Glutamatergic activation of the amygdala differentially mimics the effects of audiogenic seizure kindling in two substrains of genetically epilepsy-prone rats

Comparisons of neuronal network mechanisms in closely related inherited seizure models are providing novel insights into epileptogenic pathophysiology. Genetically epilepsy-prone rats (GEPRs) exist in two substrains that inherit long-term susceptibility to behaviorally distinct audiogenic seizures (AGS). GEPR-3s exhibit generalized clonic AGS, while GEPR-9s exhibit generalized tonic AGS. After AGS kindling the tonic AGS of GEPR-9s is followed by generalized posttonic clonus (PTC), while the generalized clonic AGS is followed by facial and forelimb (F&F) clonus in GEPR-3s. PTC and F&F clonus are very rare in GEPRs before AGS kindling. The neuronal network subserving AGS in GEPR-9s lies exclusively in brainstem sites, but amygdala (AMG) and other sites are recruited into the network after AGS kindling. The present study attempted to mimic the effects of AGS kindling by bilaterally microinjecting subconvulsive doses of N-methyl-D-aspartate (NMDA) into the AMG of nonkindled GEPRs. NMDA (10 nmol/side) microinjected into AMG reversibly induced susceptibility to F&F clonus immediately following generalized clonic AGS in most nonkindled GEPR-3s. NMDA (7.5 nmol/side), microinjected into AMG temporarily induced susceptibility to generalized PTC immediately following tonic AGS in most nonkindled GEPR-9s. No seizures were induced in normal rats by these treatments, and no seizures were seen in GEPRs with these NMDA doses except those induced by acoustic stimuli. These findings support a critical role in AGS kindling for the AMG in the neuronal networks for both forms of AGS. However, the behavioral effect of the treatment was different in the two AGS substrains, suggesting interrelated but not identical pathophysiological mechanisms in these closely related epilepsy models.     

Effect of hypertension on the integrity of blood brain and blood CSF barriers,cerebral blood flow and CSF secretion in the rat

Hypertension has been related to the development of brain damage, dementia and other CNS dysfunctions. Disruption of the blood-brain barrier (BBB) is thought to contribute to these disorders. In this study, the integrity of both blood-brain and blood-CSF barriers during chronic hypertension was investigated. For this, the entry of [14C]sucrose and of lanthanum into brain tissue, choroid plexus, and CSF was studied. Also brain regional blood flow and brain [14C]sucrose volume of distribution were measured using indicator fractionation and ventriculo-cisternal perfusion methods, respectively. The above measurements were performed in normotensive (WKY) rats and in the spontaneously hypertensive rats (SHR). Choroid plexus and CSF uptakes of [14C]sucrose were found to be significantly greater in SHR compared to WKY rats (P<0.05). Intercellular entry of lanthanum was observed in choroidal tissue of SHR but not in that of WKY rats and at the BBB. Choroid plexus blood flow was significantly greater in SHR, 2.82+/-0.21 ml g(-1) min(-1), compared to 2.4+/-0.08 ml g(-1) min(-1) in WKY (P<0.05). There were no significant differences (P>0.05) in brain % water content and extracellular fluid [14C]sucrose volume of distribution between SHR and WKY rats. However, choroid plexus showed greater % water content in SHR (85.7+/-1.9%) compared to the WKY rats (81.5+/-1.7%). These results suggest that chronic hypertension in SHR may cause more pronounced defects in the integrity of the blood-CSF barrier than in the BBB.     

Effects of a static magnetic field on audiogenic seizures in black Swiss mice

Effects of a static magnetic field (SMF) with strong gradient components were studied in black Swiss mice. Exposure to SMF (100-220 mT, 15-40 T/m for 1h) did not affect the threshold for detecting auditory brainstem responses. Serial seizures elevated the hearing threshold at some frequencies, but there was no difference between SMF-exposed and unexposed control mice. EEG changes were recorded during audiogenic seizures. Pretreatment with SMF prolonged seizure latency in response to stimulation with white noise of increasing intensity from 74 to 102 dBA (1 min interval between 2 and 4 dBA increments) without significant effects on seizure severity. Gender-related differences were not statistically significant. Stimulation with 10 min sound steps revealed prolongation of latency and reduction of seizure severity in SMF-exposed, but not unexposed, mice. Pretreatment with phenytoin (5 mg/kg) in combination with SMF had significantly greater effects on seizure latency and severity than either pretreatment alone. These findings indicate that the SMF studied here under different conditions elevated seizure threshold and had anticonvulsant properties in Black Swiss mice and increased the efficacy of a conventional anticonvulsant drug.     

