Ethanol-induced inhibition of testosterone biosynthesis in rat Leydig cells: role of mitochondrial substrate shuttles and citrate.
The mechanisms by which ethanol inhibits testicular testosterone synthesis in rats were studied in vitro using isolated rat Leydig cells. The ethanol-induced inhibition was reversed by 4-methylpyrazole, an alcohol dehydrogenase inhibitor, suggesting that ethanol metabolism was responsible for this inhibition. L-glutamate and pyruvate, when added to the Krebs-Ringer incubation medium, reversed the inhibition by ethanol. The membrane glutamate receptor agonists kainic acid and quisqualic acid had no effects, indicating metabolic mechanisms for the L-glutamate action. This was verified also by observations that the metabolic transaminase inhibitors aminooxyacetate and cycloserine inhibited testosterone synthesis. In the amino acid supplemented Krebs-Ringer, pyruvate could not fully prevent inhibition by ethanol alone, but addition of L-glutamate to this medium abolished ethanol-induced inhibition. Experiments performed using a new inhibitor of testosterone biosynthesis in intact Leydig cells, triethylcitrate, indicated that active citrate metabolism, and/or efflux from mitochondria, was essential for the steroidogenic pathway from pregnenolone to testosterone in the smooth endoplasmic reticulum. The early steps of hCG stimulation before pregnenolone formation were most sensitive to its effect. Our results indicate that the inhibition of steroidogenesis by ethanol results from decreased availability of the metabolites involved in the substrate shuttles maintaining the NAD(P)H redox states between the mitochondrial and the smooth endoplasmic reticulum compartments, and that the inhibition can be overcome by a proper selection of exogenous sources for these metabolites.
Orpana
A K
AK
Department of Clinical Chemistry, University of Helsinki, Meilahti Hospital, Finland.
Härkönen
M
M
Eriksson
C J
CJ
D000437:Alcoholism / Q000473:pathology*; D000818:Animals; D002478:Cells, Cultured; D002951:Citrates / Q000494:pharmacology*; D000431:Ethanol / Q000493:pharmacokinetics / Q000633:toxicity*; D005971:Glutamates / Q000494:pharmacology*; D018698:Glutamic Acid; D007985:Leydig Cells / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism; D011773:Pyruvates / Q000494:pharmacology*; D019289:Pyruvic Acid; D051381:Rats; D013739:Testosterone / Q000096:biosynthesis*
D002951:Citrates; D005971:Glutamates; D011773:Pyruvates; D000431:Ethanol; D018698:Glutamic Acid; D013739:Testosterone; D019289:Pyruvic Acid