Direct Action of Melatonin in Human Granulosa-Luteal Cells

Abstract

The direct involvement of melatonin in modulation of ovarian steroidogenesis, the high levels of melatonin found in human follicular fluid, and the presence of melatonin binding sites in the ovary led us to hypothesize that melatonin acts as a modulator of ovarian function. In contrast to the hypothalamus and pituitary, the mechanism of melatonin action at the level of the ovary is still poorly understood. In the present study, we investigated the gene expression of the two different forms of melatonin receptors in human granulosa-luteal cells, using RT-PCR. PCR products corresponding to the expected sizes of the melatonin receptor subtypes, mt(1)-R and MT(2)-R, were obtained from granulosa-luteal cells, and the authenticity of the PCR products was confirmed by Southern blot hybridization with cDNA probes. Subsequent cloning and sequence analysis revealed that the ovarian mt(1)-R and MT(2)-R cDNAs are identical to their brain counterparts. Because gonadotropins and GnRH acting through specific receptors in the human ovary regulate cellular functions, we investigated the role of melatonin in the regulation of FSH receptor, LH receptor, GnRH, and GnRH receptor levels. Treatment with melatonin (10 pM-100 nM) significantly increased LH receptor mRNA levels without altering the expression of the FSH receptor gene. Both GnRH and GnRH receptor mRNA levels were significantly decreased, to 61% and 45% of control levels, respectively, after melatonin treatment. Melatonin treatment alone had no effect on basal progesterone production but enhanced the effects of human CG-stimulated progesterone production. Because MAPKs are activated in response to a diverse array of extracellular stimuli leading to the regulation of cell growth, division, and differentiation, and because melatonin has been shown to modulate cellular proliferation and differentiation, in this study, we demonstrated that melatonin activated MAPK in a dose- and time-dependent manner. In summary, our studies demonstrate, for the first time, that melatonin can regulate progesterone production, LH receptor, GnRH, and GnRH receptor gene expression through melatonin receptors in human granulosa-luteal cells, which may be mediated via the MAPK pathway and activation of Elk-1. Our results support the notion that melatonin plays a direct role in regulating ovarian function.

Melatonin and the ovary: physiological and pathophysiological implications

“…Melatonin is concentrated in human ovarian FF relative to the level in plasma (20)(21)(22), and melatonin alters granulosa cell (GC) steroidogenesis and follicular function in hen (44), hamster (45), and humans (8,(46)(47)(48). Melatonin binding sites are detected in the membrane fraction of human GCs (7) and both MT1 and MT2 melatonin receptors were identified in human GC/luteal cells (8,49) and in rat ovaries (antral follicles and corpus luteum [CL]) (50).…”

“…This is supported in part by the demonstration of melatonin receptors in the suprachiasmatic nuclei in the brain and pars tuberalis of the pituitary (5,6). Also, the demonstration of melatonin receptors in the ovary indicates multiple sites at which melatonin may influence the reproductive system (7,8).…”

Role of photoperiod and melatonin in the regulation of ovarian functions in Indian carp Catla catla: basic information for future application

“…Extra-hypothalamic pathway of action of melatonin on oocyte maturation Demonstration of melatonin binding sites in mammalian reproductive organs (Yie et al 1995;Woo et al 2001) raised the possibility of direct action of this indole amine on the gonad. However, the results of previous in vitro studies on the oocytes of carp C. carpio (Popek et al 1991(Popek et al , 1996 and killifish F. heteroclitus (Cerda et al 1997) did not show any effect of melatonin on oocyte maturation.…”

Effects of melatonin on histomorphology and on the expression of steroid receptors, VEGF, and PCNA in ovaries of pinealectomized female rats

“…Some authors (10,11) have shown evidence of melatonin's direct action on the ovaries. In the reproductive system, melatonin may interact with sex steroids and interfere with ovarian function.…”