Effects of carisbamate (RWJ-333369) in two models of genetically determined generalized epilepsy, the GAERS and the audiogenic Wistar AS

PURPOSE: The antiepileptic effects of carisbamate were assessed in two models of genetic epilepsy, a model of absence seizures, the Genetic Absence Epilepsy Rat from Strasbourg (GAERS) and a model of convulsive seizures, the Wistar Audiogenic Sensitive (AS) rat. METHODS: GAERS were equipped with four cortical electrodes over the frontoparietal cortex and the duration of spike-and-wave discharges (SWD) was recorded for 20-120 min. In Wistar AS, the occurrence of, latency to, and duration of wild running and tonic seizures were recorded. RESULTS: In GAERS, carisbamate (10, 30, and 60 mg/kg) dose dependently reduced the expression of SWD that totally disappeared at the two highest doses by 40 min after injection. SWD duration returned to control levels by 100 min after the injection of 30 mg/kg carisbamate while SWDs were totally suppressed for 120 min after the injection of 60 mg/kg carisbamate. In Wistar AS, 10 mg/kg carisbamate increased the latency to the first running episode and induced the occurrence of a second running episode in three of eight rats. This episode was not present in untreated rats and was indicative of decreased sensitivity to the stimulus. This dose of carisbamate increased by 327% the latency to the tonic seizure that still occurred in the six of eight rats studied. At 20 and 30 mg/kg, no rats exhibited any wild running or tonic seizure. CONCLUSIONS: The present results support the broad spectrum of antiepileptic activity of carisbamate confirming its efficacy in animal models of primary generalized seizures of both tonic-clonic and of the absence type.     

Alterations in brain metabolism, CNS morphology and CSF dynamics in adult rats with kaolin-induced hydrocephalus

The present study describes the biochemical changes, morphological development and the cerebrospinal fluid dynamics of the kaolin-induced hydrocephalus in the adult rat. Two, 4 and 6 weeks after microsurgical kaolin instillation into the rat cisterna magna the basal intracranial pressure and the cerebrospinal fluid outflow resistance were measured. To determine possible biochemical changes in the rat cerebrum, brain stem and cerebellum the concentrations of glutamine, glutamate, glutathione, aspartate, GABA, alanine and taurine were measured by high pressure liquid chromatography. In addition, ventriculomegaly and syringomyelia were assessed, measuring the lateral ventricles and central canals by means of an image-processing computer program. It could be shown that the acute phase of kaolin-induced hydrocephalus in the first 4 weeks is characterised by a high basal intracranial pressure, a considerably increased CSF outflow resistance and a rise in brain water content in the fourth week. The changes in the concentrations of amino acids were moderate. Glutamine was increased and taurine was decreased in the cerebrum and alanine was increased in the brain stem. The chronic phase, however, is defined by normal basal pressure, declining outflow resistance, progression of ventriculomegaly and distinct changes in the biochemical parameters such as a remarkable decrease of glutamate, glutamine and taurine in the cerebellum, a decrease of taurine and alanine plus an increase in glutamine in the cerebrum and an increase of alanine in the brain stem. Moreover, cerebral metabolism in the adult rat seems to be more resistant to the effects of hydrocephalus than metabolism in neonatal and infantile rats.     

Diagnostic significance of nerve cells in human CSF with particular reference to CSF cytology in the brain death syndrome

Summary Cytological examination of the cerebrospinal fluid (CSF) in 5 cases of the brain death syndrome treated on a respirator revealed the presence of nerve cells (mostly Purkinje cells), massive pleocytosis with many polymorphonuclear leukocytes, and numerous macrophages containing erythrocytes, leukocytes, lipid droplets, and hemosiderin. Portions of connective tissue with well preserved capillaries, and debris which could not be more closely identified, were also found.Neuropathological examination in 4 cases revealed progressive autolysis of brain tissue and displacement of the autolyzed tissue into the subarachnoid space of the spinal canal, partly due to tonsillar herniation. All five patients exhibited clinical signs of brain death and had been placed on respirators for various periods of time.The cytological findings in the CSF, together with the clinical signs of brain death, constitute an intravital morphological indication of brain tissue autolysis.This paper was presented in part at the 11th Danube Symposium for Neurological Sciences in Budapest 1978     

