Epileptiform synchronization and high-frequency oscillations in brain slices comprising piriform and entorhinal cortices.

We employed field potential recordings in extended in vitro brain slices form Sprague-Dawley rats containing the piriform and entorhinal cortices (PC and EC, respectively) to identify the characteristics of epileptiform discharges and concomitant high-frequency oscillations (HFOs, ripples: 80-200Hz, fast ripples: 250-500Hz) during bath application of 4-aminopyridine (4AP, 50µM). Ictal-like discharges occurred in PC and EC either synchronously or independently of each other; synchronous ictal discharges always emerged from a synchronous "fast" interictal background whereas asynchronous ictal discharges were preceded by a "slow" interictal event. In addition, asynchronous ictal discharges had longer duration and interval of occurrence than synchronous ictal discharges, and contained a higher proportion of ripples and fast ripples. Cutting the connections between PC and EC made synchronicity disappear and increased ictal discharges duration in the EC but failed in changing HFO occurrence in both areas. Finally, antagonizing ionotropic glutamatergic receptors abolished ictal activity in all experiments, increased the duration and rate of occurrence of interictal discharges occurring in PC-EC interconnected slices while it did not influence the slow asynchronous interictal discharges in both areas. Our results identify some novel in vitro interactions between olfactory (PC) and limbic (EC) structures that presumably contribute to in vivo ictogenesis as well.

Mechanisms of epileptiform synchronization in cortical neuronal networks.

Neuronal synchronization supports different physiological states such as cognitive functions and sleep, and it is mirrored by identifiable EEG patterns ranging from gamma to delta oscillations. However, excessive neuronal synchronization is often the hallmark of epileptic activity in both generalized and partial epileptic disorders. Here, I will review the synchronizing mechanisms involved in generating epileptiform activity in the limbic system, which is closely involved in the pathophysiogenesis of temporal lobe epilepsy (TLE). TLE is often associated to a typical pattern of brain damage known as mesial temporal sclerosis, and it is one of the most refractory adult form of partial epilepsy. This epileptic disorder can be reproduced in animals by topical or systemic injection of pilocarpine or kainic acid, or by repetitive electrical stimulation; these procedures induce an initial status epilepticus and cause 1-4 weeks later a chronic condition of recurrent limbic seizures. Remarkably, a similar, seizure-free, latent period can be identified in TLE patients who suffered an initial insult in childhood and develop partial seizures in adolescence or early adulthood. Specifically, I will focus here on the neuronal mechanisms underlying three abnormal types of neuronal synchronization seen in both TLE patients and animal models mimicking this disorder: (i) interictal spikes; (ii) high frequency oscillations (80-500 Hz); and (iii) ictal (i.e., seizure) discharges. In addition, I will discuss the relationship between interictal spikes and ictal activity as well as recent evidence suggesting that specific seizure onsets in the pilocarpine model of TLE are characterized by distinctive patterns of spiking (also termed preictal) and high frequency oscillations.

Neurosteroids modulate epileptiform activity and associated high-frequency oscillations in the piriform cortex.

Allotetrahydrodeoxycorticosterone (THDOC) belongs to a class of pregnane neurosteroidal compounds that enhance brain inhibition by interacting directly with GABAA signaling, mainly through an increase in tonic inhibitory current. Here, we addressed the role of THDOC in the modulation of interictal- and ictal-like activity and associated high-frequency oscillations (HFOs, 80-500 Hz; ripples: 80-200 Hz, fast ripples: 250-500 Hz) recorded in vitro in the rat piriform cortex, a highly excitable brain structure that is implicated in seizure generation and maintenance. We found that THDOC: (i) increased the duration of interictal discharges in the anterior piriform cortex while decreasing ictal discharge duration in both anterior and posterior piriform cortices; (ii) reduced the occurrence of HFOs associated to both interictal and ictal discharges; and (iii) prolonged the duration of 4-aminopyridine-induced, glutamatergic independent synchronous field potentials that are known to mainly result from the activation of GABAA receptors. Our results indicate that THDOC can modulate epileptiform synchronization in the piriform cortex presumably by potentiating GABAA receptor-mediated signaling. This evidence supports the view that neurosteroids regulate neuronal excitability and thus control the occurrence of seizures.

