The objectives of this study are to determine whether persistent alterations in the GABAA receptor complex (GABAR) can provide a molecular explanation for the development of physical dependence on ethanol in an animal model of alcoholism. Chronic intermittent ethanol (CIE) administration to rats has many features resembling human alcohol abuse behavior, including long-lasting susceptibility to readdiction. The numerous episodes of ethanol-induced depression of the nervous system and the following rebound hyperexcitability (withdrawal) have been shown to exert a kindling-like effect, i.e., a persistent increased severity of the hyperexcitable withdrawal symptoms. Rats treated in this manner become ethanol dependent, one measure being a decreased seizure threshold to the convulsant drug pentylenetetrazol (PTZ), a blocker of the GABAR. The hyperexcitability to PTZ (kindling) lasts at least 40 days after cessation of ethanol. Neurochemical and electrophysiological studies have been focused on whether this ethanol withdrawal kindling can be associated with alterations in the molecular properties of the GABAR, and have demonstrated a significant reduction in GABAR function. In addition, several pharmacological properties of GABAR are altered in hippocampus, although tolerance to eth enhancement of GABAR by ethanol was not found. We showed that GABAR are positively regulated by protein kinase C and neurosteroids Measurements of receptor subunit mRNA by reverse-transcriptase polymerase chain reaction (RT-PCR) and in situ hybridization techniques reveal significant changes in isoform composition, with elevated levels of alpha4 and gamma2short subunits. In combination with protein biochemistry studies, we can compare naive and CIE rats in order to define the subunit isoforms present. Then we express the different hippocampal GABAR recombinant subunit isoforms in Xenopus oocytes and study what physiological consequences might arise from changing the subunit composition. We are testing initial suggestions that hypoinhibition may result from reduced positive modulation of GABAR by PKC and neurosteroids. We suggest that reduced GABAR function in the hippocampus of ethanol-dependent individuals has profound effects on various emotional and intellectual aspects of brain activity. Finding the molecular mechanisms responsible may help in treatment of withdrawal symptoms and hopefully in reduction of ethanol dependence.
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