The NMDA-receptor antagonist, MK-801, suppresses limbic kindling and kindled seizures

90

Brain Research, 463 (1988) 90-99 Elsevier

BRE 14005

The NMDA-receptor antagonist, MK-801, suppresses limbic kindling and kindled seizures M.E. Gilbert Northrop Services Inc., Environmental Sciences, Research Triangle Park, NC27709 (U.S.A.) (Accepted 11 May 1988) Key words: Kindling; Amygdala; Hippocampus; MK-801; N-Methyl-D-aspartate; Anticonvulsant

Excitatory amino acid receptors in the mammalian central nervous system (CNS) are divided into 3 receptor subtypes: kainate, quisqualate, and N-methyl-D-aspartate (NMDA). MK-801 is a selective, non-competitive antagonist of excitatory amino acid transmitters at the NMDA receptor site. The role of excitatory amino acid neurotransmission in electrical kindling was examined in animals stimulated daily in the amygdala following i.p. administration of low dosages of MK-801 (0.1 and 0.5 mg/kg). A second experiment evaluated the anticonvulsant properties of MK-801 in rats kindled in the hippocampus and amygdala, and contrasted its efficacy with the antiepileptic agents, diazepam, phenobarbital and phenytoin, and the dissociative anesthetics phencyclidine and ketamine. MK801 (0.5 mg/kg) retarded the development of amygdala kindling and reduced mean AD duration over the first 10 stimulation sessions. The low dosage reduced total AD accrued during each kindling stage but failed to alter kindling rate. MK-801 blocked motor seizures induced by stimulation of hippocampal or amygdala kindled foci, but was more effective in reducing seizure severity and AD duration resulting from stimulation of the hippocampal focus. All other drugs tested, with the exception of phenytoin, protected against amygdaloid kindled seizures. It was concluded that excitatory amino acid transmission contributes in an important, but non-critical way to amygdala kindling.

INTRODUCTION The excitatory amino acids, glutamate and aspartare, are found throughout the m a m m a l i a n central nervous system where they are believed to subserve excitatory synaptic transmission 53. T h r e e receptor subtypes have been pharmacologically and electrophysiologically identified on the basis of agonists which selectively activate them: N-methyl-D-asparrate ( N M D A ) , kainate, and quisqualate ~3,29. N M D A receptors and their associated ionic channels demonstrate the unique p r o p e r t y of being blocked by magnesium ions when the cell is at rest, depolarization acting to dislodge the magnesium in a voltage-dependent m a n n e r 1~'31. Consequently, N M D A receptors do not a p p e a r to participate in low frequency synaptic transmission, but are activated by glutamate and aspartate only following sufficient m e m b r a n e depo-

larization. Once activated, N M D A r e c e p t o r responses are characterized by slow rise times, prolonged, rhythmic depolarizations, and regular bursts of action potentials 26. G l u t a m a t e , aspartate and their agonists have been shown to induce epileptiform activity (e.g. refs. 21, 36, 37, 50). Antagonists of the N M D A - r e c e p t o r subtype have been shown to possess strong anticonvulsant properties in various acute chemical seizure models 3"61~~2"Ks'19~25, and audiogenic seizures in a genetically predisposed strain of mouse 2~. Until recently, however, little was known of the role of excitatory amino acid transmission in the chronic kindling seizure model. Kindling is a model of t e m p o r a l lobe epilepsy in which seizures are gradually induced in response to r e p e a t e d , brief, low-intensity stimulation of a n u m b e r of limbic brain sites 17. Behaviorally and electrographically, the animal progresses through

Correspondence: M.E. Gilbert, United States Environmental Protection Agency, Health Effects Research Laboratory, Neurotoxicology Division, Mail Drop 74B, Research Triangle Park, NC 27711, U.S.A. 0006-8993/88/$03.50 © 1988 Elsevier Science Publishers B.V. (Biomedical Division)

