- Email: [email protected]
Role of afferent input of subcortical origin in the genesis of bilaterally synchronous epileptic discharges of feline generalized penicillin epilepsy
EXPERIMENTAL
NEUROLOGY
64, 155- 173 (1979)
Role of Afferent Input of Subcortical Origin in the Genesis of Bilaterally Synchronous Epileptic Discharges of Feline Generalized Penicillin Epilepsy A. PELLEGRINI,
J. MUSGRAVE,AND
P. GLOOR'
Montreal Neurological Institute, 3801 University Street, Montreal, Quebec, H3A 284 Canada Received October 13. 1978 The effects of (i) neural isolation of large bilateral cortical slabs with intact callosal connections, and (ii) of large unilateral subtotal thalamectomies associated with transection of the interhemispheric commissures on the generalized epileptiform discharges of generalized penicillin epilepsy of the cat were studied. Cortex devoid of subcortical, particularly thalamic, inputs was incapable of generating the characteristic 3.5- to 5-cycles/s epileptiform bursts of feline generalized penicillin epilepsy either in response to the i.m. injection of a large dose of penicillin or to the diffuse direct cortical application of a dilute penicillin solution. Such deafferented cortex, however, produced, even without penicillin, widespread often highly synchronized epileptiform discharges of a different kind consisting at times of slow, l- to 2.5-cycles/s spike and wave complexes. This epileptiform activity increased in response to i.m. penicillin injection or to widespread direct cortical application of a dilute penicillin solution, but never assumed the characteristics of the typical discharges of feline generalized penicillin epilepsy as seen in cortex which has retained its subcortical connections. The discharges differed from those of intact cortex by being slower in frequency and less prone to become organized in bursts. In the bilateral slab preparation with preserved callosal connections, these atypical discharges occurred, however, in a bilaterally synchronous manner, as did the typical discharges in the intact cat with generalized penicillin epilepsy. This suggests that the corpus callosum is necessary for the bilateral synchrony of both types of epileptiform discharges.
Abbreviation: EEG, electroencephalogram. 1 This work was supported by Research Grant MT-3 140 of the Medical Research Council of Canada. Address reprint requests to Dr. P. Gloor. Dr. Pellegrini’s present address is Clinica Neurologica dell’ Universita, Via Giustiniani 1, 35100 Padua, Italy. 155 0014-4886/79/040155-19$02.00/O Copyright 0 1979 by Academic Press. Inc. All rights of reproduction in any form reserved.
156
PELLEGRINI,
MUSGRAVE,
AND
GLOOR
INTRODUCTION Previous investigations (7, 8, 20) suggested that the synchronous epileptic bursts occurring in generalized penicillin epilepsy of the cat are closely related to spindles and/or recruiting responses which, in normal animals, are induced by thalamocortical volleys. Much of this evidence was obtained from stimulation studies which showed that single shock or low-frequency thalamic stimulations which, in normal animals, induced spindles or recruiting responses, elicited generalized bilaterally synchronous epileptic bursts, often assuming a spike and wave form, in animals having received a large dose of intramuscular penicillin. These experiments, however, did not conclusively demonstrate that thalamocortical inputs were necessarily involved in the genesis of the spontaneously occurring epileptic discharges in this model. The present series of experiments was therefore undertaken to determine whether or not disconnection of the cortex from all or nearly all inputs arising from subcortical, particularly thalamic, structures would result in the absence of the epileptic bursts typical for this condition in cortex disconnected in this manner. Two preparations were used for this purpose: (i) large symmetrical neuronally isolated cortical slabs of both hemispheres which remained interconnected through the corpus callosum, prepared in a manner similar to that described by Marcus and Watson (16); and (ii) unilateral thalamectomy in cats with a transsected corpus callosum. Both preparations were chronic and recordings were carried out after the animals had fully recovered from the surgical trauma. METHODS Surgical procedures were done under pentobarbital (Nembutal) anesthesia using sterile techniques and the operating microscope. Pre- and postoperative care with i.v. and S.C. fluids, systemic antibiotics, and dexamethasone was given. Cortical Isolation Experiments. Four cats weighing 3 to 5 kg were used in these three-stage experiments. In the first stage, a wide symmetrical craniectomy was made which extended from the coronal suture posteriorly for about 2 cm and laterally as far as the medial border of the middle ectosylvian gyrus. The bone was partly replaced by an acrylic cranioplasty in which stainless-steel screw electrodes and a multiple-pin miniature connector were imbedded. In the fourth animal, the screws were placed in the outer table of the cranial vault, and bone flaps later were cut around them. Of the regions to be included in the isolated slab the activity of the middle suprasylvian gyrus was recorded in all animals and that of the lateral gyrus in two. The interelectrode distance was 5 mm. Additional recordings
SUBCORTICAL
INPUT IN GENERALIZED
EPILEPSY
157
FIG. 1. Schematic drawing showing bilateral neuronally isolated cortical slab with intact callosal connections.
were obtained from regions of cortex anterior, lateral, and posterior to those to be included in the isolated slab, consisting of the sigmoid, ectosylvian, and posterior parts of the lateral gyri, respectively. After the animals had recovered from the surgical procedure, control electroencephalograms (EEGs) before and after an i.m. injection of 300,000 to 400,000 IU/kg sodium penicillin G were obtained. In the second stage, the cranioplasty was elevated, and a wide dural flap was hinged to the midline. The medial portion of the middle ectosylvian gyrus was removed by suction to give working space, and the corona radiata was cut across until the lateral ventricle was entered (Fig. 1). The slab was then completed by aspirating a thin coronal trench at either end through pial windows in the suprasylvian and lateral gyri and gradually deepening these to reach the ventricle. The cingulate and retrosplenial gyri were cut across subpially, and the coronal trench was carried into the corpus callosum anteriorly, usually just above the intraventricular foramen (Fig. 1). The posterior line of section passed behind the splenium of the corpus callosum. All arteries and most veins crossing the incision line were preserved in pial bridges. In the last animal, the pillar of the fomix was severed, and the fimbria or anterior hippocampus was cut across diagonally to reach the posterior midline at the splenium. A water-tight dural closure was made with 7-O silk before the cranioplasty or bone flap was replaced. At least 2 weeks later the same procedure was carried out on the other side, thus completing the isolation of the slab of bilateral cortical tissue, the isolated regions of each
158
PELLEGRINI,
MUSGRAVE,
AND
GLOOR
hemisphere remaining interconnected by the corpus callosum (Fig. 1). At the close of the experiments, the circulation in the slab was studied by fluorescein angiography (4) and radionucleide scanning with Xe133 (11) and/or KF5 (13, 22) and was found to be normal except for some small regions of relatively slow circulation at the edge of the slab as revealed by the fluorescein angiogram. The intracortical concentration of P4-labeled penicillin G was measured in three of the four animals using the methods reported by Quesney and Gloor (19). The cortical concentration of penicillin inside and outside the slab was essentially the same. Thalamectomy. A row of five stainless-steel screw electrodes was implanted over each middle suprasylvian gyrus at 6-mm intervals, and their recording plugs were screwed to the cranial vault posteriorly in five cats weighing 3 to 4.5 kg. A reference screw electrode was inserted in the occipital bone at the midline. Control recordings of the response to intramuscularly injected penicillin at a dose of 300,000 to 400,000 IU/kg and to intraperitoneal injection of 5 to 10 mg/kg pentobarbital were then made. At a later date a craniectomy about 15 mm wide and 25 mm long was made between the two rows of screw electrodes. The dura was slit open along the left side of the superior sagittal sinus at a distance of about 2 mm from it. Care was taken to leave intact, if at all possible, all bridging veins to the sinus originating from the left hemisphere, but frequently it was necessary to divide some of the smaller ones in the interhemispheric fissure. The corpus callosum and the hippocampal commissures were then divided by suction to expose the third ventricle. The dorsal surface of the right thalamus was exposed by aspirating the fomix and by reflecting the choroid plexus of the lateral ventricle medially. The posterior commissure, massa intermedia, and anterior commissure served as landmarks to guide the removal. The right thalamus was then removed as completely as possible, commencing at the midline, extending ventrally to the inferior limit of the massa intermedia, posteriorly to the mouth of the aqueduct, and anteriorly to the intraventricular foramen. The thalamostriate vein was spared. Lateral dissection under direct vision was limited by the amount of retraction the left hemisphere would tolerate, and the most lateral removal was by necessity carried out blindly, using a suction tip with a right angle, 9 mm in length. Last, the anterior commissure was divided. After careful hemostasis in the removal cavity, the dura was closed with 7-O silk in a water-tight manner. The bone defect was bridged with dental cement over Gelfoam strips, and the skin was closed. In three cats with a unilateral thalamectomy, a terminal acute experiment was carried out, using the surgical, anesthetic, and analgesic procedures described by Quesney et al. (20). A dilute solution of sodium penicillin G was applied diffusely to the cortex of both cerebral hemispheres at a
