BINDING-SITES FOR [H-3] MELATONIN IN HUMAN PLATELETS

A number of in vitro effects of melatonin on human platelets were revealed in previous studies. In order to examine whether high affinity binding sites for [H-3]-melatonin were present in membrane preparations of human platelets, a rapid filtration procedure through Whatman GFB paper was employed. Maximal melatonin binding was attained within 3 hr at 0-degrees-C. Scatchard analysis indicated a single population of binding sites with a dissociation constant (Kd) = 4.1 +/- 0.5 nM and maximal number of binding sites (Bmax) = 24.2 +/- 1.9 fmol/mg protein (mean +/- SEM of five experiments). When various indole analogs were tested for their ability to inhibit [H-3]-melatonin binding, the following Ki (nM) were obtained: 6-chloromelatonin (11.4), 2-iodomelatonin (22.0), melatonin (24.7), 5-methoxytryptophol (49.9), N-acetylserotonin (68.9), 6-hydroxymelatonin (78.2), 5-methoxytryptamine (184). Serotonin was a potent inhibitor of [H-3]-melatonin binding with a Ki = 20.6 nM. Except for 2-methylserotonin and alpha-methylserotonin, a number of serotonin agonists and antagonists tested did not affect melatonin binding to platelet membranes. Binding experiments carried out at either 0800 or 2000 did not reveal time-dependent differences in Kd or Bmax. The results suggest that high affinity melatonin acceptors are present in human platelets.