Hypothalamic substrates for brain stimulation-induced attack, teeth-chattering and social grooming in the rat

Brain Research, 449 (1988) 311- 327

311

Elsevier

BRE 13~46

Hypothalamic substrates for brain stimulation-induced attack, teeth-chattering and social grooming in the rat J.H.C.M. Lammers, M.R. Kruk, W. Meelis a n d A . M . v a n d e r Poel Eepurtment c,f Pl2armacology of the University of Leiden, Sylvtus Laboratories. Leiden (The Netherlands) (Accepted 10 November 1987)

Key words: Rat; Hypothalamus: Aggression; Teeth-chatterir, g; Socmlgrooming; Ele,-'rlcal brain s~:lmulalton: Discrimmant analysts, Mapping

In this paper the boundaries of the hypothalamtc response areas [or brain stimulation-induced attack, social grooming and teethchattering were delimited A total of 641 hypothalamic sites in 71 male CPW/WU Wistar rats were electrically stimulated. Positive sites for any behavloural response cluster into restricted hypothalamtc areas. Dtscriminant analysis o( both pos,live and negative electrode Iocalizationsyields areas with high, intermediate and low probabdities of inducing the behavioural response concerned. Each response has its own response area where probabihttes are high Neuroanatomical correlates oF these response areas are d~scussed.The response area of attack ts ~aggested to be an integrative processing area, stimulation of which overrules some aspects of integration and directly activates the behavloural program of attack. Although some authors consider all three responses to be part of the behavioural repertoire of aggression, the response areas are not identical. Social grooming and attack are considered to be induced from different neural systems Simda, ly, attack and teeth-chattering have been shown to derive from different neural mechanisms, despite substantial overlap of both response areas. It is suggested that teeth-chattering derives from the simultaneous activation o( both attack and flight tendencies No further dlstinctit~n~with respect to threshold current intensities can be made within response areas. However, the underlying neural substrates are not homogeneous, for thresholds vary along the course of individualelectrodes.

INTRODUCTION Brain stimulation methods have widely been used to investigate the f u n c a o n a l organization of the brain. Since the pioneering work of Hess and Briigger 19 the brain, in particular the hypothalamus and brainstem, has been extensively explored z ' 32,34,40,4..,50,53. Despite the effort and the encouraging results, generally, however, the way electrical stimulation was applied has been shown to be too gross to analyze the functional organization of the brain in sufficient detail. Thick (often monopolar) electrodes, stimulating relatively large areas, prevented precise localization of the neural systems revolved in particular behavioural responses. The rapid development of modern neurochemical and neuroimmunological techntques during the past decade has enormously increased our knowledge of n e u r o a n a t o m y

and neurochemistry. Modern high resolution neuroanatomical brain maps illustrate the extreme complexity of the brain. Indeed the classical stimulation techniques were insufficient to match this high resolution. Today electrical stimulation is mainly used in other contexts. Behavioural responses can be asso~_'iated ~ activation of specific brain areas by combining electrical stimulation and injection of [~4C]deoxyglucose -~. Electrical stimulation can also be used as a tool to study the effect of physical or pharmaco!ogtcal bram manipulation. For example drug effects can be studied on reliably induced and stable responses 2~-47. lnterestingl3 the pharmacology of behavioural responses from the hypothalamus is rather different from comparable responses m spontaneous behavtour2s. Therefore tt seems worthwhile to determine from which neural system thesc responses derive,

Correspondence. J H C M Lammers, Department of Pharmacology, University of Leiden, P O Box 9503, 23i~) RA Leiden. The Netherlands. 0006-8993/88f$03.50© 1988 Elsevier Science Publishers B.V. (Biomedical Dwlston)

312 Comparison of the exact localization of the area from which a response can be induced with known distributions of neural elements may provide information about the neural system involved in a specific response. Although a number of elaborate mapping studies have been performed 3"32"34"4°'53,this coupling of a function to a substrate has not been successful for many functions. Why former studies were limited in this respect has been extensively discussed in previous papers 29"3~. We have tried to overcome the disadvantages of these studies by refining and combining a number of techniques: stimulation via thin bipolar moveable electrodes; testing subjects in an enriched environment in which animals can display most of their behavioural repertoire; discriminant analysis as a formal method to process the behavioural data; a detailed atlas of the hypothalamus to depict the results. In fact our methods are 'classical', for Hess and Brtigger 19 used essentially the same (systematic exploration of the diencephalon, induction of as many as possible responses, analysis of both positive and negative electrode Iocalizations) for the same purpose (to relate a function to a substrate). This paper is part of an extensive study in which we tried to determine the hypothalamic substrates for as many behavioural responses as we could regularly evoke by electrical brain stimulation. In previous papers 3°'31 we described response areas and discussed putative neural substrates for digging, grooming, circling, locomotion and escape jump. This last paper of the series deals with intraspecific attack behaviour and two other responses usually associated with aggression: social grooming and teeth-chattering. • . ~ . . . . . . . . . . . . v,ou, has been induced by electrical stimulation in the hypothalamus of, among others, the cat 19,37,41,51, rat 1,22,23,27,38,49,53, opossum 40, guinea pig34, dog 14 and marmoset 32. In earlier papers 25'29, based on a large number of electrode Iocalizations we described the hypothalamic response area for attack, which is the area from which attack can be induced with high probability. It was demmstrated that the hypothalamic response area for attack behaviour did not coincide with classical neuroanatomical subdivisions, thus stressmg the need for a greater resolution in delimiting functional areas. Attack responses could be evoked from the anterior hypothalamus frontal to the ventromedial

nucleus, the central part of the ventromedial nucleus and its dorsolateral capsule, the perifornical hypothalamus and the adjacent part of the lateral hypothalamus. However, the exact boundaries of the response area could not be delimited because only a part of the hypothalamic area was studied. In this study we tried to evenly distribute the electrodes over the hypothalamus. Teeth-chattering is considered to be part of the aggressive repertoire 2.4s, in parhcular offensive behaviour 6. However, teeth-chattering can also be heard in other disturbing situations. Therefore, it could also be a distress signal outside the social context. Electrically induced attack behaviour is frequently accompanied by teeth-chattering 27"3s'53. Electrically induced teeth-chattering is more often associated with attack and alarm than with any other response 53. The question arises whether or not attack and teeth-chattering are mediated by a single neural mechanism. As far as we know, social grooming induced from the hypothalamus has not been reported before. In spontaneous behaviour grooming of the partner has been considered as part of the aggressive repertoire. Some studies 1a,45 distinguished between social grooming and, as a development of it, aggressive grooming. According to Barnett 2 social grooming is seen mainly in conflict situations, but it is not confreed to them. Obviously the relation of social grooming to aggression is not quite well established. The response areas for attack, social grooming and teeth-chattering are defined by discriminant analysis of a large group of electrode placements distributed over the entire hypothalamus. Neuroanatomical correlates are discussed. All three responses appear to have different, though partially overlapping, response areas. Relations between these and other responses are discussed in this perspective. MATERIALS AND METHODS Materials and methods applied in this study are essentially the same as described in the previous paper in this volume 3°, and in others 27"29,31. Additional details are given below.

Behavioural tesung and threshold determination Rats were tested in an enriched environment con-

313 taining a variety of goal objects: sawdust on the floor, food pellets and wooden blocks of equal size scattered around, water bottles in both back corners, some hay as nesting material, a plastic Petri dish and a naive male partner. Methods of testing and threshold determination have been descnbed in a previous paper 31. Fig. 1 of that paper clearly illustrates these experimental procedures. Threshold current intensities of evoked responses were determined using the up-and-down method of Dixon and Mood tl, as modified by Wetheril152. During threshold determination a round Plexiglas cage with a 40 cm diameter was placed in the test cage, in order to keep the animals in the vicinity of the microphones. When behavio,tral responses were obtained for the first time along an electrode track, at least 5 thresholds were determined on consecutive days until threshold currents stabilized. At subsequent locations at least 3 thresholds were obtained. For every behavioural response on every location a mean threshold value was computed from the last 3 threshold determinations. Sonic as well as ultrasonic vocalizations produced by stimulated rats and partners were detected. Sonics were detected using a standard set of sound recording equipment. Ultrasound was detected by a Bruel & Kjaer type 4135 condensor microphone and amplified by a type 2633 preamplifier connected to a type 2610 measuring amplifier. The frequency range was set between 15 and 100 kHz using a programmable dual filter (Dlfa PDF3700B). After filtering tl:e signal was made audible by a 10-fold frequency dwision. Behavioural criteria If during threshold determination an' ~tttack behaviour or social grooming towards the natve partner was observed, or any teeth-chattering was heard, within the 10 s stimulatmn period the trial was scored positive. Attacks were distinguished into the following categories27: clinch fight, .lump attack, severe bite, gentle bite, skin pulling. These categories are not mutually exclusive, but compose a continuum of aggressive responses zS. In addition, the body region (head, neck, back, belly) first b~tten by the sumulated rat was recorded.

