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Lecture. Invest. Ophthalmol.

'el of free a' idI : 625-632~1~~~~~ sand myoinositol in

7vest.Ophtha/mol. Visual Sc .• 5 : 65-74.

. BioI.Mea., 32 : 971-973. 1935orphology in 126 young d' b t' '.la e IC patlent~.

on the isolated lens. Invest. Cphthalmol.

eformation of galactose induced cataracts.

fter injecling dehydroascorbic acid and

betic cataracts. Am. J. Ptrysiot., 172

anceof mature sugar cataracts. Invest;

. New York. Academic Press. 1961.Invest. Ophtha/mol. Visual Sc .• 13

Res.• 1 : 296. 1962.

taract. Nature (Lond). 184 : 194-195.

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actosemic lenses Biochem B' h. • lOp vs.s aldose reductase. Science. 188 :

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28; 1981

EXPERIMENTALSPIKE AND WAVE EEG PATTERN FROM THALAMICSTIMULATION IN ANAESTHETIZED JUVENILE MONKEYS·

J. DAVID. S. B. MARATHE. S. D. PATIL. G. P. WAGLE AND R. S. GREWAL

CIBA-GEIGY Research Centre. Goregaon, Bombev - 400063

(Received on June 30. 1981)

Summary : A bilaterally synchronous 3/sec spike and wave pattern in the EEG can be obtainedduring stimulation of the nucleus media lis dorsalis (nM D) and nucleus paracentralis (nPC) ofthe thalamus. in lightly anaesthetized juvenile monkeys. The spike and wave EEG patterns havea frequency of 2i to 3Hz and occur during low frequency stimulation. without continuing intothe post-stimulatory phase. Stimulation of the mesencephalic reticular formation (MRF) athigher voltaqes inhibits the thalamic-induced spike and wave response. This finding has heuristicsignificance. as activating stirnu li arrest the wave and spike EEG pattern of human absence (petit malepilepsy) seizures. The study also demonstrates the morphological similarity of the 3/sec spikeand wave thalamic induced response in experimental and clinical absence states as well assirnllarities in their alterauons with different levels of activation.

Key words: absence spike and wave discharge Rhesus monkey thalamic nuclei

INTRODUCTION

Spike and wave patterns in the EEG evoked by electrical stimulation of the intra-laminar thalamic nuclei of the cat. are morphologically similar to the 3/sec spike and wavepatters of patients with absence seizures (3). Previous experiments by Jasper andDroogleever-Fortuyn (1) and Pollen et al. (2) have helped to clarify many physiologicalproperties 01 spike and wave responses and their anatomical relationship to a inqle fibresystem. Jasper and Droogleever-Fortuyn (1) using anaesthetized cats. lirst showedthat electrical stimulation of a small section of the intralarninar nuclei produced an evokedpatternwith prominent 3/sec spi~e" and wave complexes. It was later demonstrated thatstimulation of this region in unanaesthetized cats produced fixation of the eyes. staringandan arrest reaction associated with the characteristic spike and wave EEG pattern (3).

'Contribution No. 619 from CIBA-GEIGY Research Centre. Bombay

202 David et al.

The present investigation was undertaken as part of a larger study for which theprimary objective was to reproduce behavioural concomitants of thalamic induced 3/secspike/wave discharges in unanaesthetized juvenile monkeys. This report describes thelocalization of areas in the nucleus medialis dorsalis CnMD) and nucleus paracentralis(nPC) from which typical spike and wave EEG patterns could be elicited by electricalstimulation in lightly anaesthetized monkeys.

MATERIALS AND METHODS

Colony reared Rhesus monkeys. 2.5 to 2.8 kg. were preferentially selected so thattheir age was about 14 to 18 months according to criteria established by Bourne (4).The monkeys were screened electroencephalographically before experimentation accordingto a method described previously (11 and 12). Ten monkeys were anaesthetized withsodium pentobarbital. 35 mg/kg i.p .. and positioned in a stereotaxic instrument. lncisionaledges and pressure points were infiltrated with 2% Novocaine* (VET. Hoechst). Additionalamounts of pentobarbital. 5 mg/kg. were given intravenously when needed. but generallya relaxed. lightly anaesthetized animal was most suitable.

