Melatonin-mediated regulation of human MT₁ melatonin receptors expressed in mammalian cells

In mammals, the pineal hormone melatonin activates G protein-coupled MT₁ and MT₂ melatonin receptors. Acute exposure of recombinant MT₁ and MT₂ melatonin receptors to supraphysiological concentrations of melatonin differentially regulates these two receptors with the MT₂, but not the MT₁, exhibiting rapid desensitization and internalization. In the present study, we sought to determine whether prolonged exposure to supraphysiological and physiological concentrations of melatonin desensitized and/or internalized the MT₁ melatonin receptor. Using a Chinese hamster ovary (CHO) cell line stably expressing MT₁-FLAG or transiently expressing MT₁-green fluorescent protein (GFP) melatonin receptors, we found that prolonged exposure (8 h) to supraphysiological concentrations of melatonin (100 nM) significantly increased the number of MT₁ melatonin receptors and decreased the affinity (K_i) of melatonin for competition for 2-[ ¹²⁵I]iodomelatonin. A similar treatment also desensitized the MT ₁ melatonin receptor-mediated stimulation of [³⁵S] GTPγS binding, but did not internalize the receptor. In contrast, prolonged exposure to a concentration of melatonin mimicking nocturnal levels (400 pM) did not affect the number of MT₁ melatonin receptors, the affinity for melatonin, or the functional sensitivity of the receptor. We conclude that in vivo endogenous melatonin does not significantly affect the functional sensitivity of MT₁ melatonin receptors, however, exogenous melatonin taken therapeutically at doses above physiological levels could desensitize the receptor thereby affecting physiological responses mediated following activation of MT₁ melatonin receptors.