In vitro ictogenesis and parahippocampal networks in a rodent model of temporal lobe epilepsy.

Temporal lobe epilepsy (TLE) is a chronic epileptic disorder involving the hippocampal formation. Details on the interactions between the hippocampus proper and parahippocampal networks during ictogenesis remain, however, unclear. In addition, recent findings have shown that epileptic limbic networks maintained in vitro are paradoxically less responsive than non-epileptic control (NEC) tissue to application of the convulsant drug 4-aminopyridine (4AP). Field potential recordings allowed us to establish here the effects of 4AP in brain slices obtained from NEC and pilocarpine-treated epileptic rats; these slices included the hippocampus and parahippocampal areas such as entorhinal and perirhinal cortices and the amygdala. First, we found that both types of tissue generate epileptiform discharges with similar electrographic characteristics. Further investigation showed that generation of robust ictal-like discharges in the epileptic rat tissue is (i) favored by decreased hippocampal output (ii) reinforced by EC-subiculum interactions and (iii) predominantly driven by amygdala networks. We propose that a functional switch to alternative synaptic routes may promote network hyperexcitability in the epileptic limbic system.

Laser-evoked potentials as a tool for assessing the efficacy of antinociceptive drugs.

Laser-evoked potentials (LEPs) are brain responses to laser radiant heat pulses and reflect the activation of Adelta nociceptors. LEPs are to date the reference standard technique for studying nociceptive pathway function in patients with neuropathic pain. To find out whether LEPs also provide a useful neurophysiological tool for assessing antinociceptive drug efficacy, in this double-blind placebo-controlled study we measured changes induced by the analgesic tramadol on LEPs in 12 healthy subjects. We found that tramadol decreased the amplitude of LEPs, whereas placebo left LEPs unchanged. The opioid antagonist naloxone partially reversed the tramadol-induced LEP amplitude decrease. We conclude that LEPs may be reliably used in clinical practice and research for assessing the efficacy of antinociceptive drugs.

Zn(2+) slows down Ca(V)3.3 gating kinetics: implications for thalamocortical activity.

We employed whole cell patch-clamp recordings to establish the effect of Zn(2+) on the gating the brain specific, T-type channel isoform Ca(V)3.3 expressed in HEK-293 cells. Zn(2+) (300 microM) modified the gating kinetics of this channel without influencing its steady-state properties. When inward Ca(2+) currents were elicited by step depolarizations at voltages above the threshold for channel opening, current inactivation was significantly slowed down while current activation was moderately affected. In addition, Zn(2+) slowed down channel deactivation but channel recovery from inactivation was only modestly changed. Zn(2+) also decreased whole cell Ca(2+) permeability to 45% of control values. In the presence of Zn(2+), Ca(2+) currents evoked by mock action potentials were more persistent than in its absence. Furthermore, computer simulation of action potential generation in thalamic reticular cells performed to model the gating effect of Zn(2+) on T-type channels (while leaving the kinetic parameters of voltage-gated Na(+) and K(+) unchanged) revealed that Zn(2+) increased the frequency and the duration of burst firing, which is known to depend on T-type channel activity. In line with this finding, we discovered that chelation of endogenous Zn(2+) decreased the frequency of occurrence of ictal-like epileptiform discharges in rat thalamocortical slices perfused with medium containing the convulsant 4-aminopyridine (50 microM). These data demonstrate that Zn(2+) modulates Ca(V)3.3 channel gating thus leading to increased neuronal excitability. We also propose that endogenous Zn(2+) may have a role in controlling thalamocortical oscillations.

Antiepileptic drugs and muscarinic receptor-dependent excitation in the rat subiculum.

