Spontaneous Epileptiform Bursts Research Articles

Early intervention with levetiracetam prevents the development of cortical hyperexcitability and spontaneous epileptiform activity in two models of neurotrauma in rats

This study examined the antiepileptogenic potential of the antiseizure drug (ASD) levetiracetam (LEV) using the in vitro traumatized-slice and in vivo controlled cortical impact (CCI) models of traumatic brain injury (TBI) in rats when administered early after the injury. For the in vitro model, acute coronal slices (400–450 μm) of rat neocortex (P21–32) were injured via a surgical cut that separated the superficial layers from the deeper regions. Persistent stimulus-evoked epileptiform activity developed within 1–2 h after trauma. In randomly selected slices, LEV (500 μM) was bath-applied for 1 h starting immediately or delayed by 30–80 min after injury. Treated and untreated slices were examined for epileptiform activity via intracellular and extracellular recordings. For the in vivo model, rats (P24–32) were subjected to a non-penetrating, focal, CCI injury targeting the neocortex (5.0 mm diameter; 2.0 mm depth). Immediately after injury, rats were given either a single dose of LEV (60–150 mg/kg, i.p.) or the saline vehicle. At 2–3 weeks after the injury, ex vivo cortical slices were examined for epileptiform activity. The results from the traumatized-slice experiments showed that in vitro treatment with LEV within 60 min of injury significantly reduced (> 50%) the proportion of slices that exhibited stimulus-evoked epileptiform activity. LEV treatment also increased the stimulus intensity required to trigger epileptiform bursts in injured slices by 2–4 fold. Consistent with these findings, LEV treatment of CCI-injured rats (n = 15) significantly reduced the proportion of animals that exhibited spontaneous and stimulus-evoked epileptiform bursts in ex vivo cortical slices compared to saline-treated controls (n = 15 rats), and also significantly increased the stimulus intensity required to evoke epileptiform bursts. These results suggest that early administration of LEV has the potential to prevent or reduce posttraumatic epileptogenesis and that there may be a narrow therapeutic window for successful prophylactic intervention.

Glycolytic inhibition by 2-deoxy-d-glucose abolishes both neuronal and network bursts in an in vitro seizure model

Neuronal activity is energy demanding and coupled to cellular metabolism. In this study, we investigated the effects of glycolytic inhibition with 2-deoxy-d-glucose (2-DG) on basal membrane properties, spontaneous neuronal firing, and epileptiform network bursts in hippocampal slices. The effect of glycolytic inhibition on basal membrane properties was examined in hippocampal CA1 neurons, which are not ordinarily active spontaneously. Intracellular application of 2-DG did not significantly alter the membrane input resistance, action-potential threshold, firing pattern, or input-output relationship of these neurons compared with simultaneously recorded neighboring neurons without intracellular 2-DG. The effect of glycolytic inhibition on neuronal firing was tested in spontaneously active hippocampal neurons (CA3) when synaptic transmission was left intact or blocked with AMPA, NMDA, and GABAA receptor antagonists (DNQX, APV, and bicuculline, respectively). Under both conditions (synaptic activity intact or blocked), bath application of 2-DG (2 mM) blocked spontaneous firing in ~2/3 (67 and 71%, respectively) of CA3 pyramidal neurons. In contrast, neuronal firing of CA3 neurons persisted when 2-DG was applied intracellularly, suggesting that glycolytic inhibition of individual neurons is not sufficient to stop neuronal firing. The effects of 2-DG on epileptiform network bursts in area CA3 were tested in Mg2+-free medium containing 50 µM 4-aminopyridine. Bath application of 2-DG abolished these epileptiform bursts in a dose-dependent and all-or-none manner. Taken together, these data suggest that altered glucose metabolism profoundly affects cellular and network hyperexcitability and that glycolytic inhibition by 2-DG can effectively abrogate epileptiform activity.NEW & NOTEWORTHY Neuronal activity is highly energy demanding and coupled to cellular metabolism. In this study, we demonstrate that glycolytic inhibition with 2-deoxy-d-glucose (2-DG) effectively suppresses spontaneous neuronal firing and epileptiform bursts in hippocampal slices. These data suggest that an altered metabolic state can profoundly affect cellular and network excitability, and that the glycolytic inhibitor 2-DG may hold promise as a novel treatment of drug-resistant epilepsy.

