Abstracts

RATIONALE: NeuroD (ND) is a bHLH transcription factor required for neuronal differentiation. Homozygous mice deficient for the NeuroD gene reveal a lack of granule cells in the dentate gyrus, with only a cluster of precursor cells remaining in the cap region and a neurological phenotype of spontaneous limbic epilepsy. Since seizures activate granule cell neurogenesis, we wondered whether the granule cell precursors remaining in NeuroD knockout mice could undergo mitosis after prolonged seizure activity induced by pilocarpine (PILO), an experimental model for temporal lobe epilepsy. METHODS: Male NeuroD knock out mice (2 months old)(n=7) and unaffected control mice (n=5) were used. Methyl scopolamine s.c.(1 mg/kg) was given to the animals prior to 350 mg/kg and 360 mg/kg of pilocarpine i.p. The animals received i.p. injections of BrdU (50 mg/kg) 2 weeks after pilocarpine induction. BrdU immunohistochemistry was performed according to Parent et al., 1997. Microelectrodes were implanted in the cortex and digital recordings were performed with simultaneous videotaping. RESULTS: Unexpectedly, ND mutants showed a higher threshold for PILO seizures. After one PILO injection of 350 mg/kg i.p. (a dose which can reliably induce prolonged status epilepticus for 4-6 hours in wild type mice), no behavioral or EEG disturbances were observed; at higher doses (360 mg/kg) the ND mice had a brief seizure (1 minute) and died. Despite the lack of visible seizures in surviving mice, we injected BrdU in 2 ND mice treated with pilocarpine and no labeled cells were seen, while unaffected mice after seizures showed extensive BrdU labeling in the subgranular layer when compared to saline-treated control mice. CONCLUSIONS: Dentate granule cells receive a dense cholinergic input, and these receptors may be an important target for PILO-induced activation underlying sustained seizures. While the organization of the mutant hippocampal circuitry remains to be characterized, our results suggest that granule cells are crucial for the establishment of status epilepticus induced by pilocarpine. This work was supported by FAPESP (MM) and NS29709 (JLN).