- Email: [email protected]
Experimental Kuru in the rhesus monkey: A study of EEG modifications in the waking state and during sleep
Electroencephalography and Clinical Neurophysiology, 1978, 4 5 : 6 1 1 - - 6 2 0 © Elsevier/North-Holland Scientific Publishers, Ltd.
611
E X P E R I M E N T A L K U R U IN THE R H E S U S MONKEY: A STUDY OF EEG MODIFICATIONS IN THE WAKING STATE AND D U R I N G SLEEP * J. B E R T a, G. V U I L L O N - C A C C I U T T O L O a, E. BALZAMO a, p. DE MICCO J. T A M A L E T b and H. G A S T A U T a
b, D.
G A M B A R E L L I c,
a Service de Neurophysiologie Clinique et C.R.P. 444, C.N.R.S., b Laboratoire de Virologie, and c Laboratoire de Neuropathologie, C.H.U. Timone, Marseilles (France) (Accepted for p u b l i c a t i o n : April 4, 1978)
Kuru, a degenerative disease of the central nervous system (CNS) occurring among a certain population in New Guinea, was shown to be transmissible to subhuman primates by Gajdusek and Gibbs in 1966. In 1969, Gibbs and Gajdusek similarly established the transmissibility of Creutzfeldt--Jakob disease which, like Kuru, is caused b y unconventional infectious agents. These authors thus opened up a new chapter in the aetiology of diseases affecting the human CNS. The determination of the aetiology is difficult: the transmissibility to animals, in fact, constitutes the only formal criterion of it. This problem emphasizes the practical interest of studying experimental diseases in primates, making it possible: (1) to list all the clinical, electrophysiological and neuropathological effects of the disease agents, and (2) to analyse all the evolutionary phases o f pathological processes which are most often perceived only in the terminal phases in human neurology. This work on experimental Kuru in the rhesus m o n k e y was carried o u t from this viewpoint.
Material and m e t h o d Early in 1975, 13 rhesus m o n k e y s (Macaca mulatta) were subjected to surgical implantation of electrodes (4 mm diameter stainless steel screws) at the surface of the dura mater * This work was in part supported by a grant from the F r e n c h F o u n d a t i o n for Medical Research.
for EEG recording and at the orbital peripheries for eye m o v e m e n t recording. Eight monkeys were inoculated with a strain of Kuru passaged in rhesus m o n k e y s in Dr. Gajdusek's laboratory at Bethesda (ENAGE strain rhesus L6 56): intracerebral inoculations (0.2 ml of 10 -2 dilution of a 20% cerebral solution in P.B.S., pH 7.4, introduced in the left frontal lobe through the r o o f of the orbit) and intravenous inoculations (0.2 ml of 10 -2 cerebral suspension) were performed. Six m o n k e y s were inoculated in May and June 1975: three (Mm 03, 06, 14) with a strain unfrozen for the first time less than I h prior to inoculation and 3 others (Mm 04, 05, 07) with a strain unfrozen at the time of the inoculation of Mm 03, 06, 14, then refrozen and thawed again. Two animals (Mm 23, 24) were inoculated in December 1975, with a strain which had been unfrozen during each of the previous inoculations. The first 6 animals were adult at the time of inoculation, while the latter 2 were between 8 and 12 months old in December 1975. A control group of 5 m o n k e y s was kept under identical conditions. These animals showed no abnormalities at any time during several recording sessions after implantation and up to July 1977. EEG records were made every 4 months from all of the inoculated monkeys. The animals were placed in restraining chairs several days prior to the session in order to familiarize them with the experimental conditions. Each recording began at a b o u t 5.45 pm
612 in order to obtain an EEG sample for the waking state in lit surroundings. The lights were switched off at 6.00 pm until 6.00 am the following day, at which time recording was terminated. Moreover, at least one continuous 24-h record was made for each animal to establish the normal daily amounts of sleep. With the appearance of the initial symptoms, the recording intervals were shortened, first to monthly EEG sessions and then, according to the development of the disease, every 2 weeks. A total of 80 records was made in this way. The recording intervals no doubt led to a loss of data; nevertheless, the evolution of the disease was monitored with sufficient accuracy. The records were analysed in 15-sec periods according to the criteria adopted in this laboratory for the stages of sleep among Cercopithecinae (Bert et al. 1970). The results were then entered on punch cards for computer processing. When physiological criteria became impractical, new criteria were defined, and 3 stages of Slow Wave Sleep (SWS) were identified (a, b, c). Two monkeys (Mm 04, 14) have shown no formal signs of illness to date, and have been disregarded for the purpose of this study. The 6 remaining animals were killed as late as possible in view of their clinical condition: on average 700 ± 40 days after inoculation (between the 645th and the 762nd postinoculatory day). The data of serial clinical examinations (Comte-Devolx et al., in preparation) and neuropathological findings (Gambarelli et al., in preparation) will be presented independently. Results
In view of the variability of the initial symptomatology following the incubation period, two evolutionary phases must be discriminated for Kuru: an initial prodromal phase and a mature phase.
J. BERT ET AL.
(1) The initial phase includes isolated or associated clinical and EEG signs. (a) In 4 monkeys, the first isolated clinical signs appeared after about 15 months. Two animals (Mm 03, 06) showed behavioural troubles (disinterest, inertia, modified feeding habits). The other 2 monkeys presented slight (Mm 23) or grave (Mm 24) myoclonic syndromes with no behavioural troubles. The remaining two animals (Mm 05, 07) showed no precocious clinical troubles. (b) Throughout the prodromal phase, the EEG recorded in the waking state remained normal for 5 monkeys (Mm 03, 05, 06, 23, 24). In the sixth (Mm 07), bursts of spike and wave complexes at 3--4 c/sec were recorded in the frontal regions shortly before the onset of the mature phase of the disease (20th postinoculatory month). (c) Sleep pattern modifications began to appear after 16 months. The physiological stages could be differentiated without difficulty. Nevertheless during stage 1 the alpha frequency was slowed by 1--2 c/sec and the amplitude of the beta activity was considerably reduced. The sleep spindles disappeared during stage 2. The transition from the waking state to sleep was seriously perturbed, presenting a long initial period of light sleep (stages 1 and 2), interrupted by frequent awakenings. The latency of the first episode of stage 4 and of the first episode of Rapid Eye Movement (REM) sleep increased considerably, and the total duration of these stages gradually decreased. Thus the stage 4 sleep normally occurring in the first few hours of sleep was retarded until the middle of the night. Furthermore, spontaneous transient awakening reactions, lasting 2--5sec and often preceded by a high voltage wave, were noted. Such awakening reactions are occasionally observed in normal rhesus monkeys, but are rather rare. These modified sleep patterns, which were discussed in a previous paper (Bert et al. 1977), were not observed in one of the monkeys (Mm 23).