Field and intracellular recordings were made in an in vitro slice preparation to establish whether the antiepileptic drugs topiramate and lamotrigine modulate cholinergic excitation in the rat subiculum. Bath application of carbachol (CCh, 70-100microM) induced: (i) spontaneous and synchronous field oscillations (duration=up to 7s) that were mirrored by intracellular depolarizations with rhythmic action potential bursts; and (ii) depolarizing plateau potentials (DPPs, duration=up to 2.5s) associated with action potential discharge in response to brief (50-100ms) intracellular depolarizing current pulses. Ionotropic glutamatergic receptor antagonists abolished the field oscillations without influencing DPPs, while atropine (1microM) markedly reduced both types of activity. Topiramate (10-100microM, n=8-13 slices) or lamotrigine (50-400microM, n=3-12) decreased in a dose-dependent manner, and eventually abolished, CCh-induced field oscillations. During topiramate application, these effects were accompanied by marked DPP reduction. When these antiepileptic drugs were tested on DPPs recorded in the presence of CCh+ionotropic glutamatergic and GABA receptor antagonists, only topiramate reduced DPPs (n=5-19/dose; IC(50)=18microM, n=48). Similar effects were induced by topiramate during metabotropic glutamate receptor antagonism (n=5), which did not influence DPPs. Thus, topiramate and lamotrigine reduce CCh-induced epileptiform synchronization in the rat subiculum but only topiramate is effective in controlling DPPs. We propose that muscarinic receptor-mediated excitation represents a target for the action of some antiepileptic drugs such as topiramate.

Reduced GABAB receptor subunit expression and paired-pulse depression in a genetic model of absence seizures.

Neocortical networks play a major role in the genesis of generalized spike-and-wave (SW) discharges associated with absence seizures in humans and in animal models, including genetically predisposed WAG/Rij rats. Here, we tested the hypothesis that alterations in GABA(B) receptors contribute to neocortical hyperexcitability in these animals. By using Real-Time PCR we found that mRNA levels for most GABA(B(1)) subunits are diminished in epileptic WAG/Rij neocortex as compared with age-matched non-epileptic controls (NEC), whereas GABA(B(2)) mRNA is unchanged. Next, we investigated the cellular distribution of GABA(B(1)) and GABA(B(2)) subunits by confocal microscopy and discovered that GABA(B(1)) subunits fail to localize in the distal dendrites of WAG/Rij neocortical pyramidal cells. Intracellular recordings from neocortical cells in an in vitro slice preparation demonstrated reduced paired-pulse depression of pharmacologically isolated excitatory and inhibitory responses in epileptic WAG/Rij rats as compared with NECs; moreover, paired-pulse depression in NEC slices was diminished by a GABA(B) receptor antagonist to a greater extent than in WAG/Rij rats further suggesting GABA(B) receptor dysfunction. In conclusion, our data identify changes in GABA(B) receptor subunit expression and distribution along with decreased paired-pulse depression in epileptic WAG/Rij rat neocortex. We propose that these alterations may contribute to neocortical hyperexcitability and thus to SW generation in absence epilepsy.

Effects of gap junction blockers on human neocortical synchronization.

Field potentials and intracellular recordings were obtained from human neocortical slices to study the role of gap junctions (GJ) in neuronal network synchronization. First, we examined the effects of GJ blockers (i.e., carbenoxolone, octanol, quinine, and quinidine) on the spontaneous synchronous events (duration = 0.2-1.1 s; intervals of occurrence = 3-27 s) generated by neocortical slices obtained from temporal lobe epileptic patients during application of 4-aminopyridine (4AP, 50 muM) and glutamatergic receptor antagonists. The synchronicity of these potentials (recorded at distances up to 5 mm) was decreased by GJ blockers within 20 min of application, while prolonged GJ blockers treatment at higher doses made them disappear with different time courses. Second, we found that slices from patients with focal cortical dysplasia (FCD) could generate in normal medium spontaneous synchronous discharges (duration = 0.4-8 s; intervals of occurrence = 0.5-90 s) that were (i) abolished by NMDA receptor antagonists and (ii) slowed down by carbenoxolone. Finally, octanol or carbenoxolone blocked 4AP-induced ictal-like discharges (duration = up to 35 s) in FCD slices. These data indicate that GJ play a role in synchronizing human neocortical networks and may implement epileptiform activity in FCD.