91 several stages of increasingly more severe seizure activity, to culminate eventually in a fully generalized convulsion. The underlying change in brain function, occurring in the absence of overt neuropathology, is permanent, the animal maintaining the enhanced propensity for seizures 4°. Glutamate release, as measured in perfusate obtained from the lateral ventricle, is enhanced following generalized kindled seizures 36. Cain et al. 5 have recently demonstrated that microinfusion of the competitive antagonist Dt.-2-amino-5-phosphonovaleric acid (APV) directly into the amygdala suppresses development of amygdala kindling. Intraventricular 3~' and intra-amygdaloid 5 administration of APV also antagonized the expression of generalized seizures in animals previously kindled in the amygdala. MK-80 l is a non-competitive NMDA-receptor antagonist s'54 whose potency for blocking N M D A responses exceeds that of APV a4. Unlike APV, non-competitive antagonists such as MK-801, phencyclidine, and ketamine do not block the NMDA receptor itself, but act at a site within the ion channel 22'27. Their strong use dependency (increased block in the presence of the agonist), coupled with their highly lipophilic nature (permitting easy penetrability of the bloodbrain barrier) have recently led to recognition of their therapeutic potential 4"22. The present experiment was designed to investigate the role of excitatory amino acid neurotransmission in the development of kindling using MK-801 to antagonize glutaminergic neurotransmission. Since the mechanisms that underlie the development versus the maintenance of the kindled response appear to be distinct 3<4° the prophylactic properties of MK801 during kindling development and its anticonvulsant efficacy in fully kindled animals were evaluated. The anticonvulsant properties of MK-801 in kindled animals were compared and contrasted with the antiepileptics, diazepam, phenobarbital and phenytoin. Two other drugs, ketamine and phencyclidine, were included in the analysis because of their NMDA-antagonistic properties 2. A preliminary report of these data has been presented 2s. MATERIALS AND METHODS

Subjects Adult male Long-Evans rats obtained from the

Charles River Breeding Co. served as subjects. Animals were housed individually in plastic cages with wood chip bedding in a colony room maintained on a 12 h/12 h light-dark schedule and a constant room temperature of 24 °C. Animals were permitted free access to food and water at all times. At 90-120 days of age, animals were anesthetized with 60 mg/kg sodium pentobarbital, administered 0.2 mg/kg of atropine sulphate, and were implanted stereotaxically with electrodes placed in basolateral amygdala (n = 33) or dorsal hippocampus (n = 9). With the incisor bar set at 3.3 mm below the interaural plane, the coordinates for the bilateral amygdala placements were 2.5 mm posterior to bregma, 4.0 mm lateral to the midline, 7.5 mm ventral to the dura. For hippocampal kindling, electrodes were placed in the dorsal dentate gyrus (3.5 mm posterior to bregma, 2.2 mm lateral to middle, 3.5 mm posterior to dura) and ipsilateral perforant path (7.9 mm posterior to bregma, 4.4 mm lateral to midline, 3.0 mm ventral to dura). Electrodes were constructed of twisted strands of nichrome or stainless steel wire (each strand 125 urn), insulated except for the tips, and crimped onto gold-plated male Amphenol pins. The tips of the electrodes were separated vertically by 0,5 ram. The Amphenol pins were inserted into a 9-pin connector, which was cemented to the animal's skull and anchored with stainless steel screws (0.90 x 1/16 inch). The animals were grounded through a screw inserted in the skull overlying the anterior neocortex (approximately 3.0 mm anterior to bregma and 2.5 mm lateral to the midline). Immediately following surgery all animals received an administration of penicillin G (100,000 units).

