Abstracts

RATIONALE: Seizure-induced plasticity and injury proceed through several temporal stages: short-term effects are due to post-translational modifications of existing proteins and immediate early gene expression, while long-lasting changes in synaptic efficacy, structural modifications, and cell death require a coordinated program of changes in gene transcription in response to these early molecular events. Certain inbred strains of mice show significant differences in seizure-induced plasticity and vulnerability to cell death. By isolating the genes that are differentially activated at early times following seizures in these strains, we can elucidate the mechanisms responsible for these traits.
METHODS: We used pilocarpine (PILO) to determine that C3H/HeJ mice show a lower seizure threshold and higher number of apoptotic cells in hippocampal pyramidal neurons when examined 24 hours after a 6 hour episode of [italic]status epilepticus[/italic] (SE) compared to C57BL/6J mice. Using differential display of hippocampal mRNAs extracted 12 hours after SE, followed by cloning and sequencing, we identified three genes, MCL-1 (Myeloid cell leukemia-1), SRP-9 (signal recognition protein subunit 9), and Rab3-interacting protein (Rabin3/Pat-12) that were differentially expressed in the two strains. These results were confirmed by in [italic]situ hybridization[/italic] of each gene and ribonuclease protection assay for MCL-1.
RESULTS: mRNA levels of MCL-1, SRP-9, and Rabin3 were significantly increased (up to 50%) in the hippocampus of C3H/HeJ compared to C57BL/6J mice. These genes possess diverse functions: SRP-9 is involved in the co-translational translocation of proteins into the endoplasmic reticulum as well as decentralized protein biosynthesis in dendrites; MCL-1 is homologous to Bcl-2 and may also have anti-apoptotic properties; and Rabin3 interacts with RAS-like GTPase binding proteins involved with regulation of synaptic vesicle fusion and Ca²⁺-dependent neurotransmitter release.
CONCLUSIONS: These immediate early genes are differentially upregulated by seizure activity in C3H/HEJ mice, an inbred strain showing a lower threshold to pilocarpine induced [italic]status epilepticus[/italic] and more pronounced hippocampal cell death than C57BL/6J mice. These genes identify novel regulatory sites within pathways regulating protein trafficking, apoptosis, and synaptic neurotransmitter release that may influence strain differences in seizure-induced downstream gene expression, synaptic plasticity, and cell death. The differential analysis of gene expression in inbred strains provides an experimental starting point for dissecting complex genetic influences involved in human epilepsies.
Support: NS29709 (JLN)and FAPESP (MM)