been elaborately described by Kruk et al. ~'27. The attack patterns resemble those observed in spontaneous aggression 6. Within the response area for attack (see below), site dependent differences in the attack response were observed. Although not quantitatively determined, the attack response gradually changes over a mediolateral continuum. Upon stimulation of lateral sites the rat moves around in search for its opponent and attacks without hesitation. Attacks are often directed to the back of the partner, but also to the head. At medial sites the stimulated animal does not look for its opponent, but when confronted to it a fierce attack is launched. At these sites more vigorous forms of attack, like attack jumps, are seen at threshold current intensity. They are mostly directed to the head and neck region of the opponent. Electrically induced social grooming appears similar to naturally occurring social t8:5 or mutual: grooming. The rat licks and chews the fur of any part of the skin of the partner, mostly with its forepaws placed on the back or the neck of the partner. Skin pulling, as part of aggressive grooming TM is constdered to be an aggressive act and is excluded from soctal grooming. Stimulation-induced teeth-chattering resembles spontaneous teeth-chattering-' 45. The intensity varies depending on the site of stimulation. Sometimes it is clearly audtble, sometimes it can only be heard through the recording equipment. At termination of the stimulation all 3 responses immediately stopped. Hardly any vocalizations produced by the experimental animal were detected during stimulation. Loud vocalizations were produced by only two animals, 3 electrode sites being located at the mediolateral junction (frontal 8.25 ram), and one site located near the opttc tract. Ultrasonic vocalizations by the stimulated rats were only detected after convulsions which occasionally emerged from stimulation. These ammals produced long 22 kHz calls for some minutes after the convulsion ceased, whde the animals did not move. Partners produced considerable numbers of 22 kHz bouts after being defeated. However, these vocalizations did not restrain the stimulated animals from attacking over again.

RESULTS Behavioural responses The topology of elecmcally reduced attacks has

Response areas In 71 animals a total number of 641 electrode local-

s.o

,

. -L-,..~-TqL-----

]];.

I-

'

'

'

'

:

, .-

,

.,

. ,.

Z.O

]

L;

1.0

~.0

F]

'"

''"

3.0 Z.O

5,0 iiii~ 4.0 •

i

,

_

,,/

3.~ ~

'

;m

Z.O

m

1.6

Z,.D

O O

1.0

Z,O

0.o

I 0

[' fl

0.0

I..

~.0

O.O

1.0

~' 0

315 izations were tested (Fig. l). Attack behaviour (squares) was evoked from 48 electrode locations, 39 sites yielded social grooming (triangles), while at 54 locations teeth-chattering (diamonds) was induced. Only sites at which a threshold current intensity could be determined for a specific response were considered positive. Sites yielding n o n e of the responses are indicated by small plus signs. As can be se,.n from Fig. 1, and from correspondmg figures in previous papers 3°31, different responses can rehably be induced at the same electrode site, depending on the stimulation current intenstty and the presence of adequate external sttmuh Dlscrtminant analysis of both posttive and negative locations provides areas with high, intermediate or low probabilities of ehcitmg each behavioural response (Figs. 2-4). Black, heavy and light squares depict high, intermediate and low probabilities respectively. The area with high probability grid points is considered to be the response area for the behavtoural response concerned. No symbol is drawn at grid points with too low density for any group. Grid points computed outside the brain, or located in the third ventricle or the optic tract are functionally insignificant. Fig. 2 shows the response area for attack. It is located in the anteroposterlor midsection of the hypothalamus, ventral to the fornix. The anterior part of the response area is extended to the hypothalamic midline. Somewhat posterior the response area comes to occupy part of the far lateral hypothalamus_ The attack area does not seem to be hmited to a specific, well-defined nucleus or area. Attack sit,"-s are located in the medial as well as in the lateral hypothalamus, partly inside and outside the ventromedial hypothalamlc nucleus. For an analysts to he valid the area with high posteric probabilities for a response should contain a substantial number of placements mducmg that response. The results of allocation for attack are giveo in Table I. There is a significant departure from independence (Z2 = 367, P < 0.001, df = 3). Signtflcantb

TABLE 1 Allocation of attack and non-attack sites

~; = 367, d[= 3. P < 0 00l. Posterior probabrht.v for the reduction elattack

Actual behaviour induced No attack

Attack

Probabthty < 02 Probabthty > 0.8 02 ~~0.8 Too low density

497* 27 53 16

l 46** 1 0

Total per group

593

48

*

** Significantcontribution to departure from independence: adjusted residualsof cells "13 08, **19 15; * **r'< 0.00l.

more placements than expected are correctly allocated to attack and non-attack groups (adjusted residuals: 13.08, 19.15; P < 0.001). The response area for social grooming is shown in Fig. 3. High probability grid points are found over the e~ltire anteroposterior extent of the hypothalamus. The response area is located almost entirely in the lateral hypothalamus. Some electrode sites are undoubtedly located in the medial foreorain bundle. One positive site was located in the optic tract (frontal 8.1 mm), preceded dorsally by two negative sites. The functional significance of this site is questiona-

TABLE I! AIlot llflOII Ol ~01"1[11groolllll|[~ alld tlon-~ot'tal-groommg slit's

Z~= 189.01 = 3, P < I | 1~1 Posterior probablhty for the mducnon of soctal groontmg

Actual bchavmur reduced

Probability < 11.2 Probabd~tv > 0 8 0 2 ~(I 8 Too low density

437* 39 112 14

0 29** 10 0

Total per group

6112

39

No groommg Grooming

* ** Significantcontribution to departure from independence; adju,,ted residuals of ~ells- *9 43. ** 13_34. " **P < 0.001

I_

Fig I Electrode loc,thzatto,ls plotted m the aria,, of the h.vpothalamus Squares, site,, at which attack ~,ls induced, triangles, site', for so,AM grooming, dlam~mds,sites that yielded teeth-chattering, small plus signs, non-responding st'.es Ior an~ el the responses. ,,mall symbuls, high thresholds: large s.',ntbols, low threqtolds, filled synlbol,,, sites with the Iov.cst threshold ,,aluc m ,in electrode track. Arrows indicate bcgm and end el part ~,ol electrode t~,tcks located on frontal 6 6 ram. but for the purpose el ilhl,,traltondraw n on Irontal 6 75 II"1111 d r . : t r c u a t e I1Ll¢tel.IS, Cal, n l t e r n a [ c a p s u l e , l, f o r n l x , hpv. paraventrlcular n t i c i e u s , h d . dorsomcdtal h.vpotl't,ilatrttc nuclei.P.,. hvm, vcn,, omedtal hx poihdlanlic nucleus, to. optic tracl, v. third ventricle

-'C ~-7

~;-'

i

_ 2C0~_-Z_2~C2C ~O00OorqoooooOOO

~.r.

r

...... , ~ m ~ , ~ - T~BNWBDW(3g,qWccD ,~'~F~E,O~o~r~

L

O

i

C I"

_Kooomoom~oooo_

i

m =-o~tc'< 9 -,

1

I

,-<

,

l,

s.,

\

,

k

~~ ' LW~BW~,BWWWDO ' OO~PC20C~O0~OO j ~O~OOOOOOOOOOO ~2ooor~o000000

0

I

I

~COD~'

Frontot

~

95

. . . . . . . .

. . . .

+

I

I

I

l--

DOODI~OO~E OOOOO~O~

II

;ooc

J O O O O O O~;~ZlO~F--DO'C]O,',Oooo

i

I

I

I'i///L /

Front,

.

.

.....

i

i

B.~5

u

i

.

.

t 5 . 5 5 m,

.

,'

OOOOOOJ~ ODOOU~nODDDI~D/" DODOO(~ODODOE30~O DDOOOronnnnooODO OO00OO~[~nooq~oo DE]O.ODDODDOOO01~]O

-

OO00FI]]~DOOI~}~]OOO

oooooVoo.o~anoDn OOOOOOOOOOiOnool~ DDOO[~I~OOOO~3DOOF~

0ooOOOlllmOOnOO

Frontol

i

,m

.

I~W~Biiiboo_~

Frontot frontol B.t, U.~ mm u

.

OOOO0~O0~0OOD~OnL OOl~10~3 O l ~ O o o n l ~

DOOOOO~O0 Co ]ooooomn~r~co3 ~ooann=u

i

, 5., .

~O~O0

I~nnmmlmmmmDno/----iDe~Diillilll llnOllllliHl_.~

1,0

I

OOOOOO00OOO OO00OL~OOOOOC~DOO

IDOODODOOO~OL-'~r~CF~ "I[lD O D D ~ E ]] O~~ O O ~ L~G

113DDOOOO[3DODODO 1 ~DODOOOODDDOD iI D O D D O D D m D O D O D WnDOnOOHm~oo

~ommllaaooo

/

OOOOmOOOOO DOODOI~F~DO~

1100ooonDooonoo0

OOO~OOO0

w ~ u u m m m O O ~ O O O O

omnp~em~o.

