Regulation of seizure-induced MeCP2 Ser421 phosphorylation in the developing brain.

Neonatal seizures disrupt normal synaptic maturation and often lead to later-life epilepsy and cognitive deficits. During early life, the brain exhibits heightened synaptic plasticity, in part due to a developmental overabundance of CaV1.2 L-type voltage gated calcium (Ca2+) channels (LT-VGCCs) and Ca2+-permeable AMPARs (CP-AMPARs) lacking GluA2 subunits. We hypothesized that early-life seizures overactivate these channels, in turn dysregulating Ca2+-dependent signaling pathways including that of methyl CPG binding protein 2 (MeCP2), a transcription factor implicated in the autism spectrum disorder (ASD) Rett Syndrome. Here, we show that in vivo hypoxia-induced seizures (HS) in postnatal day (P)10 rats acutely induced phosphorylation of the neuronal-specific target of activity-dependent MeCP2 phosphorylation, S421, as well as its upstream activator CaMKII T286. We next identified mechanisms by which activity-dependent Ca2+ influx induced MeCP2 phosphorylation using in vitro cortical and hippocampal neuronal cultures at embryonic day (E)18 + 10 days in vitro (DIV). In contrast to the prevalent role of NMDARs in the adult brain, we found that both CP-AMPARs and LT-VGCCs mediated MeCP2 S421 and CaMKII T286 phosphorylation induced by kainic acid (KA) or high potassium chloride (KCl) stimulation. Furthermore, in vivo post-seizure treatment with the broad-spectrum AMPAR antagonist NBQX, the CP-AMPAR blocker IEM-1460, or the LT-VGCC antagonist nimodipine blocked seizure-induced MeCP2 phosphorylation. Collectively, these results demonstrate that early-life seizures dysregulate critical activity-dependent developmental signaling pathways, in part via CP-AMPAR and LT-VGCC activation, providing novel age-specific therapeutic targets for convergent pathways underlying epilepsy and ASDs.

Development of later life spontaneous seizures in a rodent model of hypoxia-induced neonatal seizures.

PURPOSE: To study the development of epilepsy following hypoxia-induced neonatal seizures in Long-Evans rats and to establish the presence of spontaneous seizures in this model of early life seizures. METHODS: Long-Evans rat pups were subjected to hypoxia-induced neonatal seizures at postnatal day 10 (P10). Epidural cortical electroencephalography (EEG) and hippocampal depth electrodes were used to detect the presence of seizures in later adulthood (> P60). In addition, subdermal wire electrode recordings were used to monitor age at onset and progression of seizures in the juvenile period, at intervals between P10 and P60. Timm staining was performed to evaluate mossy fiber sprouting in the hippocampi of P100 adult rats that had experienced neonatal seizures. KEY FINDINGS: In recordings made from adult rats (P60-180), the prevalence of epilepsy in cortical and hippocampal EEG recordings was 94.4% following early life hypoxic seizures. These spontaneous seizures were identified by characteristic spike and wave activity on EEG accompanied by behavioral arrest and facial automatisms (electroclinical seizures). Phenobarbital injection transiently abolished spontaneous seizures. EEG in the juvenile period (P10-60) showed that spontaneous seizures first occurred approximately 2 weeks after the initial episode of hypoxic seizures. Following this period, spontaneous seizure frequency and duration increased progressively with time. Furthermore, significantly increased sprouting of mossy fibers was observed in the CA3 pyramidal cell layer of the hippocampus in adult animals following hypoxia-induced neonatal seizures. Notably, Fluoro-Jade B staining confirmed that hypoxic seizures at P10 did not induce acute neuronal death. SIGNIFICANCE: The rodent model of hypoxia-induced neonatal seizures leads to the development of epilepsy in later life, accompanied by increased mossy fiber sprouting. In addition, this model appears to exhibit a seizure-free latent period, following which there is a progressive increase in the frequency of electroclinical seizures.