Melatonin MT1 and MT2 receptors display different molecular pharmacologies only in the G‐protein coupled state

Background and Purpose Melatonin receptors have been extensively characterized regarding their affinity and pharmacology, mostly using 2-[125I]-melatonin as a radioligand. Although [3H]-melatonin has the advantage of corresponding to the endogenous ligand of the receptor, its binding has not been well described. Experimental Approach We characterized [3H]-melatonin binding to the hMT1 and hMT2 receptors expressed in a range of cell lines and obtained new insights into the molecular pharmacology of melatonin receptors. Key Results The binding of [3H]-melatonin to the hMT1 and hMT2 receptors displayed two sites on the saturation curves. These two binding sites were observed on cell membranes expressing recombinant receptors from various species as well as on whole cells. Furthermore, our GTPγS/NaCl results suggest that these sites on the saturation curves correspond to the G-protein coupled and uncoupled states of the receptors, whose pharmacology was extensively characterized. Conclusions and Implications hMT1 and hMT2 receptors spontaneously exist in two states when expressed in cell lines; these states can be probed by [3H]-melatonin binding. Overall, our results suggest that physiological regulation of the melatonin receptors may result from complex and subtle mechanisms, a small difference in affinity between the active and inactive states of the receptor, and spontaneous coupling to G-proteins.