)

7 OOOO~1~O00~OO 0 ~ 0

l

~DOO

1-101"I O O E 3 O O O _ 0 0 D

aoqo

f ~

1

~l O O O O O ~ O O 0 1 O O O O 0 ll o o 0 ~ o o o n o II D D D ~ D ~ D D D [ 3 m D D ~ J 13OOO~OC~OO00OOOO, 1I D O O O ~ O n o o o o o

" ~OOOOOOO~OOOOO qOOO OOODOOOOO E~O 00,_-I uOO~oormo --

~O~C--CO,~OO-~CCC DOIDOOOOE0rmDoooo O[3[~OOF~O~OOO00 'OOOOOOqOOnOOO

L o~oooomnoÙoooo

r

}

1300000E~OOODOOD~ I~C3OOODO~DOOOO

rr,o C o ~ O O ~ - 2 c c - ~

DOD00~OIOOOOOOOQ

l

I ~|

l

OOE]~D[~D~DDODOOCI~ DOIODODI~DnODODO 3OOOOO01~l~OOOOOO ~OOOOa~g~DODO

OO~OOE~l~O~nOO000 OOOOOOl3BiaOOO00 ooo

i-

o...oooooo_

)Billllllllb l~mlliliilii l~Ollllll!lll

m~

FronLaI

i

I

5.1

I

mm

1

FronLoi 7.g5 mm i

l

i

i

i

8.0

J

3.0

'

/

001300 / / DODDDDOI30 I mnOODO~OOO~]2~O I / DODDDD~ODDCI~I~On I -40nDoonnGLODOO~OD L I Dnooo~3~)E~onrm 1 L-~OO~]GO OIEE]t3O O,E]OC~ I II 3DOD~D~DODO[~LOO ] -I 3 n o . c I G q U D O D O O D D ~ - ~ ~ODOI~O~]DDODnO DF~DImDDD~OOODDO 3D~QOOIINO[3Don ~no,',-mmmmmmm~

3~OOililiiiHO .~Oilllli~30

2.0

Front~l 7.5 z,0

"i

5.0

4.6

3.0

2.0

'

~O~I:3iBH6HBH~O~,~ ~I3 D CIm U munrom.'nD G~,O m . L ~ n l m mmm I ~ n

mm

I

i

i

i

i

I

I

I

I

I

OOODDDD[S]DD nDDDj0000~DO . -" DDO~O~OOOODI~" / ODDDOIDOOOUODO/ DDD~DnDO~3~ ,DDO0.(~ ~O L~DrlomO~E3niDrm InnDD~]tDOOODI-I~O lnrqrqDoonnoÙo[~O D D O D D n D D D m , CDD~O

i

I

Frontal

7.35

mm

]-- .... /

I

I

OD'

OOOOOOOOOOOOEl~

~i

3noo~ooooo~o~no 3PC]On D O ~ ~DC~Oorm ~[3.01~[~0 D O D O q O ~ .'mO O,0 n O n D O Dlm_qlDO 0 3OODODOOIODOE~3mO D O rmD C ~ C ~ O O O O D O 1 ~ ormooODIO0OonoDooo/

lnDOrln~nDnnDn InnDDOE~ODDDDD / ~I~oDDnDDDDm / IODOO13ooonDno~.~ ]Olil~ODD~ ;tomminuiinD~ ~,l~nm e n ~ D [ 2 1 ~ o

~ r q 0 o o onrqoooo,~ ~D1~r'Im~nDonO[3~O DCIODDr~oD(3~O D~OnDolmnD~ I

Fron[aL 7.2 mm I.(3

't

DODOD[~onDn /I D D D D D D D ~ [ D D D D ~ " JnDDQIDDIDDDDE~ ~/ rmDDO~[~O~DDDn D~3" nnODDDDOODO~rm rCIDDQ]0QonOLoDrm DI~O ]~3~-OD D O 0 E~O D D DO~O ]OE]/~O~DDnDD OO~ ~DDODDnOmDDDDDO I~ D O D D D E ~ O O O O D O m ~DODDD~n[~DonnoD I~I~ODDDDnDOOrmoooo ~IDDDDDDOI31~mG D D O D / 13/~Dn DI~ 13013r~130r',DO / ~DODmmOOnOODDD/

Frontal 6.g mm

i E

o.o

i.o

i

i

z o

I

o o

1.o

z.o

o.o

i

l.o

I

i

~ o

o.o

i.o

~.n

F=g 2. Representatmon of the posterior probabilit=es (PP) for the mnduct,on of attack responses, according to allocauon with d n u b t Black square,,. PP > (I.8: hcavv ~,quares 0 2 ~ PP <~11.8, light squares, PP < 0 2 G r i d pomnts wroth too low d e n s i t i e s are not p l o t t e d

,,

,

, ~\

\ - ' _,

,'I

~

',

'

~

'

n

~BOOOODOOOOIODCC ~OOOOOOOOOIIDCO ~OOOODCDOIIIOCC

~OOOOOOOO IIOOO q~OOOOOOOO IIOOO l~OOOOOOOOI~IOOO I~OODDOOOOIIIOO0

s.0

I

e'

From

~ ]233C~C00~03C.~ w~TZE22Z~ODIIDCO

,

/ ]

r i ,

I~OO_EOE_OIIIOOC ~zoso]oIIIOOC ~Z,Z~COC~C~OCO! ~G~OOC~OO IIOOO

~

]2-~OCE '~llOO ~WSm-ooq~amlOO '~OCZ~ZZ~O~B~ ~O~COO~O~D.

G~ZZZZZZ_.ZOIIOC] ~ l~CO~cocmomm~ooc

nuoool

~OODODC00

~

s~mscooooc r

' [

mm

F-n-'e

i

t

1.0

°.~5

-~

~

' I~CZZZD~O~OOCO 1000£oooOollooo

i~CZZ2Z~ZO~O3 llOO~oooczonmoo

~-

I

ro~t

~

,

~ r

,

nm

S.

,

, p I I

~ ~ooo ..... ' ~OCC 00~0~003 ]_~OOOO~OOOOO ~_]] CZC000IO0 I~0~ 30~OOOlOO i~OOE_O00~O010 ~¢oozsoooOOII I~DOCDOOO~OII ~OOOOOOOOOI~-

, I I

] r / i

i

7

i

.

.

.

1

.

5.0 ODD

O

DD~D

DODqDDDDD~OD

"

DDDDDD~

""

40

3.0 300DDOD~]O0 DODDDD~n~o

\

DDSODDDDDmm~ IDODDODDO~IO )DDDDOOOD~I~

~

3DDDDDDDDI~O~O ~OOOI~ODDDW~DD DGD~DDDDD~O O O D D D O D D ~

~

2.0 DO FronteI 1.0

I

DO B

me

(

I

~

fronlel I

--

O000DO~DDQ~ O~DDDODOOO~OO~D

~.0

I

I

7.g5

mm

I

I

OODDO£~3ODDDD~ DODD~QDDDDG~D

DDDDC~D~DC~CC~ F~OB[~]tOIWD~DOOD ~D~OWIU~DO00

Q~OOD~O~ODCD~O O0~IODODO~ ~DDDDDBIIODODDO ~DDDOIIWIIODDOD

]OO.~O~IIIIIOOIIO

~O~OUIImOOOOO/

~ODODDIIII~DDOD

3~O~OOII,III~II~/ 3~Oo00IOn~UlOO~/

~OOOOOI!IIIIOIlO, BOODDDIW~II~O/

~OOO~OIIIIIIODI OO000OIIDOIIOO/

~O~OQDD~QDO~-~

)~0DDDODOOODJ~.~-~

~ODDOIIDO~D~

3C~000~O~I~OU

i

i

i

i

,

!

" L I / l

OOOOOOO~E~O~O OO~OOOO~O~O O~OO00~OO000 3ODOOOII~UOOL-~ : ]O00OO|H|IIOO0~ 3ooooOiIIIOooo

|

I

2OCX~OOIIIIIIOD/' SO00~II~IIIO~.~OOO~I~Iii~

,,-om'~ ~

~,~

-

T

l

" l

I~

/

~OODDOII~IIO~/

Frontml 7 65 ~m

Frontal 7,5 mm I

q

i

~

I

I

]

l

~

I

I

I

~ & _

--

|

OOOOO00~OOO~OO OO~OOOOIIO~OojqCIO~[~OaOOO~0~ OOODOODIIIIO(3OO OOOO~IIIIOO~, OOOOD~OIIIIDOOO/

Frontal

00

i

7.05 i

1.0

I~OOODDOIIIOIE~ 3OOOODOOIIIt~O 3DDCOODD~I/O

mm

~.~

Fromtsl

i

I

~.0

0.0

OODII~DI~,~",

I

1

l~

i

--

I

;

I

I

I

I

I

DO'

-

OOOO~OO~O~OO )O00~Oa~Ooo]OOOOOOai~00~OO DOE~OOINIIIO~OO Oooo~o~IIIIO~ . ooooo~!IIIIOO~.

i

,

Frontal 7.35 ~m

OOD~C~ODDDDO~30

r

~

~ODOIIDOI~O

I

O000~DGDDOOD ODOCCDDDOOOO~

m~

2~0

I

'