EXPERIMENTAL KURU IN RHESUS MONKEY
(2) The mature phase commenced from 600 to 700 days after inoculation. It was characterized by major EEG alterations and by the development of a true dementia syndrome with generalized myoclonic jerks in 4 animals (Mm 03, 05, 06, 07) and an extrapyramidal syndrome with myoclonic jerks and minor behavioural troubles in the other 2 (Mm 23, 24). (a) The waking EEG recorded with eyes open was abnormal. It was characterized by irregular 5--7 c/sec activity of moderate amplitude (50--100~V), which was replaced by continuous slower activity (4--5 c/sec) of larger amplitude (100--150 ~V) as the animal's attention diminished. This was associated with frequent spikes, bilateral and generalized or multifocal (the majority of these spikes were recorded from the electrodes implanted along the midline, but independent spikes were noted in the temporal regions). Monkey Mm 07 showed major frontal bursts of 3 c/ sec spike~nd-wave complexes. In general, these irritative phenomena became increasingly pronounced when the animal's vigilance diminished, as measured by the drop in blinking frequency. The irritative signs were more significant in the left temporal region in 3 monkeys (Mm 03, 06, 07). (b) Although the total duration of sleep remained constant, its nature was so profoundly altered that it became impossible to refer to the classification according to physiological stages, for which the major identifying criteria had disappeared. The sleep was nevertheless polyphasic and SWS continued to present three easily identifiable stages, which persisted until the terminal phase of the disease without modification. (1) Stage 'a' appeared as soon as the animal closed its eyes. The activity recorded on the waking EEG became slower (3--4 c/sec) and of larger amplitude (100--200 ~V), and spiking became more important. In one animal (Mm 07) the EEG pattern at the terminal phase resembled an almost continuous discharge of spikes or spike-and-wave complexes (Fig. 1).
613
M.m. 07 14
_06
-
77
....9 100pV
Fig. 1. Complex slow pattern with spikes or spike-andwave discharges, in some cases bilateral and synchronous on both hemispheres, while asynchronous in other cases.
(2) Stage 'b' was characterized by mediumamplitude (75--150lzV) activity at 7--8c/ sec with occasional sporadic waves or short bursts of waves at 2--3 c/sec. Neither spindles nor V waves were observed. The spikes became much rarer, and most often then recorded from midline electrodes. Nevertheless, in the terminal phase Mm 07 presented generalized 2c/sec spiking discharges (Fig. 2). (3) Stage 'c' rather closely resembled stages 3 and 4 of normal sleep. The EEG showed increasing 1--3 c/sec waves of large amplitude (150--200 ~V) which rapidly became continuous. The irritative phenomena had practically disappeared. (4) REM sleep persisted in rare and very short episodes of no more than 20 sec. A few isolated eye movements occurred, but the bursts ceased. REM sleep completely disM.m. 07 24 - 05
- 77
I Set.
Fig. 2. Generalized bursts of spike-and-wave complexes recorded during stage 'b'.
614
J. BERT ET AL. M . m. 0 6
s th
20
month
month
th
Iil, '
21 t h
month
REM
o I 6 p.m.
6 o.m.
Fig. 3. Modified organization of sleep during the evolution of experimental Kuru. Six months after inoculation, the organization is typical of normal sleep in the rhesus monkey. During the 20th month, the physiological stages remain identifiable but are characterized by very long periods of light sleep and awakenings, with stage 4 retarded until the second half of the night. During the 21st month the sleep pattern is morphologically abnormal. It is not possible to use the classification according to physiological stages. SWS presents 3 abnormal patterns, classified as stages a, b, c, and its organization is severely perturbed.
appeared between the 615th and 708th day after inoculation. This morphological transformation of the sleeping EEG was accompanied by a general disorganization of sleep itself and the blurring of its cyclic pattern (Fig. 3). The various stages followed one another interspersed by periods of wakening, but no longer constituting veritable cycles. The stage 'c' periods tended to occur throughout the night, and not simply in the first few hours of sleep as with stages 3 and 4 of physiological sleep. However, the REM sleep episodes occurred primarily in the second half of the night, and represented the final aspects of the temporal
19_10_77
~ lOO~v 1 Sec.
6-7~
~
~
~
~
9-10~
Fig. 4. Brief awakening reaction after a massive m y o -
clonic jerk accompanied by generalized artifacts but with no repercussions in the EEG.
EXPERIMENTAL K U R U IN RHESUS MONKEY
organization of true physiological sleep. These 3 stages of SWS persisted throughout the remaining development of the disease. In the last record of one monkey (Mm 05, 729th day after inoculation), the amplitude of all activities tended to drop; the stage 'c' episodes became more widely spaced and the delta waves less abundant. Moreover, 3types of transient phenomena were observed during sleep: (1) Brief awakening reactions, less frequent than during the prodromal phase. These reactions sometimes followed a massive myoclonic jerk (Fig. 4). (2) Brief (2--4 sec) episodes of flattening resulting from the suppression * throughout the cortex of all theta and alpha activity, followed immediately by the unaltered resumption of the preceding pattern (Fig. 5A and B). When these theta and alpha activities were superimposed on lower frequency waves, the latter sometimes persisted or were suppressed as well. Such episodes might or might not be accompanied by a tonic contraction of the facial muscles. (3) 'Tonic seizures' involving a more complex event sequence (Fig. 5C and D, and Fig. 6). Occasionally preceded by more or less generalized discharge of a spike or spike burst, the 'seizure' itself began by the flattening of baseline activity for 2--6 sec, followed by a generalized discharge of sharp waves or sharp-and-slow wave complexes at 1--2 c/sec. A tonic contraction of the facial muscles occurred simultaneously, as shown by electromyographic activity from the electrodes implanted around the eyes. Such 'seizures' were not all identical. In some cases the sharp wave discharges appeared simultaneously with the flattening of background activity. Both forms might occur in the same animal during a single night. The number of ictal episodes (episodes of flattening and 'tonic seizures') increased very * In this paper the word 'suppression' is used with a merely descriptive meaning without any physiological connotation.
615 M.m. 05 5_05_77
~
loo~
1 $~.
Fig. 5. Brief episodes during which background activity is suppressed during stage 'b'. Such episodes may be isolated (A) or followed by a pseudorhythmic sharp wave discharge of variable duration (B--C--D). Example D shows a brief reinforcement of the EMG activity on the oculogram lead.
rapidly, reaching or exceeding 250 per night (Mm 05) in the terminal phase. The total duration of the 'seizures' then represented 18% of the sleeping time. Sensory stimuli (e.g. noise) could result in a brief awakening reaction or in a 'tonic seizure'. These ictal episodes appeared only after a certain minimal period of sleep, and did not occur during stage 'a'. Rather, they appeared essentially during stage 'b', while the stage 'c' periods most often ended in a 'tonic seizure'. 'Tonic seizures' of short or moderate duration did not interrupt the stage 'b' episodes, and occurred repeatedly separated by interictal tracings of a few seconds to nearly 1 min. Longer seizures were terminated by the transition to stage 'a' or by awakening of the
616
J. BERT ET AL.
M.m. 06 3 - 0 5 - 77
~ IOO~v 1 Sec.
A ,-2
.
.
.
.
.
.
,.
,,.~,7,:
~A~'~ '~-'~
....
I
,-~ ~
~, . . . . .
'""':""'""~'~~'~
I
,
~
~
~
~
,
~
,
"'"
........