Synaptic hyperexcitability of deep layer neocortical cells in a genetic model of absence seizures.

We used sharp-electrode, intracellular recordings in an in vitro brain slice preparation to study the excitability of neocortical neurons located in the deep layers (>900 microm from the pia) of epileptic (180-210-days old) Wistar Albino Glaxo/Rijswijk (WAG/Rij) and age-matched, non-epileptic control (NEC) rats. Wistar Albino Glaxo/Rijswijk rats represent a genetic model of absence seizures associated with generalized spike and wave (SW) discharges in vivo. When filled with neurobiotin, these neurons had a typical pyramidal shape with extensive apical and basal dendritic trees; moreover, WAG/Rij and NEC cells had similar fundamental electrophysiological and repetitive firing properties. Sequences of excitatory postsynaptic potentials (EPSPs) and hyperpolarizing inhibitory postsynaptic potentials (IPSPs) were induced in both the strains by electrical stimuli delivered to the underlying white matter or within the neocortex; however, in 24 of 55 regularly firing WAG/Rij cells but only in 2 of 25 NEC neurons, we identified a late EPSP that (1) led to action potential discharge and (2) was abolished by the N-methyl-D-aspartate (NMDA) receptor antagonist 3,3-(2-carboxypiperazine-4-yl)-propyl-1-phosphonate (20 microM; n = 8/8 WAG/Rij cells). Finally, we found that the fast and slow components of the stimulus-induced IPSPs recorded during the application of glutamatergic receptor antagonists had similar reversal potentials in the two strains, while the peak conductance of the fast IPSP was significantly reduced in WAG/Rij cells. These findings document an increase in synaptic excitability that is mediated by NMDA receptors, in epileptic WAG/Rij rat neurons located in neocortical deep layers. We propose that this mechanism may be instrumental for initiating and maintaining generalized SW discharges in vivo.

Repetitive low-frequency stimulation reduces epileptiform synchronization in limbic neuronal networks.

Deep-brain electrical or transcranial magnetic stimulation may represent a therapeutic tool for controlling seizures in patients presenting with epileptic disorders resistant to antiepileptic drugs. In keeping with this clinical evidence, we have reported that repetitive electrical stimuli delivered at approximately 1 Hz in mouse hippocampus-entorhinal cortex (EC) slices depress the EC ability to generate ictal activity induced by the application of 4-aminopyridine (4AP) or Mg(2+)-free medium (Barbarosie, M., Avoli, M., 1997. CA3-driven hippocampal-entorhinal loop controls rather than sustains in vitro limbic seizures. J. Neurosci. 17, 9308-9314.). Here, we confirmed a similar control mechanism in rat brain slices analyzed with field potential recordings during 4AP (50 microM) treatment. In addition, we used intrinsic optical signal (IOS) recordings to quantify the intensity and spatial characteristics of this inhibitory influence. IOSs reflect the changes in light transmittance throughout the entire extent of the slice, and are thus reliable markers of limbic network epileptiform synchronization. First, we found that in the presence of 4AP, the IOS increases, induced by a train of electrical stimuli (10 Hz for 1 s) or by recurrent, single-shock stimulation delivered at 0.05 Hz in the deep EC layers, are reduced in intensity and area size by low-frequency (1 Hz), repetitive stimulation of the subiculum; these effects were observed in all limbic areas contained in the slice. Second, by testing the effects induced by repetitive subicular stimulation at 0.2-10 Hz, we identified maximal efficacy when repetitive stimuli are delivered at 1 Hz. Finally, we discovered that similar, but slightly less pronounced, inhibitory effects occur when repetitive stimuli at 1 Hz are delivered in the EC, suggesting that the reduction of IOSs seen during repetitive stimulation is pathway dependent as well as activity dependent. Thus, the activation of limbic networks at low frequency reduces the intensity and spatial extent of the IOS changes that accompany ictal synchronization in an in vitro slice preparation. This conclusion supports the view that repetitive stimulation may represent a potential therapeutic tool for controlling seizures in patients with pharmaco-resistant epileptic disorders.