Aberrant excitatory rewiring of layer V pyramidal neurons early after neocortical trauma

Lesioned neuronal circuits form new functional connections after a traumatic brain injury (TBI). In humans and animal models, aberrant excitatory connections that form after TBI may contribute to the pathogenesis of post-traumatic epilepsy. Partial neocortical isolation (“undercut” or “UC”) leads to altered neuronal circuitry and network hyperexcitability recorded in vivo and in brain slices from chronically lesioned neocortex. Recent data suggest a critical period for maladaptive excitatory circuit formation within the first 3days post UC injury (Graber and Prince 1999, 2004; Li et al. 2011, 2012b). The present study focuses on alterations in excitatory connectivity within this critical period. Immunoreactivity (IR) for growth-associated protein (GAP)-43 was increased in the UC cortex 3days after injury. Some GAP-43-expressing excitatory terminals targeted the somata of layer V pyramidal (Pyr) neurons, a domain usually innervated predominantly by inhibitory terminals. Immunocytochemical analysis of pre- and postsynaptic markers showed that putative excitatory synapses were present on somata of these neurons in UC neocortex. Excitatory postsynaptic currents from UC layer V Pyr cells displayed properties consistent with perisomatic inputs and also reflected an increase in the number of synaptic contacts. Laser scanning photostimulation (LSPS) experiments demonstrated reorganized excitatory connectivity after injury within the UC. Concurrent with these changes, spontaneous epileptiform bursts developed in UC slices. Results suggest that aberrant reorganization of excitatory connectivity contributes to early neocortical hyperexcitability in this model. The findings are relevant for understanding the pathophysiology of neocortical post-traumatic epileptogenesis and are important in terms of the timing of potential prophylactic treatments.

Critical role of canonical transient receptor potential channel 7 in initiation of seizures.

Status epilepticus (SE) is a life-threatening disease that has been recognized since antiquity but still causes over 50,000 deaths annually in the United States. The prevailing view on the pathophysiology of SE is that it is sustained by a loss of normal inhibitory mechanisms of neuronal activity. However, the early process leading to the initiation of SE is not well understood. Here, we show that, as seen in electroencephalograms, SE induced by the muscarinic agonist pilocarpine in mice is preceded by a specific increase in the gamma wave, and genetic ablation of canonical transient receptor potential channel (TRPC) 7 significantly reduces this pilocarpine-induced increase of gamma wave activity, preventing the occurrence of SE. At the cellular level, TRPC7 plays a critical role in the generation of spontaneous epileptiform burst firing in cornu ammonis (CA) 3 pyramidal neurons in brain slices. At the synaptic level, TRPC7 plays a significant role in the long-term potentiation at the CA3 recurrent collateral synapses and Schaffer collateral-CA1 synapses, but not at the mossy fiber-CA3 synapses. Taken together, our data suggest that epileptiform burst firing generated in the CA3 region by activity-dependent enhancement of recurrent collateral synapses may be an early event in the initiation process of SE and that TRPC7 plays a critical role in this cellular event. Our findings reveal that TRPC7 is intimately involved in the initiation of seizures both in vitro and in vivo. To our knowledge, this contribution to initiation of seizures is the first identified functional role for the TRPC7 ion channel.

Organic and inorganic calcium antagonists inhibit veratridine-induced epileptiform activity in CA3 neurons of the guinea pig

Veratridine is believed to cause epileptiform discharges via its effects on sodium channels. We addressed the question whether calcium currents, known to contribute to the generation of paroxysmal depolarization shifts (PDS) in most models of epilepsies, also contribute to veratridine-induced epileptiform activity. Therefore, we recorded from CA3 neurons (n=50) of veratridine-treated hippocampal slices and analyzed the effects of two calcium antagonists. Veratridine (0.5-1.0 microM) elicited spontaneous epileptiform bursts, paroxysmal depolarization shifts (PDS) lasting 100-300 ms, and depolarizations (LD) lasting up to several minutes. Most often PDS directly preceded LD which resulted in typical composite depolarizations termed veratridine-induced complexes (VC). VC persisted even in the presence of CNQX and APV (25 micromol/l, both), or in nominally calcium-free saline, revealing the non-synaptic nature of these potentials. Cobalt (1-2mM) abolished VC within minutes, but allowed LD type-like potentials to be elicited by depolarizing current pulses. Verapamil (50 microM) also diminished or abolished amplitudes of VC. All inhibitory effects of cobalt and verapamil were at least partly reversible. Due to the effects of both calcium antagonists we conclude that veratridine-induced epileptiform activity depends not only on sodium, but also on calcium currents.

