Abstracts

Rationale: Altered function of hyperpolarization and cyclic nucleotide-gated (HCN) channels has been reported in cortical and hippocampal pyramidal neurons in multiple animal seizure models (Chen, et al. 1999; Shah, et al. 2004; Zhang, et al. 2006), and may contribute to epileptogenesis. However, seizure-associated HCN channel dysregulation has not been examined in inhibitory hippocampal interneurons, where hyperpolarization-activated currents (I_H) can promote spontaneous firing to sustain a basal rate of synaptic inhibition (Macaferri and McBain 1996; Lupica, et al. 2001). Hypoxia is the most common cause of neonatal seizures, and we previously observed decreased I_H in CA1 pyramidal neurons 1-3 days after hypoxia-induced seizures in neonatal rat. We further observed decreases in the frequency of action potential-dependent spontaneous inhibitory postsynaptic currents (sIPSCs) in pyramidal neurons during this period. Given that I_H can promote spontaneous firing of inhibitory CA1 interneurons, we hypothesized that decreased I_H in inhibitory interneurons that are presynaptic to pyramidal neurons mediated the decreased sIPSC frequency in pyramidal neurons. Therefore, we aimed to determine if I_H is altered in CA1 inhibitory interneurons after neonatal hypoxia-induced seizures.Methods: Seizures were induced in postnatal day 10 rat pups by 14-16 minute exposure to 5-7% O₂, and whole-cell voltage-clamp recordings were obtained from visualized CA1 pyramidal neurons and non-pyramidal neurons in hippocampal slices taken 1-3 days later. I_H was isolated by applying hyperpolarizing voltage steps from a holding potential of -40 mV with tetrodotoxin, TEA, 4-AP, and CdCl₂ in the bath. Recorded cells were filled with biocytin and recovered histologically to confirm their morphology and location.Results: Consistent with our previous findings, maximum steady-state I_H was significantly decreased in pyramidal neurons from the hypoxia-treated group (p<0.0001), and normalized tail currents showed an approximate l0 mV negative shift in the voltage-dependence of activation. In contrast, no differences between groups were observed in I_H recorded from CA1 stratum oriens interneurons (p=0.99). Although I_H was expressed only in a subset of interneurons, there was no difference between groups in the proportions of cells that expressed I_H. Conclusions: These data indicate that hypoxia-induced seizures modulate I_H in a cell type-specific manner, and further suggest that the decreased basal synaptic inhibition of CA1 pyramidal neurons after seizure-inducing hypoxia is not consequent to dysregulation of I_H in presynaptic inhibitory interneurons. Future work will determine if the selective dysregulation of HCN channels across cell types may be due to HCN subunit-specific susceptibility to seizure-induced modulation, or cell-specificity in the prevailing modulatory pathways or their regulation by seizures. Elucidation of endogenous mechanisms of HCN channel regulation will aid the development of strategies to reverse their potentially epileptogenic dysregulation. (Supported by NINDS R01 NS047385.)