Periodic oscillatory activity in parahippocampal slices maintained in vitro.

Brain slices maintained in vitro have been extensively used for studying neuronal synchronization. However, the validity of this approach may be questioned since pharmacological procedures are usually required to elicit spontaneous events similar to the EEG activity recorded in vivo. Here, we report that when superfused with control medium, rat brain slices comprising the entorhinal and perirhinal cortices along with a portion of the basolateral/lateral nuclei of the amygdala can synchronously generate periodic oscillatory activity at 5-11 Hz every 5-30 s. The periodic events: (i) correspond intracellularly to synaptic depolarizations in regularly firing neurons analyzed in the three areas; (ii) have no fixed site of onset; (iii) spread with time lags of 8-20 ms; and (iv) continue to occur asynchronously after their surgical isolation. NMDA receptor antagonism reduced the duration of the oscillatory events, while glutamatergic non-NMDA receptor antagonism abolished them. Activation of mu-opioid receptors, a procedure that hyperpolarizes interneurons thus decreasing GABA release, reversibly decreased the rate of occurrence of periodic oscillatory activity (POA). However, periodic events continued to occur during application of GABA(A) or GABA(B) receptor antagonists as well as in the presence of the cholinergic agent carbachol. We also found that POA was abolished by baclofen and irreversibly reduced by the gap junction decoupler carbenoxolone. These findings demonstrate that parahippocampal networks in a brain slice preparation can generate periodic, synchronous activity under quasi-physiological conditions. These network oscillations (i) reflect the activation of ionotropic glutamatergic and GABAergic receptors, (ii) are contributed by gap-junction interactions, and (iii) are controlled by GABA(B) receptors that are presumably located presynaptically.

GABAA receptor-dependent synchronization leads to ictogenesis in the human dysplastic cortex.

Patients with Taylor's type focal cortical dysplasia (FCD) present with seizures that are often medically intractable. Here, we attempted to identify the cellular and pharmacological mechanisms responsible for this epileptogenic state by using field potential and K+-selective recordings in neocortical slices obtained from epileptic patients with FCD and, for purposes of comparison, with mesial temporal lobe epilepsy (MTLE), an epileptic disorder that, at least in the neocortex, is not characterized by any obvious structural aberration of neuronal networks. Spontaneous epileptiform activity was induced in vitro by applying 4-aminopyridine (4AP)-containing medium. Under these conditions, we could identify in FCD slices a close temporal relationship between ictal activity onset and the occurrence of slow interictal-like events that were mainly contributed by GABAA receptor activation. We also found that in FCD slices, pharmacological procedures capable of decreasing or increasing GABAA receptor function abolished or potentiated ictal discharges, respectively. In addition, the initiation of ictal events in FCD tissue coincided with the occurrence of GABAA receptor-dependent interictal events leading to [K+]o elevations that were larger than those seen during the interictal period. Finally, by testing the effects induced by baclofen on epileptiform events generated by FCD and MTLE slices, we discovered that the function of GABAB receptors (presumably located at presynaptic inhibitory terminals) was markedly decreased in FCD tissue. Thus, epileptiform synchronization leading to in vitro ictal activity in the human FCD tissue is initiated by a synchronizing mechanism that paradoxically relies on GABAA receptor activation causing sizeable increases in [K+]o. This mechanism may be facilitated by the decreased ability of GABAB receptors to control GABA release from interneuron terminals.

Initiation of electrographic seizures by neuronal networks in entorhinal and perirhinal cortices in vitro.