,
~
~¢~ ~ , , ~ ~ v ~ 2 ~ , ¢
~
B
Fig. 6. A a n d B: ' T o n i c seizures' w i t h initial f l a t t e n i n g of b a c k g r o u n d activity f o l l o w e d by p s e u d o r h y t h m i c s h a r p waves a c c o m p a n i e d b y t o n i c c o n t r a c t i o n o f t h e facial muscles. I n e x a m p l e B, a s e c o n d ' s e i z u r e ' a p p e a r s a l m o s t i m m e d i a t e l y a f t e r t h e e n d o f t h e first episode.
animal. That the onset of such seizures required a minimal a m o u n t of stage 'b' sleep explains their relative absence during the day, when the EEG tended to oscillate between the waking state and stage 'a', with a few rare episodes of stage 'b' sleep. The maximum observed involved 11 daytime seizures with a burst of seizures after 8 min of continuous sleep (Mm 06, on the 691st day after inoculation). Only a single animal (Mm 23) presented no tonic seizures.
Discussion
The development of experimental Kuru in rhesus m o n k e y s is accompanied by peculiar
EEG modifications in the waking state and during sleep. The dominant character of these alterations concerns the importance of irritative signs and the occurrence of multiple nocturnal tonic seizures. These characters appear to be peculiar to the experimental disease in the rhesus m o n k e y . They were n o t detected in the disease in man {Cobb et al. 1973) or in the experimental disease in the chimpanzee (Niedermeyer et al. 1972; Asher et al. 1973). Cobb et al. (1973) found only minor abnormalities in man (slowing of the alpha frequency, increased theta wave occurrence) and --in only 2 cases -- significant delta anomalies. They emphasize the almost normal character of the EEG recorded in 2 patients during the
EXPERIMENTAL KURU IN RHESUS MONKEY terminal phase of the disease (1 m o n t h before the death of one, and 2 days prior to the death of the second). In the chimpanzee, Niedermeyer et al. (1972) and Asher et al. (1973) observed only a disorganized and moderately slow EEG pattern with no irritative elements. In fact, from the disease in man to the disease in the chimpanzee and to that in the rhesus m o n k e y , there seems to be a gradual displacement of the zone of maximal lesions from the cerebellum to the cerebral cortex. Beck and Daniel (1975) showed that, in man 'in fact it is the cerebellum and in particular its phylogenetically older parts that bear the brunt of the pathological process'. In the chimpanzee, however, the cerebellar lesions are less serious and tend to be eclipsed by the lesions affecting the cerebral cortex. A clinical study in the rhesus m o n k e y (Comte-Devolx et al., in preparation) showed that cerebellar signs were discrete or completely absent; this was confirmed by neuropathological examination (Gambarelli et al., in preparation). These differences in the target of the pathological process explain the diversity of the EEG anomalies recorded in man, in the chimpanzee and in the rhesus m o n k e y . The electrographic character of Kuru in the rhesus m o n k e y is also very different from that of other human or experimental spongiform encephalopathies. A periodic paroxysmal activity is frequently observed in Creutzfeldt--Jakob disease (70%, Traub et al. 1977), b u t was never recorded in rhesus monkeys. It should be noted, however, that Capon et al. (1976), referring to a case of Creutzfeldt--Jakob disease, describe a sleeping state p h e n o m e n o n caused by noxious stimulation, which appears to be very similar, if n o t identical, tb the 'tonic seizures' characterizing Kuru in the rhesus m o n k e y . During the evolution of experimental Creutzfeldt--Jakob disease in the chimpanzee, Court et al. (1975) describe the development of slow activity with the appearance of periodic complexes in the waking state, while sleep becomes increasingly disorganized with the development of slow abnormalities. However, they did n o t ob-
617 serve the irritative phenomena characterizing the EEG of Kuru in the rhesus monkey. In work on squirrel monkeys, some of which were inoculated with a strain of transmissible mink encephalopathy (TME) and others with a strain of Creutzfeldt--Jakob disease (CJD), Grabow et al. (1973, 1976) observed the development of slow abnormalities with the appearance of periodic paroxysmal activity in a few monkeys (2 TME-inoculated m o n k e y s and 1 CJD-inoculated monkey). Moreover, they noted the occurrence of epileptiform activity, m y o c l o n u s and seizures in several TME-inoculated monkeys. This shows the diversity of the EEG anomalies attributable to slow virus diseases of the CNS. This diversity results in part from the different topographies of the lesions, b u t certainly involves other less obvious factors. The EEG modifications during the evolution of experimental Kuru develop in welldefined steps rather than as a continuous, gradual process. The EEG pattern remains normal during the 16-months incubation period, then shows the modified sleep patterns of the prodromal phase, followed by the very rapid full development of the abnormalities characterizing the mature phase. Then it remains unchanged until the terminal phase of the disease. This t y p e of evolution suggests the existence of thresholds for lesions -- or for metabolic disturbances -- b e y o n d which certain types of functions can no longer be normally assured. In scrapie, an animal affection transmitted by a similar infectious agent, Kimberlin (1976) notes that 'the clinical disease develops when the compensatory mechanism can no longer cope with the increasing a m o u n t of agent-induced damage'. In fact, the troubles observed cannot be directly explained by a definite structural lesion. The initial phase is characterized by appearance of sleep disturbances (lighter sleep, with diminished REM). Aside from the above-mentioned morphological changes, these sleep disturbances are n o t specific, and may be observed in a variety of conditions
618
under the effect of widely differing factors. They may be interpreted from the standpoint of the current monoaminergic theories of sleep (Jouvet, 1972). Although neuropathological examination reveals no lesions of the regulatory structures of the brain stem, the role of these mechanisms cannot be ruled out. The neurochemical activity of these structures may be perturbed by the development of biochemical or metabolic processes which remain as yet unidentified. Dickinson and Fraser (1977) postulate that, in scrapie: 'Undetermined biochemical lesions produce behavioral changes from an early stage of incubation, and these lesions are the most likely cause of death.' It is nevertheless possible that the developing cortical and diencephalic lesions constitute another factor in determining this sleep trouble. The major disruption of sleep in the mature phase appears to be much more complex. The morphological characteristics and cyclic organization of normal sleep are no longer present, but have been superseded by a new, more simple, EEG organization. This pathological form of sleep seems to be considerably more resistant and persists unmodified until the death of the animal. No definite lesion can account for this disruption. McGinty et al. (1974) emphasize that 'the control of SWS appears to involve the integration of neural processes at several levels of the neuraxis'. This generalized alteration of sleep during the mature phase is a result of extensive lesions affecting the cortex, thalamus, basal ganglia, and corresponds to the demential disorganization of the behavioural sphere. A similar problem is raised by the development of the irritative signs. Kuru in the rhesus monkey is an epileptogenic encephalopathy and constitutes an EEG model -- of considerable realism in at least one animal (Mm 0 7 ) - - of a serious form of human epilepsy, the Lennox--Gastaut syndrome (Bert et al. in preparation). A number of basic differences distinguish this experimental disease" from human epilepsy, and extreme
J. B E R T E T AL.
caution is required in formulating similitude criteria. Nevertheless one aspect may be emphasized. The full development of the experimental model of this epilepsy (characteristic EEG abnormalities, nocturnal tonic seizures, behavioural deterioration) occurs during the mature phase of the disease. Therefore, the experimental reproduction of this complex of clinical and EEG phenomena requires very widespread lesions. This is partly in agreement with computerized tomography findings in patients with the Lennox--Gastaut syndrome (Gastaut and Gastaut 1976). Experimental Kuru in the rhesus monkey constitutes an extremely complex model of functional disorganization of the CNS and the problems raised by it extend well beyond the scope of transmissible dementia.
