Abstracts

Rationale: Distinct mechanisms may underlie the establishment of epileptic state during different stages of brain development. Rapid kindling is a model of epileptogenesis that allows preclinical evaluation of antiepileptic drugs in experimental animals of different ages. We examined effects of two drugs on the progression of rapid kindling in rats: ganaxolone, which augments tonic GABA inhibition via binding to extrasynaptic GABA receptors which contain a δ subunit; and bumetanide, which blocks Na⁺-K⁺-2Cl^- cotransporter (NKCC1), and thus reverses the excitatory effect of GABA during early development. Methods: The experiments were performed in Wistar rats. At the time of kindling, animals' ages were postnatal days 11 (P11, neonatal); 14 (P14, post-neonatal), 21 (P21, pre-adolescent), or 35 (P35, adolescent). Under Isoflurane anesthesia, animals were implanted with a stimulating electrode into ventral hippocampus and a recording electrode into the neocortex. One day after surgery, animals were subjected to rapid kindling: sixty 10 seconds-long trains, of bipolar 20 Hz square wave pulses delivered every 5 min (total duration 5 hours). Ganaxolone (20 mg/kg) and bumetanide (0.2 mg/kg) were injected intraperitoneally 20 minutes prior to kindling procedure. Control animals were treated with vehicles (dymethyl sulfoxide for ganaxolone and saline for bumetanide). Effects of the drugs were evaluated by quantifying both baseline excitability (afterdischarge threshold and duration) and the progression of kindling seizures. Results: Ganaxolone did not change baseline afterdischarge threshold and duration in both P21 and P35 rats. During kindling, ganaxolone accelerated the occurrence of the first stage 1 seizure, but delayed the development of the first stage 4 convulsion. Furthermore, after ganaxolone treatment animals developed fewer full kindled seizures (stages 4 and 5); however, when such seizure occurred, their duration was significantly longer, as compared with controls. Bumetanide had no effects on both afterdischarge properties and kindling progression in P14 animals. In contrast, in P11 rats bumetanide increased baseline afterdischarge threshold, delayed the development, and decreased the frequency of stage 4 and 5 seizures. Conclusions: Our data showed that the augmentation of tonic GABA inhibition by ganaxolone had dual influence on rapid kindling, which included both antiepileptic (judging by the latency and frequency of full kindled seizures) and facilitatory (judging by the latency of stage 1 seizure and seizure duration) effects. While anticonvulsant properties of ganaxolone were in line with previously reported experimental data, facilitation of certain parameters of rapid kindling was likely due to the enhanced neuronal synchronization, which may be attributed to tonic GABA inhibition. Antiepileptic efficacy of bumetanide was limited to P11 animals, thus emphasizing the importance of NKCC1 - regulated Cl^- - induced depolarization and excitatory effects of GABA in the neonatal brain. Supported by research grants from NIH/NINDS NS059505 and NS046516.