The hippocampus is often considered to play a major role in the pathophysiology of mesial temporal lobe epilepsy. However, emerging clinical and experimental evidence suggests that parahippocampal areas may contribute to a greater extent to limbic seizure initiation, and perhaps epileptogenesis. To date, little is known about the participation of entorhinal and perirhinal networks to epileptiform synchronization. Here, we addressed this issue by using simultaneous field potential recordings in horizontal rat brain slices containing interconnected limbic structures that included the hippocampus proper. Epileptiform discharges were disclosed by bath applying the convulsant drug 4-aminopyridine (50 microM) or by superfusing Mg(2+)-free medium. In the presence of 4-aminopyridine, slow interictal- (duration=2.34+/-0.29 s; interval of occurrence=25.75+/-2.11 s, n=16) and ictal-like (duration=31.25+/-3.34 s; interval of occurrence=196.96+/-21.56 s, n=17) discharges were recorded in entorhinal and perirhinal cortices after abating the propagation of CA3-driven interictal activity to these areas following extended hippocampal knife cuts. Simultaneous recordings obtained from the medial and lateral entorhinal cortex, and from the perirhinal cortex revealed that interictal and ictal discharges could initiate from any of these areas and propagate to the neighboring structure with delays of 8-66 ms. However, slow interictal- and ictal-like events more often originated in the medial entorhinal cortex and perirhinal cortex, respectively. Cutting the connections between entorhinal and perirhinal cortices (n=10), or functional inactivation of cortical areas by local application of a glutamatergic receptor antagonist (n=11) made independent epileptiform activity occur in all areas. These procedures also shortened ictal discharge duration in the entorhinal cortices, but not in the perirhinal area. Similar results could be obtained by applying Mg(2+)-free medium (n=7). These findings indicate that parahippocampal networks provide independent epileptiform synchronization sufficient to sustain limbic seizures as well as that the perirhinal cortex plays a preferential role in in vitro ictogenesis.

Involvement of amygdala networks in epileptiform synchronization in vitro.

We used field potential and intracellular recordings in rat brain slices that included the hippocampus, a portion of the basolateral/lateral nuclei of the amygdala (BLA) and the entorhinal cortex (EC). Bath application of the convulsant 4-aminopyridine (50 microM) to slices (n=12) with reciprocally connected areas, induced short-lasting interictal-like epileptiform discharges that (i) occurred at intervals of 1.2-2.8 s, (ii) originated in CA3, and (iii) spread to EC and BLA. Cutting the Schaffer collaterals abolished them in both parahippocampal areas where slower interictal-like (interval of occurrence=4-17 s) and prolonged ictal-like discharges (duration=15+/-6.9 s, mean+/-S.D., n=7) appeared. These new types of epileptiform activity originated in either EC or BLA. Similar findings were obtained in slices (n=19) in which the hippocampus outputs were not connected with the EC and BLA under control conditions. Cutting the EC-BLA connections made independent slow interictal- and ictal-like activities appear in both areas (n=5). NMDA receptor antagonism (n=6) abolished ictal-like discharges and reduced the duration of the slow interictal-like events. Repetitive stimulation of BLA at 0.5-1 Hz in Schaffer collateral cut slices, induced interictal-like epileptiform depolarizations in EC and reversibly blocked ictal-like activity (n=14). Thus, CA3 outputs in intact slices entrain EC and BLA networks into an interictal-like pattern that inhibits the propensity of these parahippocampal areas to generate prolonged ictal-like paroxysms. Accordingly, NMDA receptor-dependent ictal-like events are initiated in BLA or EC once the propagation of CA3-driven interictal-like discharges to these areas is abated by cutting the Schaffer collaterals. Similar inhibitory effects also occur by activating BLA outputs directed to EC at rates that mimic the CA3-driven interictal-like pattern.

Involvement of cholinergic and gabaergic systems in the fragile X knockout mice.

