Regulation of gonadotropin-releasing hormone and its receptor gene expression by 17 beta-estradiol in cultured human granulosa-luteal cells

There is evidence that GnRH and its binding sites are expressed in numerous extrapituitary tissues, including the primate ovary. However, the factors that regulate ovarian GnRH and its receptor (GnRH-R) remain poorly characte rized. Since gonadal steroids are key regulators of ovarian functions, the present study investigated the role of 17 beta-estradiol (E-2) in regulatin g GnRH and GnRH-R messenger RNA (mRNA) from human granulosa-luteal cells (h GLCs). RT-PCR was used to isolate the ovarian GnRH-R transcript equivalent to the full-length coding region in the pituitary from hGLCs. Sequence anal ysis revealed that the ovarian GnRH-R mRNA is identical to its pituitary co unterpart. Basal expression studies indicated that GnRH and GnRH-R mRNA lev els significantly increased with time in vitro, reaching levels of 160% and 170% on day 8 and 10 of culture, respectively (P < 0.05). Treatment with v arious concentrations of estradiol (1-100 nM) for 24 h resulted in a dose-d ependent decrease (P < 0.05) in GnRH and GnRH-R mRNA levels. Time course st udies indicated that short-term treatment (6 h) with E-2 (1 nM) had no sign ificant effect on GnRH mRNA levels, while long-term treatment (48 h) with E -2 resulted in a 40% decrease (P < 0.001) in GnRH mRNA levels. In contrast, GnRH-R mRNA levels exhibited a biphasic pattern, such that a short-term tr eatment (6 h) with E-2 increased GnRH-R mRNA levels by 20% (P < 0.05), wher eas long-term treatment (48 h) resulted in a 60% decrease (P < 0.001) in Gn RH-R expression in hGLCs. Cotreatment of estradiol and tamoxifen blocked th e E-2 induced-regulation of GnRH and its receptor mRNAs, indicating that th e E-2 effect was mediated through its receptor. Tn summary, our studies dem onstrate that the ovary possesses an intrinsic GnRH axis that is regulated during luteinization in vitro, and that E-2 is capable of regulating GnRH a nd its receptor in the human ovary.