Summary EEG patterns recorded in the waking state and during sleep were studied in 6 rhesus monkeys inoculated with a strain of Kuru previously passaged in rhesus monkey (ENAGE strain, rhesus L6 56). The onset of the disease was confirmed by the appearance of various clinical signs in 4 monkeys 15 months after inoculation. At the 16th month, the first EEG modifications appeared during sleep, which became lighter. The waking EEG was abnormal during the mature phase of the disease; it was characterized by slow anomalies and scattered spikes. The sleep EEG still presented 3 stages of Slow Wave Sleep which, however, were totally unlike the physiological stages. REM sleep rapidly disappeared, as did the cyclic organization pattern. Irritative phenomena became very significant and, in particular, very frequent 'tonic seizures' were observed. Experimental Kuru thus appears, in the rhesus monkey, as an epileptogenic encephalopathy, which is differentiated from both the human disease and the experimental disease in the chimpanzee.
EXPERIMENTAL KURU IN RHESUS MONKEY
R~sum~ Le Kuru expdrimental chez le rhesus. E t u d e des altdrations E E G p e n d a n t la veille et au cours du s o mmeil L ' E E G de veille et de s o m m e i l est ~tudi~ chez 6 singes rhesus inocul~s avec u n e s o u c h e de K u r u d~j~ pass~e sur singe rhesus ( E N A G E strain, rhesus L6 5~3). L ' a p p a r i t i o n de signes cliniques de t y p e divers a n n o n c e le d ~ b u t de la maladie 15 m o i s apr~s l ' i n o c u l a t i o n chez 4 singes. A u 1 6 ~ m e mois, les premieres m o d i fications E E G apparaissent d u r a n t le s o m m e i l qui d e v i e n t plus l~ger. A la phase d ' ~ t a t de la maladie, I ' E E G de veille est a n o r m a l ; il se caract~rise par des a n o m a l i e s lentes et des p o i n t e s diss~min~es. L ' E E G de s o m m e i l pr~sente e n c o r e trois stades de s o m m e i l ~ o n d e s lentes mais t o t a lement diff~rents des stades physiologiques. Le s o m m e i l p a r a d o x a l dispara~t rapidem e n t . L ' o r g a n i s a t i o n c y c l i q u e n ' e x i s t e plus. Les signes irritatifs s o n t tr~s i m p o r t a n t s et o n observe, en particulier, de tr~s n o m b r e u s e s crises t o n i q u e s . Le K u r u e x p e r i m e n t a l apparai't d o n c chez le singe rhesus c o m m e u n e e n c ~ p h a l o p a t h i e ~pileptog~ne. I1 se diff~rencie aussi bien de la maladie h u m a i n e que de la maladie exp~rim e n t a l e du c h i m p a n z ~ . Our thanks are due to Drs. Fran~oise Cathala, Carleton Gajdusek, Clarence Gibbs and Robert Naquet, for advice and profitable discussions. Dr. Fran~oise Cathala kindly provided us with the strain of Kuru. References Asher, D.M., Gibbs, C.J., David, E., Alpers, M.P. and Gajdusek, D.C. Experimental kuru in the chimpanzee. Non-human primates and human diseases (Symp. IVth Int. Cong. Primat.). Karger, Basel, 1973: 43--90. Beck, E. and Daniel, P.M. Neuropathological studies in primates suffering from experimental kuru or Creutzfeldt--Jakob disease. In B.S. Meldrum and C.D. Marsden (Eds.), Advances in Neurology,
619 Vol. 10. Raven, New York, 1975: 341--346. Bert, J., Pegram, V., Rhodes, J.M., Balzamo, E. and Naquet, R. A comparative sleep study of two cercopithecinae. Electroenceph. clin. Neurophysiol., 1970, 28: 32--40. Bert, J., Vuillon-Cacciuttolo, G., Balzamo, E., De Mico, P., Gambarelli, D. and Tamalet, J. Precocious modifications in the evolution of experimental kuru in rhesus monkeys. Neuroscience Letters, 1977, 6: 333--338. Capon, A., Colin, F., Deltenre, P., Hubert, J.P. et Flament-Durand, J. Etude ~lectrophysiologique de deux cas de maladie de Creutzfeldt--Jakob. Rev. EEG Neurophysiol., 1976, 6 : 130--136. Cobb, W.A., Hornabrook, R.W. and Sanders, S. The EEG of kuru. Electroenceph. clin. Neurophysiol., 1973, 34: 419--427. Court, C., Cathala, F., Gajdusek, D.C. et Rohmer, F. Modifications du comportement, de la vigilance et des activit~s ~lectriques c~r~brales au cours d'une maladie de Creutzfeldt--Jakob exp~rimentale du chimpanz~. Rev. EEG Neurophysiol., 1975, 5: 335--343. Dickinson, A.G. and Fraser, H. Scrapie. Pathogenesis in inbred mice: an assessment of host control and response involving many strains of agent. In V. ter Meulen and M. Katz (Eds.), Slow Virus Infections of the Central Nervous System. Springer, New York, 1977, 258 p. Gajdusek, D.C., Gibbs, C.J. and Alpers, M. Experimental transmission of a kuruqike syndrome to chimpanzees. Nature, 1966, 209: 794--796. Gibbs, C.J. and Gajdusek, D.C. Infection as the etiology of spongiform encephalopathy (Creutzfeldt-Jakob disease). Science, 1969, 165: 1023--1025. Gastaut, H. and Gastaut, J.L. Computerized transverse axial tomography in epilepsy. Epilepsia, 1976, 17: 325--336. Grabow, J.D., Zurhein, G., Eckroade, R.J., Zollman, P.E. and Hanson, R.P. Transmissible mink encephalopathy agent in squirrel monkeys. Neurology (Minneap.), 1973, 23: 820--832. Grabow, J.D., Campbell, R.J., Okazaki, H., Schut, L., Zollman, P.E. and Kurland, L.T. A transmissible subacute spongiform encephalopathY in a visitor to the Eastern Highlands of New Guinea. Brain, 1976, 99: 637--658. Jouvet, M. The role of monoamines and acetylcholine containing neurons in the regulation of the sleep--waking cycle. Ergebn. Physiol., 1972, 64: 166--307. Kimberlin, R.H. Biochemical and behavioral changes in scrapie. In R.H. Kimberlin, (Ed.), Slow Virus Diseases of Animals and Man. North-Holland, Amsterdam, 1976: 307--323. McGinty, D.J., Harper, R.M. and Fairbanks, M.K. Neuronal unit activity and the control of sleep
620 states. In E.D. Weitzman (Ed.), Advances in Sleep Research, Vol. 1. Spectrum, New York, 1974, 424 p. Niedermeyer, E., Gibbs, C.J. and Marsh, R. EEG studies in subacute and degenerative neurological diseases experimentally produced in ferrets and chimpanzee. Electroenceph. clin. Neurophysiol.,
J. BERT ET AL. 1972, 33: 351--352. Traub, R., Gajdusek, D.C. and Gibbs, C.J. Transmissible virus dementia: the relation of transmissible spongiform encephalopathy to Creutzfeldt--Jakob disease. In M. Kinsbourme and L. Smith (Eds.), Aging, Dementia, and Cerebral Functions. Spectrum, New York, 1977: 91--172.