SUBCORTICAL
INPUT
IN GENERALIZED
EPILEPSY
159
concentration delivering about 50 to 100 IU per hemisphere, as described by Gloor et al. (8). EEGs in both experimental groups were made between 2 weeks or 3 months after the last surgical procedure, using either an 8- or 16-channel Elema-Schoenander Mingograf or a Model T Offner g-channel EEG instrument. Bipolar and monopolar montages were used. In the latter case, either the frontal bone, the occipital bone, or the spinous process of the C, vertebra served as reference. At the completion of the experiment, the animals were killed under general anesthesia. The brain was removed, perfused with 10% formalin solution, fixed, and studied both macro- and microscopically (cresyl violet, 1~x01 fast blue, and Heidenhain stains). RESULTS Bilateral Cortical Slab Experiments. Preoperatively all four animals showed the typical response to the injection of 300,000 to 400,000 IU/kg penicillin in the form of bilaterally synchronous bursts of spike and wave or polyspike or multiple sharp-wave activity with a frequency of about 3.5 to 5 cycles/s, as described in previous publications (5, 9, 18, 20). The bursts involved all cortical regions from which records were taken, although they were often less prominent in the sigmoid and the posterior parts of the lateral gyri. The regions of the suprasylvian and lateral gyri from which the slab would ultimately be created participated early and prominently in these bursts. All animals survived the two-stage cortical isolation procedure and recovered well. The first two showed mild spastic hemiparesis after one or both procedures, presumably from edema of the internal capsule, but this quickly improved and their gait and agility returned to normal. The most constant neurologic deficit was impairment of downward vision, manifested in stumbling over feeding trays and occasional forays into space off the edge of a bench, but the animals seemed to adapt to this and thrived. Chronic infections around the recording plugs were treated by daily cleansing but never transgressed the dura. Anatomical Veri$cation of the Lesions. Although the isolation was nearly complete in all instances, some residual bridges of tissue remained in all animals. Aside from the pial bridges, these consisted of the attachment of the fomices and fimbriae to the underside of the corpus callosum, about 1 mm of corpus callosum anteriorly where the cuts from either side did not quite meet, and the retrosplenial gyri connecting the cingulate and the parahippocampal gyri on both sides in one animal. Postoperative Electroencephalogram. EEGs were taken only from the second postoperative week onward. In general, these large bilateral
160
PELLEGRINI,MUSGRAVE,ANDGLOOR
cortical slabs showed very little of the “bursting” activity alternating with periods of electrographic silence described by Grafstein (10) and Marcus and Watson (16) in acute preparations of the same type and by Ingvar (12) and Gloor et al. (6) for smaller slabs. Some brief isoelectric periods lasting 0.5 to 6 s were seen from time to time in all slabs. The activity resembled instead that of the acutely isolated hemisphere described by Kellaway er al. (14). In referential recordings, the background activity consisted of fairly low-voltage, fast rhythms with superimposed delta waves and slow sharp waves associated with occasional spikes and spike and slow wave complexes. Some brief runs of theta activity were occasionally seen. Epileptiform activity was usually present diffusely throughout the slab before any penicillin was given. It tended to be bilaterally synchronous, though usually with a unilateral predominance which shifted from side to side (Figs. 2B and 3A). Sometimes the spikes and slow waves occurred concurrently in very brief runs producing waveforms reminiscent of slow spike and wave activity at about 1 to 2 cycles/s (Fig. 3A). The synchronization of background activity and of epileptiform discharges within the slab was very striking, to the extent that bipolar recordings taken from it showed much cancellation effect by equipotentiality, especially of the slower wave forms. That this activity was indeed originating from the slab and not from the reference electrode was clear from its absence in channels recording from the intact cortex outside the slab. In the early postoperative period some focal epileptiform abnormality was occasionally recorded from nonisolated cortex, but this tended to disappear with time. Response
of Normal
and Isolated
Cortex
to Intramuscular
Penicillin.
After the intramuscular administration of penicillin, epileptiform bursts of 3.5 to 5 cycles/s, typical of those seen in generalized penicillin epilepsy of the intact cat, were recorded from the nonisolated cortex of the slabs in all animals. Interhemispheric synchrony of the epileptic bursts in the nonisolated cortex was disrupted (Figs. 2 and 3), except for one cat in which some bilateral synchrony persisted. In contrast, the isolated cortical regions within the slab never again produced the characteristic 3.5 to 5-cycles/s epileptic burst activity seen in them before isolation. However, the epileptiform abnormalities which had been present within the slab before penicillin were enhanced in amplitude, sharpness, and numbers, and slow sharp waves occurred more continuously, sometimes progressing to form spikes, the spikes becoming faster and of higher amplitude. Epileptiform features were more pronounced the higher the dose of penicillin given. Well-formed spike and wave activity was sometimes seen; it occurred in long runs, and the frequency of the spike and wave complexes was low (1 to 2.5 cycles/s), much lower than that
SUBCORTICAL
A
INPUT IN GENERALIZED
B
EPILEPSY
161 CAT M 21
1
FIG. 2. EEG from bilateral isolated cortical stab preparation. Electrodes 3 through 6 recorded from inside the slab. R-position of reference electrode. A-before cortical isolation, after i.m. penicillin. B-after cortical isolation without penicillin. Note the sharp waves within the slab. C-after cortical isolation, after i.m. penicillin. Note typical bursts outside the slab and their absence within it. Some activation of the preexisting sharp wave activity within the slab.
typical of bursts of generalized feline penicillin epilepsy (Fig. 3B) which range from about 3.5 to 5 cycles/s. Unilateral Thalamectomy in Split-brain Cats. Preoperatively the animals exhibited the typical EEG response to intramuscular penicillin described above, and with intravenous injection of 5 to 10 mg pentobarbital, bilateral spindle activity was induced in them (Fig. 7A).
162
PELLEGRINI,
MUSGRAVE,
h I
AND
GLOOR
SUBCORTICAL
INPUT IN GENERALIZED
163
EPILEPSY
During the first days after the unilateral thalamectomy the cats showed a mild contralateral spastic hemiparesis. The head was deviated to the side of the lesion. These neurological deficits gradually improved until hardly any neurologic impairment was noticeable on casual observation at the time the recording sessions were begun. Anatomical Verification of the Lesions. A complete unilateral ablation of VPA
17
FIG. 4. Extent of lesion in one cat with a maximal subtotal right thalamectomy. Abbreviations as in Jasper, H., and C. Ajmone-Marsan, A Stereotaxic Atlas of the Diencephalon of the Cat. Ottawa, National Research Council of Canada. 1954.
164
PELLEGRINI.
MUSGRAVE,
AND
GLOOR
the thalamus was never achieved. However, most of the thalamus was always destroyed. The regions which were spared to a greater or lesser degree were the reticular nucleus, expecially its most lateral and posterior parts, the lateral and medial geniculate bodies, as well as the superolateral portions of pulvinar and N. lateralis posterior and N. lateralis dorsalis. In three animals a very large subtotal right-side thalamectomy was achieved as shown in Fig. 4. In the other two cats the thalamectomy was less
TALCRO
FIG, 5.
Extent
of lesion
in one cat with
a less extensive
right
thalamectomy.