Effect of diet-induced obesity and experimental hyperinsulinemia on insulin uptake into CSF of the rat

We examined the hypothesis that the uptake of plasma insulin into cerebrospinal fluid (CSF) is saturable in two rat models. Dietary obese and control female Osborne Mendel rats received 24-h infusions of vehicle or insulin. CSF insulin levels in cafeteria- and chow-fed rats were comparable at all levels of plasma insulin (4.5 ± 2.8, 7.6 ± 2.4, and 23.9 ± 6.4 μU/ml in cafeteria diet vs. 4.5 ± 0.9, 6.8 ± 1.1, and 17.0 ± 4.0 μU/ml in chow rats). CSF insulin uptake as a percentage of plasma insulin decreased with increased plasma insulin in both groups. A similar relationship was observed in Wistar rats receiving 6-day infusions of vehicle or insulin (plasma insulin = 55 ± 12 vs. 365 ± 98 μU/ml; CSF/plasma insulin ratio = 0 ± .007 vs. 0.013 ± .006, respectively). Hyperinsulinemic Wistar rats did not demonstrate decreased brain capillary insulin binding vs. vehicle-infused controls. The results suggest that a saturable transport process contributes insulin transport into CSF in normal rats and that this process is not altered by moderate diet-induced obesity or hyperinsulinemia per se.     

Intractable seizures associated with brain tumor in childhood: lesionectomy and seizure outcome

The authors retrospectively reviewed ten pediatric brain tumor patients with intractable seizures who underwent lesionectomy without intentional identification and resection of the epileptogenic region to assess the clinical features and seizure outcome after lesionectomy in such patients. Seizures were complex partial in seven cases and simple partial, absence, and generalized tonic-clonic in one case each. Tumors mors were located at the medial temporal lobe in four cases, at the frontal lobe in four cases, at the parietooccipital and the suprasellar areas in one case each. The most common pathology was benign oligodendroglioma (five cases) followed by ganglioglioma (two cases). Others were pleomorphic xanthoastrocytoma, hamartoma, and primitive neuroectodermal tumor (one case each). In four cases, complete removal of the tumor was feasible. Postoperatively nine of the ten patients showed favorable seizure control (Engel's classification 1 and 2) and of these, six were seizure-free during the follow-up period (mean duration: 40 months). Therefore, lesionectomy can be an appropriate initial treatment for patients with brain tumor and medically intractable seizures.     

Biochemical brain markers and purinergic parameters in rat CSF after seizure induced by pentylenetetrazol

Cellular and molecular mechanisms involved in the generation of seizures and the magnitude of neural cells injury are not fully understood. We evaluated astrocyte and/or neuronal injury in rats in the pentylenetetrazol model of acute seizures by measuring S100B and NSE levels in cerebrospinal fluid. Additionally, we determined ADP and GDP hydrolysis by soluble nucleoside triphosphate diphosphohydrolase in the cerebrospinal fluid, and the concentration of nucleosides adenosine, inosine and guanosine as putative markers of brain injury. After pentylenetetrazol-induced seizures: (i) S100B values increased from 10 to 30 min, returning to control levels at 24 h; NSE levels presented a biphasic increase: an increase at 10 to 30 min returning to control levels, and again at 240 min followed by a decline at 24 h; (ii) nucleotidase activities increased from 10 min, returning to control levels at 240 min; (iii) guanosine and inosine levels increased exclusively after 30 min. In summary, this study showed biochemical changes in the cerebrospinal fluid occurring after seizures induced by pentylenetetrazol. Such events may have a modulating effect upon seizure expression, particularly nucleoside triphosphate diphosphohydrolase activities and nucleoside concentrations, but are nevertheless followed by neural death as evidenced by the increase in NSE and S100B levels.     

