P-207 Molecular mechanisms of melatonin action in human reproductive system – implications for assisted reproduction

Abstract

Abstract Study question What is the potential role of melatonin and melatonin receptors activation on the reproductive function in humans? Summary answer The presence of membrane melatonin receptors in male gametes and female hGLCs suggests a role for melatonin on motility performance of mature spermatozoa and oogenesis. What is known already The interest in the influence of the external environment on the reproductive performance in humans facilitates reevaluation of its main mediators in the body. One of the most important among them is melatonin circadian secretion by the pineal gland, a major synchronizer of the diurnal rest-activity cycle. Activation of melatonin-linked GPCR receptors by melatonin is coupled to inhibition of cAMP accumulation in target cells, which makes research on membrane receptor-mediated signaling in human reproductive system very attractive. Study design, size, duration Pilot study on the presence of MT1 and MT2 receptors in human spermatozoa and human granulosa luteinized cells (hGLCs) and the presence and localization of α- and ß-tubulin, melatonin receptors (MT1, MT2) and P450 aromatase in hGLCs. Participants/materials, setting, methods Human donor samples (spermatozoa and granulosa lutein cells (hGLC) were collected after informed consent signed from patients recruited at IVF clinics in Sofia. Two-step quantitative real time RT-PCR (qRT-PCR) was used to compare individual variations between donors. Selected proteins in samples and derived cell cultures like α- and ß-tubulin, melatonin receptors (MT1, MT2) and P450 aromatase were studied by Western blotting and localization visualized by confocal immune fluorescence. Main results and the role of chance PCR amplification of cDNA in the samples resulted in products of the expected size of 516 bp corresponding to the MTNR1A gene. In purified spermatozoa, intense expression of a MT1 receptor was found in the post-acrosomal region and the neck. In hGLC cultures up-regulation of melatonin receptors after in-vitro melatonin treatment was observed both by protein expression and densitometry of immune fuorescence. Our results demonstrate the presence in situ of membrane melatonin receptor in competent human male gametes and female hGLCs. The observed localization of the receptor protein suggests a role for melatonin on motility performance of mature spermatozoa, which may have several important implications to assisted reproductive techniques. Limitations, reasons for caution The discrete functional responses and the transduction pathways linked to specific receptor activation by melatonin in human reproductive system have further to be elucidated. Wider implications of the findings Future studies could reveal the potential therapeutic role of Melatonin in infertility patients. Trial registration number Not applicable