1
SUBCORTICAL
MS
EPILEPSY
165
(P) (
PSS -
L
INPUT IN GENERALIZED
-
e
ASSMSS (A) [
MSS (P) i PSS -
FIG. 6. Cat with maximal subtotal right thalamectomy (lesion shown in Fig. 4). A-EEG of alert cat without penicillin. B-EEG of the same cat without penicillin, when drowsy. Note the sharp waves on the thalamectomized side. Abbreviations for this and the subsequent figures: ASS-anterior suprasylvian gyrus, MES-middle ectosylvian gyrus, PSSposterior suprasylvian gyrus, OCC REF-occipital reference electrode, (A)-anterior, (P)--posterior, R-right, L-left.
complete, sparing large parts of the posterior lateral thalamus. An example of such a lesion is shown in Fig. 5. Electroencephalographic findings. EEGs were recorded between 3 weeks and 3 months after thalamectomy and commissural section. The samples in Figs. 6-8 were taken from the cat with the large right thalamic lesion illustrated in Fig. 4. When the cat was awake, there was no obvious difference in the electrical activity of the cortex between the two sides (Fig. 6A). However, when the animal became drowsy and went to sleep (Fig. 6B), slow waves predominated on the side of the lesion. Interspersed with these were sharp waves which tended to be synchronized throughout the cortex of the hemisphere on the side of the thalamectomy. Small sleep spindles which appeared on the normal side were not detected on the thalamectomized side. This unilateral absence of spindles was even more
166
PELLEGRINI,
RASS-OCC
MUSGRAVE, A
AND GLOOR WAR
REF ‘*“rruhl*r*u~~c.*yru
MSS (A) a MS
(P)
PSSA 1 ASSMS
(A) *
FIG. 7. Cat with maximal subtotal right thalamectomy. Same animal as Fig. 6 (lesion shown in Fig. 4). A-EEG after i.v. injection of pentobarbital before thalamic lesion was made. Note bilateral barbiturate spindles. B-EEG after i.v. injection of pentobarbital after thalamic lesion. Note absence of spindles on the thalamectomized side.
clearly shown after the intravenous injection of pentobarbital: Whereas bilateral spindles had been elicited by pentobarbital before the thalamic lesion had been made (Fig. 7A), spindles appeared only on the normal side when such an injection was repeated after right-side thalamectomy (Fig. 7B). Figure 8 shows the effect of penicillin on the EEG of this cat. (A) is a control record before thalamectomy; typical bilaterally synchronous Scycle/s epileptiform bursts are present. (B), taken in the awake animal, shows after right-side thalamectomy a complete absence of these bursts on the thalamectomized side; however, when the animal became drowsy (C), epileptic activity in the form of sharp waves and some slow 2-cycle/s spike and wave complexes now appeared on the side of the thalamectomy . These discharges looked like an activation of the milder sharp and slow wave disturbance which had been present on that side before penicillin was given (Fig. 6). This activity always appeared very different from the typical discharges of generalized penicillin epilepsy; it was never well-organized in bursts, always much slower in frequency, and somewhat different in wave form. On the intact side, typical bursts continued to occur.
SUBCORTICAL
INPUT IN GENERALIZED
EPILEPSY
167
FIG. 8. Cat with maximal subtotal right thalamectomy. Same animal as in Figs. 6 and 7. (lesion shown in Fig. 4). A-epileptic activity after i.m. penicillin before thalamic lesion, typical epileptic bursts on both sides. B and C-epileptic activity after i.m. penicillin after thalamic lesion; typical epileptic bursts absent on thalamectomized side. B-cat alert, no epileptiform activities on thalamectomized side, C-cat drowsy; typical epileptic bursts on the intact side, slow spike-and-waves and sharp waves on the thalamectomized side.
Figure 9 shows the EEG of a cat with a less complete thalamectomy (Fig. 5). The control EEG before the lesion (Fig. 9A) exhibited typical bilaterally synchronous epileptiform bursts at about 5 cycles/s. After the right-sided incomplete thalamectomy, the EEG shown in Fig. 9B demonstrated that the bursts remained unchanged on the normal side, while they were markedly reduced in amplitude and slightly also in frequency on the side with the incomplete thalamectomy. The bursts no longer occurred synchronously on the two sides, because of the transection of the interhemispheric commissures (Musgrave and Gloor, in preparation). In the two cats with less complete thalamic lesions, intraperitoneal
168
1ASS
PELLEGRINI,
MUSGRAVE,
AND GLOOR
-I,--,/
MS5 (A)------
-
FIG. 9. Cat with less extensive right thalamectomy (lesion shown in Fig. 5). A-epileptic activity after i.m. penicillin before thalamic lesion; typical epileptic bursts on both sides. B-epileptic activity after i.m. penicillin after thalamic lesion; epileptic bursts unilateral on normal side, reduced in amplitude and slightly reduced in frequency on the thalamectomized side (asynchrony between the two sides attributable to cahosal transection).
pentobarbital induced some spindle activity on the side of the lesion, although compared to the normal side, it was definitely reduced. In three animals with maximal subtotal thalamectomy, a dilute penicillin solution (50 to 100 IU per hemisphere) was applied to the cortex of both hemispheres in an acute terminal experiment. Epileptiform discharges appeared on both sides, but differed greatly on the thalamectomized side
SUBCORTICAL
INPUT
IN GENERALIZED
EPILEPSY
169
from those seen on the intact side (Fig. 10). On the intact side epileptiform bursts at 3 to 5 cycles/s were recorded which resembled those seen after intramuscular penicillin injection; on the thalamectomized side the discharges consisted of slow l- to 2-cycle/s spike and wave complexes. In the animal whose EEG is shown in Fig. 10, this activity was practically continuous, in another cat rather infrequent, and in a third animal an intermediate amount was seen, but many of the discharges consisted only or paroxysmal slow waves. DISCUSSION The experiments reported here demonstrated that subcortical inputs to the cerebral cortex were required for the production of the characteristic bursts of generalized feline penicillin epilepsy which usually exhibited a frequency range of 3.5 to 5 cycles/s and consist of spike and wave or multiple spike or sharp wave discharges. The experiments with unilateral submaximal thalamectomy, associated with transection of the interhemispheric commissures, indicated that the important afferent inputs to the cortex responsible for the precipitation of these characteristic bursts originated in or passed through the thalamus. The absence of spindles on the thalamectomized side in these animals confirms similar findings obtained by Naquet et al. (17), by Andersen et al. (l), and by Lanoir (15) and are in agreement with the hypothesis that thalamocortical volleys are responsible for evoking spindles as well as the generalized epileptic bursts of feline generalized penicillin epilepsy (7, 8, 20). This hypothesis receives further support from the observation that when spindle activity is merely reduced, but not totally abolished by a less extensive thalamic lesion, epileptiform bursts in response to penicillin are also reduced on the side of the lesion, but not abolished. The present experiments add the important information that the spontaneous epileptiform bursts in this model are as dependent on the thalamocortical spindle-generating mechanism as those elicited in response to single-shock and low-frequency subcortical stimulation (8, 20). However, cortex deprived of all subcortical inputs or only of thalamocortical inputs still produces epileptic discharges, even without penicillin. These are therefore not dependent on the action of this drug, although they become more active under its influence. They indicate an increased tendency of deafferented cortex to produce epileptiform discharge (2, 3, 21). The discharges in this deafferented cortex differ, however, from those characteristic of feline generalized penicillin epilepsy by their slower frequency and by their tendency to occur in a more isolated fashion rather than in well-organized bursts. They share with them,
170
PELLEGRINI,
MUSGRAVE,
AND
GLOOR
SUBCORTICAL
INPUT
IN GENERALIZED
EPILEPSY
171
