Abstracts

Rationale: Hypoxic encephalopathy is one of the most common causes of neonatal seizures (Volpe 2001; Jensen 2006). Early life seizures render the brain susceptible to later epilepsy and cognitive deficits (Ben-Ari and Holms 2006; Silverstein and Jensen 2007). Recently, epigenetic mechanisms have been reported to contribute to adult seizures, but evidence for epigenetic regulation in neonatal seizures is limited. One epigenetic regulator reported to be involved in seizures is REST (RE1 silencing transcription factor). REST is a transcription factor widely expressed in embryogenesis and significant regulator of epigenetic neuronal differentiation. Given our previous data that GluR2 subunit of AMPAR (Sanchez 2001), an established target of REST silencing, was downregulated after neonatal hypoxia-induced seizures in rats, we hypothesized that neonatal seizures might regulate REST activity in the developing brain. Importantly, REST has been shown to be modified in a febrile seizure model in immature rats (McClelland et al 2010), and in adult status (Spence et al 2006). We thus investigated the developmental expression of REST in neonatal rats and examined changes in REST expression following hypoxia-induced seizures.Methods: We have previously established that neonatal rats exposed to transient global hypoxia on postnatal day 10 (P10) exhibit electrographic and behavioral seizures that result in increased seizure susceptibility thus mimicking clinical aspects of neonatal hypoxic encephalopathy (Jensen et al. 1992). Thus, at P10, rat pups were subjected to hypoxia-induced seizures (8 min 7% O2, 6 min 5% O2, 1 min 4% O2). Age-and weight-matched littermates not exposed to hypoxic conditions served as controls. Only animals that had more than 5 tonic-clonic seizures were included in the study. At 4, 24, 48, and 72h following hypoxia, brains were dissected into cortex and hippocampus and processed for Western blot analyses (n=5/group). Results: Acutely following hypoxia-induced seizures, REST protein expression is increased in the hippocampus and cortex compared to controls. Specifically REST is mildly increased in the cortex at 4 and 24h (11 and 15% increases respectively), followed by modest increases in REST at 48 and 72h (42% and 66% respectively). In the hippocampus, REST is similarly increased at 4 and 24h (12 and 10% respectively) with a modest increase at 48h (27%). In contrast to the cortex, REST was decreased by 27% in the hippocampus at 72h compared to controls. At these timepoints examined in both regions, GluR2 was decreased as compared to controls.Conclusions: This is one of the first studies describing REST protein expression in the neonatal rat brain and alterations following hypoxia-induced seizures. REST is increased in brain regions vulnerable to seizures in the neonatal period. The increase in REST may be responsible for the downregulation of the AMPAR GluR2 subunit, and may represent an epigenetic effector of injury in the neonatal brain.