Differential regulation of gonadotropin subunit gene promoter activity by pulsatile gonadotropin-releasing hormone (GnRH) in perifused L beta T2 cells: role of GnRH receptor concentration.

The pulsatile release of GnRH by the hypothalamus is required to stimulate the pituitary-gonadal axis, and variations in GnRH pulse frequency are associated with differential synthesis and release of LH and FSH by pituitary gonadotropes. How gonadotropes differentiate between GnRH pulse frequencies and subsequently differentially regulate the expression of the LH beta and FSH beta genes remains to be determined. In the present study, using a perifusion system that allows us to replicate the GnRH pulsatility occurring in vivo, we have systematically characterized the effects of varying GnRH pulse frequencies on LH beta, FSH beta, alpha, and GnRH receptor (GnRHR) gene promoter stimulation in L beta T2 cells. We demonstrate that LH beta gene promoter activity is stimulated to the greatest extent at higher GnRH pulse frequencies, whereas the FSH beta gene promoter is preferentially stimulated at lower GnRH pulse frequencies, reflecting previous observations in primary rat pituitary cells in vivo and in vitro. By measuring GnRH binding, we demonstrate that cell-surface GnRHR number is increased at higher frequencies of pulsatile GnRH and that this increase precedes the differential regulation of LH beta and FSH beta gene promoter activity. To test the role of GnRHR number in mediating the differential effects of pulsatile GnRH, the rat GnRHR was overexpressed in L beta T2 cells, and the response to pulsatile GnRH was again assessed. Interestingly, although overexpression of GnRHR had no effect on the frequency-dependent regulation of LH beta, the induction of FSH beta gene promoter activity by pulsatile GnRH was reduced, and frequency dependence was abrogated. Our results demonstrate that L beta T2 cells represent a suitable model for the study of the differential regulation of gonadotropin subunit gene expression by pulsatile GnRH. Furthermore, our studies indicate that cell-surface GnRHR density is a critical mediator of this differential regulation.