This proposal will employ electrophysiological and immunocytochemical techniques in the hippocampal slice to investigate the effect of neurosteroids in altering tonic GABAergic inhibition and hippocampal excitability. The neurosteroid 3alpha-5alpha[beta]-THP (THP) is well-known as a positive modulator of the GABA-A receptor, with anxiolytic and sedative effects. However, there are clinical conditions where THP may possess paradoxical excitatory effects (i.e., the peri-pubertal period and PMDD). In this study, we have developed a mouse model of THP withdrawal (Wd) which results in excitatory effects of THP. THP Wd increases expression of alpha-4 and delta GABAR subunits in CA1 hippocampus, associated with the pharmacological responses of alpha4-beta-delta GABAR. In this proposal, we seek to examine the electrophysiological and anatomical correlates of altered GABAR subunit composition in CA1 hippocampus after steroid withdrawal. The delta GABAR subunit will be localized to dendritic regions which we expect will co-localize with alpha-4. Preliminary results suggest that increases in extrasynaptic delta subunit, perisynaptic to inhibitory synapses, result from THP Wd. Because THP can attenuate current gated by alpha4-beta-delta GABAR when Cl flux is inward, we will test the hypothesis that local dendritic application of THP will decrease tonic GABAergic current recorded from CA1 hippocampal pyramidal cells following THP Wd, resulting in benzodiazepine insensitivity (recorded with whole cell patch clamp techniques). THP effects on neuronal excitability will be assessed using current clamp, cell-attached patch, paired-pulse inhibition and EPSP-spike coupling. Because the delta subunit is also localized perisynaptic to putative NMDAR synapses, we will also test whether THP facilitates the induction of long-term potentiation (LTP) and long-term depression (LTD) following THP Wd as a result of reduction in shunting inhibition. Because alterations in synaptic plasticity suggest a role in cognition, we will test the effect of THP administration on performance in a hippocampal-dependent spatial task. These results may be relevant for alterations in mood, cognition and seizure susceptibility (i.e., catamenial epilepsy) reported during puberty and across the menstrual cycle.
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