Procedure Development of arnygdala kindling. The objective of the first experiment was to assess the effects of two dosages (0.1 and 0.5 mg/kg) of MK-801 on development of amygdala kindling. Dosages between 0.3 and 1.0 mg/kg have been shown to be effective in blocking bicuculline-induced seizures 7 and protecting against ischemia-induced cell death 16. One to two weeks following surgery, animals with electrodes implanted in the amygdala were individually placed in a large Plexiglas testing chamber and connected to the stimulating/recording apparatus via a low-noise shielded cable. The electroencephalogram (EEG)

92 was recorded on a Grass polygraph before and after stimulation in order to record a prestimulation baseline and evoked afterdischarge (AD). An AD was defined as rhythmic spiking, at least 4 times the amplitude of the baseline E E G , sustained for a minimum of 5 s. AD was monitored from the amygdala by recording between the tips of the bipolar electrode. Stimulation consisted of a 1-s train of 60-Hz biphasic square-waves, each 1 ms in duration, from a Grass S88 stimulator and PSIU6 constant-current converters. AD threshold testing began 30 min following the initial i.p. injection of 0.0, 0.1 or 0.5 mg/kg of MK801 in a 1-ml volume of saline. Thresholds were determined by administering a series of stimulations beginning at 20 ~tA and increasing in 20-/~A steps at 1min intervals until an AD was observed. Following threshold determination, stimulation was delivered once daily, 30 min following injection, at a standard 200 ~tA intensity until 3 Stage-5 (ref. 39) generalized seizures were observed. Behavioral seizures were scored according to the following scheme39: initial stimulation sessions result in motor arrest, facial automatisms and chewing (Stage 1). Continued stimulation produces chewing and head-nodding (Stage 2), followed by forelimb clonus contralateral to the side of stimulation (Stage 3). Rearing and bilateral forelimb clonus (Stage 4), and finally rearing with bilateral clonus and loss of postural control (Stage 5) occur with additional stimulation as the seizure discharge spreads to the hindlimbs. Severity of the behavioral seizure, duration of the evoked AD, and duration of clonic seizure activity were recorded following each stimulation. AD thresholds and the duration of clonic seizure activity were analyzed using one-way ANOVAs with the a-level set at 0.05. Kindling rate and AD development were analyzed using repeated measures ANOVA using the Geisser-Greenhouse correction factor. Paired comparisons were performed using Tukey's test with a-level set at 0.05. Loss of headcaps prior to completion of kindling removed some animals from the kindling rate dataset. AD duration data from these subjects, however, were included in the analysis if the animal had sustained a minimum of 10 ADs. Amygdala and hippocampal kindled seizures. The objectives of the second experiment were to assess and compare the anticonvulsant properties of MK-

801: (1) following the establishment of stable kindled seizures and thresholds; (2) between foci established in the amygdala or hippocampus: and (3) to other known anticonvulsants and less specific N M D A antagonists. Eight to 10 animals previously kindled in the amygdala and 5 animals previously kindled in the dentate gyrus of the dorsal hippocampus were stimulated 3 times daily, at 2-h intervals, until a minimum of 20 generalized seizures were observed. At this point, animals had stable total AD durations and duration of clonic seizure activity, and stimulation always evoked a generalized (Stage 4 or 5) seizure. Thresholds were determined, and to ensure their stability, subjects were then stimulated, on alternate days, at 10gA above and 10BA below threshold for a minimum of 6 days. Suprathreshold stimulation always provoked and subthreshold stimulation always failed to provoke a generalized seizure for animals to be included in the drug testing phase of the experiment. Drugs were administered i.p., in dosages that produced a range of effects from no obvious behavioral change to signs of overt toxicity or sedation. The dosages, time of testing postdosing, time between dosings, and stimulation sites are summarized in Table I. Subjects were stimulated at 10 etA above threshold. If an AD was not evoked, intensity was increased in 10 g A steps at 2-min intervals until an AD was observed. The failure to evoke an AD at 30 ~A above threshold resulted in the entry of 0 for seizure stage and AD duration parameters. All animals received all dosages in a completely counterbalanced fashion. Barring subject attrition due to headcap loss or the development of unstable thresholds or seizure responses, all animals received all drugs in a randomized fashion, with a minimum of 1 week between the end of one drug regimen and the beginning of the next. During that week, threshold was redetermined, and the 10uA above and 10/~A below procedure was reinstated. This provided a new baseline prior to each drug test to control for repeated testing and multiple drug administrations. Seizure stage, AD duration and duration of clonic seizure activity were analyzed with repeated measure ANOVA using the Geisser-Greenhouse correction factor, and where significant, paired comparisons were performed using Tukey's test (a-level set at 0.05).

