The effects of selective glutamate receptor antagonists on synchronized firing bursts in layer III of rat visual cortex.

In the rat visual cortex in vitro, single-shock stimulations applied to the border between layer VI and the white matter evoke synchronized burst-firing by units in layer III. We have examined the effects of glutamate receptor antagonists on this activity, with antagonists applied via the bath to allow correlation of effects with concentrations. All synaptically driven components (recorded extracellularly as field potential 'S2' spikes, dipoles 'W1' and 'W2', and coinciding single-unit spikes) were inhibited by greater than 90% in 1.0 mM kynurenic acid and in 3 or 10 microM 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, which selectively blocks AMPA/kainate receptors). S2 spike amplitudes were reduced by half in 0.7 microM CNQX. 2-Amino-5-phosphonovalerate (APV), a specific blocker of NMDA receptors, did not prevent S2 spike burst or horizontal spread of bursting within layer III. However, APV reduced the duration of synchronized bursts and the slower potentials which followed. In Mg(2+)-free medium, new components appeared which were APV-sensitive: (1) low amplitude spikes, distributed spatially like S2 spike, but recurring more slowly, and (2) slow potentials, distributed spatially like W1 and W2 potentials, but lasting for hundreds of milliseconds. The amplitudes of these spikes were reduced by half in 3 microM D-APV. Our data imply that: (1) glutamate receptors play a major role in mediating local, excitatory neurotransmission in the supragranular layers of neocortex, with NMDA and AMPA/kainate subtypes each subserving somewhat different functions; (2) AMPA/kainate receptors mediate rapid excitatory transmission between layer III neurons, responsible for driving the first 15 ms of synchronized bursts; (3) currents gated by NMDA receptors determine the duration of coherent firing bursts, and drive asynchronous neuronal firing following bursts; and (4) under conditions which circumvent block by extracellular Mg2+, activation of NMDA receptors greatly enhances and prolongs the response to single-shock stimulations. In vivo, activation of layer III neurons is likely to depend significantly upon currents gated by NMDA receptors whenever repetitively firing excitatory inputs summed over several tens of milliseconds provide enough depolarization to lift block by extracellular Mg2+.     

Epileptiform bursts elicited in CA3 hippocampal neurons by a variety of convulsants are not blocked by N-methyl-d-aspartate antagonists

Intracellular and extracellular recordings from CA₃ hippocampal neurons in vitro were used to study the ability of several NMDA (N-methyl-d-aspartate) receptor antagonists to suppress epileptiform bursts induced by NMDA and convulsants not thought to act at NMDA receptors. The antagonists, APV (d-2-amino-5-phosphonovalerate), AP-7 (d,l-2-amino-7-phosphonoheptanoate) and CPP (d,l-3-[(±)-2-car☐ypiperazin-4-yl-]-propyl-1-phosphonic acid), blocked the spontaneous and evoked bursts induced by NMDA. CPP, but not APV or AP-7, prevented the development of bursts induced by Mg-free medium. The NMDA antagonists failed to block bursting induced by kainate, 7 mM K⁺, mast cell degranulating peptide, anoxia or spontaneous bursting. In some cases the NMDA antagonists induced spontaneous bursts or enhanced burst frequency, a proconvulsant effect. It is concluded that activation of NMDA receptors is sufficient but not necessary for burst generation in the CA₃ region.     

An experimental study of the effect of isoflurane on epileptiform bursts.

The effect of isoflurane on penicillin- and picrotoxin-induced epileptiform activity was tested using hippocampal slice preparations. Isoflurane reduced both the frequency of spontaneous epileptiform bursts and the number of population spikes within each burst in a dose-dependent manner. The last population spikes in the burst were most sensitive to the anesthetic, whereas the first 4-6 spikes were quite resistant and persisted until spontaneous activity was abolished at 3% isoflurane. Isoflurane increased the stimulus current required to evoke epileptiform bursts and shifted the relationship between stimulus current and population spike amplitude to the right. At 3% isoflurane, a dose that usually causes iso-electric EEG and abolishes all spontaneous epileptiform activity, responses could still be evoked, and then invariably had an epileptiform pattern. The maximum response was reduced compared to control and 1.5% isoflurane. With isoflurane there was a reduced tendency for activity to be transmitted from one region within the hippocampus to the other. This effect was also dose-dependent. However, transmitted activity always retained a typical epileptiform character, although the number of population spikes within a train to some extent decreased with increasing concentrations of isoflurane.     