611
E X P E R I M E N T A L K U R U IN THE R H E S U S MONKEY: A STUDY OF EEG MODIFICATIONS IN THE WAKING STATE AND D U R I N G SLEEP * J. B E R T a, G. V U I L L O N - C A C C I U T T O L O a, E. BALZAMO a, p. DE MICCO J. T A M A L E T b and H. G A S T A U T a
b, D.
G A M B A R E L L I c,
a Service de Neurophysiologie Clinique et C.R.P. 444, C.N.R.S., b Laboratoire de Virologie, and c Laboratoire de Neuropathologie, C.H.U. Timone, Marseilles (France) (Accepted for p u b l i c a t i o n : April 4, 1978)
Kuru, a degenerative disease of the central nervous system (CNS) occurring among a certain population in New Guinea, was shown to be transmissible to subhuman primates by Gajdusek and Gibbs in 1966. In 1969, Gibbs and Gajdusek similarly established the transmissibility of Creutzfeldt--Jakob disease which, like Kuru, is caused b y unconventional infectious agents. These authors thus opened up a new chapter in the aetiology of diseases affecting the human CNS. The determination of the aetiology is difficult: the transmissibility to animals, in fact, constitutes the only formal criterion of it. This problem emphasizes the practical interest of studying experimental diseases in primates, making it possible: (1) to list all the clinical, electrophysiological and neuropathological effects of the disease agents, and (2) to analyse all the evolutionary phases o f pathological processes which are most often perceived only in the terminal phases in human neurology. This work on experimental Kuru in the rhesus m o n k e y was carried o u t from this viewpoint.
Material and m e t h o d Early in 1975, 13 rhesus m o n k e y s (Macaca mulatta) were subjected to surgical implantation of electrodes (4 mm diameter stainless steel screws) at the surface of the dura mater * This work was in part supported by a grant from the F r e n c h F o u n d a t i o n for Medical Research.
for EEG recording and at the orbital peripheries for eye m o v e m e n t recording. Eight monkeys were inoculated with a strain of Kuru passaged in rhesus m o n k e y s in Dr. Gajdusek's laboratory at Bethesda (ENAGE strain rhesus L6 56): intracerebral inoculations (0.2 ml of 10 -2 dilution of a 20% cerebral solution in P.B.S., pH 7.4, introduced in the left frontal lobe through the r o o f of the orbit) and intravenous inoculations (0.2 ml of 10 -2 cerebral suspension) were performed. Six m o n k e y s were inoculated in May and June 1975: three (Mm 03, 06, 14) with a strain unfrozen for the first time less than I h prior to inoculation and 3 others (Mm 04, 05, 07) with a strain unfrozen at the time of the inoculation of Mm 03, 06, 14, then refrozen and thawed again. Two animals (Mm 23, 24) were inoculated in December 1975, with a strain which had been unfrozen during each of the previous inoculations. The first 6 animals were adult at the time of inoculation, while the latter 2 were between 8 and 12 months old in December 1975. A control group of 5 m o n k e y s was kept under identical conditions. These animals showed no abnormalities at any time during several recording sessions after implantation and up to July 1977. EEG records were made every 4 months from all of the inoculated monkeys. The animals were placed in restraining chairs several days prior to the session in order to familiarize them with the experimental conditions. Each recording began at a b o u t 5.45 pm
612 in order to obtain an EEG sample for the waking state in lit surroundings. The lights were switched off at 6.00 pm until 6.00 am the following day, at which time recording was terminated. Moreover, at least one continuous 24-h record was made for each animal to establish the normal daily amounts of sleep. With the appearance of the initial symptoms, the recording intervals were shortened, first to monthly EEG sessions and then, according to the development of the disease, every 2 weeks. A total of 80 records was made in this way. The recording intervals no doubt led to a loss of data; nevertheless, the evolution of the disease was monitored with sufficient accuracy. The records were analysed in 15-sec periods according to the criteria adopted in this laboratory for the stages of sleep among Cercopithecinae (Bert et al. 1970). The results were then entered on punch cards for computer processing. When physiological criteria became impractical, new criteria were defined, and 3 stages of Slow Wave Sleep (SWS) were identified (a, b, c). Two monkeys (Mm 04, 14) have shown no formal signs of illness to date, and have been disregarded for the purpose of this study. The 6 remaining animals were killed as late as possible in view of their clinical condition: on average 700 ± 40 days after inoculation (between the 645th and the 762nd postinoculatory day). The data of serial clinical examinations (Comte-Devolx et al., in preparation) and neuropathological findings (Gambarelli et al., in preparation) will be presented independently. Results
In view of the variability of the initial symptomatology following the incubation period, two evolutionary phases must be discriminated for Kuru: an initial prodromal phase and a mature phase.
J. BERT ET AL.
(1) The initial phase includes isolated or associated clinical and EEG signs. (a) In 4 monkeys, the first isolated clinical signs appeared after about 15 months. Two animals (Mm 03, 06) showed behavioural troubles (disinterest, inertia, modified feeding habits). The other 2 monkeys presented slight (Mm 23) or grave (Mm 24) myoclonic syndromes with no behavioural troubles. The remaining two animals (Mm 05, 07) showed no precocious clinical troubles. (b) Throughout the prodromal phase, the EEG recorded in the waking state remained normal for 5 monkeys (Mm 03, 05, 06, 23, 24). In the sixth (Mm 07), bursts of spike and wave complexes at 3--4 c/sec were recorded in the frontal regions shortly before the onset of the mature phase of the disease (20th postinoculatory month). (c) Sleep pattern modifications began to appear after 16 months. The physiological stages could be differentiated without difficulty. Nevertheless during stage 1 the alpha frequency was slowed by 1--2 c/sec and the amplitude of the beta activity was considerably reduced. The sleep spindles disappeared during stage 2. The transition from the waking state to sleep was seriously perturbed, presenting a long initial period of light sleep (stages 1 and 2), interrupted by frequent awakenings. The latency of the first episode of stage 4 and of the first episode of Rapid Eye Movement (REM) sleep increased considerably, and the total duration of these stages gradually decreased. Thus the stage 4 sleep normally occurring in the first few hours of sleep was retarded until the middle of the night. Furthermore, spontaneous transient awakening reactions, lasting 2--5sec and often preceded by a high voltage wave, were noted. Such awakening reactions are occasionally observed in normal rhesus monkeys, but are rather rare. These modified sleep patterns, which were discussed in a previous paper (Bert et al. 1977), were not observed in one of the monkeys (Mm 23).