Dentate Filter Function Is Altered in a Proepileptic Fashion during Aging

The elderly have an increased incidence and prevalence for seizure disorders. Further, since up to 50% of these cases have no identifiable antecedent, it has been hypothesized that aging of the central nervous system itself may be epileptogenic. Aged rats, compared to adults, exhibit a greater susceptibility to and severity of seizures associated with hippocampal activation. Whether this aging-related change reflects proconvulsive changes in limbic circuitry is unknown and thus was the focus of this study. Hippocampal slices from adult and aged Fischer 344 rats were examined using electrophysiological techniques. The dentate gyrus was our model region since it is involved with both wet-dog shakes and limbic seizures, and it is affected preferentially with age. No differences were noted between groups in field potential activity elicited with low frequency stimulation. In contrast, 5-Hz molecular layer stimulation could evoke multiple population spikes in approximately 40% of aged versus 0% of adult slices. Further, recording in CA3 revealed that this stimulation paradigm could elicit multiple spikes in aged, but not adult, slices that frequently evolved into spontaneous epileptiform bursts. This change in the capacity of the dentate to respond to and filter afferent input was associated with an aging-related decrease in the frequency of spontaneous IPSPs and an increased propensity for large amplitude prolonged EPSPs following disinhibition. These epileptogenic changes in dentate function and circuitry could contribute to the exacerbated susceptibility for hippocampal seizures in aged rodents, as well as the aging-related decline in spatial learning and memory.

α2AAdrenergic Receptor Activation Inhibits Epileptiform Activity in the Rat Hippocampal CA3 Region

Norepinephrine has potent antiepileptic properties, the pharmacology of which is unclear. Under conditions in which GABAergic inhibition is blocked, norepinephrine reduces hippocampal cornu ammonis 3 (CA3) epileptiform activity through alpha(2) adrenergic receptor (AR) activation on pyramidal cells. In this study, we investigated which alpha(2)AR subtype(s) mediates this effect. First, alpha(2)AR genomic expression patterns of 25 rat CA3 pyramidal cells were determined using real-time single-cell reverse transcription-polymerase chain reaction, demonstrating that 12 cells expressed alpha(2A)AR transcript; 3 of the 12 cells additionally expressed mRNA for alpha(2C)AR subtype and no cells possessing alpha(2B)AR mRNA. Hippocampal CA3 epileptiform activity was then examined using field potential recordings in brain slices. The selective alphaAR agonist 6-fluoronorepinephrine caused a reduction of CA3 epileptiform activity, as measured by decreased frequency of spontaneous epileptiform bursts. In the presence of betaAR blockade, concentration-response curves for AR agonists suggest that an alpha(2)AR mediates this response, as the rank order of potency was 5-bromo-N-(4,5-dihydro-1H-imidazol-2-yl)-6-quinoxalinamine (UK-14304) >or= epinephrine >6-fluoronorepinephrine > norepinephrine >>> phenylephrine. Finally, equilibrium dissociation constants (K(b)) of selective alphaAR antagonists were functionally determined to confirm the specific alpha(2)AR subtype inhibiting CA3 epileptiform activity. Apparent K(b) values calculated for atipamezole (1.7 nM), MK-912 (4.8 nM), BRL-44408 (15 nM), yohimbine (63 nM), ARC-239 (540 nM), prazosin (4900 nM), and terazosin (5000 nM) correlated best with affinities previously determined for the alpha(2A)AR subtype (r = 0.99, slope = 1.0). These results suggest that, under conditions of impaired GABAergic inhibition, activation of alpha(2A)ARs is primarily responsible for the antiepileptic actions of norepinephrine in the rat hippocampal CA3 region.