Fragile X syndrome is an inherited cause of mental retardation. We used extra- and intracellular recordings in brain slices obtained from wild type and fragile X knockout mice to establish whether bath application of the cholinergic agent carbachol (5 microM) induces different responses in neurons of the subiculum, a limbic structure involved in learning and memory. We found that carbachol diminished excitatory post-synaptic responses induced by CA1 stratum radiatum stimulation in wild type mice, but caused an unexpected increase in knockout animals. Moreover, these responses augmented in knockout mice after carbachol washout, a phenomenon that resembled the muscarinic long-term potentiation seen in wild type mice during application of carbachol and GABA(A) receptor antagonists. We also used paired-pulse stimulation to determine whether the changes in synaptic excitability induced by carbachol were caused by pre- or post-synaptic mechanism. Under control conditions, this protocol induced facilitation in both wild type and knockout mice; in contrast, during carbachol application, this facilitatory effect was seen in wild type mice only. In conclusion, our data highlight for the first time differences in cholinergic and GABA-ergic mechanisms that may contribute to the phenotype of fragile X patients.

Intrinsic optical signals and electrographic seizures in the rat limbic system.

We measured the intrinsic optical signals (IOSs) generated by rat hippocampus-entorhinal cortex (EC) slices in response to single shock electrical stimuli delivered in the EC deep layers during application of the convulsant drug 4-aminopyridine (50 microM). With field potential recordings the stimulus-induced responses had duration = 35 +/- 6.3 s mean +/- SEM, n = 7 slices) and characteristics resembling electrographic seizures. IOS changes reflecting an increase in light transmittance occurred in the EC and hippocampus following similar stimuli (n = 45). IOSs increased progressively to reach peak values 20-30 s after the stimulus and returned slowly to prestimulus values within 100 s, thus outlasting the field potential discharge. IOS changes initiated in the medial EC, near to the stimulation site, and spread to the lateral EC, the dentate, and the CA3/CA1 areas. IOS spread from EC to hippocampus was not seen after perforant path cut (n = 5). Moreover, field potential and IOS responses were markedly decreased by excitatory amino acid receptor antagonists (n = 12). The antiepileptic drugs topiramate (10-100 microM, n = 16) or lamotrigine (100-400 microM, n = 12) reduced the IOS changes in the EC and their spread to distant areas. These effects were reversible and dose-dependent (IC50 = 48 microM and 210 microM for topiramate and lamotrigine, respectively). Thus, in 4AP-treated hippocampus-EC slices, IOS changes accompany and outlast the field potential epileptiform responses, depend on glutamatergic transmission and are characterized by temporal and spatial distributions consistent with propagation through established anatomical pathways. We also propose that IOSs may represent a reliable tool for screening the effects of neuroactive compounds such as antiepileptic drugs.

GABA-mediated synchronization in the human neocortex: elevations in extracellular potassium and presynaptic mechanisms.

Field potential and extracellular [K(+)] ([K(+)](o)) recordings were made in the human neocortex in an in vitro slice preparation to study the synchronous activity that occurs in the presence of 4-aminopyridine (50 microM) and ionotropic excitatory amino acid receptor antagonists. Under these experimental conditions, negative or negative-positive field potentials accompanied by rises in [K(+)](o) (up to 4.1 mM from a baseline of 3.25 mM) occurred spontaneously at intervals of 3-27 s. Both field potentials and [K(+)](o) elevations were largest at approximately 1000 microm from the pia. Similar events were induced by neocortical electrical stimuli. Application of medium containing low [Ca(2+)]/high [Mg(2+)] (n=3 slices), antagonism of the GABA(A) receptor (n=7) or mu-opioid receptor activation (n=4) abolished these events. Hence, they represented network, GABA-mediated potentials mainly reflecting the activation of type A receptors following GABA release from interneurons. The GABA(B) receptor agonist baclofen (10-100 microM, n=11) reduced and abolished the GABA-mediated potentials (ID(50)=18 microM). Baclofen effects were antagonized by the GABA(B) receptor antagonist CGP 35348 (0.1-1 mM, n=6; ID(50)=0.19 mM). CGP 38345 application to control medium increased the amplitude of the GABA-mediated potentials and the concomitant [K(+)](o) rises without modifying their rate of occurrence. The GABA-mediated potentials were not influenced by the broad-spectrum metabotropic glutamate agonist (+/-)-1-aminocyclopentane-trans-1,3-dicarboxylic acid (100 microM, n=10), but decreased in rate with the group I receptor agonist (S)-3,5-dihydroxyphenylglycine (10-100 microM, n=9). Our data indicate that human neocortical networks challenged with 4-aminopyridine generate glutamatergic-independent, GABA-mediated potentials that are modulated by mu-opioid and GABA(B) receptors presumably located on interneuron terminals. These events are associated with [K(+)](o) elevations that may contribute to interneuron synchronization in the absence of ionotropic excitatory synaptic transmission.