EXPERIMENTAL KURU IN RHESUS MONKEY
(2) The mature phase commenced from 600 to 700 days after inoculation. It was characterized by major EEG alterations and by the development of a true dementia syndrome with generalized myoclonic jerks in 4 animals (Mm 03, 05, 06, 07) and an extrapyramidal syndrome with myoclonic jerks and minor behavioural troubles in the other 2 (Mm 23, 24). (a) The waking EEG recorded with eyes open was abnormal. It was characterized by irregular 5--7 c/sec activity of moderate amplitude (50--100~V), which was replaced by continuous slower activity (4--5 c/sec) of larger amplitude (100--150 ~V) as the animal's attention diminished. This was associated with frequent spikes, bilateral and generalized or multifocal (the majority of these spikes were recorded from the electrodes implanted along the midline, but independent spikes were noted in the temporal regions). Monkey Mm 07 showed major frontal bursts of 3 c/ sec spike~nd-wave complexes. In general, these irritative phenomena became increasingly pronounced when the animal's vigilance diminished, as measured by the drop in blinking frequency. The irritative signs were more significant in the left temporal region in 3 monkeys (Mm 03, 06, 07). (b) Although the total duration of sleep remained constant, its nature was so profoundly altered that it became impossible to refer to the classification according to physiological stages, for which the major identifying criteria had disappeared. The sleep was nevertheless polyphasic and SWS continued to present three easily identifiable stages, which persisted until the terminal phase of the disease without modification. (1) Stage 'a' appeared as soon as the animal closed its eyes. The activity recorded on the waking EEG became slower (3--4 c/sec) and of larger amplitude (100--200 ~V), and spiking became more important. In one animal (Mm 07) the EEG pattern at the terminal phase resembled an almost continuous discharge of spikes or spike-and-wave complexes (Fig. 1).
613
M.m. 07 14
_06
-
77
....9 100pV
Fig. 1. Complex slow pattern with spikes or spike-andwave discharges, in some cases bilateral and synchronous on both hemispheres, while asynchronous in other cases.
(2) Stage 'b' was characterized by mediumamplitude (75--150lzV) activity at 7--8c/ sec with occasional sporadic waves or short bursts of waves at 2--3 c/sec. Neither spindles nor V waves were observed. The spikes became much rarer, and most often then recorded from midline electrodes. Nevertheless, in the terminal phase Mm 07 presented generalized 2c/sec spiking discharges (Fig. 2). (3) Stage 'c' rather closely resembled stages 3 and 4 of normal sleep. The EEG showed increasing 1--3 c/sec waves of large amplitude (150--200 ~V) which rapidly became continuous. The irritative phenomena had practically disappeared. (4) REM sleep persisted in rare and very short episodes of no more than 20 sec. A few isolated eye movements occurred, but the bursts ceased. REM sleep completely disM.m. 07 24 - 05
- 77
I Set.
Fig. 2. Generalized bursts of spike-and-wave complexes recorded during stage 'b'.
614
J. BERT ET AL. M . m. 0 6
s th
20
month
month
th
Iil, '
21 t h
month
REM
o I 6 p.m.
6 o.m.
Fig. 3. Modified organization of sleep during the evolution of experimental Kuru. Six months after inoculation, the organization is typical of normal sleep in the rhesus monkey. During the 20th month, the physiological stages remain identifiable but are characterized by very long periods of light sleep and awakenings, with stage 4 retarded until the second half of the night. During the 21st month the sleep pattern is morphologically abnormal. It is not possible to use the classification according to physiological stages. SWS presents 3 abnormal patterns, classified as stages a, b, c, and its organization is severely perturbed.
appeared between the 615th and 708th day after inoculation. This morphological transformation of the sleeping EEG was accompanied by a general disorganization of sleep itself and the blurring of its cyclic pattern (Fig. 3). The various stages followed one another interspersed by periods of wakening, but no longer constituting veritable cycles. The stage 'c' periods tended to occur throughout the night, and not simply in the first few hours of sleep as with stages 3 and 4 of physiological sleep. However, the REM sleep episodes occurred primarily in the second half of the night, and represented the final aspects of the temporal
19_10_77
~ lOO~v 1 Sec.
6-7~
~
~
~
~
9-10~
Fig. 4. Brief awakening reaction after a massive m y o -
clonic jerk accompanied by generalized artifacts but with no repercussions in the EEG.
EXPERIMENTAL K U R U IN RHESUS MONKEY
organization of true physiological sleep. These 3 stages of SWS persisted throughout the remaining development of the disease. In the last record of one monkey (Mm 05, 729th day after inoculation), the amplitude of all activities tended to drop; the stage 'c' episodes became more widely spaced and the delta waves less abundant. Moreover, 3types of transient phenomena were observed during sleep: (1) Brief awakening reactions, less frequent than during the prodromal phase. These reactions sometimes followed a massive myoclonic jerk (Fig. 4). (2) Brief (2--4 sec) episodes of flattening resulting from the suppression * throughout the cortex of all theta and alpha activity, followed immediately by the unaltered resumption of the preceding pattern (Fig. 5A and B). When these theta and alpha activities were superimposed on lower frequency waves, the latter sometimes persisted or were suppressed as well. Such episodes might or might not be accompanied by a tonic contraction of the facial muscles. (3) 'Tonic seizures' involving a more complex event sequence (Fig. 5C and D, and Fig. 6). Occasionally preceded by more or less generalized discharge of a spike or spike burst, the 'seizure' itself began by the flattening of baseline activity for 2--6 sec, followed by a generalized discharge of sharp waves or sharp-and-slow wave complexes at 1--2 c/sec. A tonic contraction of the facial muscles occurred simultaneously, as shown by electromyographic activity from the electrodes implanted around the eyes. Such 'seizures' were not all identical. In some cases the sharp wave discharges appeared simultaneously with the flattening of background activity. Both forms might occur in the same animal during a single night. The number of ictal episodes (episodes of flattening and 'tonic seizures') increased very * In this paper the word 'suppression' is used with a merely descriptive meaning without any physiological connotation.
615 M.m. 05 5_05_77
~
loo~
1 $~.
Fig. 5. Brief episodes during which background activity is suppressed during stage 'b'. Such episodes may be isolated (A) or followed by a pseudorhythmic sharp wave discharge of variable duration (B--C--D). Example D shows a brief reinforcement of the EMG activity on the oculogram lead.
rapidly, reaching or exceeding 250 per night (Mm 05) in the terminal phase. The total duration of the 'seizures' then represented 18% of the sleeping time. Sensory stimuli (e.g. noise) could result in a brief awakening reaction or in a 'tonic seizure'. These ictal episodes appeared only after a certain minimal period of sleep, and did not occur during stage 'a'. Rather, they appeared essentially during stage 'b', while the stage 'c' periods most often ended in a 'tonic seizure'. 'Tonic seizures' of short or moderate duration did not interrupt the stage 'b' episodes, and occurred repeatedly separated by interictal tracings of a few seconds to nearly 1 min. Longer seizures were terminated by the transition to stage 'a' or by awakening of the
616
J. BERT ET AL.
M.m. 06 3 - 0 5 - 77
~ IOO~v 1 Sec.
A ,-2
.
.
.
.
.
.
,.
,,.~,7,:
~A~'~ '~-'~
....
I
,-~ ~
~, . . . . .
'""':""'""~'~~'~
I
,
~
~
~
~
,
~
,
"'"
........