The NMDA receptor-mediated components of responses evoked by patterned stimulation are not increased by long-term potentiation

The participation of N-methyl-D-aspartate (NMDA) receptors in synaptic transmission before and after induction of long-term potentiation (LTP) was studied in field CA1 of hippocampal slices. NMDA receptor-mediated postsynaptic responses were determined by comparing responses recorded in the presence and absence of the selective antagonist, D-2-amino-5-phosphonopentanoate (D-AP5, 50 microM). In the presence of physiological magnesium concentrations (1 mM), robust D-AP5-sensitive responses could be evoked by high frequency bursts (4 pulses, 100 Hz) when burst stimulation was preceded 200 ms earlier by 'priming' stimulation (2 pulses, 15 ms apart) of a separate input. Induction of LTP resulted in a substantial potentiation (35%) of non-NMDA-mediated responses to primed bursts but not of NMDA-mediated responses. These results suggest that long-term postsynaptic modifications are at least partly responsible for the expression of LTP.     

Antagonism of the aconitine-induced inexcitability by the structurally related Aconitum alkaloids, lappaconitine and ajacine.

Aconitine, lappaconitine and ajacine are structurally related alkaloids occurring in several species of the Aconitum genus. While aconitine is known to activate the voltage-dependent sodium channel, lappaconitine has been reported to block this channel. To investigate a possible antagonism of the aconitine action on neuronal activity by lappaconitine and the closely related alkaloid ajacine, we have performed extracellular recordings of stimulus evoked population spikes and field excitatory postsynaptic potential (EPSP) in rat hippocampal slices. Aconitine (10-100 nM) diminished the amplitude of the orthodromic population spike in a concentration-dependent manner. When aconitine was applied in presence of 10 microM lappaconitine, the concentration-response curve was shifted to the right. Furthermore, the complete suppression of the population spike evoked by 100 nM aconitine was reversed by 10 microM lappaconitine. The action of lappaconitine was mimicked by ajacine, however, the latter alkaloid was less potent. Both lappaconitine and ajacine shifted the input-output relationship of the presynaptic fiber spike as function of the stimulation intensity and of the field EPSP as function of the presynaptic fiber spike to the right. After pharmacological isolation, the presynaptic fiber spike was decreased by both compounds in a frequency-dependent manner indicative for a use-dependent action. Thus, electrophysiologically these alkaloids seem to inhibit predominantly the excitability of the afferent fibres and, in consequence, neurotransmission between Schaffer collaterals and the CA1 neurons, thereby suppressing the firing of the latter. Ajacine and lappaconitine inhibited stimulus-triggered epileptiform population bursts in area CA1 elicited by omission of Mg(2+) as well as spontaneously occurring epileptiform discharges in area CA3 elicited by omission of Mg(2+) and elevation of K(+). It is concluded that the inhibitory and antiepileptiform effect of ajacine and lappaconitine is mediated by a frequency-dependent inhibition of the voltage-dependent sodium channel, thereby decreasing the excitability which might be important for filtering high frequency bursts of action potentials characteristic for epileptiform activity in the hippocampus. Moreover, these alkaloids are naturally occurring antagonists of the sodium channel activator aconitine.     

