MELATONIN AND TESTICULAR FUNCTION - CHARACTERIZATION OF BINDING-SITESFOR 2-[I-125]-IODOMELATONIN IN IMMATURE RAT TESTES

Melatonin-binding sites in membrane preparation of immature rat testes were demonstrated by utilizing 2-[I-125]-iodomelatonin as a radioliga nd. Binding at these sites was found to be reversible, saturable, spec ific and of, high affinity. Scatchard analysis of the specific binding revealed an equilibrium binding constant (k(d)) of 215 +/- 23 pmol/L and a total number of binding sites (B-max) of 0.94 +/- 0.1 fmol/mg pr otein. The Hill coefficient of 1.0 suggests a single class of 2-[I-125 ]-iodomelatonin-binding site in the rat testes. The K-d value determin ed from kinetic analysis was 179 pmol/L, which is in close agreement w ith the value determined from equilibrium studies. In competition stud ies, the order of pharmacological affinity for 2-[I-125]-iodomelatonin binding sites in the rat membrane testes was: melatonin > 6-hydroxyme latonin > N-acetylserotonin > 5-hydroxyindole-3-acetic acid > 5-hydrox ytryptamine > 5-hydroxy-L-tryptophan > tryptamine much greater than 5- methoxytryptamine, 5-methoxyl-DL-tryptophan, D-L-tryptophan. The 2-[I- 125]-iodomelatonin binding was markedly reduced by guanine nucleotides ; treatment with nonhydrolyzable GTP analog guanosine 5'-O-(3-thiotrip hosphate) caused a 10-fold decrease in receptor affinity. In this pape r, we report evidence indicating the presence of binding sites in imma ture rat testes, suggesting a possible direct role of melatonin on tes ticular steroidogenesis. (C) 1997 by Elsevier Science Inc.