,
~
~¢~ ~ , , ~ ~ v ~ 2 ~ , ¢
~
B
Fig. 6. A a n d B: ' T o n i c seizures' w i t h initial f l a t t e n i n g of b a c k g r o u n d activity f o l l o w e d by p s e u d o r h y t h m i c s h a r p waves a c c o m p a n i e d b y t o n i c c o n t r a c t i o n o f t h e facial muscles. I n e x a m p l e B, a s e c o n d ' s e i z u r e ' a p p e a r s a l m o s t i m m e d i a t e l y a f t e r t h e e n d o f t h e first episode.
animal. That the onset of such seizures required a minimal a m o u n t of stage 'b' sleep explains their relative absence during the day, when the EEG tended to oscillate between the waking state and stage 'a', with a few rare episodes of stage 'b' sleep. The maximum observed involved 11 daytime seizures with a burst of seizures after 8 min of continuous sleep (Mm 06, on the 691st day after inoculation). Only a single animal (Mm 23) presented no tonic seizures.
Discussion
The development of experimental Kuru in rhesus m o n k e y s is accompanied by peculiar
EEG modifications in the waking state and during sleep. The dominant character of these alterations concerns the importance of irritative signs and the occurrence of multiple nocturnal tonic seizures. These characters appear to be peculiar to the experimental disease in the rhesus m o n k e y . They were n o t detected in the disease in man {Cobb et al. 1973) or in the experimental disease in the chimpanzee (Niedermeyer et al. 1972; Asher et al. 1973). Cobb et al. (1973) found only minor abnormalities in man (slowing of the alpha frequency, increased theta wave occurrence) and --in only 2 cases -- significant delta anomalies. They emphasize the almost normal character of the EEG recorded in 2 patients during the
EXPERIMENTAL KURU IN RHESUS MONKEY terminal phase of the disease (1 m o n t h before the death of one, and 2 days prior to the death of the second). In the chimpanzee, Niedermeyer et al. (1972) and Asher et al. (1973) observed only a disorganized and moderately slow EEG pattern with no irritative elements. In fact, from the disease in man to the disease in the chimpanzee and to that in the rhesus m o n k e y , there seems to be a gradual displacement of the zone of maximal lesions from the cerebellum to the cerebral cortex. Beck and Daniel (1975) showed that, in man 'in fact it is the cerebellum and in particular its phylogenetically older parts that bear the brunt of the pathological process'. In the chimpanzee, however, the cerebellar lesions are less serious and tend to be eclipsed by the lesions affecting the cerebral cortex. A clinical study in the rhesus m o n k e y (Comte-Devolx et al., in preparation) showed that cerebellar signs were discrete or completely absent; this was confirmed by neuropathological examination (Gambarelli et al., in preparation). These differences in the target of the pathological process explain the diversity of the EEG anomalies recorded in man, in the chimpanzee and in the rhesus m o n k e y . The electrographic character of Kuru in the rhesus m o n k e y is also very different from that of other human or experimental spongiform encephalopathies. A periodic paroxysmal activity is frequently observed in Creutzfeldt--Jakob disease (70%, Traub et al. 1977), b u t was never recorded in rhesus monkeys. It should be noted, however, that Capon et al. (1976), referring to a case of Creutzfeldt--Jakob disease, describe a sleeping state p h e n o m e n o n caused by noxious stimulation, which appears to be very similar, if n o t identical, tb the 'tonic seizures' characterizing Kuru in the rhesus m o n k e y . During the evolution of experimental Creutzfeldt--Jakob disease in the chimpanzee, Court et al. (1975) describe the development of slow activity with the appearance of periodic complexes in the waking state, while sleep becomes increasingly disorganized with the development of slow abnormalities. However, they did n o t ob-
617 serve the irritative phenomena characterizing the EEG of Kuru in the rhesus monkey. In work on squirrel monkeys, some of which were inoculated with a strain of transmissible mink encephalopathy (TME) and others with a strain of Creutzfeldt--Jakob disease (CJD), Grabow et al. (1973, 1976) observed the development of slow abnormalities with the appearance of periodic paroxysmal activity in a few monkeys (2 TME-inoculated m o n k e y s and 1 CJD-inoculated monkey). Moreover, they noted the occurrence of epileptiform activity, m y o c l o n u s and seizures in several TME-inoculated monkeys. This shows the diversity of the EEG anomalies attributable to slow virus diseases of the CNS. This diversity results in part from the different topographies of the lesions, b u t certainly involves other less obvious factors. The EEG modifications during the evolution of experimental Kuru develop in welldefined steps rather than as a continuous, gradual process. The EEG pattern remains normal during the 16-months incubation period, then shows the modified sleep patterns of the prodromal phase, followed by the very rapid full development of the abnormalities characterizing the mature phase. Then it remains unchanged until the terminal phase of the disease. This t y p e of evolution suggests the existence of thresholds for lesions -- or for metabolic disturbances -- b e y o n d which certain types of functions can no longer be normally assured. In scrapie, an animal affection transmitted by a similar infectious agent, Kimberlin (1976) notes that 'the clinical disease develops when the compensatory mechanism can no longer cope with the increasing a m o u n t of agent-induced damage'. In fact, the troubles observed cannot be directly explained by a definite structural lesion. The initial phase is characterized by appearance of sleep disturbances (lighter sleep, with diminished REM). Aside from the above-mentioned morphological changes, these sleep disturbances are n o t specific, and may be observed in a variety of conditions
618
under the effect of widely differing factors. They may be interpreted from the standpoint of the current monoaminergic theories of sleep (Jouvet, 1972). Although neuropathological examination reveals no lesions of the regulatory structures of the brain stem, the role of these mechanisms cannot be ruled out. The neurochemical activity of these structures may be perturbed by the development of biochemical or metabolic processes which remain as yet unidentified. Dickinson and Fraser (1977) postulate that, in scrapie: 'Undetermined biochemical lesions produce behavioral changes from an early stage of incubation, and these lesions are the most likely cause of death.' It is nevertheless possible that the developing cortical and diencephalic lesions constitute another factor in determining this sleep trouble. The major disruption of sleep in the mature phase appears to be much more complex. The morphological characteristics and cyclic organization of normal sleep are no longer present, but have been superseded by a new, more simple, EEG organization. This pathological form of sleep seems to be considerably more resistant and persists unmodified until the death of the animal. No definite lesion can account for this disruption. McGinty et al. (1974) emphasize that 'the control of SWS appears to involve the integration of neural processes at several levels of the neuraxis'. This generalized alteration of sleep during the mature phase is a result of extensive lesions affecting the cortex, thalamus, basal ganglia, and corresponds to the demential disorganization of the behavioural sphere. A similar problem is raised by the development of the irritative signs. Kuru in the rhesus monkey is an epileptogenic encephalopathy and constitutes an EEG model -- of considerable realism in at least one animal (Mm 0 7 ) - - of a serious form of human epilepsy, the Lennox--Gastaut syndrome (Bert et al. in preparation). A number of basic differences distinguish this experimental disease" from human epilepsy, and extreme
J. B E R T E T AL.
caution is required in formulating similitude criteria. Nevertheless one aspect may be emphasized. The full development of the experimental model of this epilepsy (characteristic EEG abnormalities, nocturnal tonic seizures, behavioural deterioration) occurs during the mature phase of the disease. Therefore, the experimental reproduction of this complex of clinical and EEG phenomena requires very widespread lesions. This is partly in agreement with computerized tomography findings in patients with the Lennox--Gastaut syndrome (Gastaut and Gastaut 1976). Experimental Kuru in the rhesus monkey constitutes an extremely complex model of functional disorganization of the CNS and the problems raised by it extend well beyond the scope of transmissible dementia.
