Abstract

The hormone melatonin mediates a variety of physiological functions in mammals through activation of pharmacologically distinct MT₁ and MT₂ G protein-coupled melatonin receptors. We therefore sought to investigate how the receptors were regulated in response to short melatonin exposure. Using 2-[¹²⁵I]iodomelatonin binding, cAMP functional assays, and confocal microscopy, we demonstrated robust differences in specific 2-[¹²⁵I]iodomelatonin binding, receptor desensitization, and cellular trafficking of hMT₁ and hMT₂melatonin receptors expressed in Chinese hamster ovary (CHO) cells after short (10-min) exposure to melatonin. Exposure to melatonin decreased specific 2-[¹²⁵I]iodomelatonin binding to CHO-MT₂ cells (70.3 ± 7.6%, n = 3) compared with vehicle controls. The robust decreases in specific binding to the hMT₂ melatonin receptors correlated both with the observed functional desensitization of melatonin to inhibit forskolin-stimulated cAMP formation in CHO-MT₂ cells pretreated with 10 nM melatonin (EC₅₀ of 159.8 ± 17.8 nM, n = 3, p < 0.05) versus vehicle (EC₅₀ of 6.0 ± 1.2 nM, n= 3), and with the arrestin-dependent internalization of the receptor. In contrast, short exposure of CHO-MT₁ cells to melatonin induced a small decrease in specific 2-[¹²⁵I]iodomelatonin binding (34.2 ± 13.0%,n = 5) without either desensitization or receptor internalization. We conclude that differential regulation of the hMT₁ and hMT₂ melatonin receptors by the hormone melatonin could underlie temporally regulated signal transduction events mediated by the hormone in vivo.