Pavel I. Ortinski

Cognitive side effects of antiepileptic drugs

Antiepileptic drugs produce global changes in the excitation levels in the central nervous system and often lead to cognitive and behavioral deficits. These deficits vary and must be considered independently in every patient. A number of consistent risk factors have been established. Polypharmacy and high blood levels of an antiepileptic drug (AED) increase the risk of cognitive side effects. Different effects have been demonstrated for some AEDs, but comparative data are incomplete across all of them. Other factors such as patient age and type/frequency of seizures may also be important contributors to the patients cognitive state. AEDs can have positive or negative effects on mood, providing another consideration in choosing the course of treatment.

Termination of epileptiform activity by cooling in rat hippocampal slice epilepsy models

Cooling has been shown to terminate experimentally induced epileptiform activity in models of epilepsy without causing injury to the cooled brain, suggesting that cooling could represent an approach to seizure control in intractable focal epilepsies. Here we sought to determine the most effective way to apply cooling to abort spontaneous epileptiform discharges in in vitro brain slice models. We induced spontaneous epileptiform activity in rat brain slices by exposure to 4-aminopyridine (4-AP), 4-AP plus bicuculline, and Mg(2+)-free artificial CSF (aCSF) at 28-34 degrees C. Extracellular field recordings were made at hippocampal or neocortical sites. Slice temperature was reduced by perfusion with cold aCSF. Rapid cooling at rates of 2-5 degrees C/s was compared to cooling at slower rates of 0.1-1 degrees C/s. Cooling at both rates reversibly aborted epileptiform discharges in all three models and at all recording sites. With rapid cooling, small temperature drops were highly effective in terminating discharges, an effect that was sustained for as long as the reduced temperature level was maintained. In contrast, slow cooling required much larger temperature drops to inhibit discharges. With slow cooling, absolute temperature drops to 21-22 degrees C caused a 90% reduction in event frequency, but cooling to 14-15 degrees C was required to terminate discharges. We conclude that rapid cooling as effectively aborts discharges in in vitro epilepsy models as does slow cooling, but the magnitude of the temperature change required is less. Practical devices to inhibit seizure activity may only need to induce small temperature drops, if the cooling can be applied sufficiently rapidly.

Desensitization and binding properties determine distinct alpha1beta2gamma2 and alpha3beta2gamma2 GABA(A) receptor-channel kinetic behavior.

GABAA receptor subtypes comprising the α1 and α3 subunits change with development and have a specific anatomical localization in the adult brain. These receptor subtypes have been previously demonstrated to greatly differ in deactivation kinetics but the underlying gating mechanisms have not been fully elucidated. Therefore, we expressed rat α1β2γ2 and α3β2γ2 receptors in human embryonic kidneyu2003293 cells and recorded current responses to ultrafast GABA applications at macroscopic and single‐channel levels. We found that the slow deactivation of α3β2γ2‐mediated currents is associated with a relatively small rate and extent of apparent desensitization. In contrast, responses mediated by α1β2γ2 receptors had faster deactivation and stronger desensitization. α3β2γ2 receptors had faster recovery in the paired‐pulse agonist applications than α1β2γ2 channels. The onset of currents mediated by α3β2γ2 receptors was slower than that of α1β2γ2 for a wide range of GABA concentrations. Single‐channel analysis did not reveal differences in the opening/closing kinetics of α1β2γ2 and α3β2γ2 channels but burst durations were longer in α3β2γ2 receptors. Simulation with a previously reported kinetic model was used to explore the differences in respective rate constants. Reproduction of major kinetic differences required a smaller desensitization rate as well as smaller binding and unbinding rates in α3β2γ2 compared with α1β2γ2 receptors. Our work describes the mechanisms underlying the kinetic differences between two major GABAA receptor subtypes and provides a framework to interpret data from native GABA receptors.