Effects of micromolar and nanomolar calcium concentrations on non-synaptic bursting in the hippocampal slice

To determine how [Ca²⁺]₀ affects non-synaptic epileptogenesis in the CA1 area of hippocampal slices, we compared the extracellularly recorded hyperactivity induced by ACSF containing either micromolar (‘low’-Ca²⁺, LC-ACSF) or nanomolar concentrations of Ca²⁺ (‘zero’-Ca²⁺, ZC-ACSF). Both solutions effectively blocked chemical synaptic transmission but spontaneous bursts developed more quickly and consistently in ZC-ACSF and were longer in duration and more frequent than those recorded in LC-ACSF. Antidromically evoked bursts were less epileptiform, i.e., they exhibited fewer population spikes (PSs), in ZC-ACSF. Increasing [Mg²⁺]₀ or decreasing [K⁺]₀ suppressed spontaneous LC-ACSF bursting but only decreased the intensity and frequency of bursting in ZC-ACSF. Either manipulation increased the epileptiform nature of the antidromically evoked field potential, thereby mimicking the effect of increasing [Ca²⁺]₀ from nanomolar to micromolar levels. Bath application of 250–500 μM GABA commonly arrested spontaneous bursting in LC-ACSF. In ZC-ACSF, GABA decreased the burst frequency but paradoxically superimposed high amplitude PSs on each burst. These effects were reversed by the GABA_A receptor antagonists bicuculline methiodide or picrotoxin (50–100 μM). These results indicate that simply lowering [Ca²⁺]₀ from micromolar to nanomolar concentrations increases the burst propensity and intensity of the CA1 population and can dramatically alter responses to pharmacological agents.     

The epileptiform activity induced by 4-aminopyridine in rat amygdala slices: antagonism by non-N-methyl-D-aspartate receptor antagonists

The effects of excitatory amino acid receptor antagonists kynuretic acid and 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) on epileptiform activity induced by 4-aminopyridine (4-AP) were studied in rat amygdala slices using intracellular recording techniques. Five to 10 min after switching to 4-AP-containing solution, spontaneous epileptiform bursts were observed in 35 out of 45 slices studied. The spontaneous epileptiform events consisted of an initial burst followed by a number of afterdischarges. Superfusion with kynuretic acid, a broad-spectrum excitatory amino acid receptor antagonist, reversibly reduced the duration of the spontaneous bursting discharges in a dose-dependent manner. The frequency of spontaneous bursting was also decreased. The IC50, estimated from the graph of the concentration-response relationship, was approximately 130 microM. In addition, CNQX which is a specific non-N-methyl-D-aspartate (NMDA) receptor antagonist blocked the spontaneous and evoked epileptiform bursting. In 11 out of 15 neurons tested, there was a residual depolarizing component remained. This depolarizing component was reversibly blocked by specific NMDA receptor antagonist, D,L-2-amino-5-phosphonovaleate (D,L-APV). Relative to the CNQX-sensitive component, the D,L-APV-sensitive component is much smaller in amplitude and shorter in duration indicating that NMDA receptor plays only a minor role in this process. These data suggest that the generation or propagation of spontaneous epileptiform events induced by 4-AP in the neurons of basolateral amygdala is mediated by excitatory amino acids and that activation of non-NMDA receptors is of primary importance.     

Dependence of calcium influx in neocortical cells on temporal structure of depolarization, number of spikes, and blockade of NMDA receptors

Increase of intracellular [Ca(2+)] evoked by action potentials in a cell can induce long-term synaptic plasticity even without concomitant presynaptic stimulation. We used optical recording of the fluorescence of a Ca(2+)-indicator Oregon Green to investigate whether differences in results obtained with modifications of that purely postsynaptic induction protocol could be due to differential Ca(2+) influx. We compared changes of the somatic [Ca(2+)] in layer II-III pyramidal cells in slices of rat visual cortex evoked by bursts of depolarization pulses and long depolarizing steps. During weak depolarizations, the Ca(2+) influx was proportional to the amplitude and duration of the depolarization. With suprathreshold depolarizations, the Ca(2+) influx was proportional to the number of action potentials. Because the burst depolarizations evoked more spikes than did the long duration steps, this burst protocol led to a larger Ca(2+) influx. With all stimulation protocols, the spike-induced Ca(2+) influx was reduced during blockade of N-methyl-D-aspartate (NMDA) receptors. Differences in intracellular [Ca(2+)] increases thus may be one reason for differential effects of purely postsynaptic challenges on synaptic transmission.     

NMDA receptor activation during epileptiform responses in the dentate gyrus of epileptic patients.