however, the tendency to be widely synchronized. In the case of a unilateral thalamectomy with split corpus callosum, the discharges appear over the thalamectomized side only and are synchronized within the cortical regions from which recordings are obtained. In the case of the bilateral cortical slab with intact callosal connections, the discharges are almost always bilaterally synchronous. These findings indicate that powerful cortico-cortical synchronizing pathways exist within each hemisphere, and that the corpus callosum is an important anatomic substrate for establishing bilateral synchrony. The corpus callosum is also responsible for the bilateral synchrony of the typical discharges of generalized penicillin epilepsy in the intact animal (Musgrave and Gloor, in preparation). This is also suggested by the observations made in the bilateral cortical slab preparation in which the typical bursts of feline generalized penicillin epilepsy in cortex lateral to the bilateral cortical slabs occurred asynchronously on the two sides, presumably because the callosal connections of these cortical regions had been severed in the course of the surgical procedures used to produce the bilateral cortical slabs (Fig. 1). The ability of thalamically deafferented cortex to produce widespread epileptiform discharge of a slow spike and wave type was also demonstrated by the fact that such discharges could be elicited by the topical application of a dilute penicillin solution to the cortex of both hemispheres in animals with a unilateral subtotal thalamectomy. The discharges on the two sides, however, differed from each other in the same manner as in animals with an unilateral thalamectomy in which pencillin had been injected intramuscularly. An interesting observation made in cats with a unilateral thalamic lesion was that the amount of epileptic activity occurring on that side both before and after penicillin was dependent on the level of arousal, epileptic discharges being present only when the animal was drowsy or asleep. This suggests that, in spite of the removal of the thalamus, connections from the brain stem mediating arousal responses remained, at least in part, intact and thus must have by-passed the thalamus (23). It is of interest that moderate arousal compatible with the presence of the typical epileptic discharges of feline generalized penicillin epilepsy on the intact side was sufficient to suppress the “atypical” cortical epileptic activity on the thalamectomized side, even under the influence of systemic penicillin. These experiments thus demonstrate that the occurrence of the typical spontaneous generalized bilaterally synchronous 3.5 to Scycle/s epileptiform bursts of feline generalized penicillin epilepsy depend on an intact thalamocortical input which normally most likely is involved in spindle genesis (7,8,20). Deafferented cortex, however, even without penicillin, is
172
PELLEGRINI,
MUSGRAVE,
AND
GLOOR
capable of producing widespread epileptiform discharges which are slow, but often assume a spike and wave form and exhibit prominent intrahemispheric or interhemispheric synchronization, the latter being dependent on an intact corpus callosum. The mechanism of this widespread synchronization thus, does not depend on subcortical structures, and thus confirms earlier findings obtained in a different model by Marcus and Watson (16). The slow spike and wave form occurring in widely synchronized fashion in these animals with deafferented cortex raises the intriguing question of whether slow generalized or widespread spike and wave discharge in human epilepsy seen in association with large cerebral destructive lesions (as, for example, in porencephalic cysts) could be caused by the same mechanism. REFERENCES 1. ANDERSEN, P., S. A. ANDERSSON, AND T. LIMO. 1967. Some factors involved in the thalamic control of spontaneous barbiturate spindles. J. Physiol. (London) 192: 257-281. 2. ECHLIN, F. A. 1959. The supersensitivity of chronically “isolated” cerebral cortex as a mechanism in focal epilepsy. Electroenceph. C/in. Neurophysiol. 11: 697-722. 3. ECHUN, F. A., AND A. BATTISTA. 1963. Epileptiform seizures from chronic isolated cortex. Arch. Neurol. 9: 154-170. 4. FEINDEL, W., Y. L. YAMAMOTO, AND C. P. HODGE. 1967. Intracarotid fluorescein angiography: a new method for examination of the epicerebral circulation in man. Can. Med. Assoc. J. 96: l-7. 5. FISHER, R. S., AND D. A. PRINCE. 1977. Spike-wave rhythms in cat cortex induced by parenteral penicillin. I. Electroencephalographic features. Electroenceph. C/in. Neurophsyiol. 42: 608-624. 6. GLOOR, P., G. BALL, AND N. SCHAUL. 1977. Brain lesions that produce delta waves in the EEG. Neurology (Minneapolis) 21: 326-333. 7. GLOOR, P., A. PELLEGRINI, AND G. K. KOSTOPOULOS. 1979. Effects of changes in cortical excitability upon the epileptic bursts in generalized penicillin epilepsy of the cat. Electroenceph. C’lin. Neurophysiol., in press. 8. GLOOR, P., L. F. QUESNEY, AND H. ZUMSTEIN. 1977. Pathophysiology of generalized penicillin epilepsy in the cat: the role of cortical and subcortical structures. II. Topical application of penicillin to the cerebral cortex and to subcortical structures. Electroenceph. Clin. Neurophysiol. 43: 79-94. 9. GLOOR, P., AND G. TESTA. 1974. Generalized penicillin epilepsy in the cat: Effects of intracarotid and intravertebral pentylenetetrazol and amobarbital injections. Electroenceph. Clin. Neurophysiol. 36: 499-515. 10. GRAFSTEIN, B. 1959. Organization of callosal connections in the suprasylvian gyrus of cat. .I. Neurophysiol. 22: 504-515. 11. HOHBERGER,~. P.,Y. L. YAMAMOTO,~. J.THOMPSON,AND W. FEINDEL. 1975. On-line computer measurement of microregional cerebral blood flow by xenon-133 clearance. Int. J. Nucl. Med. Biol. 2: 153-158. 12. INGVAR, D. H. 1955. Electrical activity of isolated cortex in the unanaesthetized cat with intact brainstem. Acra Physiol. Stand. 33: 151-168.
SUBCORTICAL
INPUT IN GENERALIZED
EPILEPSY
I73
13. INGVAR, D. H., AND N. A. LASSEN. 1962. Regional blood flow of the cerebral cortex determined by Krypton 85. Acta Physiol. Stand. 54: 325-338. 14. KELLAWAY, P., A. GOL, AND M. PROLER. 1966. Electricalactivity ofthe isolated cerebral hemisphere and isolated tha.lamus. Exp. Neural. 14: 281-304. 15. LANOIR, J. 1972. Etude tilectrocorticographique de la veille et du sommeil chez le chat. Organisation du cycle nyctht!mtral. R81e du thalamus. Thesis. Universitt de Provence Aix-Marseille, France. 16. MARCUS, E. M., AND C. W. WATSON. 1%6. Bilateral synchronous spike wave electrographic patterns in the cat. Interaction of bilateral cortico-callosal and adiencephalic preparation. Arch. Neural. 14: 601-610. 17. NAQUET, R., M. DENAVIT, J. LANOIR, AND D. ALB~-FESSARD. 1965. AltCrations transitoires ou definitives des zones diencephaliques chez le chat. Leurs effets sur l’activite electrique corticale et le sommeil. Pages 107-131 in Aspects Anatomo-fonctionnels de la Physiologie du Sommeil. Editions du Centre National de la Recherche Scientifique, Paris. 18. PRINCE, D. A., AND D. FARRELL. 1%9. “Centrencephalic” spike wave discharges following parenteral penicillin injection in the cat. Neurology (Minneapolis) 19: 309-310. 19. QUESNEY, L. F., AND P. GLOOR. 1978. Generalized penicillin epilepsy in the cat: correlation between electrophysiological data and distribution of r4C-penicillin in the brain. Epilepsia 19: 35-45. 20. QUESNEY, L. F., P. GLOOR, E. KRATZENBERG, AND H. ZUMSTEIN. 1977. Pathophysiology of generalized penicillin epilepsy in the cat: the role of cortical and subcortical structures. I. Systemic application of penicillin. Etectroenceph. C/in. Neurophysiol. 42: 640-655. 21.
SHARPLESS, S. K., AND L. M. HALPERN. 1962. The electrical excitability of chronically isolated cortex studied by means ofpermanently implanted electrodes. Electroenceph. Clin.
Neurophysiol.
14: 244-255.
22. SOEJIMA, T.. Y. L. YA~AMOTO, E. MEYER, AND W. FEINDEL. 1979. Effects of steroids on early microcirculatory changes following focal cold injury of cerebral cortex. J. Neurosurg., in press. 23. STARZL, T. E., C. W. TAYLOR, AND H. W. MAGOUN. 1951. Ascending conduction in reticular activating systems with special reference to the diencephalon. J. Neurophysiol.
