 fall, conditioning, 205-210 bradycardia, 203,204 hypertensive effects, 196, 197, 204 localization, 197-201 prolonged duration, 204 and respiration, 189-205 visceral cortex, interoceptive conditioned reflexes, 293, 294 Brain stem, electrical stimulation, and affective behavior, 103-125 and flight pattern, 105 flight zones, 116122 threat zones, 116-122 Bardycardia, and septal area, stimulation, 203, 204 Cardiac response, and sound stimulation, 314, 315 Cerebellum, activity, and hippocampus, 254-279 and limbic system, 254-279 click stimulation, response, 258,275, evoked responses, latency, 257, 258, 263 and hippocampus, 265-269 and light flash, 267-269,273 and spike latency, 276-279 and hippocampus, stimulation, 261-265, 272, 273 latency, 274-279 light stimulation, responses, 256-274 sensory interaction, mechanisms, 274-279 stimulation, intramodal interaction, 258-261, 273 Cholinergic agonists-antagonists, and awake-sleep cycle, 340-351 Cholinergic mechanisms, and limbic system, 337-362 and neocortex, 337-362 Conditioned reflex, blood pressure, and subcortical structures, 210

486

SUBJECT I N D E X

Cortex, evoked responses, and amygdala, 269-272 and hippocampus, 265-269 Discrimination, rate, and hippocampus, 307,308 reversal, and hippocampus, lesions, 357459 Feeding, and hypothalamus, activity, 1-33 dual function, 1 neuronal mechanisms, 1-33 Feeding center, and hypothalamus, lateral area, 1,2 Flight behavior, active areas, 116-122 adaptation, and environmental conditions, 112-114 and cardiac activity, 106-1 11 and postural tonus, 106-1 11 Flight pattern, and brain level, 105, 111 Flight responses, and amygdala, zones, 119-122 Food intake, and hypothalamus, lesions, 1, 2 Forebrain, electrical stimulation, and affective behavior, 103-125 and mewing, 114,122,123 and threat pattern, 103-105 flight zones, 116-122 threat zones, 116-122 Fornix, radio stimulation, evoked effects, 57 responses, 57-60 stimulation, responses, central integration, 65 Frequency, potentiation, and hippocampus, 400-41 1 cell discharges, 407-409 mechanism, 404,405 recurrent inhibition, 411 time course, 4 0 5 4 7 Habituation, and hippocampus, 329-333 Hemicholinium, and acetylcholine levels, 354 and limbic system, 351-354 and neocortex, 351-354 Hippocampus, ablation, and alternation learning, 311

and cardiac response, 314, 315 and discrimination rate, 307, 308 and disinhibition, 307 and learning, 305-3 16 flexibility, 312 and performance, 308, 309 and response alternation, 312, 313 activity, and labyrinth, stimulation, 193-188 after-discharge, and chlorpromazine, 186 and labyrinth, stimulation, 185-187 and spontaneous acitivity, 442-446 and attention, 228-243 cerebral processes, 242,243 cell discharges, and frequency potentiation, 407409 and cerebellum activity, 254-279 evoked responses, 265-269,273 and cortical activity, 254-279 discharges, and labyrinth, stimulation, 185 EEG activity, and behavior, 229-231 and discriminative behavior, 235,237 and estrus cycle, 76-81 and hallucinogenic agents, 235 impedance responses, 238-242 and orientating behavior, 235, 237 and ovulation, 81-83,98, 99 and performance capability, 232 and psychomimetic agents, 234-236 and seizure discharge, 232 and sex hormones, 71-76 slow-wave -, 218-227 and subthalamus, lesions, 232-234 electrical activity, regional aspects, 229-234 electrical stimulation, behavioral effects, 435-437 and conditioned reflex, 248-251 and estrus cycle, ARC potentials, 78-80 frequency potentiation, 400411 and behavior, 400,401,410 and learning, 400,410 mechanism, 404,405 pathways, 402,403 recordings, 401,402 and recurrent inhibition, 411 synaptic activity, 403,404 time course, 405407 function, and endocrine regulation, 72-14 properties, 442460 and habituation, 329-333 hypoxia, and behavior, 466,467 and lesions, histology, 468

