Endocrine Abstracts

The pulsatile release of GnRH regulates the synthesis and secretion of pituitary FSH and LH. Two transcription factors, the cAMP response element binding protein (CREB) and inducible cAMP early repressor (ICER), have been implicated in the regulation of rat FSHβ gene expression. We hypothesized that CREB and ICER are activated by distinct signaling pathways in response to pulsatile GnRH to modulate FSHβ gene expression, which is preferentially stimulated at low (every 120 min) vs high (every 30 min) pulse frequencies. Using the LβT2 gonadotrope-derived cell line, we have shown previously that at low GnRH pulse frequencies, PKA is activated, leading to CREB phosphorylation and subsequent FSHβ transcription. In contrast, ICER expression is preferentially stimulated by GnRH at high pulse frequencies. In static culture, GnRH stimulation of LβT2 cells resulted in a time-dependent increase in ICER mRNA at 12 and 24 h, as measured by qRT-PCR. Pharmacological blockade of MEKI/II with two selective inhibitors, U0126 and PD0325901, significantly attenuated GnRH-stimulated ICER induction in a dose-dependent fashion, whereas PKC (GF109203X), CamKII (KN-93) and PKA (H89) inhibitors had minimal effects. Similarly, MEKI/II inhibition attenuated GnRH induction of ICER protein as determined by western blot analysis. Importantly, MEKI/II inhibition with PD0325901 in perifused LβT2 cells stimulated with pulsatile GnRH abrogated ICER induction at high GnRH pulse frequency, whereas stimulation of ICER at low GnRH pulse frequency was unaffected. MEKI/II inhibition abrogated GnRH stimulation of FSHβ expression at both high and low pulse frequencies, suggesting that MEK signaling pathways have additional effects on GnRH regulation of FSHβ, beyond those mediated by ICER. Taken together, we conclude that the signaling pathways mediating GnRH activation of CREB and ICER are distinct. The PKA pathway mediates GnRH-stimulated CREB phosphorylation, whereas GnRH induction of ICER occurs via MAPK pathways, contributing to the decoding of the pulsatile GnRH to regulate FSHβ expression.