Melatonin and testicular function: characterization of binding sites for 2-[125I]-iodomelatonin in immature rat testes.

Melatonin-binding sites in membrane preparation immature rat testes were demonstrated by utilizing 2-[125I]-iodomelatonin as a radioligand. Binding at these sites was found to be reversible, saturable, specific and of, high affinity. Scatchard analysis of the specific binding revealed an equilibrium binding constant (kd) of 215 +/- 23 pmol/L and a total number of binding sites (Bmax) of 0.94 +/- 0.1 fmol/mg protein. The Hill coefficient of 1.0 suggests a single class of 2-[125I]-iodomelatonin-binding site in the rat testes. The Kd value determined from kinetic analysis was 179 pmol/L, which is in close agreement with the value determined from equilibrium studies. In competition studies, the order of pharmacological affinity for 2-[125I]-iodomelatonin binding sites in the rat membrane testes was: melatonin > 6-hydroxymelatonin > N-acetylserotonin > 5-hydroxyindole-3-acetic acid > 5-hydroxytryptamine > 5-hydroxy-L-tryptophan > tryptamine > > 5-methoxytryptamine, 5-methoxyl-DL-tryptophan, D-L-tryptophan. The 2-[125I]-iodomelatonin binding was markedly reduced by guanine nucleotides; treatment with nonhydrolyzable GTP analog guanosine 5'-O-(3-thiotriphosphate) caused a 10-fold decrease in receptor affinity. In this paper, we report evidence indicating the presence of binding sites in immature rat tests, suggesting a possible direct role of melatonin on testicular steroidogenesis.