We previously showed that a low frequency (1 Hz) train of perforant path stimulation evokes burst discharges in the dentate gyrus of hippocampal slices obtained from patients surgically treated for intractable temporal lobe epilepsy. We report here that multiple population spikes that characterize the burst discharge are blocked reversibly by the specific NMDA receptor antagonist, D-(-)-2-amino-5-phosphonovaleric acid (D-APV). The epileptiform discharge evoked in human dentate gyrus by stimulation trains of 1 Hz could be reproduced in the rat dentate gyrus in vitro by the same stimulation protocol but required the presence of low concentrations (0.2-0.6 mM) of extracellular magnesium. We suggest that low frequency orthodromic stimulation of dentate granule cells through the perforant path progressively evokes an increase in the activation of NMDA receptors resulting in burst discharges in tissue from epileptic patients.     

The epileptiform activity induced by 4-aminopyridine in rat amygdala slices: antagonism by non-N-methyl-d-aspartate receptor anatagonists

The effects of excitatory amino acid receptor antagonists kynuretic acid and 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) on epileptiform activity induced by 4-aminopyridine (4-AP) were studied in rat amygdala slices using intracellular recording techniques. Five to 10 min after switching to 4-AP-containing solution, spontaneous epileptiform bursts were observed in 35 out of 45 slices studied. The spontaneous epileptiform events consisted of an initial burst followed by a number of afterdischarges. Superfusion with kynuretic acid, a broad-spectrum excitatory amino acid receptor antagonist, reversibly reduced the duration of the spontaneous bursting discharges in a dose-dependent manner. The frequency of spontaneous bursting was also decreased. The IC₅₀, estimated from the graph of the concentration-response relationship, was approximately 130 μM. In addition, CNQX which is a specificnon-N-methyl-d-aspartate (NMDA) receptor antagonist blocked the spontaneous and evoked epileptiform bursting. In 11 out of 15 neurons tested, there was a residual depolarizing component remained. This depolarizing component was reversibly blocked by specific NMDA receptor antagonist,d,l-2-amino-5-phosphonovaleate (d,l-APV). Relative to the CNQX-sensitive component, thed,l-APV-sensitive component is much smaller in amplitude and shorter in duration indicating that NMDA receptor plays only a minor role in this process. These data suggest that the generation of propagation of spontaneous epileptiform events induced by 4-AP in the neurons of basolateral amygdala is mediated by excitatory amino acids and that activation of non-NMDA receptors is of primary importance.     

The contribution of AMPA and NMDA receptors to graded bursting activity in the hippocampal CA1 region in an acute in vitro model of epilepsy.

The AMPA/KA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) and the NMDA receptor antagonist 2-amino-5-phosphonovalerate (D-APV) were used to investigate the contribution of excitatory amino acid (EAA) receptors to graded bursting activity recorded in the CA1 region of the rat hippocampal slice following bath application of the convulsant drug bicuculline methiodide (BIC, 2-3 microM). CNQX (5-9 microM) significantly antagonised the burst in a reversible, concentration-dependent manner (n = 5). The effect involved a reduction in the amplitude but not the number of population spikes of the burst and also a depression of the underlying burst excitatory post-synaptic potential (EPSP). D-APV (5-25 microM), in contrast, reduced the amplitude and number of spikes in the burst but had no effect on the burst EPSP (n = 5). Following a single concentration of CNQX (5 microM), applied in the presence of bicuculline, it was observed that the components of epileptiform response which remained could be completely abolished with D-APV (10 microM; n = 10). It was also shown that, following elimination of synaptic transmission with CNQX (5 microM), application of bicuculline (2-3 microM) induced a small burst that could be reversibly antagonised with D-APV (10 microM). These results show that evoked epileptiform activity witnessed in the presence of bicuculline involves the activation of both AMPA and NMDA receptors, the AMPA receptor activation making the major contribution. The burst mediated by NMDA receptors is not dependent on prior activation of AMPA receptors.     

Excitatory synaptic potentials in kainic acid-denervated rat CA1 pyramidal neurons.