Repeated administration of citalopram and imipramine alters the responsiveness of rat hippocampal circuitry to the activation of 5-HT 7 receptors

The effects of a selective serotonin reuptake inhibitor, citalopram, and a tricyclic antidepressant drug, imipramine, administered repetitively for 14 days, were investigated ex vivo in rat hippocampal slices. Spontaneous epileptiform bursts were recorded from the CA3 area in nominally Mg 2+-free incubation conditions. 5-carboxamidotryptamine (5-CT) dose-dependently increased bursting frequency in the presence of N-[2-[4-(2-methoxyphenyl)-1 piperazinyl]ethyl]- N-2-pyridinylcyclohexanecarboxamide (WAY 100635). This effect could be dose-dependently blocked by (2 R)-1-[(3-Hydroxyphenyl)sulfonyl]-2-[2-(4-methyl-1-piperidinyl)ethyl]pyrrolidine hydrochloride (SB 269970), thus implicating the involvement of 5-HT 7 receptors. Repeated treatment with citalopram or imipramine resulted in an attenuation of the excitatory effects of the activation of hippocampal 5-HT 7 receptor.

Modulation of epileptiform burst frequency by the metabotropic glutamate receptor subtype mGluR3

Spontaneous epileptiform burst activity occurs in acute hippocampal slice dentate granule cells perfused with 10 mM potassium and 0.5 mM calcium [J. Neurophys. 68 (1992) 2016]. We report that activation of the group II metabotropic glutamate receptor subtype 3 (mGluR3) induces an increase in spontaneous burst duration, whereas inhibition of mGluR3 reversibly reduces spontaneous burst frequency. Neither activation, nor inhibition, of group II mGluR had any effects on spontaneous negative dc shifts, or the number of spikes per burst, as compared to control. We conclude that mGluR3 can modulate high potassium, low calcium-induced spontaneous epileptiform burst activity in acute rat hippocampal slice dentate granule cells.

Determination of group I metabotropic glutamate receptor subtypes involved in the frequency of epileptiform activity in vitro using mGluR1 and mGluR5 mutant mice

In mouse hippocampal slices, bicuculline elicited spontaneous epileptiform bursts with a duration of 200–300 ms and with a frequency of five to six events per minute. Application of group I metabotropic glutamate receptor agonist (RS)-3,5-dihydroxyphenylglycine ((RS)-DHPG) increased the burst frequency up to 300% at concentrations of 50 to 100 μM, while it decreased the burst duration below 100 ms. In slices of subtype I mGluR1 or subtype I mGluR5 knockout mice, bicuculline elicited spontaneous epileptiform bursts with similar duration and frequency as those measured in wild-type mice but without the previous effects seen following application of DHPG at concentrations up to 100 μM. Likewise, in slices of wild-type mice, preincubation with mGluR1 antagonist, 1-aminoindan-1,5-dicarboxylic acid (AIDA) or mGluR5 receptor antagonist, 2-methyl-6-(phenylethynyl)-pyridine (MPEP) blocked in both cases completely the increase in frequency following DHPG application. These findings suggest an interactive mechanism between mGluR1 and mGluR5 receptors in the modulation of epileptiform bursting activity by DHPG that could indicate a common intracellular signaling mechanism or possibly direct interaction between these two receptors.

Long-term change in synaptic transmission in CA3 circuits followed by spontaneous rhythmic activity in rat hippocampal slices

The relevance of long-term potentiation (LTP) at excitatory synapses in CA3 circuits to generation of spontaneous epileptiform bursts in CA3 was investigated using rat hippocampal slices. CA3 pyramidal cells were antidromically stimulated through Schaffer collaterals. Evoked field potentials were extracellularly recorded from the stratum pyramidale and the stratum radiatum in CA3. Therefore, field potentials reflecting recurrent excitatory post-synaptic potentials (EPSPs) and inhibitory post-synaptic potentials (IPSPs) were positive at the stratum pyramidale and negative at the stratum radiatum. First, we tested how the amplitude of the evoked field potentials depends on a gamma-aminobutyric acid (GABA(A)) antagonist. Both of the positive and negative field potential peaks reduced in the medium containing penicillin (2 mM) or bicuculline (20 microM). This suggests that unmasked EPSPs due to suppression of IPSPs do not result in an increase in the evoked potentials. Second, CA3 pyramidal cells were antidromically stimulated by tetanic stimulation of Schaffer collaterals in order to induce LTP at synapses in CA3 circuits. Both of the positive and negative field potentials increased, suggesting that recurrent EPSPs were enhanced by tetanic stimulation. Induction of LTP at recurrent excitatory synapses was followed by spontaneous epileptiform bursts which persisted throughout experiments (approximately 1.5 h), while LTP of afferent synaptic potential evoked by hilar test stimulation was not induced. These results suggest that LTP at the afferent synapses is not necessary to spontaneous epileptiform bursts in CA3, but LTP at excitatory synapses between CA3 pyramidal cells contribute to spontaneous epileptiform bursts.