93 TABLE I Summarv of drugs, dosages, test times and threshold elevations in amygdaloid and htppocampal kindled foci

ADT=afterdischarge threshold. Drug

n

Dosages(mg/kg)

Post-dose test time (rain)

Inter-dosetest time (h)

Mean ADT (I,A)

% Subjects with threshold increases*

7 7 6 7 7 7

0 0 0 0 0 0

1.0 1.0 60 100 80 6

30 30 90 60 15 15

48 48 72 48 48 48

68.6+7.62 65.7 + 5.61 60.8_+ 4.55 50.7 + 6.94 85.0 ± 6.36 50.7 + 6.94

0 29 50 0 28 0

5 0 0.1 0.5 1.0

30

48

45.0-+6.52

40

Amygdala

MK-801 Diazepam Phenobarbital Phenytoin Ketamine Phencyclidine

0.1 0.1 10 20 20 2

0.5 0.5 30 50 40 4

Hippocampus

MK-801

* Percentage of subjects demonstrating threshold increase in excess of 30,uA at the highest dosage level.

Upon completion of the experiment, animals were overdosed with sodium pentobarbital and perfused through the heart with 10% formalin and saline. The brains were removed and prepared for histological analysis using standard techniques. Coronal sections were cut at 60/~m on a freezing microtome, mounted and stained with Cresyl violet to verify electrode placement. Only animals with electrodes lying within the amygdaloid complex and hilus of the dentate gyrus were included in the analyses. RESULTS Dosages of 0.5-1.0 mg/kg of MK-801 produced lateral head-nodding, circling, ataxia, thigmotaxis, and catatonic posturing in most animals. These druginduced behavioral effects were attenuated over the course of repeated injections dictated by the kindling paradigm. Similar results documenting the development of tolerance to the behavioral but not the anti° convulsant properties following chronic treatment with the N M D A antagonists, ketamine, phencyclidine 24 and A P V 5, have been reported. Animals in the low dosage (0.1 mg/kg) group did not exhibit any overt signs of toxicity. Development o f kindling

Kindling development, presented as the mean number of evoked A D s at each stage of motor seizure, is presented in Fig. 1. MK-801 significantly reduced kindling rate as confirmed by significant ef-

fects of Dose (F2,22 = 10.22, P < 0.0007), and Dose x Number of ADs interaction (Fs,ss = 5.26, P < 0.0018). Daily pretreatment with 0.5 mg/kg of MK801 (n = 8) prior to electrical stimulation retarded the development of amygdaloid kindling. Animals remained in stages 1 and 2 for a mean of 19.25 (_+3.51) sessions prior to the onset of generalization (Stages 3, 4 and 5). The mean number of A D s to the first Stage 5 seizure was 31.0 (_+3.41) as compared to the control mean of 17.75 (_+3.09). Four subjects from the high dosage group were eliminated from the analysis of kindling rate. Three of these animals had experienced 24, 34 and 37 evoked ADs prior to loss of their head caps, yet had not progressed beyond Stage 2. The fourth subject failed to progress beyond Stage 2 despite 56 evoked ADs. The 0.1 mg/kg dosage of MK-801 (n = 9) was without effect on kindling rate. The mean number of A D s to Stage 5 was 13.56 (_+1.61) which did not differ from the control group (n = 8). Mean duration of clonic seizure activity did not differ among the groups. A slight increase in A D threshold was detected and found to be marginally significant (F2,22 = 3.48, P < 0.0488). Thresholds for both the 0.1 mg/kg group (100.0 _+ 10.54/~A) and the 0.5 mg/kg group (85.0 _+ 8.24/~A) were elevated relative to the control group (67.5 + 6.48/~A) (Tukey P < 0.05). The progressive development of A D duration was examined over the first 10 stimulation sessions. A dose-dependent reduction in A D duration is evident