14: 461-477.
NEUROLOGY
64, 155- 173 (1979)
Role of Afferent Input of Subcortical Origin in the Genesis of Bilaterally Synchronous Epileptic Discharges of Feline Generalized Penicillin Epilepsy A. PELLEGRINI,
J. MUSGRAVE,AND
P. GLOOR'
Montreal Neurological Institute, 3801 University Street, Montreal, Quebec, H3A 284 Canada Received October 13. 1978 The effects of (i) neural isolation of large bilateral cortical slabs with intact callosal connections, and (ii) of large unilateral subtotal thalamectomies associated with transection of the interhemispheric commissures on the generalized epileptiform discharges of generalized penicillin epilepsy of the cat were studied. Cortex devoid of subcortical, particularly thalamic, inputs was incapable of generating the characteristic 3.5- to 5-cycles/s epileptiform bursts of feline generalized penicillin epilepsy either in response to the i.m. injection of a large dose of penicillin or to the diffuse direct cortical application of a dilute penicillin solution. Such deafferented cortex, however, produced, even without penicillin, widespread often highly synchronized epileptiform discharges of a different kind consisting at times of slow, l- to 2.5-cycles/s spike and wave complexes. This epileptiform activity increased in response to i.m. penicillin injection or to widespread direct cortical application of a dilute penicillin solution, but never assumed the characteristics of the typical discharges of feline generalized penicillin epilepsy as seen in cortex which has retained its subcortical connections. The discharges differed from those of intact cortex by being slower in frequency and less prone to become organized in bursts. In the bilateral slab preparation with preserved callosal connections, these atypical discharges occurred, however, in a bilaterally synchronous manner, as did the typical discharges in the intact cat with generalized penicillin epilepsy. This suggests that the corpus callosum is necessary for the bilateral synchrony of both types of epileptiform discharges.
Abbreviation: EEG, electroencephalogram. 1 This work was supported by Research Grant MT-3 140 of the Medical Research Council of Canada. Address reprint requests to Dr. P. Gloor. Dr. Pellegrini’s present address is Clinica Neurologica dell’ Universita, Via Giustiniani 1, 35100 Padua, Italy. 155 0014-4886/79/040155-19$02.00/O Copyright 0 1979 by Academic Press. Inc. All rights of reproduction in any form reserved.
156
PELLEGRINI,
MUSGRAVE,
AND
GLOOR
INTRODUCTION Previous investigations (7, 8, 20) suggested that the synchronous epileptic bursts occurring in generalized penicillin epilepsy of the cat are closely related to spindles and/or recruiting responses which, in normal animals, are induced by thalamocortical volleys. Much of this evidence was obtained from stimulation studies which showed that single shock or low-frequency thalamic stimulations which, in normal animals, induced spindles or recruiting responses, elicited generalized bilaterally synchronous epileptic bursts, often assuming a spike and wave form, in animals having received a large dose of intramuscular penicillin. These experiments, however, did not conclusively demonstrate that thalamocortical inputs were necessarily involved in the genesis of the spontaneously occurring epileptic discharges in this model. The present series of experiments was therefore undertaken to determine whether or not disconnection of the cortex from all or nearly all inputs arising from subcortical, particularly thalamic, structures would result in the absence of the epileptic bursts typical for this condition in cortex disconnected in this manner. Two preparations were used for this purpose: (i) large symmetrical neuronally isolated cortical slabs of both hemispheres which remained interconnected through the corpus callosum, prepared in a manner similar to that described by Marcus and Watson (16); and (ii) unilateral thalamectomy in cats with a transsected corpus callosum. Both preparations were chronic and recordings were carried out after the animals had fully recovered from the surgical trauma. METHODS Surgical procedures were done under pentobarbital (Nembutal) anesthesia using sterile techniques and the operating microscope. Pre- and postoperative care with i.v. and S.C. fluids, systemic antibiotics, and dexamethasone was given. Cortical Isolation Experiments. Four cats weighing 3 to 5 kg were used in these three-stage experiments. In the first stage, a wide symmetrical craniectomy was made which extended from the coronal suture posteriorly for about 2 cm and laterally as far as the medial border of the middle ectosylvian gyrus. The bone was partly replaced by an acrylic cranioplasty in which stainless-steel screw electrodes and a multiple-pin miniature connector were imbedded. In the fourth animal, the screws were placed in the outer table of the cranial vault, and bone flaps later were cut around them. Of the regions to be included in the isolated slab the activity of the middle suprasylvian gyrus was recorded in all animals and that of the lateral gyrus in two. The interelectrode distance was 5 mm. Additional recordings
SUBCORTICAL
INPUT IN GENERALIZED
EPILEPSY
157
FIG. 1. Schematic drawing showing bilateral neuronally isolated cortical slab with intact callosal connections.
were obtained from regions of cortex anterior, lateral, and posterior to those to be included in the isolated slab, consisting of the sigmoid, ectosylvian, and posterior parts of the lateral gyri, respectively. After the animals had recovered from the surgical procedure, control electroencephalograms (EEGs) before and after an i.m. injection of 300,000 to 400,000 IU/kg sodium penicillin G were obtained. In the second stage, the cranioplasty was elevated, and a wide dural flap was hinged to the midline. The medial portion of the middle ectosylvian gyrus was removed by suction to give working space, and the corona radiata was cut across until the lateral ventricle was entered (Fig. 1). The slab was then completed by aspirating a thin coronal trench at either end through pial windows in the suprasylvian and lateral gyri and gradually deepening these to reach the ventricle. The cingulate and retrosplenial gyri were cut across subpially, and the coronal trench was carried into the corpus callosum anteriorly, usually just above the intraventricular foramen (Fig. 1). The posterior line of section passed behind the splenium of the corpus callosum. All arteries and most veins crossing the incision line were preserved in pial bridges. In the last animal, the pillar of the fomix was severed, and the fimbria or anterior hippocampus was cut across diagonally to reach the posterior midline at the splenium. A water-tight dural closure was made with 7-O silk before the cranioplasty or bone flap was replaced. At least 2 weeks later the same procedure was carried out on the other side, thus completing the isolation of the slab of bilateral cortical tissue, the isolated regions of each
158
PELLEGRINI,
MUSGRAVE,
AND
GLOOR
hemisphere remaining interconnected by the corpus callosum (Fig. 1). At the close of the experiments, the circulation in the slab was studied by fluorescein angiography (4) and radionucleide scanning with Xe133 (11) and/or KF5 (13, 22) and was found to be normal except for some small regions of relatively slow circulation at the edge of the slab as revealed by the fluorescein angiogram. The intracortical concentration of P4-labeled penicillin G was measured in three of the four animals using the methods reported by Quesney and Gloor (19). The cortical concentration of penicillin inside and outside the slab was essentially the same. Thalamectomy. A row of five stainless-steel screw electrodes was implanted over each middle suprasylvian gyrus at 6-mm intervals, and their recording plugs were screwed to the cranial vault posteriorly in five cats weighing 3 to 4.5 kg. A reference screw electrode was inserted in the occipital bone at the midline. Control recordings of the response to intramuscularly injected penicillin at a dose of 300,000 to 400,000 IU/kg and to intraperitoneal injection of 5 to 10 mg/kg pentobarbital were then made. At a later date a craniectomy about 15 mm wide and 25 mm long was made between the two rows of screw electrodes. The dura was slit open along the left side of the superior sagittal sinus at a distance of about 2 mm from it. Care was taken to leave intact, if at all possible, all bridging veins to the sinus originating from the left hemisphere, but frequently it was necessary to divide some of the smaller ones in the interhemispheric fissure. The corpus callosum and the hippocampal commissures were then divided by suction to expose the third ventricle. The dorsal surface of the right thalamus was exposed by aspirating the fomix and by reflecting the choroid plexus of the lateral ventricle medially. The posterior commissure, massa intermedia, and anterior commissure served as landmarks to guide the removal. The right thalamus was then removed as completely as possible, commencing at the midline, extending ventrally to the inferior limit of the massa intermedia, posteriorly to the mouth of the aqueduct, and anteriorly to the intraventricular foramen. The thalamostriate vein was spared. Lateral dissection under direct vision was limited by the amount of retraction the left hemisphere would tolerate, and the most lateral removal was by necessity carried out blindly, using a suction tip with a right angle, 9 mm in length. Last, the anterior commissure was divided. After careful hemostasis in the removal cavity, the dura was closed with 7-O silk in a water-tight manner. The bone defect was bridged with dental cement over Gelfoam strips, and the skin was closed. In three cats with a unilateral thalamectomy, a terminal acute experiment was carried out, using the surgical, anesthetic, and analgesic procedures described by Quesney et al. (20). A dilute solution of sodium penicillin G was applied diffusely to the cortex of both cerebral hemispheres at a