SUBJECT INDEX

neonatal period, 469471 impedance measurement, and learning, 237-239 isohippocampal rhythm, see IHR and learning, 228-243 cerebral processes, 242, 243 lesion, and discrimination, reversal, 457-459 and hypoxia, 462-474 and learning set, performance, 457459 potential responses, and single unit activities, 4 4 W 5 2 and reticular formation, interaction, 43-32, 436

slow rhythm, and behavior, 218-220 and orienting reaction, 219-222 slow-wave, activity, and behavior, 218-227 stimulation, and amygdala, activity, 81, 85, 99 and estrogen formation, 84-87, 92, 93 and ovulation, 84-86, 99 photic responses, 447 and progesterone formation, 84-87, 92, 93, 99

projection pathways, 450,451 subcortical effects, 452457 theta-rhythm, and arousal, 415419 and auditory responses, 422427 behavioral responses, 445 functional significance, 413-437 and hypoxia, 465, 466 and neocortex, 419-425 and reticular formation, 41W19, 436 suppression, and activated sleep, 433-435 behavioral effects, 433435 and thalamus, 425430 theta-waves, and cortex, 281-291 and IHR, 282-290 HVM activity, and anesthesia, 3-6, 27-29 anesthesia, and discharge pattern, 4-6,27-29 electrical stimulation and SUDs, 14-25 latency, 18-24 and feeding center, 1, 2 and LH activity, 3-16 crosscorrelation, 6-14$ 28, 29 SUDs (spontaneous unitary discharges), and anesthesia, depth, 3-6 Hyperphagia, and amygdala, lesions, 1, 2 and hypothalamus, lesions, 1, 2

487

Hypotension, conditioned -, and septa1 area, stimulation, 205-210 Hypothalamus, and amygdala, functional interaction, 16-21, 29 electrical stimulation, and conditioning responses, 56, 57, 62 evoked effects, 55, 56 and pupillary response, 52-57 EEG activity, and estrus cycle, 76-81 lesions, and aphagia, 1, 2 and hyperphagia, 1,2 radio stimulation, and pupillary response, 52-57 and sleep induction, 35, 37 stimulation, and attack behavior, latency, 172 Hypoxia, and hippocampus, reactions, 462474 theta-rhythm, 465466 IHR and brain stem, stimulation, 282 cortical -, EEG, frequency analysis, 282, 283 general features, 282-284 and hippocampus, synchrony, 282-284 induction, 284, 285, 290 cortical distribution, 284, 285 and cortical excitability, 295-297 and hippocampus, theta-waves, 282-290 Inhibition, collatera type, and limbic system, 319-323 loss, and hippocampus, ablation, 301 neural -, efferent control, and limbic system, 318-335 recurrent type, and limbic system, 319-324 Labyrinth, stimulation, and hippocampus, activity, 183-188 after-discharge, 185-1 87 discharges, 185 and optic nystagmus, 183, 184 Learning, cerebral processes, and hippocampus, 242,243 and hippocampus, 228-243 frequency potentiation, 400,410 impedance measurements, 237-239 studies, 305-316

488

SUBJECT INDEX

Learning set, performance, and hippocampus, lesion, 457-459 LH, activity, and anesthesia, 3-6,27-29 anesthesia, and discharge pattern, 4-6,27-29 electrical stimulation, and SUDS, 14-25 latency, 18-24 and feeding center, 1 , 2 and HVM, activity, 3-16 crosscorrelation, 6-14, 28, 29 SUDS, and anesthesia, depth, 3-6 Limbic system, activation, and neurovegetative conditioning, 205 and avoidance, conditioned reflex, 247, 248 and behavior, conditioned reflex, 247, 248 efferent control, 318-335 free -, 48-66 mechanisms, 295 recordings, 50-52 and reticular formation, 295 and behavioral functions, 189-215 and behavioral reinforcement, 144-163 classical conditioning, 151-153 correlated behaviors, 161-163 covariation, 153-156 histological findings, 149 interspike intervals, 156458 operant conditioning, 153-1 55 special tests, 157-161 spike patterns, 149, 150 studies, 144-148 variability, 150-152 and blood sugar, studies, 296 and cerebellar activity, 254-279 cholinergic mechanism, 337-362 and conditioning motivation, 246-252 cortex, conditions, 293-302 functional significance, 295 and respiratory system, 296,297 visceral analyzers, 293-302 and cortical activity, 254-279 EEG, and chemical concentration, 297,298 EEG activity, and estrus cycle, 76-81 and sex hormones, 71-76 electrical activity, and drugs, 365-385 and sinomenine, 365-385 electrical stimulation, experiments, 48,49