Summary EEG patterns recorded in the waking state and during sleep were studied in 6 rhesus monkeys inoculated with a strain of Kuru previously passaged in rhesus monkey (ENAGE strain, rhesus L6 56). The onset of the disease was confirmed by the appearance of various clinical signs in 4 monkeys 15 months after inoculation. At the 16th month, the first EEG modifications appeared during sleep, which became lighter. The waking EEG was abnormal during the mature phase of the disease; it was characterized by slow anomalies and scattered spikes. The sleep EEG still presented 3 stages of Slow Wave Sleep which, however, were totally unlike the physiological stages. REM sleep rapidly disappeared, as did the cyclic organization pattern. Irritative phenomena became very significant and, in particular, very frequent 'tonic seizures' were observed. Experimental Kuru thus appears, in the rhesus monkey, as an epileptogenic encephalopathy, which is differentiated from both the human disease and the experimental disease in the chimpanzee.
EXPERIMENTAL KURU IN RHESUS MONKEY
R~sum~ Le Kuru expdrimental chez le rhesus. E t u d e des altdrations E E G p e n d a n t la veille et au cours du s o mmeil L ' E E G de veille et de s o m m e i l est ~tudi~ chez 6 singes rhesus inocul~s avec u n e s o u c h e de K u r u d~j~ pass~e sur singe rhesus ( E N A G E strain, rhesus L6 5~3). L ' a p p a r i t i o n de signes cliniques de t y p e divers a n n o n c e le d ~ b u t de la maladie 15 m o i s apr~s l ' i n o c u l a t i o n chez 4 singes. A u 1 6 ~ m e mois, les premieres m o d i fications E E G apparaissent d u r a n t le s o m m e i l qui d e v i e n t plus l~ger. A la phase d ' ~ t a t de la maladie, I ' E E G de veille est a n o r m a l ; il se caract~rise par des a n o m a l i e s lentes et des p o i n t e s diss~min~es. L ' E E G de s o m m e i l pr~sente e n c o r e trois stades de s o m m e i l ~ o n d e s lentes mais t o t a lement diff~rents des stades physiologiques. Le s o m m e i l p a r a d o x a l dispara~t rapidem e n t . L ' o r g a n i s a t i o n c y c l i q u e n ' e x i s t e plus. Les signes irritatifs s o n t tr~s i m p o r t a n t s et o n observe, en particulier, de tr~s n o m b r e u s e s crises t o n i q u e s . Le K u r u e x p e r i m e n t a l apparai't d o n c chez le singe rhesus c o m m e u n e e n c ~ p h a l o p a t h i e ~pileptog~ne. I1 se diff~rencie aussi bien de la maladie h u m a i n e que de la maladie exp~rim e n t a l e du c h i m p a n z ~ . Our thanks are due to Drs. Fran~oise Cathala, Carleton Gajdusek, Clarence Gibbs and Robert Naquet, for advice and profitable discussions. Dr. Fran~oise Cathala kindly provided us with the strain of Kuru. References Asher, D.M., Gibbs, C.J., David, E., Alpers, M.P. and Gajdusek, D.C. Experimental kuru in the chimpanzee. Non-human primates and human diseases (Symp. IVth Int. Cong. Primat.). Karger, Basel, 1973: 43--90. Beck, E. and Daniel, P.M. Neuropathological studies in primates suffering from experimental kuru or Creutzfeldt--Jakob disease. In B.S. Meldrum and C.D. Marsden (Eds.), Advances in Neurology,
619 Vol. 10. Raven, New York, 1975: 341--346. Bert, J., Pegram, V., Rhodes, J.M., Balzamo, E. and Naquet, R. A comparative sleep study of two cercopithecinae. Electroenceph. clin. Neurophysiol., 1970, 28: 32--40. Bert, J., Vuillon-Cacciuttolo, G., Balzamo, E., De Mico, P., Gambarelli, D. and Tamalet, J. Precocious modifications in the evolution of experimental kuru in rhesus monkeys. Neuroscience Letters, 1977, 6: 333--338. Capon, A., Colin, F., Deltenre, P., Hubert, J.P. et Flament-Durand, J. Etude ~lectrophysiologique de deux cas de maladie de Creutzfeldt--Jakob. Rev. EEG Neurophysiol., 1976, 6 : 130--136. Cobb, W.A., Hornabrook, R.W. and Sanders, S. The EEG of kuru. Electroenceph. clin. Neurophysiol., 1973, 34: 419--427. Court, C., Cathala, F., Gajdusek, D.C. et Rohmer, F. Modifications du comportement, de la vigilance et des activit~s ~lectriques c~r~brales au cours d'une maladie de Creutzfeldt--Jakob exp~rimentale du chimpanz~. Rev. EEG Neurophysiol., 1975, 5: 335--343. Dickinson, A.G. and Fraser, H. Scrapie. Pathogenesis in inbred mice: an assessment of host control and response involving many strains of agent. In V. ter Meulen and M. Katz (Eds.), Slow Virus Infections of the Central Nervous System. Springer, New York, 1977, 258 p. Gajdusek, D.C., Gibbs, C.J. and Alpers, M. Experimental transmission of a kuruqike syndrome to chimpanzees. Nature, 1966, 209: 794--796. Gibbs, C.J. and Gajdusek, D.C. Infection as the etiology of spongiform encephalopathy (Creutzfeldt-Jakob disease). Science, 1969, 165: 1023--1025. Gastaut, H. and Gastaut, J.L. Computerized transverse axial tomography in epilepsy. Epilepsia, 1976, 17: 325--336. Grabow, J.D., Zurhein, G., Eckroade, R.J., Zollman, P.E. and Hanson, R.P. Transmissible mink encephalopathy agent in squirrel monkeys. Neurology (Minneap.), 1973, 23: 820--832. Grabow, J.D., Campbell, R.J., Okazaki, H., Schut, L., Zollman, P.E. and Kurland, L.T. A transmissible subacute spongiform encephalopathY in a visitor to the Eastern Highlands of New Guinea. Brain, 1976, 99: 637--658. Jouvet, M. The role of monoamines and acetylcholine containing neurons in the regulation of the sleep--waking cycle. Ergebn. Physiol., 1972, 64: 166--307. Kimberlin, R.H. Biochemical and behavioral changes in scrapie. In R.H. Kimberlin, (Ed.), Slow Virus Diseases of Animals and Man. North-Holland, Amsterdam, 1976: 307--323. McGinty, D.J., Harper, R.M. and Fairbanks, M.K. Neuronal unit activity and the control of sleep
620 states. In E.D. Weitzman (Ed.), Advances in Sleep Research, Vol. 1. Spectrum, New York, 1974, 424 p. Niedermeyer, E., Gibbs, C.J. and Marsh, R. EEG studies in subacute and degenerative neurological diseases experimentally produced in ferrets and chimpanzee. Electroenceph. clin. Neurophysiol.,
J. BERT ET AL. 1972, 33: 351--352. Traub, R., Gajdusek, D.C. and Gibbs, C.J. Transmissible virus dementia: the relation of transmissible spongiform encephalopathy to Creutzfeldt--Jakob disease. In M. Kinsbourme and L. Smith (Eds.), Aging, Dementia, and Cerebral Functions. Spectrum, New York, 1977: 91--172.