OO'Di~ O I I I m U I O O / ~O000OIIIIIIO~V"-0000OIIl~I/

~0

_

i

I

O~OE~DDODO0 OOOO~ODO~O/

'

FrgrtaI I

ODODODOO3~D~D~ ODDCDDD3~O0~

,o[~NIw~uoqooo

__I

~

,

3~I3,~_~OOIIIIIIII

__

u

] mm

B

i

~oo~Nn~wooono~

Frontal 7.B mm 0

Frontal I

OO~OOI~O~IIDIII/ O0~DOD~O~OIOD~ D D ~ O O ~ I O ~

.

mm

I

DOO~DCDD~O~ODO~ 3DODDODO~DOO~OO 3D~D~0~OOODDO

2~o~o[~lmmwmmm

I

~.Z5

I

O000DC~r]CD~OC O00OGO~CG03~C

DOOE]OOO~OOOOD~ 3ODO~OI~OOOO~OO 3ooIs~S~I~i~OIm[]I~

].o

_

I O 0 0 O D D D D O ~ IOOO OOOO O O O ~ O ~ O O~OO~OOC~OO nor-tan O ~ OD D ~ O oO~ oOu u aoOoOotaIImI~0oo ~ o o O ~ l l ~ O . 8OOOOO~IIIOO~ lOtODO~OOOIIIO0~ OOOOOOO~ ~0

mm

Fronte~

1

1

~0

0.~

~.~5

~

m~

l

1o

~.o

Fig, 3, Represeniationoftheposterlorprobabihiies(PP) ~rtheinductionofsocialgrooming, accordingtoallocati°n wmthdouht. Blacksquares, P P > 0 8 ; h e a v y s q u a r e s , 0 2 ~ PP ~ 0 . 8 , hghtsquares, PP <0.2. G n d p o i m s w , h t o o l o w d e n s t t i e s a r e n o t p l o t t c d

I ~30 ODOE~OOOOD~D, ~moooooooooooooo ~OOOOOO~OOOOOO ~OOOOOOnOOOOOOO ~®ooonoOOooonnoo ! Immmooooooooooo ~~ m i i o O O ~ O O O O O O O O I~ n n n o n o o o o o o o o o

~.0

/ , F

, I h "

Ljmmo=oOOOOOOOOO

~

-°°

3.0

l.O

~-B

3-0

'

OOO [] :L~C~- I ql D O O O - O ~ D O O ~ O O O E ~ I I| O O O ~ O C ~ r = ~ O O O ~ O , l~OOO~O~OOOOOOO~ I 4~ooc~oO~OOO~,Oo..q l~OOOO~OOOOOOO ] I ~OOOOOOF~Or, o o o o [ ~OOOOO~OOO~~ I~ n o o o o n i n ~ n O O O I I:]OoooommDoooc I mg=ommmmmoooo i ~mmmmmmmmK]~DO ~ llmmmmmmm!QnOF_~L~ , lulnuiiulo~o~ \, ~ mmilimii~DD ~ '~

i

i

i

n

i

i

~ ~ ~ Q ~ 8

/

'

'

'~-

~--

n

n

I

',_ " -

I

~OODOO~OOOCJC ~[~G~-OOOO~OOOOO 3 ; #~Ol310rsnnrs~ rsrn~_D 0 :" . ~I °mm~=~~°°°°°~/I o ommm B B ~ / ~

=Immmmummm~o~z~_.~ umlmmmmmmmml~nD

unnmnnun~o~ -m=mmulum[]mm;;~

OOO

i

i

i i

i

3.0

ODDDDDD~DO DDDD.~;~DDOIqnD DDDD~EN3DDODI~" DDDDDDDODD~DDD/ CDDD~DODDC~B~ ODDD~D~Q~D EjE~ODDOQ~ODDD OQODDI~ODDD~D UDDDDDDDDDDDD~D DDODDOODDODDI~O ]DODOD~DDDDDDQ

~- 0

] ~ o o o 0 ]inumnm~l:3,~

......

i

~/I

f

--

i

i

i

i

i

n

n

0.0

i.O

4. Representation squares,

l

PP

i

I

I

I

I

|p

~mmmmno~onO--

::',' /

'

"

"/1 L

O000OO~36 _/ q OOOOU~OOOOO~O/ I DDDDO~ODOOD[~O~O I OOOOOOOOOOOO~OO 4ODDDOO~3BODD~OO kB~ODOODODDO0~3O ~OOOmO~O~ODOOOO0~ ~DOODDO~OOOOOOO ~OOOOOr~OOOOOO II~K]onnoooooonono

/

/

&II~nDoooonmmoODDDD @l~ll01~oommm(]oDDo~

umonoummmmoonoo~ Jinimmnmmoooo~ I~mmRBm'mnmmouO~O ~ H l _mflnil r'n~-...~ iDimmmmmmmm~

I

OO~

'

,

~

I

I

I

ODOOOD~OOOo~OOODDNOOOO.,

OOOOOOOOOOOOOO OOOOOOOOO~OOO~ OOOOOOOO~DOE~O OOOOOOODDOE~OD OOOOOOO~OO~OOO ~E~3~3~O00

OOO~OO~OOmETO/ DDOE~DOODO~ OOODOOOOOO~E~30 3OOOOOOOO~OO~O ~E~30 OOO~OOOO0~o 3OE~3~DOODOOD~

3OO~OOO ~3OOOOOOO~OOE~gE~-~ ~ O ~ o ~ O O O O O O / ~O~o~OOO00~ ~ O O m = ~ O O O O ~ O~ m O ~ O / ~onmmmmmm~ o o o ~

,nnnoo~OOOOO~

~U~L~uunmunnr.~o~ ~Nmmmmmm,=mo~

:-mmmmmmoooo~3

,,~=o.....~.o~

| Jl

I i

I I OOOO~OOOO~OOO I IDDOOO~DOF~D~DOOO _LOOODDE~OOO~ODOO ] I ~ 3 O~D Do E o~ DnD Oo o ooo n o / ~DDE530~3O@ODOOO~ -~ E~3D~OO~OODODD 1:110r ~ O O o O ~ 3 ~ n O O o O O noooOo[~mmmoDnnDo I~mmoooommmmoooooo IIl~ml~mommmmoon nDD mmnmmmnm~onoo~fgilimlilUr'no~ Ogini~O~DO ~mlmminmme~ooo ~

[]

o~o

,

/q oO nO oO oO' oO oO ~O OO' oO oO O E'0~o' ~3OO

L

i ~r~o O m n ~ r~OOO -I~ D i i n m n ~ n o o o I~ m m m m m o ~ o o o o

I

/

, ; ~ ( ' I

I ~0oooooooooooo

q Ooooooo~oooooo

[

I

i

OOOOCOOOnnaO~_

C~OCS~::

_

nooooooo©ooooOC nooooo~O~OOOOOC O~mOO~OOOOOE 3OMr~mm¢~nDOOOOO, -~onmmmmm~noooo/

~.....oono~ :~mmmmiuoou~D

Frontal 7.5 mm I

,

I

Frontal 7.35 mm i

i

i

i

i

i

I

DDD~DDDD I ~DQDDDDBODDDD I I DDDDDD~ODODD -/4 IDDQDDDDDODDD.E~D" J ~DDDD~I~DDDGSE~ JDDDD~D~D13~C~3~O ~DDD~DDDD~3~DD ~D~ODDDDDDDDI~DDD ~CDCII~DGE]ODDDDDC ~I~ODOCDDD~D ~DODDDD~3-[31ODO~D~/ ~ O O ~ O O E ~

Frontal 7.2 mm 1.0

UOO~

Frontal 7.65 mm

n

~

~

I

]nOOOD~DOOODO ]OO0~mi=ono ODD/, °oo°n~mm~oooo~/ mm'~,~mmmmmmmnooG~G~ i~OOOni==OOOOOO

~.o

Black

'

OOOOqOOOOOOOO ~ODODOE~3O~000~ WODDDODOOO~D~D~ I|ODODO~DODOO~O, 4|DODO~DE~OO~O0~00 l~3OOOO~O~O~O~ $~QE~3DDDE~OODODO ~OE~3ODO~DOOOO ~. rmOOoor~n~nooO m~loK]ooooi~t]ooo llllllmmoommommmmmmlooOO I~mmmmmmmmm~DDO Ui=mi~iiin~oo~ lllmmmmmnmu~.ooc~3 @mmmmimmiiiQ~O0 ununmnuu-u~o_---.~

OE~OO~OOOOn%OO

__=sc-~,-zccc:_

!i

Front@l 7.@ mm

Fig

,

"

-

~---m~-~ ..... P-SP-AP~ P~----,f: ~ ~II ~:,~D~CCO:~ ~ /'==mmnno~3OOO ~ I] ==mn _ooD~zooo~o ~

I~l~,~,~:-,

OOOOOOOi3OOO~ OOOOOO~OOOOC~J OOOOOO~OOOO~P OOODOD~OOO~OOC O~OOOOOOOO~OO ~ o o o r ~ o o o o o o o O~JE'

1.0

4.O

I~_~ J

OOOOOO~O ' / OOOOOOOOOO~ OnOOOO~OO~OO 0000000000001:300 OOOOO~O~OOO B~E~3OOOBOO~OD 3OO0~D~DODDE~OD

,.,.=..=..ooooo

2.0

4 l~c~co~3o5ooccc

~OOOO-~_O_~OOOO, ~ Q ~ ~ . o.OcE2 .: ~OOOOOO~OOOOOOO ] ~OOOOOO~,OOO~O03 ~ l==O=O~OOOOOOOOO I rhlllr~OO0OOO~OOO ~ ~n~OOO.OOOOOOO ~ I~ = d m n o n o O o o o o o o o i

f1

~°~' '&/.~o, ~.o

o [

Frontal 7 05 mm I

I

2.0

0.0

of the > 0 8; heavy

posterior squares,

n

n

probabilities 0 2 ~<

l

l

l.O

F_ CI

(PP)

PP <~0 . 8 ;

0.0

for the induction

light squares,

PP

1 0

?.G

of teetb-chattermB, < 0 2. Grid

points

~.~ according

with

too low

I n to allocatnon denslt.es

?.n with

doubt.

are not plotted.