SUBCORTICAL
INPUT
IN GENERALIZED
EPILEPSY
159
concentration delivering about 50 to 100 IU per hemisphere, as described by Gloor et al. (8). EEGs in both experimental groups were made between 2 weeks or 3 months after the last surgical procedure, using either an 8- or 16-channel Elema-Schoenander Mingograf or a Model T Offner g-channel EEG instrument. Bipolar and monopolar montages were used. In the latter case, either the frontal bone, the occipital bone, or the spinous process of the C, vertebra served as reference. At the completion of the experiment, the animals were killed under general anesthesia. The brain was removed, perfused with 10% formalin solution, fixed, and studied both macro- and microscopically (cresyl violet, 1~x01 fast blue, and Heidenhain stains). RESULTS Bilateral Cortical Slab Experiments. Preoperatively all four animals showed the typical response to the injection of 300,000 to 400,000 IU/kg penicillin in the form of bilaterally synchronous bursts of spike and wave or polyspike or multiple sharp-wave activity with a frequency of about 3.5 to 5 cycles/s, as described in previous publications (5, 9, 18, 20). The bursts involved all cortical regions from which records were taken, although they were often less prominent in the sigmoid and the posterior parts of the lateral gyri. The regions of the suprasylvian and lateral gyri from which the slab would ultimately be created participated early and prominently in these bursts. All animals survived the two-stage cortical isolation procedure and recovered well. The first two showed mild spastic hemiparesis after one or both procedures, presumably from edema of the internal capsule, but this quickly improved and their gait and agility returned to normal. The most constant neurologic deficit was impairment of downward vision, manifested in stumbling over feeding trays and occasional forays into space off the edge of a bench, but the animals seemed to adapt to this and thrived. Chronic infections around the recording plugs were treated by daily cleansing but never transgressed the dura. Anatomical Veri$cation of the Lesions. Although the isolation was nearly complete in all instances, some residual bridges of tissue remained in all animals. Aside from the pial bridges, these consisted of the attachment of the fomices and fimbriae to the underside of the corpus callosum, about 1 mm of corpus callosum anteriorly where the cuts from either side did not quite meet, and the retrosplenial gyri connecting the cingulate and the parahippocampal gyri on both sides in one animal. Postoperative Electroencephalogram. EEGs were taken only from the second postoperative week onward. In general, these large bilateral
160
PELLEGRINI,MUSGRAVE,ANDGLOOR
cortical slabs showed very little of the “bursting” activity alternating with periods of electrographic silence described by Grafstein (10) and Marcus and Watson (16) in acute preparations of the same type and by Ingvar (12) and Gloor et al. (6) for smaller slabs. Some brief isoelectric periods lasting 0.5 to 6 s were seen from time to time in all slabs. The activity resembled instead that of the acutely isolated hemisphere described by Kellaway er al. (14). In referential recordings, the background activity consisted of fairly low-voltage, fast rhythms with superimposed delta waves and slow sharp waves associated with occasional spikes and spike and slow wave complexes. Some brief runs of theta activity were occasionally seen. Epileptiform activity was usually present diffusely throughout the slab before any penicillin was given. It tended to be bilaterally synchronous, though usually with a unilateral predominance which shifted from side to side (Figs. 2B and 3A). Sometimes the spikes and slow waves occurred concurrently in very brief runs producing waveforms reminiscent of slow spike and wave activity at about 1 to 2 cycles/s (Fig. 3A). The synchronization of background activity and of epileptiform discharges within the slab was very striking, to the extent that bipolar recordings taken from it showed much cancellation effect by equipotentiality, especially of the slower wave forms. That this activity was indeed originating from the slab and not from the reference electrode was clear from its absence in channels recording from the intact cortex outside the slab. In the early postoperative period some focal epileptiform abnormality was occasionally recorded from nonisolated cortex, but this tended to disappear with time. Response
of Normal
and Isolated
Cortex
to Intramuscular
Penicillin.
After the intramuscular administration of penicillin, epileptiform bursts of 3.5 to 5 cycles/s, typical of those seen in generalized penicillin epilepsy of the intact cat, were recorded from the nonisolated cortex of the slabs in all animals. Interhemispheric synchrony of the epileptic bursts in the nonisolated cortex was disrupted (Figs. 2 and 3), except for one cat in which some bilateral synchrony persisted. In contrast, the isolated cortical regions within the slab never again produced the characteristic 3.5 to 5-cycles/s epileptic burst activity seen in them before isolation. However, the epileptiform abnormalities which had been present within the slab before penicillin were enhanced in amplitude, sharpness, and numbers, and slow sharp waves occurred more continuously, sometimes progressing to form spikes, the spikes becoming faster and of higher amplitude. Epileptiform features were more pronounced the higher the dose of penicillin given. Well-formed spike and wave activity was sometimes seen; it occurred in long runs, and the frequency of the spike and wave complexes was low (1 to 2.5 cycles/s), much lower than that
SUBCORTICAL
A
INPUT IN GENERALIZED
B
EPILEPSY
161 CAT M 21
1
FIG. 2. EEG from bilateral isolated cortical stab preparation. Electrodes 3 through 6 recorded from inside the slab. R-position of reference electrode. A-before cortical isolation, after i.m. penicillin. B-after cortical isolation without penicillin. Note the sharp waves within the slab. C-after cortical isolation, after i.m. penicillin. Note typical bursts outside the slab and their absence within it. Some activation of the preexisting sharp wave activity within the slab.
typical of bursts of generalized feline penicillin epilepsy (Fig. 3B) which range from about 3.5 to 5 cycles/s. Unilateral Thalamectomy in Split-brain Cats. Preoperatively the animals exhibited the typical EEG response to intramuscular penicillin described above, and with intravenous injection of 5 to 10 mg pentobarbital, bilateral spindle activity was induced in them (Fig. 7A).
162
PELLEGRINI,
MUSGRAVE,
h I
AND
GLOOR
SUBCORTICAL
INPUT IN GENERALIZED
163
EPILEPSY
During the first days after the unilateral thalamectomy the cats showed a mild contralateral spastic hemiparesis. The head was deviated to the side of the lesion. These neurological deficits gradually improved until hardly any neurologic impairment was noticeable on casual observation at the time the recording sessions were begun. Anatomical Verification of the Lesions. A complete unilateral ablation of VPA
17
FIG. 4. Extent of lesion in one cat with a maximal subtotal right thalamectomy. Abbreviations as in Jasper, H., and C. Ajmone-Marsan, A Stereotaxic Atlas of the Diencephalon of the Cat. Ottawa, National Research Council of Canada. 1954.
164
PELLEGRINI.
MUSGRAVE,
AND
GLOOR
the thalamus was never achieved. However, most of the thalamus was always destroyed. The regions which were spared to a greater or lesser degree were the reticular nucleus, expecially its most lateral and posterior parts, the lateral and medial geniculate bodies, as well as the superolateral portions of pulvinar and N. lateralis posterior and N. lateralis dorsalis. In three animals a very large subtotal right-side thalamectomy was achieved as shown in Fig. 4. In the other two cats the thalamectomy was less
TALCRO
FIG, 5.
Extent
of lesion
in one cat with
a less extensive
right
thalamectomy.