94 NUMBER OF SESSIONS PER STAGE OF SEIZURE I

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Fig. 1. A: mean (+ S.E.) number of evoked afterdischarges (AD) to each stage of motor seizure for amygdala kindling following treatmerit with 0.0, 0.1 or 0.5 mg/kg MK-801 delivered i.p. B: mean (+_ S.E.) afterdischarge (AD) durations (in seconds) during the first 10 amygdala stimulation sessions in control (0.(/mg/kg) and treated (0.1 and 0.5 mg/kg) animals. C: mean (_+ S.E.) cumulative AD at each stage of motor seizure. (* Indicates a significant difference from 0.0 mg/kg, Tukey P < 0.05.)

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Fig. 2. Mean (+ S.E.) seizure stage, afterdischarge (AD) duration, and duration of clonic seizure activity in (A) amygdala (n = 7), and (B) hippocampus (n = 5) kindled rats at baseline (B) and 4 dosage levels of MK-801, (C) phencyclidine (n - 7), (D) ketamine (n = 7), (E) phenobarbital (n = 6), (F) diazepam (n = 7), and (G) phenytoin (n = 7). Paired comparisons (Tukey, P < 0.05) were performed (significant differences from baseline indicated by *) if repeated-measures ANOVA revealed a significant effect of Dose, P < 0.05.

96 in Fig. lB. A repeated-measures ANOVA confirmed significant Dose (F2,30 = i4.62, P < 0.0001 and Dose x AD Session interaction (F~8,270 = 3.11, P < 0.0026) effects. The high dosage group (n = 15) differed significantly from controls (n = 9) at all but the first of ten AD sessions. AD duration was also suppressed in the low dosage group and differed significantly from controls at the third through fifth AD sessions (Tukey, P < 0.05). The total cumulative AD at each stage of motor seizure is presented in Fig. 1C. Cumulative AD was significantly shorter in the low dosage group as evidenced by a significant Dose effect (~.22 = 3.80, P < 0.0382). The greater number of stimulation sessions required to kindle animals in the high dosage group relative to controls equated the total AD accrued in these two groups. There was no Dose x AD Duration interaction (F~.ss = 1.98, P > 0.1316).

Amygdala and hippocampal kindled seizures The stage of motor seizure, AD duration, and duration of clonic seizure activity, at baseline and each of the 4 dosage levels for amygdala and hippocampal stimulation are presented in Fig. 2. Phenytoin was without effect on any parameters in amygdala kindled rats (Fig. 2G). All other drugs produced a significant reduction in seizure stage and duration of clonic seizure activity at the two high dosage levels. All drugs except MK-801 (Fig. 2A) and phenytoin (Fig. 2G) reduced AD duration at the two high dosage levels. MK-801 failed to reduce the focal AD, in spite of reducing the maximal seizure stage. Repeated-measures ANOVA were performed and significant differences relative to baseline levels (Tukey, P < 0.05) are indicated by an asterisk in Fig. 2. In animals kindled in the dorsal dentate gyrus, MK-801 effectively reduced the stage of motor seizure, duration of clonus, and the AD duration at the two high dosage levels (Fig. 2B). MK-801 appeared to be more efficacious in reducing motor seizure stage in the dentate gyrus (mean seizure stage at 1.0 mg/kg = 1.40 _+ 0.87) than in the amygdala focus (mean seizure stage at 1.0 mg/kg = 2.57 + 1.40) (compare Fig. 2A and B). A similar pattern was seen with AD duration in amygdala versus hippocampus. The maximal decrease from mean saline AD duration was 71% for the hippocampal group, ,and only 28% for the amygdala group (Fig. 2A,B).