319 ble, as stimulation at this site evoked aberrant response patterns. For example, mounting was indm-ed at low current intensity. The allocation matrix for social grooming (Table II) shows a significant departure from independence ( ~ = 189, P < 0.001). The cells for correctly allocated groonlhig and nongrooming placement° contain significantly more placements than expected (adjusted residuals: 9.43, 13.34; P < 0.001), Fig. 4 shows the response area for teeth-chattering. It is located almost entirely in the medial hypothalamus, ventral to the fornix, from the anterior hypothalamus just caudal to the anterior commissure to a posterior level at the caudal end of the ventromedial nucleus. The majority of positive sites is located in the anteropasterior midsection of the hypothalamus, as is t~:e case for attack sites. Again the allocation ,~atrix (Table III) shows a significant departure from independence (Z~ = 270, P < 0.001). Sigmflcantly more correctly allocated positive and negative sites than expected are observed (adjusted residuals: 12.29, 15.97; P < 0.001). So far only the information on the spatial distribution of the locations in which a specific response was induced has been used. However, threshold currents provide additional information on the relative responsweness of sites, and hence on the internal structure of the response areas. The frequency diagram of attack thresholds has one maximum. Apparently all attack sites belong to a single distribution of sites. Four outlying sites had high thresholds above 230 /~A. Two of these are in the ventral hypothalamus surrounded by sites with much lower thresholds

TABLE Ill Allocatton of teeth-chattering and non.teeth-chattering sttes

= 270, df = 3, P < 0 001. Postertor probabdtty for the md,,~tton of teeth-chattering

Actual behaviour induced No teethTeethchattering chattering

Probability < 0.2 Probability > 0 8 0.2 ~~0 8 Too low density

461* 34 76 16

0 43** 11 0

Total per group

587

54

* ** S]gmflcant contribution to departure from independence, adjusted residualsof cells. *12 29, *'15 97, * **P < 0.001.

(frontal 8.25 ram). One site is located near the hypothalamic midline (frontal 8.4), one is really outlying in the optic tract. In order to differentiate within the response area the positive attack sites have been split into two groups of equal size, containing the sites with high and low threshold values respectively. Split point was set at 160#A. High threshold sites are indicated by small symbols (Fig. 1). The large symbols for sites with low thresholds are drawn to the scale of the electrode diameter (150/zm). The lowest threshold value in an electrode track is represented by a filled symbol (Fig. 1), provided that at least two positive sites are present in this track. Lowest threshold sites for attack do not seem to cluster, although in some cases (frontal 7.8 and 7.35 ram) they are close to each other in adjacent tracks. The allocation matrix resulting from the discriminant analysis of high and low threshold sites reveals no significant departure from independence ( ~ = 3, P > 0.2). The distributions of high and low threshold sites do not significantly differ. Most of the elements from both groups are allocated to the doubt groups. The frequency diagram of social grooming sites has one maximum too. The range in threshold values is somewhat wider tnan for attack, The positive sites were split into two groups, with split point set at 122 /~A (so that 19 sites were in the low threshold group and 20 sites in the high threshold group). The lowest threshold sites for social grooming do not cluster (Fig. 1). The allocation matrix of the discriminamt analysis of high and low threshold sites shows no significant departure from independence ( ~ = 7, P > 0.05). The distributions for high and low social grooming threshold sites do not significantly differ. The frequency diagram of teeth.chattering thresholds shows one maximum, although a small group of 6 sites with thresholds above 250/~A might be distinguished. Of this group, 3 sites are located in the paraventricular nt,cleus (frontal 8.1 mm), two sites are in the arcuate nucleus (frontal 7.65 mm) and one site m the suprachiasmatlc nucleus (frontal 8.85 ram). All of these sites are neg.~twe for attack. No clear clustering of lowest threshold sttes for teeth-chattering is observed, although in some adjacent tracks they are near each other. Also for teeth-chattering two groups of equal size containing high and low threshold sites were distinguished. Split point was set at 97.uA. The allocation matrix (Table IV) from the disctiminant

i

3. C

Iii ,l a- g ]DK2.ON

1.0

3.0

IromL~L g ~ m

il,-,

;

~ODI:]OUlO

Fronlal B.N5 mm

/

n

,

_._

I

._

]OnOODDO~3D

A

'.

Frontal 8 7 mm

n

I ; ml'a n

DD[3[~ 131313~ra

,IDDDO O[3DI3D~

3DODDODD[3D 3OOOOOOODD j--~iis. 3OOOHDO~D~ "-~',

[3DOO(3ODon~J ~

~--~

'-

/

3 DEIOODDD[Z3r7 _~ 3QrOo~or~o[~DD / 3OOOOODDDD /

~-

3OrloomIIIIm~

1

OD [ ~ r ~ o r a ~ r a D DDr~oOI~OUDDD

//

//

~)10l:]l~D []1[l,'a O[3 D

L.7~o,'n~Oim~j--"~

L lFronL@L 8.4 mm

u

|.D

n

4

'/

Frontal 8-25 mm

n

n

FI

i

Front~l 61.] mm

n

I

~j

i

i

~--

1 n

Frontal 7 g5 mm

¢

T

i

.-

a

i

,

.I

--t ~o

i;;!:q ~:~

.6oo

........

IDDOOIOOOODD ~]{~OOOO[~OODD

//

3OoDInmOnnD 3D~OOliOC]~on ~OOOOOOOD ~noomtl~IDq ~

//

// i~DOOOOOt]OOD~,.~.~DOBOOODO,,~ I~]G[3MMMM~3DDD

Z.~

FronLol 7.8 mm ] .0

I

n

n

i

0 I 0

n

/

I

// _ ~ I I I

Frontal 712 1.0

i

]

i

0

/"/ // / / .- .~.C.~

.---III

', mmlIIa '.OllKdN

Zi 0

n

;'::", ii~il .} m a m m a

~

°°I ~

n

i

0 10

Frontei 7.5 mm a

i

I

n

•

~

Front~t 7.35 mm n

~

7

| I

/

. Frontal B.g mm

~

i

] .0

,

O ommII

FronL~l 7.05 m,rl i

/

3[31=nmm~

fronLel 7.65 mm n

•" " - ' I i l l

~", ni l m lll l ~rulnII

l

: ...... ! ,'" - ' -I"I I D t ~ D°D"

i

i

ZI 0

0 10

i

110

i

Frontal 6.75 mm 1

~'O

~

0 10

~

l . 0

F~ I -

2

0

FIZZ. 5. Representation of Áhe posterior probabilitnes (PP) for the reduction o( tecth-chattenn~ at low threshold levels, accordinB to allocation with doubl Black squares, PP > 0 8, h,'avy s4uar,'s, 0.2 ~ PP ~ 0 8, h[ht squares, PP ~: 0.2. Orld points with too low dcnsitxcs are not plotted

s0

I

,

7J!:!i....':, :: • °- -" x

,"

~.0

,

'

l'doe

H

'

V~7

xTv

11_9o9 1"9 _o~oo o . -

' vvV.

I

0.7

......--.'ii~" - -.

2.0

I l.O

F,-o,~

~ ,,m

Fr~nt~t

I F, Jl~ ---'F

mm

I

I I

,

~

i

i

I

I

I

I

5.0

,

I I

&.O

...........'\

i

ont~'

B.5 ~_ ~m

i

~

I

..........

