Abstracts

Rationale: Recurrent seizures in neonatal and adult cortical structures have long been known to cause acute, activity-dependent alterations in neuronal ion concentrations. Recent data suggest that not only the concentrations but also the transport of ions can be changed by particular patterns of activity. Persistent alterations in chloride transport during recurrent seizures are of particular interest because the first-line medical treatment for neonatal seizures is comprised of drugs that potentiate GABA(A) receptor-mediated chloride currents. Neonatal seizures may induce neuronal chloride accumulation and a consequent excitatory shift in GABA action which may preclude the efficacy of GABAergic anticonvulsants. Methods: Simultaneous high resolution two-photon fluorescence chloride imaging and extracellular field potential recordings were performed during recurrent seizure activity in the in vitro whole hippocampus prepared from neonatal transgenic mice expressing the chloride sensitive dye Clomeleon. Results: We found that in the intact neonatal hippocampus recurrent seizures progressively increase the intracellular chloride concentration ([Cl-]i) and invert the net effect of GABA(A) receptor activation from inhibition to excitation. The quantity of seizure activity was strongly correlated with both the increase in [Cl-]i assayed by Clomeleon imaging and the degree of GABA(A) receptor-mediated excitation assayed by the frequency of action potentials and reduction in phenobarbital efficacy. These excitatory effects persisted in sub-population of neurons after cessation of seizure activity, indicating that they were due to transport rather than activity-dependent ionic accumulation. Further, NKCC1 activity was necessary for persistent, activity-dependent chloride accumulation. Conclusions: Our results demonstrate a novel mechanism by which seizure activity increases the probability of subsequent seizures and precludes the efficacy GABAergic anticonvulsants, providing a potential mechanism for the early crescendo phase of neonatal seizures.