Trimethylolpropane Phosphate Induces Epileptiform Discharges in the CA1 Region of the Rat Hippocampus

The actions of trimethylolpropane phosphate (TMPP), an ethyl bicyclophosphate convulsant produced during the partial pyrolysis of some phosphate ester-based lubricants, were tested on CA1 neurons of rat hippocampal slices using intracellular recording techniques. Bath application of TMPP (0.1–100 μM) induced spontaneous paroxysmal depolarizing shifts and the associated spontaneous epileptiform bursts followed by after-hyperpolarizations in 63% of neurons tested. The TMPP-induced epileptiform bursts were blocked by muscimol, a γ-aminobutyric acid A (GABAA) receptor agonist, diazepam (DZP), a GABAA-benzodiazepine ionophore complex agonist, or baclofen, a GABAB receptor agonist. While bath application of muscimol, DZP, or baclofen suppressed spontaneous activity in CA1 neurons not previously exposed to TMPP, subsequent application of TMPP (10 μM) reversed the actions of muscimol and diazepam, but not baclofen. TMPP (0.1–100 μM) also induced membrane hyperpolarization associated with an increase in peak input resistance and inward rectification in 33% of neurons tested or membrane depolarization associated with an increase in input resistance in 17% of neurons tested. In summary, TMPP induced epileptiform activities in hippocampal CA1 neurons. The epileptogenic effects of TMPP are consistent with its interaction with GABAA–benzodiazepine receptors.

Ca 2+ entry through l-type Ca 2+ channels helps terminate epileptiform activity by activation of a Ca 2+ dependent afterhyperpolarisation in hippocampal CA3.

In CA3 neurons of disinhibited hippocampal slice cultures the slow afterhyperpolarisation, following spontaneous epileptiform burst events, was confirmed to be Ca 2+ dependent and mediated by K + ions. Apamin, a selective blocker of the SK channels responsible for part of the slow afterhyperpolarisation reduced, but did not abolish, the amplitude of the post-burst afterhyperpolarisation. The result was an increased excitability of individual CA3 cells and the whole CA3 network, as measured by burst duration and burst frequency. Increases in excitability could also be achieved by strongly buffering intracellular Ca 2+ or by minimising Ca 2+ influx into the cell, specifically through l-type (but not N-type) voltage operated Ca 2+ channels. Notably the l-type Ca 2+ channel antagonist, nifedipine, was more effective than apamin at reducing the post-burst afterhyperpolarisation. Nifedipine also caused a greater increase in network excitability as determined from measurements of burst duration and frequency from whole cell and extracellular recordings. N-methyl d-aspartate receptor activation contributed to the depolarisations associated with the epileptiform activity but Ca 2+ entry via this route did not contribute to the activation of the post-burst afterhyperpolarisation. We suggest that Ca 2+ entry through l-type channels during an epileptiform event is selectively coupled to both apamin-sensitive and -insensitive Ca 2+ activated K + channels. Our findings have implications for how the route of Ca 2+ entry and subsequent Ca 2+ dynamics can influence network excitability during epileptiform discharges.

1PA131 CA3錐体細胞相間の興奮性シナプスのLTPと自発性のてんかん様バースト

1PA131 CA3錐体細胞相間の興奮性シナプスのLTPと自発性のてんかん様バースト

The anticonvulsant actions of sigma receptor ligands in the Mg2+-free model of epileptiform activity in rat hippocampal slices.