{ 1

>oo~u~D/~

o

oo

Da~vvvv~

¢1o ~tmvvvvv o o 9 omQt~t~r~vvv~zs

~/oo o t3~m~r~', v v v v

t

I

5.0

B.4

i

mm

I

,

I

i

I

O O O~ ~3~300~Z0~..~

> C~ O ~ [ ] ~ D ~ @ Z D D

I I

I

Fronlml

8.25

mm

Fromt~t

_

ooo~O[]O~R~-~

8.1

Fronla{

mm

I~ I I

I

//I

//~

t

" o ~ oo~o~o0~/

- - - " - - .Vl~r,21~

VV

:-.--".VVVV

VVV

.... ,

z

' o ~ , v v I/ . .. e>o~ou::~v // >' ~' O O K 3 ~ O K 3 ~ V /I ,

~v~/

If,' oov~16a~vv / , -II, o~ow3Dvv/ / I|", K~Z~K3113DG.VV / 1%-, O D O Q O g C ] O ~ ' L~ / L x~:>~~ ~ [] pC3 D CJ, ) - . ' - - -

- - 0 (> ~ ~ I ~ I I ~ I : 3

FronlaL

7,8

Fromto

mm

i.O

----

5.0

I

I

I

,,

I

'

l--

_I~

I

x .

V

v~v

Frontal I

I

/

"(~OVVVVVV

",¢,DK3aoirn

7 65 mm

i

l

7 . 9 5 mm

~

3.D

/

•

k_/"-:!i(~

j/"/

,l.O

~"

I

~," 6 ~

Frontol

VV

VVV VVVVVVV ~V~VVVV!

2.0

1.0

%

.'

3.0

>o o t 3 u r * ~ o j ~

/

."

r~

I

/

V~"

7.35 mm I

I

I

",7;

4.0

3 0

2.D

Frontal

I.O

I

0.0

I

1.0

7.Z mm I

I

~.0

f L~ 0

,

-

,

,

l.O

2.0

I (J.O

I

0

2 0

0-0

1.0

2 0

Fig. 6. Overlap 6etween the response areas for attack (squares}, social grooming (triangles) and teeth-chattering (diamonds) Onl.~ high probability (PP > 0 8) grid points are drawn

322 TABLE IV Allocatton o f teeth-chattering sites wtth high and low thresholds

= 21, df = 3, P < 0.091. Posterior probabdity for the induction o f teeth.chattering at low threshold values

Actual behavtour induced Htgh Low threshold threshold

Probability < 0.2 Probability > 0.8 0.2 ~0.8 Too low density

15" 2 l0 0

l 11** 13 2

Total per group

27

27

*-** Significant contribunon to departure from independence; adjusted residuals of cells: "4.17, **2.86; *P < 0.001, **P < 0.005

analysis of these groups show a significant departure from independence (Z2 = 21, P < 0.001). The number of correctly allocated high and low threshold sites exceed those expected (adjusted residuals: 4.17, 2.86; P < 0.001, P < 0.005). Note that the doubt groups contain considerable numbers of sites, especially the low threshold group. The results of the discriminant analysis are visualized in Fig. 5. Especially ;,u the posterior part of the response area teeth-chattering can be induced at low current intensities. High current intensities are needed to evoke teeth-chattering in the periventricular part and the dorsal part of the midsection of the response area. The response areas presented in this study are partially overlapping. In Fig. 6 only grid points with high probability for attack (squares), social grooming (triangles) and teeth-chattering (diamonds) are drawn. The overlap of the areas for attack and social groom" ;o .,,~ ...m . .;.~.;.. .. ..~.t. . . . . D. .~.,.k responses are induced al on ly 3 sites (Fig. 1). One unusual site is located in the optic tract, At one of the remaining sites the attack threshold is higher, at the other it is lower than the threshold for social grooming. This lower threshold for attack can be explained by the fact that directly upon displacement of the electrode to the concerned site social grooming was induced, whereas a few days later attack behaviour emerged. Some time may be needed for the brain tissue to rearrange around the electrode after it has beer, pushed down by the displaced electrode. The response areas for attack and teeth-chattering show considerable overlap. At 28 sites both responses were induced. At 20 of them the

attack threshold was higher than the threshold for teeth-chattering, at ~ sites the threshold for teethchattering was higher. Teeth-chattering and social grooming hardly overlap. Only two sites yielded both responses, thresholds being about equal. DISCUSSION It appears that electrical brain stimulation is still a useful tool in establishing the function of neuroanatomical entities. The results indicate that the electrode localization and the stimulation intensity are important determinants of the induced behavioural responses. Sites yielding a specific response more or less cluster into a well definable response area. Different behavioural responses have different response areas within the hypothalamus, although they may considerably overlap. A number of responses can often be evoked from the same electrode site. Differences in threshold current intensities of a specific response within electrode tracks primarily reflect local differences in density of activated ~,eural elements. This study shows a major methodological improvement over previous studies from our lab 25"29. Instead of selecting a limited set of target coordinates for electrode implantation, the electrodes in this study were evenly distributed over the entire hypothalamus. This may decrease the percentage of sites inducing a specific response, but increases the area with sufficient electrode densitv. In previous studies 25'29exact boundaries of response areas could only be defined in a limited area. However, comparison of the response areas provides obvious similarities, although entirely different sets of electrode placements have been analyzed. Except for the perifornical aspect of the initial response area 29 the response areas from previous studies 25.29 are included in the response area described here. The response area for attack encompasses: (1) Part of the anterior hypothalamus rostral to the ventromedial nucleus. (2) The ventromedial hypothalamus at the level of the dorsal supraoptic commissure. (3) The ventrolateral part of the ventromedial nucleus. (4) The subfornical hypothalamus. (5) The area between the ventromedial hypothalamus and the medial forebrain bundle. The ventromedial extension of the response area possibly reflects fibers taking part in the dorsal supraoptic commissure. Ef-

323 ferent fibers from the ventromedial nucleus have been shown to project rostrally through the commissure to the contralaterai amygdala 42. The lateral part of the response area does not overlap with the medial forebrain bundle. Except for one electrode site, aggression was not induced at sites yielding digging, of which the response area is shown to be located in the medial forebrain bundle 31. At this single site the threshold for aggression appeared much higher than the threshold for digging. The distributions of sites from other studies inducing quiet biting and affective attack 3s, mouse-killing 1'22"49or intraspecific attack 2327"53 fit well into the response area, taken into account the electrode size and the stimulation parameters. These different responses may derive from the activation of a single neural system. An elaborate discussion of the neural system of attack behaviour is given by Kruk et ai. 25'2s. They hypothesize that information from various parts of the brain is processed in the hypothalamus in order to 'decide' whether or not to attack. Stimulation in the response area for attack overrules this integrative processing and directly activates the behavioural program of attack, resulting in a well-coordinated behaviourai response. The response is not stereotyped, but continuously influenced by external and internal sensory information. The shape of the response area suggests that intrinsic nerve cells and related fibers are involved rather than fibers of passage. The area is completely surrounded by non-attack zones. Apparently stimulation of the afferent and efferent pathways of the attack area does not give rise to the expression of complete attack patterns. For example, stimulation of the periventricular area through which fibers from the attack area are shown to project to the midbrain central gray 35 does not induce attack responses. Obviously the output of the attack response area takes route to a number of brain structures via different pathways, which all contribute to the final expression of the attack response. In some of these projection areas attack can be reduced (e.g. midbrain central gray35). Nevertheless, these projection areas themselves may not be a prerequisite for attack, for lesioning these areas does not prevent the induction of the response from the hypothalamus 3°. Some anatomical evidence favours these considerations. The response

area coincides well with a number of nerve cell areas described by Geeraedts et al.'7: the anterior part encloses the intermediate hypothalamic area 1, the lateral anterior hypothalamic nucleus and the anterior hypothalamic subarea 1. Posteriorly the intermediate hypothalamic area 1 remains, joined by part of the ventcomedial nucleus. Fiber connections within the response area have been shown to exist between the ventromedial hypothalamus (VMH) and the medial anterior hypothalamus a-~°'42. Afferents and efferents from and to this part of the hypothalamus have been described to some extent. Degenerating fibers from electrolytic lesions placed at the t~p of attack inducing electrodes have been traced to the septurn, the mammillary bodies and the mldbrain central gray 35. Afferent fibers from the medial amygdala project to the core and capsule of the VMH 4"24. Since the response area is extended to areas frontal to the ventromedial nucleus, as was suggested 29, these amygdala projections fit well to the response area. Frequent convergence of lateral septum and amygdala afferents onto medial hypothalamic neurons has been shown 7. Luiten et al. 33 described reciprocal connections between the medial amygdala and amygdalohippocampal area on one side, and the ventrolateral part of the VMH and the perifornical area on the other side. Cells projecting to the medial amygdala were located in a continuous layer in the floor of the hypothalamus, which resembled our attack area conspicuously well. Anterograde tracer injections in the VMH revealed efferents to the medial amygdala, the midbrain periaquaductal gray, and the ventral premammillary nucleus. Lesions in the anterior part of the attack area gave rise to efferent projections to the lateral septum, the premammillary and supramammiliary nuclei, the bed nucleus of the stria terminalis and the arcuate nucleus ~°. In the cat the neural mechanism of aggression has been studied more thoroughly than in the rat ~'t2"13:5. Recently Siegel and co-workers L~'la:3:4 investigated the neural orcuitry of affective defense and quiet bitmg attack The hypothalamlc pathways mediating affectlve defense behaviour involve first-order projections from the ventromedlal nucleus to the antertor medial hypothalamus 15~'. From there sec,3nd-order neurons run caudally into the periaqueductal gray, and to a number of other brain regions. Similar circuitry could be present in the ra~ hypothalamus, for