Audiogenic seizures in Wistar rats before and after repeated auditory stimuli: clinical,pharmacological, and electroencephalographic studies

Summary A strain of Wistar rats was inbred for susceptibility to audiogenic seizures characterized by one or two wild running fits followed by tonic dorsiflexion with open mouth and then a catatonic state. During the tonic phase, the cortical EEG was flat for 1 to 2 sec, then changed to a slow, regular lowamplitude discharge, 9 to 12c/s, for 25 to 60 sec. In these rats exposed to 40 daily 90-sec auditory stimuli, behavior and EEG changed. The wild running became disorganized by myoclonic jerks of the limbs and body. In some animals, the tonic extension disappeared and a myoclonic seizure developed progressively, with facial and forelimb clonus, and rearing and falling. In others, the tonic phase was followed by a generalized clonic phase. The EEG during the myoclonic and tonic-clonic seizures showed high-amplitude rhythmic spikes, polyspikes and spike-waves, 1 to 10 c/s, for 40 to 120 sec, often outlasting the sound stimulus. The effects of ethosuximide, carbamazepine and phenytoin were the same on primary and modified audiogenic seizures. The progressive behavioral and EEG modifications of audiogenic seizures following repeated auditory stimuli suggest that kindling had developed, the seizures being propagated from the brain stem to forebrain structures.     

C-Fos mapping and EEG characteristics of multiple mice brain regions in pentylenetetrazol-induced seizure mice model

ABSTRACT

Purpose: To confirm different local brain activities characterized in pentylenetetrazol (PTZ)-induced seizure model.

Methods: we induced seizure response by a single dose of PTZ injection (45 mg/kg, i.p.). Local activity was recorded in different brain regions by EEG in time and c-Fos staining at different time points (0.5 h, 1 h, 2 h, 4 h) after PTZ treatment.

Results: EEG recordings showed distinctive features of activation in different brain areas. With the aggravation of behavioral manifestations of seizures, the frequency and amplitude of the discharges on EEG were increasing gradually. The epileptic response on EEG immediately ended after reaching the maximum stage of seizures, followed by a short period of suppression. The labeling of c-Fos was enhanced in the medial prefrontal cortex, the piriform cortex, the amygdala, hippocampal CA1, CA3 and dentate gyrus, but inapparent in the striatum. The most potent changes in c-Fos were observed in cortex, amygdala nuclei, and dentate gyrus. EEG and c-Fos immunolabeling in neuronal activation showed discrepancies in the striatum. For each brain region, the maximum c-Fos labeling was observed at 2 h after injection and diminished at 4 h. The level of c-Fos immunoreactivity was even lower than the control group, which was accompanied by increased labeling of parvalbumin neurons (PVNs).

Conclusions: These findings validated PTZ-induced seizure as a seizure model with a specific spatial-temporal profile. Neuronal activity was enhanced and then subsequently inhibited during seizure evolution.

Abbreviations: AEDs: anti-epileptic drugs; AF: Alexa Fluor; CA1: Cornu Ammonis area 1; CA3: Cornu Ammonis area 3; DAB, 3: 3P-diaminobenzidine; DAPI: 4‘,6-diamidino-2-phenylindole; DG: dentate gyrus; EEG: electroencephalogram; GABA: gamma-aminobutyric acid; IEG: immediate early gene; mPFC: medial prefrontal cortex; NAc: nucleus accumbens; PB: phosphate buffer; PBS: phosphate buffered saline; PBST: phosphate buffered saline with Tween; PFA, paraformaldehyde; PTZ: pentylenetetrazol; PVN: parvalbumin neuron; ROI: regions of interest; SE: status epilepticus.     

Influence of Audiogenic Seizures on Synaptic Facilitation in Mouse Hippocampal Slices Is Mediated by N-Methyl-d-Aspartate Receptor

The influence of audiogenic seizures (AGS) on synaptic facilitation was studied in DBA/2J (D2) and C57BL/6J (B6) mice. AGS susceptibility is inherited in D2 mice, but can be acquired in AGS-resistant B6 mice through acoustic priming. The experiments were per formed on hippocampal slices obtained from D2 and B6 mice both with and without seizures. Long-term potenti ation (LTP) and low-Mg^{2 +}-induced synaptic facilitation (LMISF) were evaluated after stimulation of Schaffer col laterals. The magnitude of LTP and LMISF was signifi cantly greater in slices obtained from mice with seizures than from mice without seizures in both strains. Seizure induced enhancement of LTP and LMISF was markedly reduced by the JV-methyl-D-aspartate (NMDA) receptor antagonist AP-5. The noncompetitive NMDA receptor antagonist MK 801 reduced the efficiency of priming in B6 mice and abolished AGS in D2 mice and primed B6 mice, the data suggest that audiogenic seizures can en hance synaptic facilitation through activation of the NMDA receptor.     