Intracellular recordings were performed in the CA1 region of the rat hippocampus following an ipsilateral intraventricular injection of kainic acid. Seven days postlesion, graded bursts of up to four action potentials could be evoked by stimulation of the stratum radiatum. The evoked EPSPs underlying these bursts showed a prolonged 10-90% rise time and half-width compared to control EPSPs, an absence of a significant inhibitory phase, and an increase in magnitude and duration at depolarized resting levels. The evoked EPSPs also exhibited a significant decrease in amplitude and time course in response to D-APV (D-2-amino-5-phosphonovaleric acid; 1-20 microM), though this effect was variable from cell to cell. The prolonged time course, voltage sensitivity, and response to a selective NMDA antagonist confirmed that the major component of the EPSP in neurons from lesioned slices was mediated by NMDA receptors. The partial denervation of the CA1 area induced by the kainic acid led to both an enhanced NMDA-mediated excitatory phase and a decrease in postsynaptic inhibition, resulting in the pronounced hyperexcitability noted in the lesioned slices.     

Antagonism of the aconitine-induced inexcitability by the structurally related Aconitum alkaloids, lappaconitine and ajacine

Aconitine, lappaconitine and ajacine are structurally related alkaloids occurring in several species of the Aconitum genus. While aconitine is known to activate the voltage-dependent sodium channel, lappaconitine has been reported to block this channel. To investigate a possible antagonism of the aconitine action on neuronal activity by lappaconitine and the closely related alkaloid ajacine, we have performed extracellular recordings of stimulus evoked population spikes and field excitatory postsynaptic potential (EPSP) in rat hippocampal slices. Aconitine (10–100 nM) diminished the amplitude of the orthodromic population spike in a concentration-dependent manner. When aconitine was applied in presence of 10 μM lappaconitine, the concentration–response curve was shifted to the right. Furthermore, the complete suppression of the population spike evoked by 100 nM aconitine was reversed by 10 μM lappaconitine. The action of lappaconitine was mimicked by ajacine, however, the latter alkaloid was less potent. Both lappaconitine and ajacine shifted the input–output relationship of the presynaptic fiber spike as function of the stimulation intensity and of the field EPSP as function of the presynaptic fiber spike to the right. After pharmacological isolation, the presynaptic fiber spike was decreased by both compounds in a frequency-dependent manner indicative for a use-dependent action. Thus, electrophysiologically these alkaloids seem to inhibit predominantly the excitability of the afferent fibres and, in consequence, neurotransmission between Schaffer collaterals and the CA1 neurons, thereby suppressing the firing of the latter. Ajacine and lappaconitine inhibited stimulus-triggered epileptiform population bursts in area CA1 elicited by omission of Mg²⁺ as well as spontaneously occurring epileptiform discharges in area CA3 elicited by omission of Mg²⁺ and elevation of K⁺. It is concluded that the inhibitory and antiepileptiform effect of ajacine and lappaconitine is mediated by a frequency-dependent inhibition of the voltage-dependent sodium channel, thereby decreasing the excitability which might be important for filtering high frequency bursts of action potentials characteristic for epileptiform activity in the hippocampus. Moreover, these alkaloids are naturally occurring antagonists of the sodium channel activator aconitine.     

Do N-methyl-D-aspartate receptors mediate synaptic responses in the mudpuppy retina?

Whole-cell recordings of amacrine and ganglion cells in the superfused retina-eyecup preparation of the mudpuppy were obtained in order to determine which excitatory amino acid receptor (EAAR) subtype mediates the synaptic responses of these neurons. All third-order retinal neurons tested were depolarized by kainic acid (KA, N-methyl-D-aspartate (NMDA), and quisqualate (QQ). The responses evoked by NMDA were blocked by the addition of D-2-amino-5-phosphonovaleric acid (D-AP5) and D-2-amino-7 phosphonoheptonoic acid (D-AP7) to the perfusate. When the actions of exogenously applied NMDA were completely blocked by D-AP5 and D-AP7, the light-evoked responses of inner retinal neurons persisted without any apparent reduction or, alternatively, a slight enhancement of the response was observed. Light-evoked responses of bipolar, amacrine, and ganglion cells associated with the On pathway were attenuated by L-AP5 in a manner similar to its lower-order homolog L-2-amino-4-phosphonobutyrate (AP4); nevertheless, L-AP5 was not an effective NMDA antagonist. Although synaptic transmission between retinal second- and third-order neurons appears to be mediated by EAARs, the NMDA receptor does not appear to play a prominent role under our experimental conditions. Nevertheless, our results suggest that the racemic mixture of AP5 should not be used as an NMDA antagonist in retinal research, due to the AP4-like actions of its L-enantiomer.     