Olszewski's (5) stereotaxic atlas. prepared especially for exploration of the thalamusof Macaca rnulatta was used for orienting the electrodes to the site of stimulation. Con.centric bipolar steel stimulating electrodes (or rnonopolar Hess electrodes insulated exceptfor 1 mm above the tip) with a tip separation of 0.5 to 1 mm were used for stimulating thenMD and nPC. They were directed vertically to the stimulation site. Stimulation wascarried out with a Grass S8 stimulator through a stimulus isolation unit. Biphasic pulses.1 to 2 msec in duration at frequencies of 3 or 6 Hz were used for all experimentsHigher frequencies of 30 Hz gave rise to clonic seizures. Voltages varied from 1 to 15vand both single pulses or 5 to 10 second trains were employed. Electrocorticogramswere recorded on a Grass Model 78 8-channel EEG Polygraph with the help of silver ballelectrodes on the exposed pial surface. The method used in these experiments consistedof electrically stimulating the thalamic nuclei. recording the electrocorticographic responsesand determining the site and electrical parameters which produced optimal 3/sec spike andwave discharge. The exposed cortical surface was kept covered with warm (3rC)liquid paraffin. The frame of the stereotaxic instrument was used as the reference point.All cortical EEG records were obtained 6 to 7 mm from the midline in the pre-frontal. parietaland ore-occipital areas.

In order to varify that the stimulating current did not spread more than 0.5 mmfrom the tip of the electrode. the method described by Jasper and Droogleever-Fortuyn

"Registered Trade Mark

Volume 25Number 3

(1) was employed. Histolopoints of stimulation were mnMD and nPC regions were

About 400 sites weretures such as nMD and nPCgra ph ic effects. structures WI

few sites were also stimulatecephal ic reticular formation. c

Nucleus medialis dorsalis (nThe stimulating electr

PC. Those areas which w:ave forms are shown in .Fianterior 9.5 and 1.5. tapennlimits of this nucleus do notvertical extent is between cfrom anterior 4 to 8 and overof 3 Hz. 1 to 2 rnsec pulse dbest bilaterally synchronousL 0.5 to 3 and H 8 to 10 abo

In all the monkeys eselectively produced well sha. (F'lg 2) No spike antion .'

threshold for spike/wave prinitially surface negative powas lowered from the cornclarge amplitude spikes andappoached. stimulation re.sulampl itude of the spikes (Fig.395 /LV respectively. The mearespectively (Table I). Piaa rostro-caudal manner at Awave forms. Irregular spikthe stimulated hemisphere.electrodes in each hernisphe

July-September 1981Ind. J. Physiol. Pharmac

t of a larger study for which theitants of thalamic induced 3/seckeys This report describes the

MD) and nucleus paracentraliscould be elicited by electrical

Dsre preferentially selected so thateria established by Bourne (4).efore experimentation accordingkeys were anaesthetized withreotax.c instrument. Incisional* (VET,Hoechst). Additional

sly when needed. but generally

for exploration of the thalamus~the site of stimulation. Con-esselectrodes insulated except

were used for stimulating theulation site. Stimulation wasalation unit. Biphasic pulses.re used for all experiments.Voltages varied from 1 to 15 vmployed. Electrocorticogramsh with the help of silver ballthese experiments consisted

lectrocorticographic responsesuced optimal 3/sec spike andcovered with warm (3rC)used as the reference point.

line in the pre-frontal, parietal

spread more than 0.5 mmer and Droogleever-Fortuyn

3/Sec Spike and Wave Response in Monkeys 203

(1) was employed. Histological verification' of electrode placement was done. Thepoints of stimulation were marked by electrolytic deposit of iron and serial sections of thenMD and nPC regions were examined (6).

About 400 sites were stimulated which covered fairly extensively subcortical struc-tures such as nMD and nPC In attempts to obtain bilaterallv synchronous electrocorti-graphic effects, structures within 4mm from the midline were specifically explored. Afew sites were also stimulated in nucleus reuniens, nucleus centralis intermedialis, mesen-cephalic reticular formation. central gray and the centrum medianum.

