Matthew C. Walker

Extrasynaptic GABAA Receptors: Form, Pharmacology, and Function

GABA is the principal inhibitory neurotransmitter in the CNS and acts via GABAA and GABAB receptors. Recently, a novel form of GABAA receptor-mediated inhibition, termed “tonic” inhibition, has been described. Whereas synaptic GABAA receptors underlie classical “phasic” GABAA receptor-mediated inhibition (inhibitory postsynaptic currents), tonic GABAA receptor-mediated inhibition results from the activation of extrasynaptic receptors by low concentrations of ambient GABA. Extrasynaptic GABAA receptors are composed of receptor subunits that convey biophysical properties ideally suited to the generation of persistent inhibition and are pharmacologically and functionally distinct from their synaptic counterparts. This mini-symposium review highlights ongoing work examining the properties of recombinant and native extrasynaptic GABAA receptors and their preferential targeting by endogenous and clinically relevant agents. In addition, it emphasizes the important role of extrasynaptic GABAA receptors in GABAergic inhibition throughout the CNS and identifies them as a major player in both physiological and pathophysiological processes.

Multiple and Plastic Receptors Mediate Tonic GABAA Receptor Currents in the Hippocampus

Persistent activation of GABAA receptors by extracellular GABA (tonic inhibition) plays a critical role in signal processing and network excitability in the brain. In hippocampal principal cells, tonic inhibition has been reported to be mediated by α5-subunit-containing GABAA receptors (α5GABAARs). Pharmacological or genetic disruption of these receptors improves cognitive performance, suggesting that tonic inhibition has an adverse effect on information processing. Here, we show that α5GABAARs contribute to tonic currents in pyramidal cells only when ambient GABA concentrations increase (as may occur during increased brain activity). At low ambient GABA concentrations, activation of δ-subunit-containing GABAA receptors predominates. In epileptic tissue, α5GABAARs are downregulated and no longer contribute to tonic currents under conditions of raised extracellular GABA concentrations. Under these conditions, however, the tonic current is greater in pyramidal cells from epileptic tissue than in pyramidal cells from nonepileptic tissue, implying substitution of α5GABAARs by other GABAA receptor subtypes. These results reveal multiple components of tonic GABAA receptor-mediated conductance that are activated by low GABA concentrations. The relative contribution of these components changes after the induction of epilepsy, implying an adaptive plasticity of the tonic current in the presence of spontaneous seizures.

Monosynaptic GABAergic Signaling from Dentate to CA3 with a Pharmacological and Physiological Profile Typical of Mossy Fiber Synapses

Mossy fibers are the sole excitatory projection from dentate gyrus granule cells to the hippocampus, where they release glutamate, dynorphin, and zinc. In addition, mossy fiber terminals show intense immunoreactivity for the inhibitory neurotransmitter GABA. Fast inhibitory transmission at mossy fiber synapses, however, has not previously been reported. Here, we show that electrical or chemical stimuli that recruit dentate granule cells elicit monosynaptic GABA(A) receptor-mediated synaptic signals in CA3 pyramidal neurons. These inhibitory signals satisfy the criteria that distinguish mossy fiber-CA3 synapses: high sensitivity to metabotropic glutamate receptor agonists, facilitation during repetitive stimulation, and NMDA receptor-independent long-term potentiation. GABAergic transmission from the dentate gyrus to CA3 has major implications not only for information flow into the hippocampus but also for developmental and pathological processes involving the hippocampus.

GABAA Receptors at Hippocampal Mossy Fibers

Presynaptic GABAA receptors modulate synaptic transmission in several areas of the CNS but are not known to have this action in the cerebral cortex. We report that GABAA receptor activation reduces hippocampal mossy fibers excitability but has the opposite effect when intracellular Cl- is experimentally elevated. Synaptically released GABA mimics the effect of exogenous agonists. GABAA receptors modulating axonal excitability are tonically active in the absence of evoked GABA release or exogenous agonist application. Presynaptic action potential-dependent Ca2+ transients in individual mossy fiber varicosities exhibit a biphasic dependence on membrane potential and are altered by GABAA receptors. Antibodies against the alpha2 subunit of GABAA receptors stain mossy fibers. Axonal GABAA receptors thus play a potentially important role in tonic and activity-dependent heterosynaptic modulation of information flow to the hippocampus.