Precipitous induction of audiogenic kindling by activation of adenylyl cyclase in the amygdala

Purpose: Kindling of audiogenic seizure (AGS) involves ≥14 AGS over 1–2 weeks in genetically epilepsy‐prone rats (GEPR‐9s) and induces gradual seizure duration increases, epileptiform electroencephalography (EEG), and emergence of post tonic clonus (PTC), which are long‐lasting. N‐methyl‐d ‐aspartate (NMDA)–receptor activation in lateral amygdala (LA) is implicated in AGS kindling initiation. However, the persistence of AGS kindling appears to be dependent on molecular mechanisms initiated by NMDA‐receptor activation, which may involve adenylyl cyclase (AC). This study attempted to mimic AGS kindling persistently in nonkindled GEPR‐9s by one‐time activation of AC in LA. Methods: The effects of a single focal bilateral microinjection into LA of an AC activator, MPB forskolin {7‐Deacetyl‐7‐[O‐(N‐methylpiperazino)‐γ‐butyryl]‐forskolin dihydrochloride} (25–100 pmol/side), on seizure behavior in GEPR‐9s were evaluated. Results: One‐time bilateral microinjection of MPB forskolin in GEPR‐9s precipitously induced an AGS kindling–like effect, which involved significant increases in seizure duration and long‐lasting susceptibility to AGS that culminates in PTC. This effect occurred at 24 h after MPB forskolin microinjection and lasted ≥5 weeks. The effect was seen when AGS was initiated at 1 and 12 h after microinjection, but not if AGS was induced only at 24 h, indicating the importance of the temporal proximity of AGS induction to the MPB forskolin microinjection. Discussion: These findings indicate that one‐time activation of AC within the NMDA receptor–mediated molecular cascade results in precipitous induction of AGS kindling. These data further suggest that AC activation in the LA may be an important epileptogenic mechanism that subserves the long‐lasting persistence of AGS kindling.     

Cortical Aquaporin-4 in relation to brain oedema and neurological function of cortical cryo-injured mice

To estimate the spatial and temporal expression of Aquaporin-4 (AQP-4) in a murine model of automated cerebral cryoinjury and correlate AQP-4 expression with development of brain oedema and neurological function. AQP-4 levels were determined quantitatively by Western blots at site of injury and at sites adjacent to and distant from injury in brains of cryoinjured (experimental) (n = 18), sham injured (n = 18) & normal mice at 24, 48, 72 h post injury. AQP-4 expression was correlated with percentage water content of brain, Neurological Severity Score (NSS) and rotarod scores. We found a 1.4-fold increase in expression of AQP-4 at the site of injury and at sites distant from injury at 24 h when compared to normal mice (p = 0.05). The increase in expression of AQP-4 24 h post injury was significantly higher in experimental group at the site of injury and at the site adjacent to the injury in the ipsilateral hemisphere when compared to the sham injured mice (p = 0.05). At 24 h post injury the median NSS score in the experimental group was 9 (interquartile range 7.25–10) and that in the sham group was 0.5 (interquartile range 0.0–1.0) (p < 0.001).At 48 and 72 h, AQP-4 expression remained elevated in the experimental group when compared to normal brain, but the levels were not significantly different from that in sham group. AQP-4 expression was significantly elevated in the ipsilateral hemisphere in the first 24 h following cerebral cortical injury in mice and this could be correlated with worsening of neurological function. Over the next 48 h, there was a trend towards decrease in AQP-4 expression that was associated with partial recovery of neurological function.     

Acoustic priming lowers the threshold for electrically induced seizures in mice inferior colliculus, but not in the deep layers of superior colliculus

Mice become highly susceptible to audiogenic seizures (AGS) after exposing them to an intense noise in their early life (priming). To elucidate the brain mechanisms for this priming effect of AGS, we compared the threshold current intensities inducing AGS syndromes between primed (n=88) and non-primed (n=84) mice by electrically stimulating the central nucleus and external cortex of the inferior colliculus (CIC and ECIC), and the deep layers of the superior colliculus (DLSC). The threshold for wild running was significantly lower for the primed mice than for the control mice in the case of the CIC and ECIC, but not the DLSC. The current intensity for inducing clonic seizure was lower for the primed mice than for the control mice in the case of the ECIC. These results show that the inferior colliculus (IC) plays an important role in the priming effect of AGS in mice, but that the DLSC does not.     