324 the response area is to a large extent located anterior to the VMH. Similar to our distinction between qualitatively different responses, different modes of attack were observed by Woodworth 53. Attack that was evoked without concomitant alarm differed from attack which was accompanied by alarm, in the same way as described above. Despite minor differences in the distributions of alarm sites from this study and sites from our study 3° yielding flight-directed locomotion or escape jumps (resembling alarm), the same relation between flight responses and attack was observed in this study. Sites from which 'affective' attack responses could be evoked were located in the area with high or intermediate probabilities for flight responses 3°. At 20 medial sites attack was induced in combination with flight-directed locomotion and/or escape jump. The attack threshold was 9 out of 11 times lower than the locomotion threshold. We suggest that the attack response induced from the medial hypothalamus is influenced by simultaneous (subthreshold) activation of other behavioural mechanisms represented in this area. The response area for social grooming is located dorsal and lateral to the attack area. It is extended in anteroposterior direction throughout the hypothalamus. It is not clear whether cells or fibres are primarily involved. Undoubtedly some sites are located in the medial forebrain bundle. In the anterior hypothalamus the response area coincides with the medial preoptic cell area 17. Somewhat posteriorly it fits into the lateral preoptic subarea 4, the lateral hypothalamic nucleus 1, the anterolateral subarea of the lateral hypothalamic area, and the ventrolateral subarea I of the lateral hypotha!amic area. More caudally it comes to occupy the anterolateral and the first part of the ventrolateral subareas of the lateral hypothalamic area, the lateral hypothalamic nucleus parts 1, 2 and 4, the first part of the intermediate hypothalamic area, and the first part of the perifornical nucleus The response area covers the C, C1 and G components of the medial forebrain bundle as described by Veenlng et al. 4s. It is yet impossible to determine which neural elements are involved in social grooming. This part of the hypothalamus contains numerous projections to and from many brain structures 5,48, subserving many behavioural functions. In a previous paper 31 digging

was reported to be induced from this area. Although response areas for social grooming and digging substantially overlap, both responses have been induced from different neural substrates. At 14 sites both social grooming and digging are induced. Sometimes social grooming thresholds are higher, sometimes they are lower than digging thresholds. This ~ggests different neural elements mediating these responses. In contrast to digging, the response area for social grooming cannot be further distinguished into low and high threshold areas, although thresholds vary along electrode tracks probably due to local variation in densities of neural elements involved in social grooming. If fibers of passage running through the medial forebrain bundle were involved in this response, threshold variability would be much smaller in anteroposterior direction than dorsoventrally. No evidence can be given for this. In this study social grooming as defined by Grant and MacKintosh Is was considered a separate response, while aggressive grooming, including skin pulling, was considered to be part of the aggressive response (although rarely observed). Some evidence emerges from the data to consider attack and social grooming different behavioural categories mediated by different mechanisms. The response areas for both responses are to a large extent separated. Only 3 electrode sites yielded both responses, as has been mentioned before. If social grooming is to be considered a weak aggressive response, it should turn into more vigorous attack behaviour when the stimulation current intensity is raised, which was never obser:ed. On the other hand, at electrodes yielding attack social grooming was never induced at subthreshold current intensities. Moreover, in tracks in which both responses appeared thresholds for social grooming are increased before attack is evoked from subsequent sites. The response areas for attack and social grooming cannot be further distinguished into subdivisions according to threshold current intensities. However, the neural substrates are not homogeneous, because in mdlvidual electrode tracks thresholds gradually increase and decrease around the minimum threshold site, suggesting a subarea with high density of neural elements exists. Probably the distinct groups o~-Iectrode sites are just too small to discriminate between subareas. The response area for teeth-chattering is almost

325 entirely located in the medial hypothala_mus. Rostrally it coincides with the anterior hypothalamic subareas 1-4, the first part of the intermediate hypothalamic area, and the lateral anterior hypothalamic nucleus. More caudally, it is related to the para-arcuate hypothalamic nucleus, part of the ventromedial nucleus, and the area of the tuber cinereum t7. The response area can be divided into subareas for high and low thresholds. In the posterior part of the response area teeth-chattering can be induced at relatively low threshold intensity. The high thresholds at periventricular sites in the paraventrieular, arcuate and suprachiasmatic nuclei may be due to activation of neural elements located just outside these nuclei, meaning these nuclei do not take part in the response area. This study provides clear evidence for a prevtous suggestion 26"46 that teeth-chattering and attack do not derive from the activation of a single neural mechanism. Although substantial overlap between the distributions of attack and teeth-chattering sites is observed, a n u m b e r of attack sites as well as teethchattering sites do not yield the other response. This was also reported by others 27'3853. In addition, at dual response sites thresholds for teeth-chattering sometimes are higher, sometimes are lower than attack thresholds. Teeth-chattering is considered to signal autonomic arousal 2'46. In its turn ~..~tonomlc arousal can be indicat~,e for a behavloural conflict between opposing tendencies 2°. Since the response area for teeth-chattering ts roughly located in the me-

REFERENCES 1 Albert, D.J , Nanji, N., Brayley, K.N. and Madryga, F-J , Hyperreacnvity as well as mouse killing :s mdz,ced by electrical sUmulatlonof the lateral hypothalamus in the rat, Behay Ne lral Blol , 27 ( 1979159-71 2 Barnett. S.A, A Stud)' in Behavto,Jr. Methuen, London, 1963 3 Bellt, B M and Keesey, R.E, Hypothalamlc map of stimulation current thresholds for inhibition of feeding m rats, A m . J. Phystol . 229 (1975) 1124-1133. 4 Berk, M L and Fmkelstein, J A , Afferent prolcctions to the preoptle area and hypothalamic regions m the rat brain, Neurosclence. 6 (19811 1601-1624 5 Berk, M L. and Fmkelstemn, J A , Efferent connections of the lateral hypothalamic area of the rat an aatorad~ographlc mvesugatlon, Bratn Res Bull., 8 ( 19821511-526 6 Blanchard, R J and Blanehard, D.C., Aggressive behavior m the rat, Behav BIol, 21 (19771 197-224 7 Blume, H.W., Pittman, O.J-, Lafontame, S. and Renaud, L P , Lateral septum-medml hypothalam~cconnections: an

dial hypothalamic area where the response areas for attack and flight responses 3° overlap, it is tempting to consider the induced teeth-chattering to indicate the simultaneous activation of both attack and flight tendencies. Accordingly, Woodworth 53 found that teeth-chattering was more often associated with attack and alarm than with any other response. However, teeth-chattering can be induced in the absence of an opponent. In addition, simultaneous activation of opposing tendencies for attack and flight should evoke other ambivalent acts like lateral display~6. In hypothalamic attack behavlour lateral display is not observed at any current intensity2s. Note that the overlap of response areas not necessarily implies functional interconnections between both neural systems extst. It would be interesting to determine whether teeth-chattering can be induced by simultaneous stimulation of 'pure" attack and flight sites. Summarizing, we conclude that attack, social grooming and teeth-chattering have different, though overlapping, response areas in the hypothalamus. It depends on the site of stimulation and the stimulation intensity which behavioural response is evoked. Apparently these behavioural responses have different neural substrates. Knowledge of the exact locatton of the response areas simplifies the further charactertzatton of the underlying neural substrates. In additton, future experiments can be carried out to determme if and how the concerned neural systems interact.

electrophysiologlcalstudy in the rat. Neuroscience. 7 (19821 2783-2792 8 Chi, C C . Afferent connections to the ventromedial nucleus of the hypothalamus m the rat. Brain Research. 17 (1¢7(I) 439-445 9 Chi, C C. and Flynn, J P , Neural pathways associated with hypothalamlcally elicited attack behavior in cats. Sctence. 171 (19711703-706. 10 Conrad, L C.A and Pfaff. D.W., Efferents from medial basal forebram and hypo[halamus in the rat, J Comp N e , rol. 169 ( 19761 221-262 11 Dixon, W.J and Mood, A M , A method for obtaim,~ ,~nd analyzing sensitivity data, d Attl Star Assoc., 43 (19481 109-126

12 Flynn, J.P , Smith. D , Coleman. K and Opsahl. C A . Anatomtcal pathways for attack behavior m cats. In M yon Cranach, K. Foppa, W. Lepemis and D. Ploog (Eds), Hvman Ethology

Cla,ns and Lonits o f a N e w DJsophne.