Diazepam, phenobarbital, and ketamine increased AD thresholds in excess of 30ktA at the highest dosage level (see Table I). MK-801, phencyclidine, and phenytoin did not alter threshold in amygdala kindled rats. Interestingly, MK-801 did elevate thresholds in 2 of 5 animals kindled in the hippocampus but a ~ analysis failed to reach statistically significant levels ()~ = 3.36, P < 0.10).

DISCUSSION MK-801 produced a dramatic retardation in the development of amygdala kindling, but not at a dosage level that was without behavioral side-effects. It is possible that MK-801 retarded kindling through a restriction of spread of AD outside the amygdala focus since the seizures remained focal (Stages 1 and 2) for many sessions. The argument could be made that MK-801 retarded kindling by a simple shortening of evoked AD as is seen with anticonvulsants such as phenobarbital and diazepam. Like high dosages of MK-801, these drugs shorten the duration of each focal AD and increase in the number of ADs required to kindle the animal, but the total AD accrued in reaching the fully kindled state is not modified 35. In contrast, however, in the present experiment the low dosage of MK-801 (in the absence of overt behavioral effects), also suppressed AD for several sessions and the total cumulative AD at each seizure stage, yet failed to disrupt normal development of the motor response to kindling stimulation. Paradoxically, in amygdala kindled animals, MK-801 reduced seizure stage in the absence of a significant reduction in AD duration. These findings demonstrate complex dose-related actions of MK-801 on seizure activity and suggest a degree of independence in processes underlying the motoric and electrographic components of kindling. N M D A receptor binding sites have been localized in the amygdala, although the highest density appears within the hippocampus m'54. MK-801 has been shown to bind with high affinity to rat brain membrane and to antagonize depolarizing responses to NMDA a°. The present findings are consistent with previous reports in which the competitive NMDAantagonist APV retarded the development of an acute form of hippocampal kindling in vitro 47, and

97 chronic amygdaloid kindling in vivos. MK-801 and APV have also been shown to block the increase in synaptic efficacy induced by high frequency trains (long-term potentiation, LTP) in area CA1 of hippocampus in vitro ~'9 and dentate gyrus in vivo 1a33. The repetitive firing characteristics of epileptiform activity provides the conditions necessary for NMDA activation -~7 and induces LTP 41. Moreover, prior delivery of a series of non-AD provoking LTP-inducing tetanizing trains has been shown to facilitate subsequent kindling 42"49. Although synaptic potentiation cannot fully account for kindling 4a, its blockade may delay the initial activation of extrafocal sites. Antagonism of NMDA-induced excitation with MK-801 does prevent the induction of LTP (see above) and in this manner, may have retarded seizure spread outside of the amygdala focus. Alternatively, or perhaps additionally, MK-801 may produce its effect on kindling development by blocking the response to NMDA in other brain areas. For example, it has been demonstrated that seizures induced by the cholinergic agonist, pilocarpine, can be blocked by microinfusion of APV into the substantia nigra, and subconvulsive dosages of pilocarpine summate with intranigral microinfusions of NMDA to induce strong motor seizures 51. Recently, Cain et al. 5 demonstrated greater suppression of kindling development following joint administration of intra-amygdaloid APV and systemic scopolamine (a cholinergic antagonist), relative to APV treatment alone. Kindling rates following MK-801 in the present study were similar to those reported by Cain et al. 5 following co-administration of APV and scopolamine. Moreover, direct stimulation 32 or enhanced GABAergic activity in the substantia nigra retards the development46 and blocks the expression 3° of amygdala kindled seizures. It is possible, therefore, that MK-801 may suppress kindling development by blockade of excitatory neurotransmission at the focal amygdala site in addition to a disruption of the balance between inhibitory (e.g. GABA) and excitatory (e.g. NMDA, ACh) systems within other brain regions (e.g. the substantia nigra). MK-801 effectively reduced the stage of motor seizure and duration of clonic seizure activity induced by threshold stimulation of kindled amygdala and hippocampal foci. AD duration was significantly shorter in the hippocampus, but not the amygdala