RESULTS

Nucleusmedialis dorsalis (nMD) and nucleus paracentralis (nPC) :

The stimulating electrode was positioned in a number of sties within the nMD andnPC. Those areas which when stimulated .oroduced reproducible 2t to 3 Hz spike andwave forms are shown in Fig. 1,. In the monkey, the nMD lies between coordinatesanterior 9.5 and 1.5. tapering away as the frontal zero plane is approached, The laterallimits of this nucleus do not exceed coordinates 4 to 5 on either side of the midline and thevertical extent is between coordinates 8 to 12 above zero. nMD has the largest areafrom anterior 4 to 8 and over 300 sites in this region alone were stimulated with parametersof 3 Hz. 1 to 2 msec pulse duration and varying voltage for trains of 5 or 10 seconds. Thebestbilaterallv synchronous spike and wave responses were obtained in the nMD at A 8,LO.5 to 3 and H 8 to 10 above zero (Fig. 1).

In all the monkeys examined this region at A 8, L 0.5 to 1 and D + 8 to 1 + 10selectively produced well shaped spike and wave patterns in the EEG during 3/sec stimula-tion (Fig. 2). No spike and wave patterns were seen in the post-stimulatory period. Thethreshold for spike/wave production varied from 7 to 15 V. The spike was seen as aninitially surface negative potential followed by a slow negative wave. As the electrodewas lowered from the cortical surface it was observed that cortical stimulation resulted inlarge amplitude spikes and very low amplitude slow waves. However, as nMD wasappoached. stimulation resulted in well formed waves of larger amplitude with decreasedamplitude of the spikes (Fig. 2). The mean height of the spike and slow wave was 184 and395 wrespectively. The mean duration of the spike and slow wave was 34 and 228 msecrespectively (Table I). Placement of bipolar stimulating electrodes in one. hemisphere ina rostro-caudal manner at A 8, L 1. H + 9 failed to produce bilaterally synchronous spikewave forms. Irregular spike and wave forms appeared mainly in the frontal EEG leads ofthe stimulated hemisphere. On the other hand, placement of monopolar stimulatingelectrodes in each hemisphere, from 0.5 to 1 mm lateral on either side of the midline yielded

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3/Sec Spike and Wave Response in Monkeys 207

the best spike and wave forms. Spike and wave forms of lower amplitude were alsoobtained at 2 and 3 mm lateral. on either side of the midline. when the distance between thetips of the stimulating (monopolar) Hess electrodes was 4 and 6 mm respectively. Noparticular behavioural responses accompanied the spike and wave discharge in the EEGalthough at higher frequencies of 30 Hz. 1 msec and 10V behavioural and EEG clonic

seizureswere seen in the lightly anaesthetized animals.TABLE I : Average height and duration of thalamic induced spike and slow waves

calculated from pre-frontal and frontal EEG leads.---X Height (Il-v)·

X Duration (msec)·

Spike 184 (130-262)

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34 (20-40)

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"Ail responses were obtained as a result of stimulating nMO at A 8. L 1 and 0+9.

Nucleus reuniens. centrum medianum, central gray and mesencephalic reticulsr formation!A vertical stimulating concentric bipolar electrode lowered in the midline at frontal

plane10. from a height of 12 mm to the nucleus reuniens at 5 mm above zero. did not yieldcharacteristic spike and wave patterns at any site in the vertical track. Likewise. stimula-tion of the mesencephal ic reticular formation (M RF) at A 1.5. L 3 or 4 and H + 2 wasineffective (Fig. 3). Simulataneous stimulation of MRF (200 Hz 1 rnsec 10V) duringnMD stimulation (3Hz, 1 msec 10 v) showed marked inhibition of the nMD induced spikeandwave responses. Stimulation of centrum medianum, central gray and centralis inter-medialis resulted in poorly tormed spike and wave forms appearing only in the frontal and

anterior frontal EEG leads.

DISCUSSION

Following the classic work of Jasper and Droogleever Fortuyn (1), it has beenaccepted that the 3/sec spike and wave patterns evoked by stimulation of the intralaminarsystem of the thalamus of the cat are morphologically similar to EEG patterns found inabsence (petit mal epilepsy) in the human (1,2). The spike and wave complexes in thecat are best seen bilaterally and symmetrically in gyrus proreus. anterior and posterior sig-moid cortex and over the anterior portions of the lateral gyrus, after 3/sec stimulation ofnucleus centralis medialis and nucleus reuniens. The experiments reported here havedealt with stimulation and exploration of specific nuclei in the thalamus of the juvenilemonkey in order to obtain spike and wave responses bearing a close resemblance to that ofhuman petit mal epilepsy. The morphological similarity of the thalamic induced experi-mental spike and wave response and clinical 3/sec spike and wave EEG pattern is only

208 David et al. July-September 1981Ind J. Physiol. Pharmac.

considered as an initial step in the preparation of a chronic monkey model of absenceseizures. It does not imply a direct relationship between the experimental and clinicalforms of absences.