Optogenetic and Potassium Channel Gene Therapy in a Rodent Model of Focal Neocortical Epilepsy

Light-activated gene therapy acutely suppresses seizures, and gene therapy with a potassium channel prevents epileptogenesis and treats established epilepsy in a rodent model. Casting Light on the Shadow of Epilepsy Epilepsy affects 1% of the population and is often resistant to medication. Surgery to remove the region of the brain that generates seizures is only feasible in a minority of cases because of risks to movement, language, vision, and other essential functions. There is an urgent need for alternative treatments. In a new study, Wykes et al. used a virus to express a therapeutic gene in a small number of brain neurons in the seizure-generating zone in a rat model of drug-resistant epilepsy. They also developed new wireless technology to monitor and detect seizures using a miniaturized implanted transmitter and advanced algorithms. They took two approaches to reduce brain circuit excitability and hence epileptic seizures in the rat model. First, to suppress neuronal firing acutely, they expressed the light-sensitive chloride transporter halorhodopsin in the seizure-generating zone. When laser light was delivered via an optic fiber to this region and the halorhodopsin was activated, they observed a decrease in electrical seizure activity. The success of this “optogenetic” approach implies that a device could be developed to detect and stop seizures “on demand” akin to an implantable defibrillator for heart rhythm disturbances. For longer-term suppression of epilepsy, the investigators overexpressed a brain potassium ion channel that normally regulates both neuronal excitability and neurotransmitter release. This gene therapy treatment fully prevented epilepsy from developing in the rat model. When it was applied during established epilepsy, potassium channel gene therapy progressively diminished the frequency of seizures until they stopped after a few weeks. Neither of the gene therapy approaches tested interfered with normal behavior, most likely because only a small number of neurons were targeted. Although still in the earliest stages of study, this gene therapy approach may hold promise for treating drug-resistant epilepsy. Neocortical epilepsy is frequently drug-resistant. Surgery to remove the epileptogenic zone is only feasible in a minority of cases, leaving many patients without an effective treatment. We report the potential efficacy of gene therapy in focal neocortical epilepsy using a rodent model in which epilepsy is induced by tetanus toxin injection in the motor cortex. By applying several complementary methods that use continuous wireless electroencephalographic monitoring to quantify epileptic activity, we observed increases in high frequency activity and in the occurrence of epileptiform events. Pyramidal neurons in the epileptic focus showed enhanced intrinsic excitability consistent with seizure generation. Optogenetic inhibition of a subset of principal neurons transduced with halorhodopsin targeted to the epileptic focus by lentiviral delivery was sufficient to attenuate electroencephalographic seizures. Local lentiviral overexpression of the potassium channel Kv1.1 reduced the intrinsic excitability of transduced pyramidal neurons. Coinjection of this Kv1.1 lentivirus with tetanus toxin fully prevented the occurrence of electroencephalographic seizures. Finally, administration of the Kv1.1 lentivirus to an established epileptic focus progressively suppressed epileptic activity over several weeks without detectable behavioral side effects. Thus, gene therapy in a rodent model can be used to suppress seizures acutely, prevent their occurrence after an epileptogenic stimulus, and successfully treat established focal epilepsy.

The drug treatment of status epilepticus in Europe: consensus document from a workshop at the first London Colloquium on Status Epilepticus.

In this issue of Epilepsia, we present the next in our series of review, opinion, and commentary (pp. 1277–1288). Other such series, published in Gray Matters over the past couple of years, include discussions about: ethical dimensions of epilepsy genetics [2006, 47(10):1747–1756]; internet research [2007, 48(7):1415–1424]; drug resistance [2007, 48(12):2369–2374]; SCN1A mutation and sudden unexplained death in epilepsy (SUDEP) [2008, 49(2)360– 369]; and Wada testing [2008-49(4):715–727]. One reader has pointed out that we have not explained the format or purpose of these series, and so we have now added a section heading to clarify our intent. In these series, we publish a lead paper followed by commentaries that have been invited by the editors specifically to put forward different views. In commissioning such commentary, we hope to encourage discussion and debate that will lead to further research and elucidation of these still controversial issues. We hope this initiative is of interest to readers, and we invite readers to suggest new topic areas for debate within the pages of Epilepsia.

