Effect of inhibin on activators of protein kinase-C and calcium-mobilizing agents which stimulate secretion of gonadotropins in vitro: implication of a postgonadotropin-releasing hormone receptor effect of inhibin on gonadotropin release.
To further characterize the subcellular mechanisms by which inhibin suppresses GnRH-stimulated gonadotropin release, anterior pituitary cells from adult male Sprague-Dawley rats were treated on day 2 of culture with or without purified 31-kDa bovine inhibin (1-300 pM) for a further 3 days. On day 5, the pretreated cells were washed and incubated in the absence or presence of various secretagogues for 4 h. At the end of the stimulation, the media were saved, and cells were lysed for measurement of both extracellular and intracellular FSH and LH by specific RIAs. Released hormone was expressed as the proportion of total (released plus intracellular) hormone that was available for release in each case. This manipulation of the data corrects for the differential effect of the inhibin pretreatments to suppress intracellular FSH before the stimulation period. Pretreatment for 3 days with inhibin suppressed the proportions of FSH and LH released during 4 h in response to 1) phorbol 12-myristate 13-acetate (100 nM), an activator of protein kinase-C, by maxima of 48% and 53% with inhibin median inhibitory concentrations (IC50) of 17 and 18 pM, respectively; 2) mezerein (100 nM), another type of activator of protein kinase-C, by maxima of 49% and 50% with inhibin IC50 of 19 and 20 pM, respectively; 3) high extracellular K+ (60 mM) by 42% (P less than 0.01) and 38% (P less than 0.01), respectively, with 130 pM inhibin; 4) the calcium ionophore, A23187 (100 microM) by maxima of 54% and 56% with IC50 of 18 and 17 pM, respectively; and 5) GnRH (10 nM) by maxima of 52% and 53% with IC50 of 18 and 19 pM, respectively. However, inhibin had no effect on the proportional release of gonadotropin induced by melittin, an activator of phospholipase-A2. Finally, inhibin had no effect on ACTH release either under basal conditions or in response to CRF (10 nM), phorbol 12-myristate 13-acetate (100 nM), or A23187 (100 microM). We conclude that inhibin suppresses the stimulated release of hormones from gonadotrophs in part by a mechanism common to both gonadotropins that is independent of the previously described inhibitory effect of inhibin on the GnRH receptor. The results are consistent with an action at a site(s) beyond the GnRH receptor, such as protein kinase-C and calmodulin.
Wang
Q F
QF
Prince Henry's Institute of Medical Research, South Melbourne, Victoria, Australia.
Farnworth
P G
PG
Burger
H G
HG
Findlay
J K
JK
D000324:Adrenocorticotropic Hormone / Q000378:metabolism; D000818:Animals; D016718:Arachidonic Acid; D001095:Arachidonic Acids / Q000494:pharmacology; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D004224:Diterpenes; D004789:Enzyme Activation / Q000187:drug effects; D005640:Follicle Stimulating Hormone / Q000378:metabolism*; D007987:Gonadotropin-Releasing Hormone / Q000494:pharmacology; D007265:Inhibins / Q000494:pharmacology*; D007986:Luteinizing Hormone / Q000378:metabolism*; D008297:Male; D008555:Melitten / Q000494:pharmacology; D010903:Pituitary Gland, Anterior / Q000187:drug effects / Q000378:metabolism; D011188:Potassium / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D011966:Receptors, LHRH / Q000502:physiology*; D013729:Terpenes / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D001095:Arachidonic Acids; D004224:Diterpenes; D011966:Receptors, LHRH; D013729:Terpenes; D008555:Melitten; D016718:Arachidonic Acid; D007987:Gonadotropin-Releasing Hormone; C011309:mezerein; D000001:Calcimycin; D007265:Inhibins; D000324:Adrenocorticotropic Hormone; D007986:Luteinizing Hormone; D005640:Follicle Stimulating Hormone; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D011188:Potassium; D002118:Calcium