David E. Naylor

Trafficking of GABA(A) receptors, loss of inhibition, and a mechanism for pharmacoresistance in status epilepticus.

During status epilepticus (SE), GABAergic mechanisms fail and seizures become self-sustaining and pharmacoresistant. During lithiumpilocarpine-induced SE, our studies of postsynaptic GABAA receptors in dentate gyrus granule cells show a reduction in the amplitude of miniature IPSCs (mIPSCs). Anatomical studies show a reduction in the colocalization of the β2/β3 and γ2 subunits of GABAA receptors with the presynaptic marker synaptophysin and an increase in the proportion of those subunits in the interior of dentate granule cells and other hippocampal neurons with SE. Unlike synaptic mIPSCs, the amplitude of extrasynaptic GABAA tonic currents is augmented during SE. Mathematical modeling suggests that the change of mIPSCs with SE reflects a decrease in the number of functional postsynaptic GABAA receptors. It also suggests that increases in extracellular [GABA] during SE can account for the tonic current changes and can affect postsynaptic receptor kinetics with a loss of paired-pulse inhibition. GABA exposure mimics the effects of SE on mIPSC and tonic GABAA current amplitudes in granule cells, consistent with the model predictions. These results provide a potential mechanism for the inhibitory loss that characterizes initiation of SE and for the pharmacoresistance to benzodiazepines, as a reduction of available functional GABAA postsynaptic receptors. Novel therapies for SE might be directed toward prevention or reversal of these losses.

Rapid surface accumulation of NMDA receptors increases glutamatergic excitation during status epilepticus.

After 1h of lithium-pilocarpine status epilepticus (SE), immunocytochemical labeling of NMDA receptor NR1 subunits reveals relocation of subunits from the interior to the cell surface of dentate gyrus granule cells and CA3 pyramidal cells. Simultaneously, an increase in NMDA-miniature excitatory postsynaptic currents (mEPSC) as well as an increase in NMDA receptor-mediated tonic currents is observed in hippocampal slices after SE. Mean-variance analysis of NMDA-mEPSCs estimates that the number of functional postsynaptic NMDA receptors per synapse increases 38% during SE, and antagonism by ifenprodil suggests that an increase in the surface representation of NR2B-containing NMDA receptors is responsible for the augmentation of both the phasic and tonic excitatory currents with SE. These results provide a potential mechanism for an enhancement of glutamatergic excitation that maintains SE and may contribute to excitotoxic injury during SE. Therapies that directly antagonize NMDA receptors may be a useful therapeutic strategy during refractory SE.

Seizure-induced Neuronal Death in the Immature Brain

The response of the developing brain to epileptic seizures and to status epilepticus is highly age-specific. Neonates with their low cerebral metabolic rate and fragmentary neuronal networks can tolerate relatively prolonged seizures without suffering massive cell death, but severe seizures in experimental animals inhibit brain growth, modify neuronal circuits, and can lead to behavioral deficits and to increases in neuronal excitability. Past infancy, the developing brain is characterized by high metabolic rate, exuberant neuronal and synaptic networks and overexpression of receptors and enzymes involved in excitotxic mechanisms. The outcome of seizures is highly model-dependent. Status epilepticus may produce massive neuronal death, behavioral deficits, synaptic reorganization and chronic epilepsy in some models, little damage in others. Long-term consequences are also highly age- and model-dependent. However, we now have some models which reliably lead to spontaneous seizures and chronic epilepsy in the vast majority of animals, demonstrating that seizure-induced epileptogenesis can occur in the developing brain. The mode cell death from status epilepticus is largely (but not exclusively) necrotic in adults, while the incidence of apoptosis increases at younger ages. Seizure-induced necrosis has many of the biochemical features of apoptosis, with early cytochrome release from mitochondria and capase activation. We speculate that this form of necrosis is associated with seizure-induced energy failure.

Epileptogenesis After Self‐Sustaining Status Epilepticus

Summary:  Purpose: To describe the natural history of chronic epilepsy after experimental self‐sustaining status epilepticus (SSSE) and to correlate patterns of SSSE with ictal, interictal, and plastic changes that characterize chronic epilepsy.

Short-Term Plasticity of Hippocampal Neuropeptides and Neuronal Circuitry in Experimental Status Epilepticus

Summary:  Purpose: We used a model of self‐staining status epilepticus (SSSE), induced by brief intermittent stimulation of the perforant path in unanesthetized rats, to study the mechanism of initiation and of maintenance of SSSE and the role of neuropeptides in those processes.

Rational polytherapy in the treatment of acute seizures and status epilepticus.

We used a model of severe cholinergic status epilepticus (SE) to study polytherapy aimed at reversing the effects of seizure‐induced loss of synaptic GABAA receptors and seizure‐induced gain of synaptic NMDA receptors. Combinations of a benzodiazepine with ketamine and valproate, or with ketamine and brivaracetam, were more effective and less toxic than benzodiazepine monotherapy in this model of SE.

GABA synapses and the rapid loss of inhibition to dentate gyrus granule cells after brief perforant-path stimulation.

Summary:  Purpose: To study the pharmacologic and synaptic basis for the early loss of paired‐pulse inhibition that occurs in the perforant‐path stimulation model of status epilepticus.

Trafficking of NMDA receptors during status epilepticus: Therapeutic implications

We used two models of status epilepticus (SE) to study trafficking of N‐methyl‐d‐aspartate (NMDA) receptors. SE is associated with increased surface expression of NR1 subunits of NMDA receptors, and with an increase of NMDA synaptic and extrasynaptic currents suggesting an increase in number of functional NMDA receptors on dentate granule cells. The therapeutic implications of these results are discussed.

Molecular Basis of Self-Sustaining Seizures and Pharmacoresistance During Status Epilepticus: The Receptor Trafficking Hypothesis Revisited

During status epilepticus (SE), seizures become selfsustaining (Wasterlain, 1974), and pharmacoresistance to benzodiazepines develops progressively, and can result in a 20-fold reduction of response to diazepam after 30 min of experimental seizures (Kapur & Macdonald, 1997; Mazarati et al., 1998a,b). This time-dependent pharmacoresistance is not mediated by classical transporter mechanisms (Lçscher, 2007), but may reflect seizureinduced internalization of synaptic c-aminobutyric acid (GABA)A receptors (GABAAR) containing the b2–3 and/ or c2 subunits (Goodkin et al., 2005, 2008; Naylor et al., 2005).

Glutamate and GABA in the balance: Convergent pathways sustain seizures during status epilepticus

Seizures rapidly become self‐sustaining and pharmacoresistant to benzodiazepines during status epilepticus (SE). A decrease in the number of postsynaptic γ‐aminobutyric acid (GABA)A receptors with SE causes a loss of synaptic inhibition, whereas increases in postsynaptic glutamatergic receptors further upset the balance between excitation and inhibition. Although extracellular GABA levels may increase during SE and contribute to postsynaptic GABAA receptor desensitization, other pathways involving glutamatergic activation ultimately may be responsible for the persistent down‐regulation of postsynaptic GABAA receptors and erosion of synaptic inhibition.