1
SUBCORTICAL
MS
EPILEPSY
165
(P) (
PSS -
L
INPUT IN GENERALIZED
-
e
ASSMSS (A) [
MSS (P) i PSS -
FIG. 6. Cat with maximal subtotal right thalamectomy (lesion shown in Fig. 4). A-EEG of alert cat without penicillin. B-EEG of the same cat without penicillin, when drowsy. Note the sharp waves on the thalamectomized side. Abbreviations for this and the subsequent figures: ASS-anterior suprasylvian gyrus, MES-middle ectosylvian gyrus, PSSposterior suprasylvian gyrus, OCC REF-occipital reference electrode, (A)-anterior, (P)--posterior, R-right, L-left.
complete, sparing large parts of the posterior lateral thalamus. An example of such a lesion is shown in Fig. 5. Electroencephalographic findings. EEGs were recorded between 3 weeks and 3 months after thalamectomy and commissural section. The samples in Figs. 6-8 were taken from the cat with the large right thalamic lesion illustrated in Fig. 4. When the cat was awake, there was no obvious difference in the electrical activity of the cortex between the two sides (Fig. 6A). However, when the animal became drowsy and went to sleep (Fig. 6B), slow waves predominated on the side of the lesion. Interspersed with these were sharp waves which tended to be synchronized throughout the cortex of the hemisphere on the side of the thalamectomy. Small sleep spindles which appeared on the normal side were not detected on the thalamectomized side. This unilateral absence of spindles was even more
166
PELLEGRINI,
RASS-OCC
MUSGRAVE, A
AND GLOOR WAR
REF ‘*“rruhl*r*u~~c.*yru
MSS (A) a MS
(P)
PSSA 1 ASSMS
(A) *
FIG. 7. Cat with maximal subtotal right thalamectomy. Same animal as Fig. 6 (lesion shown in Fig. 4). A-EEG after i.v. injection of pentobarbital before thalamic lesion was made. Note bilateral barbiturate spindles. B-EEG after i.v. injection of pentobarbital after thalamic lesion. Note absence of spindles on the thalamectomized side.
clearly shown after the intravenous injection of pentobarbital: Whereas bilateral spindles had been elicited by pentobarbital before the thalamic lesion had been made (Fig. 7A), spindles appeared only on the normal side when such an injection was repeated after right-side thalamectomy (Fig. 7B). Figure 8 shows the effect of penicillin on the EEG of this cat. (A) is a control record before thalamectomy; typical bilaterally synchronous Scycle/s epileptiform bursts are present. (B), taken in the awake animal, shows after right-side thalamectomy a complete absence of these bursts on the thalamectomized side; however, when the animal became drowsy (C), epileptic activity in the form of sharp waves and some slow 2-cycle/s spike and wave complexes now appeared on the side of the thalamectomy . These discharges looked like an activation of the milder sharp and slow wave disturbance which had been present on that side before penicillin was given (Fig. 6). This activity always appeared very different from the typical discharges of generalized penicillin epilepsy; it was never well-organized in bursts, always much slower in frequency, and somewhat different in wave form. On the intact side, typical bursts continued to occur.
SUBCORTICAL
INPUT IN GENERALIZED
EPILEPSY
167
FIG. 8. Cat with maximal subtotal right thalamectomy. Same animal as in Figs. 6 and 7. (lesion shown in Fig. 4). A-epileptic activity after i.m. penicillin before thalamic lesion, typical epileptic bursts on both sides. B and C-epileptic activity after i.m. penicillin after thalamic lesion; typical epileptic bursts absent on thalamectomized side. B-cat alert, no epileptiform activities on thalamectomized side, C-cat drowsy; typical epileptic bursts on the intact side, slow spike-and-waves and sharp waves on the thalamectomized side.
Figure 9 shows the EEG of a cat with a less complete thalamectomy (Fig. 5). The control EEG before the lesion (Fig. 9A) exhibited typical bilaterally synchronous epileptiform bursts at about 5 cycles/s. After the right-sided incomplete thalamectomy, the EEG shown in Fig. 9B demonstrated that the bursts remained unchanged on the normal side, while they were markedly reduced in amplitude and slightly also in frequency on the side with the incomplete thalamectomy. The bursts no longer occurred synchronously on the two sides, because of the transection of the interhemispheric commissures (Musgrave and Gloor, in preparation). In the two cats with less complete thalamic lesions, intraperitoneal
168
1ASS
PELLEGRINI,
MUSGRAVE,
AND GLOOR
-I,--,/
MS5 (A)------
-
FIG. 9. Cat with less extensive right thalamectomy (lesion shown in Fig. 5). A-epileptic activity after i.m. penicillin before thalamic lesion; typical epileptic bursts on both sides. B-epileptic activity after i.m. penicillin after thalamic lesion; epileptic bursts unilateral on normal side, reduced in amplitude and slightly reduced in frequency on the thalamectomized side (asynchrony between the two sides attributable to cahosal transection).
pentobarbital induced some spindle activity on the side of the lesion, although compared to the normal side, it was definitely reduced. In three animals with maximal subtotal thalamectomy, a dilute penicillin solution (50 to 100 IU per hemisphere) was applied to the cortex of both hemispheres in an acute terminal experiment. Epileptiform discharges appeared on both sides, but differed greatly on the thalamectomized side
SUBCORTICAL
INPUT
IN GENERALIZED
EPILEPSY
169
from those seen on the intact side (Fig. 10). On the intact side epileptiform bursts at 3 to 5 cycles/s were recorded which resembled those seen after intramuscular penicillin injection; on the thalamectomized side the discharges consisted of slow l- to 2-cycle/s spike and wave complexes. In the animal whose EEG is shown in Fig. 10, this activity was practically continuous, in another cat rather infrequent, and in a third animal an intermediate amount was seen, but many of the discharges consisted only or paroxysmal slow waves. DISCUSSION The experiments reported here demonstrated that subcortical inputs to the cerebral cortex were required for the production of the characteristic bursts of generalized feline penicillin epilepsy which usually exhibited a frequency range of 3.5 to 5 cycles/s and consist of spike and wave or multiple spike or sharp wave discharges. The experiments with unilateral submaximal thalamectomy, associated with transection of the interhemispheric commissures, indicated that the important afferent inputs to the cortex responsible for the precipitation of these characteristic bursts originated in or passed through the thalamus. The absence of spindles on the thalamectomized side in these animals confirms similar findings obtained by Naquet et al. (17), by Andersen et al. (l), and by Lanoir (15) and are in agreement with the hypothesis that thalamocortical volleys are responsible for evoking spindles as well as the generalized epileptic bursts of feline generalized penicillin epilepsy (7, 8, 20). This hypothesis receives further support from the observation that when spindle activity is merely reduced, but not totally abolished by a less extensive thalamic lesion, epileptiform bursts in response to penicillin are also reduced on the side of the lesion, but not abolished. The present experiments add the important information that the spontaneous epileptiform bursts in this model are as dependent on the thalamocortical spindle-generating mechanism as those elicited in response to single-shock and low-frequency subcortical stimulation (8, 20). However, cortex deprived of all subcortical inputs or only of thalamocortical inputs still produces epileptic discharges, even without penicillin. These are therefore not dependent on the action of this drug, although they become more active under its influence. They indicate an increased tendency of deafferented cortex to produce epileptiform discharge (2, 3, 21). The discharges in this deafferented cortex differ, however, from those characteristic of feline generalized penicillin epilepsy by their slower frequency and by their tendency to occur in a more isolated fashion rather than in well-organized bursts. They share with them,
170
PELLEGRINI,
MUSGRAVE,
AND
GLOOR
SUBCORTICAL
INPUT
IN GENERALIZED
EPILEPSY
171