and estrus cycle,sARC potential, 76-81 and fiber degeneration, 298-301 function, experiments, 69-71, 98-100 mechanisms, 69-100 neurophysiology, 318-325 gonadal steroids, feedback effect, 71-81 and hemicholinium, 351-354 inhibition, collateral type, 319-325 neural -, efferent control, 318-335 recurrent type, 319-325 and memory, recent -, 246-252 and neurovegetative functions, 189-21 5 and progesterone feedback, 69-100 radio stimulation, and behavior, 5&52,63 stimulation, and estrogen formation, 81-91 and progesterone formation, 81-91 and sequential behavior, 65 Memory, recent -, and limbic system, 246-252 Midbrain, electrical stimulation, and pain reactions, 115, 116 Motivation, conditioning -, and limbic system, 246-252 Neocortex, cholinergic mechanisms, 337-362 and hemicholinium, 351-354 and hippocampus, theta-rhythm, 419-425 Orienting, regulation, and amygdala, 325-329 Ovulation, and amygdala, electrical stimulation, 87-91 and ARC, electrical stimulation, 89-91, 99, 100 and hippocampus, electrical stimulation, 81-91, 99 Pain perception, and brain level, 116, 124 Pain reaction, patterns, 115, 116 threshold answers, and brain level, 124 Performance, curves, and hippocampus, 308, 309 Progesterone, feedback control,

SUBJECT I N D E X

and amygdala, 69-100 and hippocampus, 69-100 and limbic system, 69-100 Psychomimetic drugs, and orienting behavior, 234, 235 Pupillary response, and hypothalamus, stimulation, 52-57, 62 Reserpine, and behavior, 395, 396 Respiration, and limbic system, cortex, 296, 297 and septa1 area, stimulation, 189-205 Reticular activating system, and sleep induction, experiments, 3440, 46 Reticular formation, electrical stimulation, and conditioned reflex, 248-251 and hippocampus, interaction, 430-433,436 theta-rhythm, 4 1 M 1 9 , 436 Rhinencephalon, activity, and drugs, 388-399 patterns, 389-393 sequential averaging, 390-394 studies, 388-399 and behavior, conditional -, 388-399 Satiety center, and hypothalamus, ventromedial nucleus (HVM), 1, 2 SHR, see Hippocampus, slow rhythm and conditioned reflex, 221-225 and orientating reaction, 219, 220, 222 and sensory information, 226 Sinomenine, effects, and CNS, 365-385 Sleep induction,

and behavior, chronic experiments, 38, 41,42 and conditioning experiments, 38, 41, 43 electrophysiological experiments, 34-37 and hypothalamus, 35, 37 mechanisms, and behavioral inhibition, 34-47 ergotropic zone, 34, 35 trophotropic zone, 34, 35 pathways, 4 4 4 7 and reticular activating system, experiments, 34-40,46 Spontaneous unitary discharge, HVM, and anesthesia, 3-6 latency, and HVM, electrical stimulation, 18-24 and LH, electrical stimulation, 18-24 LH, and anesthesia, depth, 3-6 SUD, see spontaneous unitary discharge Thalamus, and hippocampus, theta rhythm, 425-430 Threat behavior, active areas, topography, 116122 adaptation, and environmental conditions, 112-1 14 and cardiac activity, 106111 and postural tonus, 106111 Threat responses, and amygdala, zones, 119-122 Threat pattern, and brain level, 103-105 Visceral analyzers, cortex, localization, 293, 294 limbic system, studies, 295-302 subcortical formations, 295-302

489