Long-lasting potentiation and epileptiform activity produced by GABAB receptor activation in the dentate gyrus of rat hippocampal slice

Bath application of the GABAB receptor agonist baclofen produced a concentration-dependent long-lasting potentiation (LLP) of the evoked population spike in the dentate gyrus of rat hippocampal slices. A high concentration of baclofen (5 microM) also produced a loss of inhibition that was manifested as the appearance of epileptiform, multiple evoked population spikes and a decrease in paired-pulse inhibition. Both baclofen-induced potentiation and epileptiform activity could be blocked or significantly reduced in slices from pertussis toxin-treated animals (1 microgram, intradentate) or in slices pretreated with the NMDA receptor antagonist D-(-)-2-amino-5-phosphonovaleric acid (10 microM). At a concentration that had no significant effect on individual evoked responses (0.1 microM) but still produced a loss in paired-pulse inhibition, baclofen facilitated the induction of beta-adrenergic receptor-mediated LLP. LLP was induced in the dentate gyrus by bath application of 1 microM, but not 0.1 microM, isoproterenol. Coapplication of baclofen and isoproterenol, both at a concentration (0.1 microM) that individually had no effect on the population spike, produced a synergistic LLP of the population spike. We propose that baclofen produces a selective disinhibitory effect in the granule cell layer of the dentate gyrus by inhibiting the activity of GABAergic interneurons. At a low concentration, the subtle loss of inhibition can facilitate the induction of isoproterenol-induced LLP. At a high concentration, baclofen can produce an LLP that is probably induced by a loss of inhibition.     

Primed-burst potentiation occludes the potentiation phenomenon and enhances the epileptiform activity induced by transient pentylenetetrazol in the CA1 region of rat hippocampal slices

The effects of pentylenetetrazol (PTZ) following induction of long-term potentiation (LTP) on population spikes in CA1 of hippocampal slices were investigated. Population spikes were evoked by activation of Schaffer collaterals with a range of stimulation intensities. LTP was induced using θ-pattern primed burst tetanic stimulation. Changes in the population spike amplitude and number of population spikes were used as indices to quantify the effects of PTZ exposure in the control (non-tetanized) and LTP (tetanized) conditions. The amplitude of population spike was measured 20 min before, during 20 min chemical application (3 mM), and also after 30 or 60 min washout period. In non-tetanized slices, the population spike input-output curve was significantly increased 20 min after PTZ application and persisted at least for 60 min. Multiple population spikes or after potentials also appeared, but did not persist. When PTZ was applied on tetanized slices, 60 min after LTP induction, the amplitude increase produced by PTZ was smaller than the increase seen in the control condition. Also LTP induction preceding PTZ exposure increased the number of population spikes evoked by stimulation of Schaffer collaterals. It is concluded that a transient PTZ application produces a long-lasting increase in population spike amplitude. Primed burst LTP occludes PTZ-induced potentiation while also increasing the epileptogenic effect of PTZ.     