however, the tendency to be widely synchronized. In the case of a unilateral thalamectomy with split corpus callosum, the discharges appear over the thalamectomized side only and are synchronized within the cortical regions from which recordings are obtained. In the case of the bilateral cortical slab with intact callosal connections, the discharges are almost always bilaterally synchronous. These findings indicate that powerful cortico-cortical synchronizing pathways exist within each hemisphere, and that the corpus callosum is an important anatomic substrate for establishing bilateral synchrony. The corpus callosum is also responsible for the bilateral synchrony of the typical discharges of generalized penicillin epilepsy in the intact animal (Musgrave and Gloor, in preparation). This is also suggested by the observations made in the bilateral cortical slab preparation in which the typical bursts of feline generalized penicillin epilepsy in cortex lateral to the bilateral cortical slabs occurred asynchronously on the two sides, presumably because the callosal connections of these cortical regions had been severed in the course of the surgical procedures used to produce the bilateral cortical slabs (Fig. 1). The ability of thalamically deafferented cortex to produce widespread epileptiform discharge of a slow spike and wave type was also demonstrated by the fact that such discharges could be elicited by the topical application of a dilute penicillin solution to the cortex of both hemispheres in animals with a unilateral subtotal thalamectomy. The discharges on the two sides, however, differed from each other in the same manner as in animals with an unilateral thalamectomy in which pencillin had been injected intramuscularly. An interesting observation made in cats with a unilateral thalamic lesion was that the amount of epileptic activity occurring on that side both before and after penicillin was dependent on the level of arousal, epileptic discharges being present only when the animal was drowsy or asleep. This suggests that, in spite of the removal of the thalamus, connections from the brain stem mediating arousal responses remained, at least in part, intact and thus must have by-passed the thalamus (23). It is of interest that moderate arousal compatible with the presence of the typical epileptic discharges of feline generalized penicillin epilepsy on the intact side was sufficient to suppress the “atypical” cortical epileptic activity on the thalamectomized side, even under the influence of systemic penicillin. These experiments thus demonstrate that the occurrence of the typical spontaneous generalized bilaterally synchronous 3.5 to Scycle/s epileptiform bursts of feline generalized penicillin epilepsy depend on an intact thalamocortical input which normally most likely is involved in spindle genesis (7,8,20). Deafferented cortex, however, even without penicillin, is
172
PELLEGRINI,
MUSGRAVE,
AND
GLOOR
capable of producing widespread epileptiform discharges which are slow, but often assume a spike and wave form and exhibit prominent intrahemispheric or interhemispheric synchronization, the latter being dependent on an intact corpus callosum. The mechanism of this widespread synchronization thus, does not depend on subcortical structures, and thus confirms earlier findings obtained in a different model by Marcus and Watson (16). The slow spike and wave form occurring in widely synchronized fashion in these animals with deafferented cortex raises the intriguing question of whether slow generalized or widespread spike and wave discharge in human epilepsy seen in association with large cerebral destructive lesions (as, for example, in porencephalic cysts) could be caused by the same mechanism. REFERENCES 1. ANDERSEN, P., S. A. ANDERSSON, AND T. LIMO. 1967. Some factors involved in the thalamic control of spontaneous barbiturate spindles. J. Physiol. (London) 192: 257-281. 2. ECHLIN, F. A. 1959. The supersensitivity of chronically “isolated” cerebral cortex as a mechanism in focal epilepsy. Electroenceph. C/in. Neurophysiol. 11: 697-722. 3. ECHUN, F. A., AND A. BATTISTA. 1963. Epileptiform seizures from chronic isolated cortex. Arch. Neurol. 9: 154-170. 4. FEINDEL, W., Y. L. YAMAMOTO, AND C. P. HODGE. 1967. Intracarotid fluorescein angiography: a new method for examination of the epicerebral circulation in man. Can. Med. Assoc. J. 96: l-7. 5. FISHER, R. S., AND D. A. PRINCE. 1977. Spike-wave rhythms in cat cortex induced by parenteral penicillin. I. Electroencephalographic features. Electroenceph. C/in. Neurophsyiol. 42: 608-624. 6. GLOOR, P., G. BALL, AND N. SCHAUL. 1977. Brain lesions that produce delta waves in the EEG. Neurology (Minneapolis) 21: 326-333. 7. GLOOR, P., A. PELLEGRINI, AND G. K. KOSTOPOULOS. 1979. Effects of changes in cortical excitability upon the epileptic bursts in generalized penicillin epilepsy of the cat. Electroenceph. C’lin. Neurophysiol., in press. 8. GLOOR, P., L. F. QUESNEY, AND H. ZUMSTEIN. 1977. Pathophysiology of generalized penicillin epilepsy in the cat: the role of cortical and subcortical structures. II. Topical application of penicillin to the cerebral cortex and to subcortical structures. Electroenceph. Clin. Neurophysiol. 43: 79-94. 9. GLOOR, P., AND G. TESTA. 1974. Generalized penicillin epilepsy in the cat: Effects of intracarotid and intravertebral pentylenetetrazol and amobarbital injections. Electroenceph. Clin. Neurophysiol. 36: 499-515. 10. GRAFSTEIN, B. 1959. Organization of callosal connections in the suprasylvian gyrus of cat. .I. Neurophysiol. 22: 504-515. 11. HOHBERGER,~. P.,Y. L. YAMAMOTO,~. J.THOMPSON,AND W. FEINDEL. 1975. On-line computer measurement of microregional cerebral blood flow by xenon-133 clearance. Int. J. Nucl. Med. Biol. 2: 153-158. 12. INGVAR, D. H. 1955. Electrical activity of isolated cortex in the unanaesthetized cat with intact brainstem. Acra Physiol. Stand. 33: 151-168.
SUBCORTICAL
INPUT IN GENERALIZED
EPILEPSY
I73
13. INGVAR, D. H., AND N. A. LASSEN. 1962. Regional blood flow of the cerebral cortex determined by Krypton 85. Acta Physiol. Stand. 54: 325-338. 14. KELLAWAY, P., A. GOL, AND M. PROLER. 1966. Electricalactivity ofthe isolated cerebral hemisphere and isolated tha.lamus. Exp. Neural. 14: 281-304. 15. LANOIR, J. 1972. Etude tilectrocorticographique de la veille et du sommeil chez le chat. Organisation du cycle nyctht!mtral. R81e du thalamus. Thesis. Universitt de Provence Aix-Marseille, France. 16. MARCUS, E. M., AND C. W. WATSON. 1%6. Bilateral synchronous spike wave electrographic patterns in the cat. Interaction of bilateral cortico-callosal and adiencephalic preparation. Arch. Neural. 14: 601-610. 17. NAQUET, R., M. DENAVIT, J. LANOIR, AND D. ALB~-FESSARD. 1965. AltCrations transitoires ou definitives des zones diencephaliques chez le chat. Leurs effets sur l’activite electrique corticale et le sommeil. Pages 107-131 in Aspects Anatomo-fonctionnels de la Physiologie du Sommeil. Editions du Centre National de la Recherche Scientifique, Paris. 18. PRINCE, D. A., AND D. FARRELL. 1%9. “Centrencephalic” spike wave discharges following parenteral penicillin injection in the cat. Neurology (Minneapolis) 19: 309-310. 19. QUESNEY, L. F., AND P. GLOOR. 1978. Generalized penicillin epilepsy in the cat: correlation between electrophysiological data and distribution of r4C-penicillin in the brain. Epilepsia 19: 35-45. 20. QUESNEY, L. F., P. GLOOR, E. KRATZENBERG, AND H. ZUMSTEIN. 1977. Pathophysiology of generalized penicillin epilepsy in the cat: the role of cortical and subcortical structures. I. Systemic application of penicillin. Etectroenceph. C/in. Neurophysiol. 42: 640-655. 21.
SHARPLESS, S. K., AND L. M. HALPERN. 1962. The electrical excitability of chronically isolated cortex studied by means ofpermanently implanted electrodes. Electroenceph. Clin.
Neurophysiol.
14: 244-255.
22. SOEJIMA, T.. Y. L. YA~AMOTO, E. MEYER, AND W. FEINDEL. 1979. Effects of steroids on early microcirculatory changes following focal cold injury of cerebral cortex. J. Neurosurg., in press. 23. STARZL, T. E., C. W. TAYLOR, AND H. W. MAGOUN. 1951. Ascending conduction in reticular activating systems with special reference to the diencephalon. J. Neurophysiol.
14: 461-477.