1. The anticonvulsant potency of a series of structurally-dissimilar compounds which possess nanomolar affinities for high-affinity sigma binding sites was examined in the Mg2+-free model of epileptiform activity in rat hippocampal slices. Extracellular field potential recordings in the CA1 region were employed to examine the effects of test compounds on spontaneous epileptiform activity and multiple population spikes evoked by stimulation of the Schaffer collateral-commissural pathway. 2. Applied at sigma site-selective (i.e. nanomolar) concentrations, dextromethorphan, ditolylguanidine, caramiphen and opipramol failed to modify Mg2+-free epileptiform activity; neither pro- nor anticonvulsant effects were observed. However, applied at micromolar concentrations, these and additional test compounds reversibly inhibited orthodromically-evoked epileptiform field potentials with a rank order potency (IC50 values in microM): dextrorphan (1.5) > ifenprodil (6.3) > dextromethorphan (10) > ditolylguanidine (15) > loperamide (28) > carbetapentane (38) > caramiphen (46) > opipramol (52). Micromolar concentrations of the same compounds also inhibited spontaneous epileptiform bursts recorded during perfusion with Mg2+-free medium. 3. Co-application of ropizine (10 microM), an allosteric modulator of dextromethorphan binding to high-affinity sigma receptors, failed to endow dextromethorphan 10 nM with anticonvulsant properties and did not modify the anticonvulsant potency of 10 microM dextromethorphan. 4. The effects of dextrorphan (10 microM), ifenprodil (20 microM), loperamide (50 microM) and caramiphen (100 microM) were examined in the presence of external Mg2+ on field potential input/output (I/O) relationships and paired-pulse facilitation (PPF) of field excitatory postsynaptic potentials. Only caramiphen elicited effects on these parameters, affecting synaptic transmission at the point of synaptic transfer and depressing PPF ratios to below baseline values. The effects of caramiphen on I/O relationships mimicked those of the established anticonvulsant adenosine: in contrast, adenosine evoked an increase in PPF ratios. 5. Because anticonvulsant activity was observed only at micromolar concentrations of the sigma ligands tested, the results indicate that their anticonvulsant actions should not be ascribed to their occupancy, observed at nanomolar concentrations, of high-affinity sigma binding sites. Rather, anticonvulsant activity more likely reflects functional NMDA receptor antagonism and/or blockade of high voltage-activated Ca2+ channels, effects which are associated with micromolar concentrations of the test compounds. Modulation of GABAergic inhibitory mechanisms may also contribute to the anticonvulsant properties of caramiphen.

Spontaneous epileptiform bursts and long-term potentiation in rat CA3 hippocampal slices induced by chaotic stimulation of mossy fibers

The relation between long-term potentiation (LTP) and spontaneous rhythm in CA3 was investigated using rat hippocampal slices. Field potential response of CA3 to mossy fiber stimulation consisted of a mono-synaptic positive potential and subsequent poly-synaptic negative potentials. LTP of both field potentials was induced by chaotic mossy fiber stimulation. Although CA3 did not show any spontaneous rhythm before LTP induction in a normal perfusing medium, CA3 spontaneously caused epileptiform bursts after LTP induction by chaotic mossy fiber stimulation. The amplitude of those epileptiform bursts and the inter-burst interval were not uniform. After LTP induction, the cross-correlation function of spontaneous field potentials simultaneously recorded at two sites approximately 300 μm apart in CA3 showed a large central peak. This indicates that neuronal activity at two sites is synchronized. These results suggest that epileptiform bursts in CA3 are caused by synchronization of spontaneous CA3 pyramidal cell activity due to LTP induced by chaotic burst stimulation.

Enhanced propagation of epileptiform activity through the kindled dentate gyrus.

Extracellular recordings were performed in combined hippocampal-entorhinal cortex (HC-EC) slices obtained from control and commissural kindled rats to investigate the propagation of epileptiform activity from the entorhinal cortex (EC) to the hippocampus (HC) after chronic epilepsy. Lowering extracellular Mg2+ concentration in control slices induced epileptiform activity consisting of spontaneous epileptiform bursts in area CA3 and of electrographic seizures in the EC. In contrast, the CA3 region of HC-EC slices obtained from kindled rats displayed significantly longer lasting epileptiform bursts and electrographic seizures. The electrographic seizures that were absent in controls propagated from the EC because disconnecting the HC from the EC stopped their occurrence in the CA3, whereas epileptiform bursts persisted with an unaltered pattern and frequency. Thus the area CA3 is affected by kindling and contributes to the spread of epileptiform activity within the EC-HC complex. We developed a method to induce focal epileptiform activity in the EC by locally perfusing the gamma-aminobutyric acid-A (GABA) antagonist bicuculline (50 mM) in 10 mM KCl containing artificial cerebrospinal fluid. This method enabled us to investigate the propagation of epileptiform discharges from the disinhibited EC to the DG without affecting the DG with the epileptogenic medium. We show here that kindling facilitates the propagation of epileptiform activity through the DG. These data are consistent with the normal function of the DG as a filter limiting the spread of epileptiform activity within the HC-EC complex. This gating mechanism breaks down after chronic epilepsy induced by kindling.

Role of group I metabotropic glutamate receptors in the patterning of epileptiform activities in vitro.

