Specific binding of 2-[125I]melatonin by partially purified membranes of rat thymus

Melatonin binding sites were characterized in partially purified rat thymus membranes. The specific binding of 2-[125I]iodomelatonin ([125I]MEL) to thymus membranes was dependent on time and temperature, stable, saturable, and reversible. Concentration-dependent binding of [125I]MEL to thymus membranes was saturable and resulted in a linear Scatchard plot, suggesting binding to a single class of binding sites. The Kd for this single site was 0.47 nM with a binding capacity of 1.01 pM. In competition studies, the specific binding of [125I]MEL to thymus membranes was inhibited by increasing concentrations of native melatonin. Scatchard analysis showed that, unlike in saturation studies with [125I]MEL, data were compatible with the existence of two classes of binding sites: a high-affinity site with a Kd of 1.72 +/- 0.25 nM and a binding capacity of 1.40 +/- 0.18 pM, and a low-affinity site with a Kd of 1226 +/- 325 nM and a binding capacity of 460 +/- 87 pM. Interestingly, Kd and BC values of the high-affinity binding site described by competition studies are similar to those obtained by saturation studies with [125I]MEL. Binding of [125I]MEL to thymus membranes was specific as indicated by the fact no other melatonin precursor or derivative was as potent as melatonin in inhibiting the binding of [125I]MEL to membranes. Results strongly suggest that melatonin is involved in regulation of thymus activity.