The present application is a direct and natural extension of the research of the Principal Investigator over the preceding 10 years. As such, the main objective of the proposal and the overall objective of the Principal Investigator's research is to define the pathophysiologic mechanisms by which ethanol and/or its metabolism produces gonadal injury in the male. Therefore, the consequences of ethanol and acetaldehyde exposure upon Leydig cells and pituitary gonadotrophs in a tissue culture system will be evaluated. Utilizing metabolic blocking agents (4-methyl pyrazole methylene blue and penicillamine) the specific roles of ethanol, acetaldehyde, and redox changes will be assessed. In addition to the in vitro studies described above, testicular alcohol dehydrogenase will be isolated from rat testes and characterized. An antibody for the purified alcohol dehydrogenase will be produced and immunocytochemical methods will be utilized to identify the location of the enzyme within normal testicular tissues and within the testes obtained from chronic alcohol-fed rats. Finally, the consequences of ethanol exposure on testicular membranes (principally spermatazoa and their precursors) will be examined. For these experiments, the effect of chronic ethanol exposure in vivo utilizing the ethanol-fed rat model established in the Principal Investigator's laboratory will be utilized. In order to examine the effects upon Leydig cell membranes, short-term in vitro exposures of ethanol utilizing Leydig cell cultures of normal and tumor Leydig cells which are presently maintained within the laboratory of the Principal Investigator will be used for these studies as well. Particular attention will be paid to the role of peroxidation injury as a mechanism for ethanol-induced gonadal injury. Should evidence of peroxidation injury be found, the protective role of supplemental vitamin A and E in inhibiting such ethanol-induced peroxodation injury in gonadal tissues will be examined.
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