The long-term objectives of the proposed study are to elucidate the mechanisms by which alcohols exert their toxic actions on the nervous system. Specific attention is focused on the mechanisms underlying the interactions of ethanol and longer-chain alcohols with ion channels by using patch clamp techniques as applied to mammalian neurons. Our previous studies as well as those by other investigators have laid out the groundwork along this line, establishing the phenomenological aspects of alcohol interactions with certain types of ion channels including those activated by GABA, excitatory amino acid (EAA), acetylcholine (ACh) and 5-hydroxytryptamine (5-HT), and voltage-activated sodium, potassium and calcium channels. However, conflicting data have been obtained for certain types of ion channels, and in many cases no detailed mechanisms of alcohol action have been elucidated. The proposed study is aimed at solving some of these problems. Thus the proposed projects will focus on elucidation of the mechanisms of ethanol and longer-chain alcohols on the GABA/A receptor-channel complex. Both whole-cell and single-channel patch clamp techniques will be applied to rat dorsal root gang lion, hippocampal and cortical neurons, and cerebellar Purkinje and granular layer neurons.
The specific aims will be concerned with the modulation of open and/or closed channels, dependence of alcohol action on neuron type, animal age and experimental temperature, the role of intracellular components in alcohol action, and the comparison of subtypes and subunits of receptor-channel complex. For the GABA receptor subunit study, human embryonic kidney (HEK-293) cells, in which various combinations of GABA subunits have been transfected, will be used to determine the role of each subunit in alcohol action. Attention will be focused not only on changes in tee amplitude of GABA-induced currents, but also on changes in the rate of desensitization of the currents. The results of the proposed study will provide the basis for the mechanisms of alcoholism and for approaches to prevention and cure of the disorder.
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