Network and intrinsic contributions to carbachol-induced oscillations in the rat subiculum.

Low-frequency network oscillations occur in several areas of the limbic system where they contribute to synaptic plasticity and mnemonic functions that are in turn modulated by cholinergic mechanisms. Here we used slices of the rat subiculum (a limbic area involved in cognitive functions) to establish how network and single neuron (intrinsic) membrane mechanisms participate to the rhythmic oscillations elicited by the cholinergic agent carbachol (CCh, 50-100 microM). We have found that CCh-induced network oscillations (intraoscillatory frequency = 5-16 Hz) are abolished by an antagonist of non-N-methyl-D-aspartate (NMDA) glutamatergic receptors (n = 6 slices) but persist during blockade of GABA receptors (n = 16). In addition, during application of glutamate and GABA receptor antagonists, single subicular cells generate burst oscillations at 2.1-6.8 Hz when depolarized with steady current injection. These intrinsic burst oscillations disappear during application of a Ca(2+) channel blocker (n = 6 cells), intracellular Ca(2+) chelation (n = 6), or replacement of extracellular Na(+) (n = 4) but persist in recordings made with electrodes containing a blocker of voltage-gated Na(+) channels (n = 7). These procedures cause similar effects on CCh-induced depolarizing plateau potentials that are contributed by a Ca(2+)-activated nonselective cationic conductance (I(CAN)). Network and intrinsic oscillations along with depolarizing plateau potentials were abolished by the muscarinic receptor antagonist atropine. In conclusion, our findings demonstrate that low-frequency oscillations in the rat subiculum rely on the muscarinic receptor-dependent activation of an intrinsic oscillatory mechanism that is presumably contributed by I(CAN) and are integrated within the network via non-NMDA receptor-mediated transmission.

Do interictal discharges promote or control seizures? Experimental evidence from an in vitro model of epileptiform discharge.

Interictal and ictal discharges are recorded from limbic structures in temporal lobe epilepsy patients. In clinical practice, interictal spikes are used to localize the epileptogenic area, but they also are assumed to promote ictal events. Here I review data obtained from combined slices of mouse hippocampus-entorhinal cortex that indicate an inverse relation between interictal and ictal events. In this preparation, application of 4-aminopyridine or Mg2+-free medium induce (a) interictal discharges that originated from CA3 and propagate (via the Schaffer collaterals) to CA1 and entorhinal cortex, to return to the hippocampus through the dentate area; and (b) ictal discharges that initiate in the entorhinal cortex and propagate to the hippocampus via the dentate gyrus. Interictal activity occurs throughout the experiment (up to 6 h), whereas ictal discharges disappear after 1-2 h. Schaffer collateral cut abolishes interictal discharges in CA1, entorhinal cortex, and dentate and reestablishes entorhinal ictal discharges. Moreover, ictal discharge generation in the entorhinal cortex after Schaffer collateral cut is prevented by mimicking CA3 activity with rhythmic electrical stimulation of CA1 outputs. Thus hippocampal interictal activity controls the ability of the entorhinal cortex to generate seizures. It also may be proposed that Schaffer collateral cut may model the epileptic condition in which CA3 damage results in loss of hippocampal control over the entorhinal cortex. In conclusion, these experiments demonstrate that interictal activity controls rather than promotes ictal events, and functional integrity of CA3 constitutes a critical control mechanism in temporal lobe epilepsy.