kindled group. Similar profiles of anticonvulsant efficacy in amygdala kindled animals were evident with most of the reference compounds included in the analysis with the following exceptions: phenytoin was without effect on amygdaloid kindled seizures. This finding is in keeping with previous reports in the kindling literature (see ref. 52), but contrasts with that of Albright and Burnham I in kindled rats and Clineschmidt et al. 7 in acute seizure models in mice. In keeping with previous findings, all other drugs tested, phenobarbital, diazepam l, phencyclidine, and ketamine 23'45, reduced seizure stage, duration of clonic seizure activity, and AD duration. Thresholds in the amygdala were elevated by drugs with sedative or anesthetic properties (i.e. phenobarbital, diazepam, ketamine). In contrast, MK801 and phencyclidine induced similar excitatory behavioral syndromes and reduced seizure severity without altering threshold for evocation of AD. Phencyclidine has been reported to raise thresholds in amygdala kindled rats 15. The discrepancy between this report and the present findings may be attributable to procedural differences in detecting threshold shifts. Freeman et al. 15employed an ascending series of stimulus intensities, beginning at an intensity well below threshold (20/~A) and increasing until an AD was evoked (mean of 96/~A in the high dosage group to comprise a mean of more than 9 stimulations). Since massed short-interval stimulation can result in temporary AD threshold increases 3s, the absence of threshold shift with phencyclidine in the present analysis may reflect the suprathreshold intensity chosen for the initial stimulation. MK-801 did appear to increase AD threshold for hippocampally evoked seizures, but this effect failed to reach statistically significant levels. The effects of the other drugs on the hippocampal focus were not tested. Thus, MK-801 has potent anticonvulsant properties. It demonstrates higher efficacy in protecting against seizures with a hippocampal focus (seizure stage and AD duration) than those originating in the amygdala. Hippocampal kindling in vitro results in an enhancement in the sensitivity to iontophoretically administered NMDA concomitant with a reduction in the amplitude of intracellularly recorded inhibitory postsynaptic potentials and sensitivity to iontophoretically applied GABA 47'4s. Enhanced excitability and reduced inhibition may combine to pro-

98 duce the epileptiform bursting characteristic of kindling. F u r t h e r m o r e , Mody and H e i n e m a n n 31 have

ACKNOWLEDGEMENTS

recently demonstrated the participation of N M D A dentate granule cells of kindled but not control animals. Therefore, the ability of MK-801 to block exci-

The MK-801 was a generous gift from Dr. Bradley Clineschmidt of Merck, Sharp and Dohme, Inc. The author would like to acknowledge Dr. Robert Dyer for his contribution to and instigation of the study,

tation and eliminate the enhanced response to N M D A may underlie its capacity to retard kindling

and the technical assistance of Cina Mack, Shawn Acheson and Susan Lucas. Special thanks to Dr. Jim

receptors in low-frequency synaptic transmission in

development and to suppress kindled seizures. Dif-

O'Callaghan and Dr. Linda Burdette for comments

ferential effects of MK-801 on A D duration, threshold, and degree of seizure suppression in amygdala and hippocampus may reflect: (1) NMDAI°'54; and/

on earlier versions of this manuscript. The research reported here was supported by a contract from the

or (2) glycine receptor densities in these forebrain structures, since glycine has been shown to potentiate responses to N M D A 2° and to enhance MK-801 binding43; or perhaps, (3) as suggested above, differential nigral influence on seizures emanating from these two limbic sites.

U . S . E . P . A . The research described in this article has been reviewed by the Health Effects Research Laboratory, U.S. E n v i r o n m e n t a l Protection Agency and approved for publication. Approval does not signify that the contents necessarily reflect the views and policies of the Agency nor does m e n t i o n of trade names or commercial products constitute endorsement or r e c o m m e n d a t i o n for use.

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