Interactions between signaling pathways in mediating GnRH-stimulated GTH release from goldfish pituitary cells: protein kinase C, but not cyclic AMP is an important mediator of GnRH-stimulated gonadotropin secretion in goldfish.
In the goldfish, it has been proposed that gonadotropin (GTH) release induced by GTH-releasing hormone (GnRH) involves Ca2+ entry through voltage-sensitive Ca2+ channels (VSCC), protein kinase C (PKC) activation, and arachidonic acid (AA) metabolism, but not cyclic AMP (cAMP) action. However, cAMP appears to mediate GnRH action in other teleosts. In this study, the relative importance of PKC and cAMP in mediating GnRH action in goldfish was studied using primary cultures of dispersed pituitary cells. Consistent with an involvement of PKC in GnRH action, the GTH responses to the PKC activating tetradecanoyl phorbol acetate (TPA), salmon (s)GnRH, and chicken (c)GnRH-II were inhibited by two selective PKC inhibitors, calphostin C, and staurosporine. Furthermore, GTH release responses induced by sGnRH or cGnRH-II were not additive to responses stimulated by the PKC-activating diglyceride DiC8, in either long-term static incubation or acute perifusion experiments. In static incubation studies, the GTH responses to sGnRH and DiC8 were potentiated by the VSCC agonist Bay K 8644, suggesting that VSCC participates in both PKC and GnRH action. Concentrations of K+ < 100 mM did not elicit GTH secretion when tested alone, but were effective in stimulating GTH release in the presence of subthreshold doses of DiC8 or TPA. This suggests that minimal activation of PKC greatly enhances the effectiveness of Ca2+ influx to increase GTH secretion. Taken together, these results indicate that PKC is an important mediator of GnRH-induced, VSCC-dependent GTH release. In contrast to the involvement of PKG, cAMP-dependent mechanisms showed no evidence of direct participation in GnRH-induced GTH release in goldfish. In static incubation studies, the GTH responses to sGnRH and cGnRH-II were not affected by H89, a cAMP-dependent protein kinase (PKA) inhibitor. Furthermore, the GTH release stimulated by cAMP was additive to the response to sGnRH, cGnRH-II, DiC8, TPA, or AA. However, compared to the response to forskolin or TPA alone, combinations of forskolin and TPA resulted in a potentiated increase in GTH release. The acute GTH response to forskolin was also enhanced by DiC8. Thus, cAMP-dependent mechanisms may constitute an independent pathway that interacts positively with GnRH-dependent mechanisms in the regulation of GTH release.
Jobin
R M
RM
Department of Biological Sciences, University of Alberta, Edmonton, Canada.
Van Goor
F
F
Neumann
C M
CM
Chang
J P
JP
D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester / Q000494:pharmacology; D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D016718:Arachidonic Acid / Q000494:pharmacology; D002120:Calcium Channel Agonists / Q000494:pharmacology; D002478:Cells, Cultured; D002645:Chickens; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism* / Q000494:pharmacology; D004075:Diglycerides / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D006054:Goldfish / Q000378:metabolism*; D007987:Gonadotropin-Releasing Hormone / Q000378:metabolism / Q000494:pharmacology*; D006065:Gonadotropins, Pituitary / Q000096:biosynthesis / Q000378:metabolism*; D008297:Male; D008863:Microspheres; D009281:Naphthalenes / Q000494:pharmacology; D010902:Pituitary Gland / Q000166:cytology / Q000378:metabolism*; D011188:Potassium / Q000494:pharmacology; D011493:Protein Kinase C / Q000187:drug effects / Q000378:metabolism*; D012474:Salmon; D015398:Signal Transduction; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D000470:Alkaloids; D002120:Calcium Channel Agonists; D004075:Diglycerides; D004791:Enzyme Inhibitors; D006065:Gonadotropins, Pituitary; D009281:Naphthalenes; C044295:1,2-dioctanoylglycerol; D005576:Colforsin; D016718:Arachidonic Acid; D007987:Gonadotropin-Releasing Hormone; D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; D000242:Cyclic AMP; D011493:Protein Kinase C; D019311:Staurosporine; C058819:calphostin C; D013755:Tetradecanoylphorbol Acetate; D011188:Potassium