Spontaneous EEG spikes in the normal hippocampus. II. Relations to synchronous burst discharges

Spontaneous EEG spikes (SPKs) were recorded from the CA1 region of the dorsal hippocampus in normal rats during awake immobility and slow wave sleep. These SPKs were accompanied by synchronous burst discharges in the pyramidal cell layer. These discharges are called 'population bursts (PBs)' in that they seem to require a population of synchronously bursting neurons. PBs were classified into 2 forms on the basis of their morphologies. One form (mixed burst or MB) consisted of a mixture or superimposition of action potential bursts from a relatively small number of neurons. The other form (ripple) was a series of 3-13 (typically 5-8) high frequency (125-250 Hz) waves, usually waxing and waning. Unit action potentials were superimposed mainly on negative portions of these high frequency waves. The ripple was considered to represent summed activity of highly synchronized complex spike bursts from a relatively large number of pyramidal cells. The similarity in wave structure between these non-pathological ripples and multipeaked, epileptiform (interictal) field potentials recorded from the penicillin-treated hippocampus suggests that they may share some common underlying mechanisms.     

N-methyl-D-aspartate induces recurrent synchronized burst activity in immature hippocampal CA3 neurones in vitro

Slices of hippocampus prepared from rats aged 1-10 days have been used to examine the chemosensitivity of CA3 pyramidal neurones to N-methyl-D-aspartate (NMDA). Superfusion of NMDA excited all neurones tested at all ages including the first day postnatal. In the majority of neurones this excitation was associated with the induction of a period of burst firing which disappeared on removal of NMDA. These bursts took the form of paroxysmal depolarizing shifts (PDSs) with a large amplitude depolarization and a high frequency discharge of spikes. The amplitude but not the frequency of occurrence of the PDSs was influenced by changes in the membrane potential and they could be abolished by either a high divalent cation medium or tetrodotoxin. Their occurrence was synchronous with an extracellularly recorded discharge. The NMDA induced excitation and the induction of the PDSs was attenuated by selective NMDA receptor antagonists D-aminophosphonovalerate (10-50 microM) and D,L-aminophosphonoheptanoate (20-30 microM). The results indicate that chemosensitivity to NMDA develops prenatally and that activation of NMDA receptors can in immature CA3 pyramidals induce recurrent synchronized burst activity.     

Low magnesium epileptogenesis in the rat hippocampal slice: electrophysiological and pharmacological features

Extra- and intracellular recording techniques were used to study the epileptiform activity generated by rat hippocampal slices perfused with Mg2(+)-free artificial cerebrospinal fluid (ACSF). This procedure induced in both CA1 and CA3 subfields the appearance of synchronous, spontaneously occurring epileptiform discharges which consisted of extracellularly recorded 100-800 ms long, positive shifts with superimposed negative going population spikes. Simultaneous, extracellular recordings from CA1 and CA3 subfields revealed that the epileptiform discharges in CA3 preceded those occurring in CA1 by 5-25 ms. Surgical separation of the two areas led to the disappearance of spontaneous events in the CA1 but not in the CA3 subfield. In this type of experiment CA1 pyramidal cells still generated epileptiform discharges following orthodromic stimuli. The intracellular counterpart of both spontaneous and stimulus-induced epileptiform discharges in CA1 and CA3 pyramidal cells was a large amplitude depolarization with high frequency discharge of action potentials which closely resembled the paroxysmal depolarizing shift recorded in the experimental epileptogenic focus. A hyperpolarizing potential triggered by alvear stimuli was recorded in CA1 cells perfused with Mg2(+)-free ACSF. This hyperpolarization was blocked by bicuculline methiodide (BMI) indicating that it represented a GABAergic inhibitory postsynaptic potential (IPSP). BMI also caused a prolongation of both spontaneous and stimulus-induced Mg(+)-free epileptiform discharges. Perfusion of the slices with the N-methyl-D-aspartate (NMDA) receptor antagonist DL-2-amino-5-phosphono-valerate (APV) reduced and eventually abolished the Mg(+)-free epileptiform discharges. These effects were more pronounced in the CA1 than in the CA3 subfield. APV also reduced the amplitude and the duration of the alveus-induced IPSP. These data demonstrate that Mg(+)-free epileptiform activity is present in the hippocampal slice at a time when inhibitory GABAergic potentials are operant as well as that in the CA1 subfield this type of epileptiform activity is dependent upon NMDA-activated conductances. Our experiments also indicate that NMDA receptors might be involved in the neuronal circuit responsible for the hyperpolarizing IPSP generated by CA1 pyramidal neurons.