The existence of [125I]iodomelatonin-binding sites in the kidney of the chicken and other avian species has been demonstrated in the present study. [125I]Iodomelatonin-binding sites in membrane preparations of the chicken kidney were reversible, saturable, specific and of high affinity. Scatchard analysis of the specific binding revealed an equilibrium binding constant (Kd) of 30.3 +/- 5.1 pmol/l (n = 5) and a total number of binding sites (Bmax) of 2.89 +/- 0.24 fmol/mg protein (n = 5) at the middle of the light period (mid-light). The Hill coefficient approached 1.0, suggesting a single class of [125I]iodomelatonin-binding sites in the chicken kidney. The diurnal variation study showed that the Bmax was 52% higher at mid-light than that in the middle of dark period (1.90 +/- 0.33 fmol/mg protein, n = 4; p < 0.05), with no significant variation in Kd. The Kd value determined from kinetic analysis was 10.8 pmol/l in the mid-light birds, which was comparable to values determined from equilibrium studies. The competition inhibitor experiments demonstrated that [125I]iodomelatonin-binding sites were highly specific for melatonin with the following order of potency: 2-iodomelatonin > melatonin, 6-chloromelatonin > 6-hydroxymelatonin N-acetylserotonin >> 5-methoxytryptamine, 5-methoxytryptophol, 5-hydroxyindole-3-acetic acid, 5-hydroxytryptamine, 1-acetylindole-3-carboxaldehyde, 3-acetylindole, tryptamine, 5-methoxyindole-3-acetic acid, L-tryptophan, harmaline, acetylcholine, epinephrine and norepinephrine. Studies on the relative binding density of [125I]iodomelatonin to the kidney in different species of birds showed the following descending order: duck > chicken > pigeon > quail.(ABSTRACT TRUNCATED AT 250 WORDS)