Audiogenic seizure susceptibility is reduced in fragile X knockout mice after introduction of FMR1 transgenes

The Fmr1 knockout (KO) mouse is characterized by an increased audiogenic seizure (AGS) susceptibility and is considered a good animal model for epilepsy and seizures in the human fragile-X (FRAX) syndrome. Here, we tested the hypothesis that the reintroduction of the FMR1 gene is able to revert the AGS susceptibility characterizing Fmr1 KO mice. To this aim, two groups of Fmr1 KO transgenic mice, which have additional copies of the human FMR1 gene (YAC) or FMR1 cDNA (G6) were used. AGS susceptibility of these mice was examined and compared to that of Fmr1 KO, wild type, and wild-type animals in whom the FMR1gene was also introduced (over-expressed). Mice were tested at different ages because AGS susceptibility is age dependent. The intensity of response was scored and the results were analyzed by means of 2-way analysis of variance to evaluate the effects of age and genetic condition. We found that AGS susceptibility rescue is complete in the G6 mice and partial in YAC mice. Our data indicate that the introduction of the human FMR1 gene in Fmr1 KO mice is able to revert the Fmr1 KO epileptic phenotype.     

Role of the superior colliculus in the expression of acute and kindled audiogenic seizures in Wistar audiogenic rats

Purpose: The role of the superior colliculus (SC) in seizure expression is controversial and appears to be dependent upon the epilepsy model. This study shows the effect of disconnection between SC deep layers and adjacent tissues in the expression of acute and kindling seizures. Methods: Subcollicular transections, ablation of SC superficial and deep layers, and ablation of only the cerebral cortex were evaluated in the Wistar audiogenic rat (WAR) strain during acute and kindled audiogenic seizures. The audiogenic seizure kindling protocol started 4 days after surgeries, with two acoustic stimuli per day for 10 days. Acute audiogenic seizures were evaluated by a categorized seizure severity midbrain index (cSI) and kindled seizures by a severity limbic index (LI). Results: All subcollicular transections reaching the deep layers of the SC abolished audiogenic seizures or significantly decreased cSI. In the unlesioned kindled group, a reciprocal relationship between limbic and brainstem pattern of seizures was seen. The increased number of stimuli provoked an audiogenic kindling phenomenon. Ablation of the entire SC (ablation group) or of the cerebral cortex only (ctx‐operated group) hampered the acquisition of limbic behaviors. There was no difference in cSI and LI between the ctx‐operated and ablation groups, but there was a difference between ctx‐operated and the unlesioned kindled group. There was also no difference in cSI between SC deep layer transection and ablation groups. Results of histologic analyses were similar for acute and kindled audiogenic seizure groups. Conclusions: SC deep layers are involved in the expression of acute and kindled audiogenic seizure, and the cerebral cortex is essential for audiogenic kindling development.     

Identification of the requisite brain sites in the neuronal network subserving generalized clonic audiogenic seizures

Comparative studies of neuronal networks that subserve convulsions in closely-related epilepsy models are revealing instructive data about the pathophysiological mechanisms that govern these networks. Studies of audiogenic seizures (AGS) in genetically epilepsy-prone rats (GEPRs) and related forms of AGS demonstrate important network similarities and differences. Two substrains of GEPRs exist, GEPR-9s, exhibiting tonic AGS, and GEPR-3s, exhibiting clonic AGS. The neuronal network for tonic AGS resides exclusively in brainstem nuclei, but forebrain sites, including the amygdala (AMG), are recruited after repetitive AGS induction. The neuronal network for clonic AGS remains to be investigated. The present study examined the neuronal network for clonic AGS in GEPR-3s by microinjecting a competitive NMDA receptor antagonist, D,L-2-amino-7-phosphonoheptanoic acid (AP7), into the central nucleus of inferior colliculus (ICc), deep layers of superior colliculus (DLSC), periaqueductal grey (PAG), or caudal pontine reticular formation (cPRF), which are implicated in tonic AGS networks. Microinjections into AMG and perirhinal cortex (PRh), which are not implicated in AGS, were also done. AGS in GEPR-3s were blocked reversibly after microinjections into ICc, DLSC, PAG or cPRF. However, AGS were also blocked by AP7 in AMG but not PRh. The sites in which AP7 blocks AGS are implicated as requisite components of the clonic AGS network, and these data support a critical role for NMDA receptors in clonic AGS modulation. The brainstem nuclei of the clonic AGS network are identical to those subserving tonic AGS. However, the requisite involvement of AMG in the clonic AGS network, which is not seen in tonic AGS, is surprising and suggests important mechanistic differences between clonic and tonic forms of AGS.