Refractory seizures remain a major source of disability despite advances in surgical and pharmacological treatment of temporal lobe epilepsy. Status epilepticus (SE) causes chronic seizures in animal models. Transcriptional changes induced by calcium entry during SE play an important role in epileptogenesis but the mechanisms of long-term transcriptional regulation after SE have not been well characterized. As changes in DNA methylation have been shown to play an important role in activity-dependent gene expression in neurons, we wondered whether similar epigenetic mechanisms play a role in transcriptional regulation in epileptogenesis. Our preliminary results show that SE causes significant epigenetic changes to brain-derived neurotrophic factor (BDNF), a gene up-regulated by SE, and implicated in epileptogenesis. We also find that demethylation of DNA leads to dysregulation of ion channel expression, with down-regulation of the A-type potassium channel, kv4.2. Interestingly, down-regulation of dendritic kv4.2 channels after SE increases the excitability of hippocampal neurons and has been proposed to increase the probability of chronic seizures. The goal of this project is to test the hypothesis that epigenetic changes after SE are crucial for promoting epileptogenesis.
The Specific Aims are to 1) test the hypothesis that dynamic changes in DNA methylation are essential for transcriptional changes after SE, and to 2) test the hypothesis that prevention of de novo DNA methylation can modulate SE-induced transcriptional changes and epileptogenesis. We will characterize genome-wide changes in DNA methylation after SE using DNA methylation microarrays. We will ascertain if DNA methylation has a direct role in changes in transcription by performing promoter luciferase expression assays in dissociated cultures. We will test whether transgenic mice with deficient de novo DNA methylation have increased down-regulation of dendritic A-type kv4.2 channels after SE. Finally we will determine if prevention of de novo methylation promotes epileptogenesis by testing the frequency and duration of chronic seizures after SE. The training offered by mentors with laboratories with specific expertise in animal models of epilepsy, epigenetic gene regulation, and advanced electrophysiological techniques will help the candidate gain expertise in multiple scientific domains during his independent career as a clinician-scientist studying epileptogenesis.
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