Abstracts

Rationale: It has been shown that changes in ion channel function occur in animal models of epilepsy at both acute and chronic time points following status epilepticus. However, it remains unclear how ion channel dysfunction is linked to the development of the epileptic state. We thus investigated how alterations in the biophysical properties of hyperpolarization-activated cation (HCN) channels, an ion channel implicated in experimental epilepsy, correlate with the development of epilepsy. Methods: We performed video-EEG monitoring on pilocarpine-treated rats during acute (one week) and chronic (two to four weeks) periods following pilocarpine-induced status epilepticus (SE) to determine onset of spontaneous seizures. Cell-attached and whole-cell patch-clamp recordings in CA1 hippocampal pyramidal neuron dendrites were performed in the acute and chronic periods to measure changes in the properties of Ih, the current produced by HCN channels, and Western blot analysis was used to measure HCN protein expression.Results: Video-EEG monitoring of pilocarpine-treated rats showed that seizures were observed beginning as early as day 3 post-pilocarpine. Racine class 0-2 seizures were frequent at 1-2 weeks post-pilocarpine, then declined in frequency, while class 3-5 seizures had increased in frequency by 3-4 weeks post-pilocarpine. By the end of 4 weeks post-pilocarpine, 85% of rats had experienced spontaneous recurrent seizures. In the acute and chronic periods post-pilocarpine, dendritic Ih densities from pilocarpine-treated animals were significantly reduced compared to age-matched, sham-injected animals. Further, the Ih half-maximal activation voltage (V1/2) from pilocarpine-treated animals was significantly hyperpolarized during the acute period, and increasingly so during the chronic period compared to sham-injected animals. Dendritic current-clamp recordings during the chronic period showed that input resistance and action potential firing from pilocarpine-treated animals were also significantly increased compared to naive animals. Further, Western blot analysis showed that there was a significant loss of hippocampal HCN1 and HCN2 protein expression in the acute period, and persistent loss of HCN1 in the chronic period. Interestingly, rats given phenobarbital during the first week post-pilocarpine to suppress spontaneous seizures showed a similar downregulation of HCN channels (decreased Ih and HCN1/2 protein expression) as rats treated with pilocarpine alone. Conclusions: Our results show that downregulation of HCN channel function and expression in the dendrites of CA1 hippocampal pyramidal neurons occurs in the acute period following pilocarpine-induced SE and persists during the emergence of chronic, spontaneous seizures. This early and progressive HCN downregulation results in increased neuronal excitability. We have also shown that spontaneous seizure activity in the acute period does not itself cause HCN channel downregulation, suggesting that the downregulation of HCN channels results from, and may contribute to, the process of epileptogenesis.