Abstracts

RATIONALE: Hypoxia is the most common cause of neonatal seizures and can lead to epilepsy. Knowledge of the mechanisms that mediate the long-term epileptogenic effects of hypoxia would aid the development of therapies to treat the acute seizures and minimize the epilepsy risk, but these have yet to be fully elucidated. We previously demonstrated increased excitability in area CA1 in hippocampal slices obtained from rats 4-8 days after hypoxia-induced seizures. We aimed to determine if this increased excitability is associated with altered [gamma]-aminobutyric acid (GABA)-mediated synaptic inhibition of CA1 pyramidal neurons.
METHODS: Seizures were induced in rat pups on postnatal day (P) 10 by exposure to 4-6% O[sub]2[/sub] for 15 min. Hippocampal slices were prepared from hypoxia-treated and control animals at P14-18. Spontaneous and miniature inhibitory postsynaptic currents (s- and mIPSCs) were recorded from CA1 pyramidal neurons under whole-cell voltage-clamp. The holding potential was kept near the reversal potential for glutamate receptors (+10 mV) to record baseline inhibitory events without blocking glutamate-mediated excitation in the hippocampal network. 1[mu]M tetrodotoxin was added to record mIPSCs. Western blots using GABA[sub]A[/sub] receptor subunit-specific antibodies also were performed on hippocampi obtained from control and hypoxia-treated animals at P14.
RESULTS: The frequency and amplitudes of sIPSCs were significantly decreased in CA1 neurons from hypoxia-treated animals compared to controls (p[lt].0001). Additionally, the amplitudes of mIPSCs were significantly decreased in the hypoxia-treated group (p[lt].001), but the mIPSC frequency was unchanged. Western blots showed no change in total hippocampal expression of the GABA[sub]A[/sub] receptor [alpha]1 subunit, but a significant increase in [beta]2/3 expression (p[lt].02) 4 days after hypoxia.
CONCLUSIONS: The decrease in mIPSC amplitudes and unaltered mIPSC frequency indicated decreased responses of postsynaptic GABA[sub]A[/sub] receptors to GABA at inhibitory synapses on CA1 pyramidal neurons after hypoxia-induced seizures. Increased [beta]2/3 subunit expression suggested that altered mIPSC properties could have resulted from changes in the molecular composition of postsynaptic GABA[sub]A[/sub] receptors. Additionally, the decreased sIPSC frequency but unchanged mIPSC frequency indicated that the baseline activity of presynaptic GABAergic neurons had decreased following hypoxia. These alterations in mechanisms of GABA-mediated inhibition of CA1 pyramidal neurons were observed at a time when hippocampal excitability was previously found to be increased. Taken together, these results indicate that decreased inhibition of CA1 pyramidal neurons following perinatal hypoxia-induced seizures occurs by both pre- and postsynaptic mechanisms, and that these changes are likely to contribute to hippocampal hyperexcitability and epileptogenesis.
Support: NINDS NS07473 (RMS) & NS31718 (FEJ)