In guinea pig hippocampal slices, picrotoxin elicited spontaneous epileptiform bursts 300-550 ms in duration. Additional application of (R,S)-3,5-dihydroxyphenylglycine or (S)-3-hydroxyphenylglycine, agonists specific for group I metabotropic glutamate receptors (mGluRs), or (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid, a broad-spectrum mGluR agonist, converted picrotoxin-induced interictal bursts into prolonged discharges measured on the order of seconds. The prolonged discharges induced by selective group I mGluR agonist continued to be produced for hours after agonist removal. The antagonists (S)-4-carboxyphenylglycine and (+)-alpha-methyl-4-carboxyphenylglycine had no effect on the duration of picrotoxin-induced interictal bursts. However, after agonist exposure, the persistent prolonged discharges occurring in the absence of agonist were reversibly suppressed by the antagonists, suggesting that the activity is maintained via endogenous activation of group I mGluRs by synaptically released glutamate. Our results suggest that, under some conditions, activation of group I mGluRs produces long-lasting enhancement of synaptic responses, mediated at least in part by autopotentiation of the group I mGluR response itself, which may result in the production of seizure discharges and contribute to epileptogenesis.

Electrophysiological approach of the antiepileptic effect of dexamethasome on hippocampal slice culture usihg a multirecording system: The physiocard®

The in vitro antiepileptic activity of the synthetic glucocorticoid dexamethasone (DEX) was tested in rat hippocampal organotypic cultures on the field potential epileptiform activity induced by picrotoxin (PTX). Spontaneous as well as evoked electrophysiological activities have been studied through the extracellular multirecording Physiocard® system. PTX typically elicited seizure-like discharges (epileptiform bursts) in the hippocampus neurons. Those epileptiform bursts can be divided in two groups, one rhythmic which lasted 43 ± 24s (mean ± sd) at a frequency of 4.6 ± 1.9Hz and the other arhythmic composed of population spikes, which occurred during 14.3 ± 6.9min. In the presence of DEX at different concentrations, results obtained were: 1) DEX 1μM decreased the occurrence of the two different groups of spontaneous epileptiform bursts, most of the time to zero. 2) DEX 50μM prevented totally the occurrence of epileptiform bursts. 3) DEX 50μM contrarily to DEX 1μM avoided the decrease of evoked field potentials' amplitude induced by PTX 3μM on all simultaneous recorded points. Those results suggest that synthetic glucocorticoid DEX presents an acute antiepileptic effect in a dose dependent manner on the hippocampus tissue.

Role of metabotropic glutamate receptor subtypes in the patterning of epileptiform activities in vitro.

1. Epileptiform activities were elicited from the in vitro guinea pig hippocampus by the addition of picrotoxin. Modification of the picrotoxin-induced activities by agents active at metabotropic glutamate receptors (mGluRs) was examined using intracellular and extracellular recordings. 2. Picrotoxin typically elicited synchronized discharges (epileptiform bursts) in CA3 neurons. These spontaneously occurred at regular intervals. In the presence of (+)-alpha-methyl-4-carboxyphenylglycine (MCPG; 700-1,000 microM), an antagonist at multiple mGluR subtypes, the frequency of spontaneous epileptiform bursts decreased. In contrast, when the mGluR agonists (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid (ACPD; 5 microM) or (2S,3S,4S)-alpha-(carboxycyclopropyl)-glycine (L-CCG-I; 10 microM) were added to the incubating medium, the frequency of epileptiform bursts increased. No consistent change in membrane potential, burst duration, nor burst afterhyperpolarization was associated with the changes in burst frequency. 3. When spontaneous burst frequency was reduced in MCPG, stimulation at a higher frequency entrained bursts without failure. Bursts evoked in MCPG were similar in waveform and amplitude to those evoked in the control state. 4. (S)-4-carboxyphenylglycine (S-4CPG) and (R,S)-4-carboxy-3-hydroxyphenylglycine (RS-4C3HPG) are antagonists at mGluR subtypes 1 and 5 but agonists at mGluRs 2 and 3. Addition of either of these agents increased the frequency of epileptiform bursts. 5. These results suggest that sufficient glutamate is released during epileptiform activities to activate mGluRs. The overall effect is to increase the frequency of synchronized discharges. This modulatory action on burst frequency is probably mediated via the mGluR 2 and 3 receptor subclass.