The relaxed lightly anaesthetized monkey was essential for obtaining favourablespike and wave responses and this observation has also been reported in the cat and human(2. 7). Stimulation of the mesencephalic reticular formation at a sufficiently high voltageinhibited the thalamic induced wave and spike response. In the cat reticular stimulationblocks wave and spike discharges which reappear when the reticular activation is loweredbelow a critical level (2). In patients also. wave and spike discharges are arrested byalerting acoustic or nociceptive stimuli (7.8).

An application of these findings has been made in unanaesthetized juvenile mon-keys. and we have shown that the behavioural concomitants of the spike and wave EEGdischarge has certain similarities to human absence states (9). Further. this monkeymodel of absence seizures can be used for predicting anti-absence activity of potentialanticonvulsant drugs and distinguishes between anticonvulsants that are effective in absenceseizures and those that are effective in focal and grand mal seizures (10).

REFERENCES

1. Jasper. H.H. and J. Drooglever-Fortuyn. Experimental Studies on the functional anatomy of petit mal epilepsy.Res. Publ. Ass. nerv. memo ::.is.• 26 : 272-298. 1947.

2 Pollen. D.A.. P. Perot and K.H. Reid. Experimental bilateral wave spike from thalamic stimulation in relationto level of arousal. Electroenceph. Clin. Neurophysiol..15 : 1017-1028. 1963.

3. Hunter. J. and H.H. Jasper. Effects of thalamic stimulation in unanaesthetized animals. Electroencepb. Clin.Neurophysiol. 1 : 299-304. 1949.

4. Bourne. G.H. In The Rhesus Monkey. Vol. I. Anatomy & Physiology. London. Academic Press. P. 80. 1975.5. Olszewski. J. The Thalamus of the macaca rnulatta - an atlas for use with the stereotaxic instrument. Basel,

New York. S. Karger. 1952.6. Marshall. W.H. An application of the frozen section technic for cutting serial sections through the brain. Stain

Tech.• 151 : 133-138. 1940.7. Li. C.L.. H.H. Jasper and L. Henderson. The effect of arousal mechanisms on various forms of abnormality in

the electro-encephalogram. Etectroenceph. Clin. Neurophysiol .• 4 : 513-526. 1952.8. Myslobodsky. M. Factors of hypersynchronization. Wave-spike discharges: Centrenergetic or Centrasthernic?

In "Petit Mal Epilepsy: a search for the precursors of Wave-spike activity." New York. Academic Press.p. 164.1976.

9. David, J .. S.B. Marathe. S.D. Patil and R.S. Grewal. Arrest reaction with concomitant spike and wave after-discharge following thalamic stimulation in conscious juvenile monkeys with Al (OH)) focal premotor lesions.Ind. J. Physiol. Pharmac .. 25(3) : 209-218. 1981.David. J .. S.B. Marathe. S.D. Pati] and R.S. Grewal. Behavioural and electrical correlates of absence seizuresinduced by thalamic stimulation in juvenile Rhesus monkeys: a new method for pharmacological studies. (Msssubmitted for publication). RDavid. J. and R.S. Grewal. Effect of carbamazepme (Tegretol) ( ) on seizure and EEG patterns in monkeyswith alvmina-tnouced motor and hippocampal foci. Epilepsie. 17 : 415-422. 1976.Oavid. J. and R.S. Grewal. Photic and pentylenetetrazol induced seizure susceptibility in Macaca mulatta.J. Med. Primetotoqv, 6 : 337-343. 1977.

10.

11.

12.

CIBA-GElGY

RREST REACTION WITFOLLOWING THA

MONKEYS WI

J. DAVID. S.8

Summary : The behaviof a specific area in the nkeys is described. A beconcomitant 21 to 3Hzspof the nMD in monkeysanterior premotor eortica

Key words: absence

Hunter and Jasperin unanaesthetized cats resthe behavioural componena bilaterally synchronousthe EEG and was associatstudies on pentobarbitaland wave responses in re.dorsalis (nMD) (2). ThlEEG pattern following sumonkeys.

20 male colony reaanaesthetized with sodiu--------