The Pheromone Androstenol (5α-Androst-16-en-3α-ol) Is a Neurosteroid Positive Modulator of GABAA Receptors

Androstenol is a steroidal compound belonging to the group of odorous 16-androstenes, first isolated from boar testes and also found in humans. Androstenol has pheromone-like properties in both animals and humans, but the molecular targets of its pheromonal activity are unknown. Androstenol is structurally similar to endogenous A-ring reduced neurosteroids that act as positive modulators of GABAA receptors. Here we show that androstenol has neurosteroid-like activity as a GABAA receptor modulator. In whole-cell recordings from cerebellar granule cells, androstenol (but not its 3β-epimer) caused a concentration-dependent enhancement of GABA-activated currents (EC50, 0.4 μM in cultures; 1.4 μM in slices) and prolonged the duration of spontaneous and miniature inhibitory postsynaptic currents. Androstenol (0.1–1 μM) also potentiated the amplitude of GABA-activated currents in human embryonic kidney 293 cells transfected with recombinant α1β2γ2 and α2β2γ2 GABAA receptors and, at high concentrations (10–300 μM), directly activated currents in these cells. Systemic administration of androstenol (30–50 mg/kg) caused anxiolytic-like effects in mice in the open-field test and elevated zero-maze and antidepressant-like effects in the forced swim test (5–10 mg/kg). Androstenol, but not its 3β-epimer, conferred seizure protection in the 6-Hz electroshock and pentylenetetrazol models (ED50 values, 21.9 and 48.9 mg/kg, respectively). The various actions of androstenol in the whole-animal models are consistent with its activity as a GABAA receptor modulator. GABAA receptors could represent a target for androstenol as a pheromone, for which it is well suited because of high volatility and lipophilicity, or as a conventional hormonal neurosteroid.

Desensitization and binding properties determine distinct α1β2γ2 and α3β2γ2 GABAA receptor-channel kinetic behavior

GABAA receptor subtypes comprising the α1 and α3 subunits change with development and have a specific anatomical localization in the adult brain. These receptor subtypes have been previously demonstrated to greatly differ in deactivation kinetics but the underlying gating mechanisms have not been fully elucidated. Therefore, we expressed rat α1β2γ2 and α3β2γ2 receptors in human embryonic kidneyu2003293 cells and recorded current responses to ultrafast GABA applications at macroscopic and single‐channel levels. We found that the slow deactivation of α3β2γ2‐mediated currents is associated with a relatively small rate and extent of apparent desensitization. In contrast, responses mediated by α1β2γ2 receptors had faster deactivation and stronger desensitization. α3β2γ2 receptors had faster recovery in the paired‐pulse agonist applications than α1β2γ2 channels. The onset of currents mediated by α3β2γ2 receptors was slower than that of α1β2γ2 for a wide range of GABA concentrations. Single‐channel analysis did not reveal differences in the opening/closing kinetics of α1β2γ2 and α3β2γ2 channels but burst durations were longer in α3β2γ2 receptors. Simulation with a previously reported kinetic model was used to explore the differences in respective rate constants. Reproduction of major kinetic differences required a smaller desensitization rate as well as smaller binding and unbinding rates in α3β2γ2 compared with α1β2γ2 receptors. Our work describes the mechanisms underlying the kinetic differences between two major GABAA receptor subtypes and provides a framework to interpret data from native GABA receptors.

Cerebellar nicotinic cholinergic receptors are intrinsic to the cerebellum: implications for diverse functional roles.

Although recent studies have delineated the specific nicotinic subtypes present in the mammalian cerebellum, very little is known about their location or function within the cerebellum. This is of increased interest since nicotinic receptors (nAChRs) in the cerebellum have recently been implicated in the pathology of autism spectrum disorders. To begin to better understand the roles of these heteromeric nAChRs in the cerebellar circuitry and their therapeutic potential as targets for drug development, we used various chemical and stereotaxic lesion models in conjunction with slice electrophysiology to examine how specific heteromeric nAChR subtypes may influence the surrounding cerebellar circuitry. Using subunit-specific immunoprecipitation of radiolabeled nAChRs in the cerebella following N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine hydrochloride, p-chloroamphetamine, and pendunculotomy lesions, we show that most, if not all, cerebellar nicotinic receptors are present in cells within the cerebellum itself and not in extracerebellar afferents. Furthermore, we demonstrate that the β4-containing, but not the β2-containing, nAChRs intrinsic to the cerebellum can regulate inhibitory synaptic efficacy at two major classes of cerebellar neurons. These tandem findings suggest that nAChRs may present a potential drug target for disorders involving the cerebellum.