Cambridge Umv Press. Cambridge, 1979. pp 301-356 13 Flynn, J P., Vanegas, H , Foote, W and Edwards, S, Neural mechamsms revolved m a cat's attack on a rat In R.E

326 Whalen, R F. Thompson, M. Verzeano and N.M. Weinberger (Eds.), The Neural Control of Behavior, Academic, New York, 1970, pp. 135-173. 14 Fonberg, E., The motivational role of the hypothalamus in animal behaviour, Acta Biol. Exp., 27 (1967) 303-318. 15 Fuchs, S.A., Edinger, H.M. and Siegel, A., The organization of the hypothalamic pathways mediating affective defense behavior in the cat, ~,ain Research, 330 (1985) 77-92. 16 Fuchs, S.A., Edinger, H.M. and Siegel, A., The role of the anterior hypothalamus in affective defense behavior elicited from the ventromedial hypothalamus of the cat, Brain Research, 330 (1985) 93-107. 17 Geeraedts, L.M.G., Veening, J.G. and Nieuwenhuys, R., The medial forebrain bundle of the rat. IV. Cytoarehitectute ofthe caudal (hypothalamic) part of its bed nucleus, in preparation. 18 Grant, E.C. and MacKintosh, J.H., A comparison of the social postures of some common laboratory rodents, Behaviour, 21 (1963) 246-259. 19 Hess, W.R. and Brugger, M., Das subkortikale Zentrum der affectiven Abwehrreaktton, Heir. Physiol. Acta, 1 (1943) 33-52. 20 Hinde, R.A., Animal BehavIour: A Synthesis of Ethology and Comparative Psychology, McGraw-Hill, New York, 1970. 21 Jiirgens, U., The hypothalamus and behavioural patterns. In D.F. Swaab and J.P. Schad~ (Eds,), Integrative Hypo-

thalamic Acuvity, Progress in Brain Research. Vol. 41, Elsevier, Amsterdam, 1974, pp. 445-463. 22 King, M.B. and Hoebel, B.G., Killing elioted by brain stimulation in rats, Commun. Behav. Biol., A2 (1968) 173-177. 23 Koolhaas, J.M., Hy~ot~halamically reduced intraspecthc aggressive behaviour in the rat, Exp. Brain Res., 32 (1978) 365-375. 24 Krettek, J.E. and Price, J.L., Amygdalotd projections to subcortical structuros within the basal forebrain and brainstem in the rat and cat, J Comp. Neurol, 178 (1978) 225-254. 25 Kruk, M.R., Van der Laan, C.E., Meehs, W., Phillips, R . E , Mos, J. and Van der Poel, A.M., Brain-stimulation induced agonistic behavlour: a noval paradigm in ethopharmacological aggression research. In K.A Miczek, M.R. Kruk and B. Olivier (Eds.), Ethopharmacological Aggresswn Research, Alan R. Liss, New York, 1984 26 Kruk, M.R. and Van der Poel, A M., ls there evidence for a neural correlate of an aggressive behavioural system m the hypothalamus of the rat? In P S McConnell, G.J. Boer, H.J. Romiln, N E. Van de Poll and M.A. Corner (Eds.), The Adaptive Capab,l,ues of the Nervous System, Progress In Brain Research, Vol. 53, Elsevier, Amsterdam, 1980, pp. 385-390 27 Krek, M.R., Van det Poel, A.M. and De Vos-Frerichs, T.P., The induction of aggressive behavlour by electrical stimulation m the hypothalamus of male rats, Behav~our, 70 (1979) 292-321. 28 Kruk, M.R., Van der Poel, A.M., Lammers, J.H.C M , Hagg, Th., De Hey, A.M.D.M. and Oostwegel, S , Ethopharmacology of hypothalamic aggression in the rat In B. OIivler, J. Mos and P F Brain (Eds.), Ethopharmacology of Agonistic Behaviour in Animals and Humans, Ntjhoff, Dordrecht, 1987, pp. 33-45. 29 Kruk, M R., Van der Poel, A . M , Meelis, W., Hermans,

J , Mostert, P.G., Mos, J. and Lohman, A.H.M., Discriminant analysis of the localization of aggression-inducing electrode placements in the hypothalamus of male rats, Brain Research, 260 (1983) 61-79. 30 Lammers, J.H.C.M., Kruk, M.R., Meelis, W and Van der Poel, A.M., Hypothalamic substrates for brain stimulationinduced patterns of locomotion and escape jumps in the rat, Brain Research, 449 (1988) 294-310. 31 Lammers, J.H.C.M., Meehs, W., Kruk, M.R. and Van der Poel, A.M., Hypothalamie substrates for brain sumulationinduced grooming, digging and circling in the rat, Brain Research, 418 (1987) 1-19. 32 Lipp, H.P. and Hunsperger, R.W., Threat, attack and flight elicited by electrical stimulation of the ventromedial hypothalamus of the marmoset monkey Callithrix ]acchus, Brain Beh, v. Evol., 15 (1978) 260-293. 33 Luiten, P.G.M., Koolhaas, J.M., De Boer, S. and Koopmarts, S.J., The corttco-medial amygdala in the central nervous system organization of agonistic behavior, Brain Research, 332 (1985) 283-297. 34 Martin, J.R., Motivated behavmrs elicited from hypothalamus, mldbrain, and pons of the guinea pig (Carla porcelins), Z Comp. Physiol. PsychoL, 90 (1976) 1011-1034. 35 Mos, J , Kruk, M R., Van der Poel, A.M. and Meehs, W., Aggressive behaviour induced by electrical stimulation in the midbram central gray of male rats, Aggress. Behaw., 8 (1982) 261-284. 36 Mos, J., Lammers, J.H.C.M., Van der Poel, A.M., Bermond, B., Meelis, W. and Kruk, M.R., Effects of midbrain central gray lesions on spontaneous and electrically induced aggression in the rat, Aggress. Behav., 9 (1983) 133-155. 37 Nakao, H., Emotional behavior produced by hypothalamic stimulation, Am. J. Physiol., 194 (1958) 411-418. 38 Panksepp, J., Aggression elicited by electrical stimulation of the hypothalamus in albino rats, Physiol. Behav., 6 (1971) 321-329. 39 Roberts, W.W., []4C]Deoxyglucose mapping of first-order projections activated by stimulation of lateral hypothalamlc sites ehcltIng gnawing, eating, and drinking in rats, J. Comp. Neurol., 194 (1980) 617-638. 40 Roberts, W.W., Stemberg, M.L. and Means, L.W., Hypothalamic mechamsms for sexual, aggressive, and other motivaUonal behaviors in the opossum, Didelphis virgintana, J. Comp. Physiol. Psychol., 64 (1967) 1-15 41 Romaniuk, A., Representation of aggression and flight reactions in the hypothalamus of the cat, Acta Biol. Exp., 25 (i965) 177-186. 42 Saper, C.B , Swanson, L.W. and Cowan, W.M., The efferent connections of the ventromedial nucleus of the hypothalamus of the rat, J. Comp. Neurol., 169 (1976) 409-442 43 Shaikh, M B., Brutus, M., Siegel, H.E. and Siegel, A., Regulation of feline aggression by the bed nucleus of the stria terminalis, Brain Res. Bull., 16 (1986) 179-182. 44 Siegel, A and Edinger, H . M , Role of the limbic system in hypothalamically elicited attack behavior, NeuroscJ. Biobehay. Rev., 7 (1983) 395-407. 45 Timmermans, P J A., Social Behavtour In The Rat, Thesis, Nqmegen Umv., 1978 46 Van der Poel, A M , Mos, J., Kruk, M.R. and Ohvier, B., A motivational analysis of ambivalent actions m the agonistic behavlour of rats in tests used to study effects of drugs on aggression. In K.A Mlczek, M.R. Kruk and B. Olivier (Eds.), Ethopharmacologwal Aggression Research, Alan R Liss, New York, 1984, pp. 115-135

327 47 Van der Poel, A.M., Olivier, B., Mos, J., Kruk, M R. and Van Aken, J.H.M., Anti-aggressive effect of a new phenylptporazme compound (DU 27716) on hypothalamicaily induced behavioural activities, Pharmacol Biochem. Behay., 17 (1982) 147-153 48 Veening, J.G., Swanson, L.W., Cowan, W M., Nieuwenhuys, R. and Geeraedts, L.M.G., The medial forebrain bundle of the rat. II. An autoradiograph~c study of the topography of the major descending and ascending components, J. Comp Neurol, 206 (1982) 82-108. 49 Vergnes, M. and Karh, P., D6clenchement d'un comportement d'aggtession par stimulation ~lectnque de rhypothalamus m~dtan chez le rat, Physiol. Behav, 5 (1970) 1427-1430

50 Waldbdlig, R.J., Attack, eating, drinking, and gnawing elicited by electrical stimulation of rat mesencephalon and pons, J. Comp Physiol. Psychol., 89 (1975)200-212. 51 Wasman, M. and Flynn, J P., Directed attack efictted fTom hypothalamus, Arch. Neurol., 14 (1966) 408-414. 52 Wetherill, G.B., Sequential estimation of points on quantal response curves. In G.B. Wetherill (Ed.), Sequential Methods in Statisucs, Methuen, London, 1966, pp. 162-172. 53 Woodworth, C.H., Attack elicited in rats by electrical stimulation of the lateral hypothalamus, Phystol. Behav, 6 (1971) 345-353