The involvement of protein kinase C (PKC) in GnRH action is still a matter of controversy. We have conducted a comparative study of LH and FSH release in response to GnRH and to phorbol ester myristate acetate (PMA), an activator of PKC, by rat pituitary cells maintained in culture. The effect of E2 pretreatment coupled or not with PKC depletion was also studied. Different kinetics in the response of LH and FSH to GnRH were observed, suggesting that the intracellular pathways involved in the release process of the two hormones were somewhat different. Moreover, PMA (10 nM) stimulated LH release greatly and FSH release only slightly. Intracellular PKC depletion, obtained by a prolonged treatment (18 h) of the cells with PMA (1 microM), produced different results according to the endocrine status of the pituitary cells. GnRH (10 nM)-induced LH release was significantly decreased in PKC-depleted cells from proestrous females. For PKC-depleted cells from OVX females, it was decreased significantly only when cells had been pretreated by E2. These results suggest that the modulation of LH secretion by E2 involves PKC activation. FSH release was poorly stimulated by PMA; but, under any conditions, PKC depletion did not affect GnRH-induced FSH release.

No abstract

No abstract

No abstract

Although enhancement of GnRH-stimulated luteinizing hormone (LH) release by estradiol (E2) has been established, it is not known at what stages of the process of transduction E₂ acts. We investigated the release of LH in response to GnRH and to Bay K 8644, an activator of L-type calcium channels, in a culture of pituitary cells obtained from ovariectomized females, these cells having being treated or not with E₂ (OVX + E₂ and OVX). We studied the effects of D600, an antagonist of T- and L-type calcium channels, and PN 200-110, an antagonist of L-type calcium channels. The effects of the latter were studied in protein kinase C-depleted cells in order to investigate the possible phosphorylation of these channels. D600 caused a decrease in GnRH-stimulated LH release in OVX and OVX + E₂ cells. However, this decrease was greater in OVX + E₂ cells, suggesting that at least one type of calcium channels may be involved as a result of treatment with E₂. We confirmed the involvement of L-type calcium channels in the action of GnRH since the GnRH-stimulated LH release was enhanced in the presence of Bay K 8644 in OVX cells. Bay K 8644 alone increased basal LH in a dose-dependent manner only in OVX + E₂ cells. PN 200-110 induced a decrease of GnRH-stimulated LH release only in OVX + E₂ cells. These results suggest that L-type calcium channels are activated in E₂-treated cells. The dose-dependent decrease cused by PN 200-110 in OVX + E₂ cells disappeared in the OVX + E₂ PKC-depleted cells. This result was confirmed with Bay K 8644 and suggests a phosphorylation of dihydropyridine-sensitive calcium channels by protein kinase C.