Loss of Dendritic HCN1 Subunits Enhances Cortical Excitability and Epileptogenesis

Hyperpolarization-activated cation nonselective 1 (HCN1) plasticity in entorhinal cortical (EC) and hippocampal pyramidal cell dendrites is a salient feature of temporal lobe epilepsy. However, the significance remains undetermined. We demonstrate that adult HCN1 null mice are more susceptible to kainic acid-induced seizures. After termination of these with an anticonvulsant, the mice also developed spontaneous behavioral seizures at a significantly more rapid rate than their wild-type littermates. This greater seizure susceptibility was accompanied by increased spontaneous activity in HCN1−/− EC layer III neurons. Dendritic Ih in these neurons was ablated, too. Consequentially, HCN1−/− dendrites were more excitable, despite having significantly more hyperpolarized resting membrane potentials (RMPs). In addition, the integration of EPSPs was enhanced considerably such that, at normal RMP, a 50 Hz train of EPSPs produced action potentials in HCN1−/− neurons. As a result of this enhanced pyramidal cell excitability, spontaneous EPSC frequency onto HCN1−/− neurons was considerably greater than that onto wild types, causing an imbalance between normal excitatory and inhibitory synaptic activity. These results suggest that dendritic HCN channels are likely to play a critical role in regulating cortical pyramidal cell excitability. Furthermore, these findings suggest that the reduction in dendritic HCN1 subunit expression during epileptogenesis is likely to facilitate the disorder.

Enhanced QT shortening and persistent tachycardia after generalized seizures

Objective: Generalized tonic-clonic seizures (GTCS) are a major risk factor for sudden unexpected death in epilepsy (SUDEP). We investigated whether ictal/postictal cardiac features were dependent on seizure type within individual patients. Methods: ECG data from patients with medically refractory temporal lobe epilepsy (TLE) undergoing presurgical investigation who had both complex partial seizures and secondarily GTCS during video-EEG telemetry were retrospectively reviewed. Peri-ictal heart rate (HR), corrected QT interval (QTc), HR variability, and cardiac rhythm were assessed. Results: Twenty-five patients were included in this study. Secondarily GTCS led to higher ictal HR, persistent postictal tachycardia, and decreased postictal HR variability. Moreover, abnormal shortening of QTc occurred in 17 patients mainly during the early postictal phase and significantly more often in secondarily GTCS. Abnormal QTc prolongation occurred in 3 patients with no significant association with GTCS. Benign cardiac arrhythmias occurred in 14 patients and were independent of seizure type. Conclusions: Our data suggest a substantial disturbance of autonomic function following secondarily generalized tonic-clonic seizures (GTCS) in patients with medically refractory temporal lobe epilepsy. The observed alterations could potentially facilitate sudden cardiac death and might contribute to the association of sudden unexpected death in epilepsy with GTCS.

Simultaneous intracranial EEG and fMRI of interictal epileptic discharges in humans

Simultaneous scalp EEG-fMRI measurements allow the study of epileptic networks and more generally, of the coupling between neuronal activity and haemodynamic changes in the brain. Intracranial EEG (icEEG) has greater sensitivity and spatial specificity than scalp EEG but limited spatial sampling. We performed simultaneous icEEG and functional MRI recordings in epileptic patients to study the haemodynamic correlates of intracranial interictal epileptic discharges (IED). Two patients undergoing icEEG with subdural and depth electrodes as part of the presurgical assessment of their pharmaco-resistant epilepsy participated in the study. They were scanned on a 1.5 T MR scanner following a strict safety protocol. Simultaneous recordings of fMRI and icEEG were obtained at rest. IED were subsequently visually identified on icEEG and their fMRI correlates were mapped using a general linear model (GLM). On scalp EEG-fMRI recordings performed prior to the implantation, no IED were detected. icEEG-fMRI was well tolerated and no adverse health effect was observed. intra-MR icEEG was comparable to that obtained outside the scanner. In both cases, significant haemodynamic changes were revealed in relation to IED, both close to the most active electrode contacts and at distant sites. In one case, results showed an epileptic network including regions that could not be sampled by icEEG, in agreement with findings from magneto-encephalography, offering some explanation for the persistence of seizures after surgery. Hence, icEEG-fMRI allows the study of whole-brain human epileptic networks with unprecedented sensitivity and specificity. This could help improve our understanding of epileptic networks with possible implications for epilepsy surgery.

Levetiracetam: Antiepileptic properties and protective effects on mitochondrial dysfunction in experimental status epilepticus

Summary:  Purpose: To assess the anticonvulsant activity of the novel antiepileptic drug, levetiracetam (LEV) in a model of self‐sustaining limbic status epilepticus, and to measure the consequence of LEV treatment on the pattern of mitochondrial dysfunction known to occur after status epilepticus (SE).