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The receptors and actions of gonadotropin-releasing hormone (GnRH) were analyzed in cultured testicular cells from 20.5-day fetal rats, in which treatment with luteinizing hormone (LH) maintained Leydig cell steroidogenesis and gonadotropic responses for up to 2 weeks. Testicular GnRH receptors were present on the 5th postnatal day, but were not demonstrable in fetal testes or 2-day cultures thereof. However, GnRH receptors were readily detectable in 4-day cultured fetal testes and were increased by exposure to GnRH agonists. In LH-treated cultures, GnRH sites were reduced by about 50% and did not increase during incubation with GnRH agonists. In such cultures, GnRH agonists inhibited LH-dependent steroid production and abolished the acute testosterone response to human chorionic gonadotropin. The half-maximal inhibitory concentration of [D-Ala6]des-Gly10-GnRH-N-ethylamide (3 X 10(-10)M) was commensurate with its binding affinity for testis receptors (Kd = 1.4 X 10(-10)M). In contrast, GnRH agonists had no inhibitory effects in 2-day cultures prior to the detection of GnRH receptors. The expression of functional GnRH receptors during culture in the absence of gonadotropin and their suppression in LH-treated cultures suggest that pituitary gonadotropins exert a tonic inhibitory effect upon testicular GnRH receptors. The demonstrated inhibitory actions of GnRH on steroidogenesis, with the expression of GnRH receptors in cultured fetal testes and 5-day postnatal testes, indicate that GnRH agonists could influence the actions of gonadotropins upon Leydig cell function in the neonatal testis.

The inability of the fetal and immature Leydig cell to be desensitized by gonadotropin treatment, a characteristic of the adult cell, is attributed to the absence of an estrogen-mediated regulation of the androgen pathway. Cultures of fetal rat Leydig cells were employed to analyze this differential response. The fetal rat Leydig cells revealed low aromatization capacity, undetectable estradiol production, a low level of estrogen receptors, and a minimally detectable level of an estradiol-regulated protein. However, exogenous estradiol caused up-regulation of its own receptor, increase of an estradiol-regulated protein, and induction of a steroidogenic lesion at the microsomal level, resulting in decreased androgen production. This estrogen-mediated enzymatic inhibition resembles that observed in gonadotropin-desensitized adult Leydig cells. The absence of this regulation in fetal life is likely due to insufficient aromatase activity, with lack of consequent receptor-mediated estrogen action. The cultured fetal Leydig cell provides a useful model to elucidate the molecular mechanism involved in the development of estradiol-mediated desensitization.