MECHANISM OF ACTION OF GONADOTROPIN-RELEASING-HORMONE UPON GONADOTROPIN ALPHA-SUBUNIT MESSENGER-RNA LEVELS IN THE ALPHA-T3-1 CELL-LINE - ROLE OF CA2-KINASE-C( AND PROTEIN)

Addition of [D-Trp(6)]gonadotropin-releasing hormone (GnRHa) to alpha T3-1 cells induced a very rapid response upon gonadotropin alpha-subun it mRNA which was detected after 30-60 min and was abolished by pretre atment with actinomycin D. A similar response was obtained with the pr otein kinase C (PKC) activator phorbol 12-myristate 13-acetate (PMA), or the Ca2+ ionophore, ionomycin. GnRHa (10 nM) also stimulated a seco ndary rise in alpha-subunit mRNA levels between 12 and 24 h of incubat ion. No additivity was obtained (at 60 min) upon the combined addition of GnRHa and PMA, GnRHa and ionomycin, or PMA and ionomycin. The effe ct of GnRHa upon alpha-subunit mRNA was blocked by the PKC inhibitors staurosporine or GF 109203X. Down-regulation of endogenous PKC activit y resulted in inhibition of the stimulatory effect of gonadotropin-rel easing hormone (GnRH), PMA and ionomycin. Removal of extracellular Ca2 + abolished the effect of GnRHa and PMA upon alpha-subunit mRNA levels . Interestingly PMA and ionomycin had no effect on alpha-subunit mRNA. levels at 24 h of incubation; however, the combined addition of the d rugs mimicked the late phase of GnRHa (10 nM) action. The data provide evidence that PKC and Ca2+ are involved in mediating the early and th e late responses of GnRHa upon alpha-subunit mRNA elevation and that d ifferential cross-talk exists between the messengers.