ETHANOL TOLERANCE AND DEPENDENCE IN VITRO

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The locus of ethanol's pharmacological and addictive effects may be at the level of the interaction between a neurotransmitter and its receptor in the synapse. Therefore this proposal focuses on the effects of acute and chronic ethanol on the functioning of certain receptors which have cyclic AMP and cyclic GMP as putative second messengers. Cultured human skin fibroblasts with prostaglandin and Beta-adrenergic receptors mediating cyclic AMP synthesis and murine neuroblastoma cells (clone N1E-115) with several receptors (muscarinic, histamine H1, angiotensin 11, bradykinin, neurotensin and thrombin) mediating cyclic GMP synthesis are being used as model systems for this work. Studies with human skin fibroblasts will determine whether results found in animal studies (including our work with murine neuroblastoma cells) are applicable to human receptors. Studies with murine neuroblastoma cells may help elucidate the mechanism whereby ethanol depletes brain levels of cyclic GMP. The prostaglandin and Beta-adrenergic receptors of human skin fibroblasts will be thoroughly characterized by biological assay (cyclic AMP production) and by radioligand binding assay prior to the studies with ethanol. The cyclic GMP studies with neuroblastoma cells will follow-up on our data showing a mixed type of inhibition (competitive and noncompetitive) by ethanol of receptor-mediated cyclic GMP synthesis and an adaptation to this effect of ethanol upon chronic exposure of cells to this alcohol. Since receptor-mediated cyclic GMP synthesis involves the turnover of phospholipids, effects of ethanol on the release of inositol phosphates and arachidonate acid will be studied.