Prolactin and MA-10 Leydig cell steroidogenesis: biphasic effects of prolactin and signal transduction.
The present investigation was designed to study the direct role of PRL on testicular Leydig cell steroidogenesis, using the MA-10 murine Leydig tumor cell line as a model system. We have previously reported on the presence of specific PRL binding sites in those cells, and we now demonstrate the functionality of those sites and the biological responses induced by the binding of PRL. When cultured MA-10 cells were exposed for 24 h to increasing concentrations of PRL, washed, and then subjected to a 3-h human CG (hCG) stimulation test, a clear dose-dependent biphasic effect of PRL on the steroidogenic response was observed, even though PRL had no effect on MA-10 cell proliferation: at low PRL concentrations (0.1-10 ng/ml), hCG-induced steroidogenesis was stimulated (maximal stimulation by 1 ng/ml PRL being 200-250% of control); at higher concentrations, hCG-induced steroidogenesis was inhibited (60% inhibition was achieved by 1000 ng/ml PRL). When steroidogenesis was induced with various concentrations of cholera toxin, instead of hCG, no effect of the prior exposure to increasing concentrations of PRL was observed, indicating that PRL acts either at the level of the LH/hCG receptor or at some stage proximal to adenylate cyclase. Indeed, further study revealed that 24 or 72 h exposure of MA-10 cells to PRL caused a dose-dependent reduction in hCG binding. Thus, the maximal inhibition of 62% after 72 h with 500 ng/ml PRL, may explain, at least in part, the inhibitory effects of high PRL concentrations on hCG-induced progesterone secretion. Evidence demonstrating possible involvement of a pertussis toxin-(PT-)sensitive G protein in the signal transduction mechanism of PRL receptors is also presented: 1. GTP caused a dose-dependent reduction in affinity (Ka) of PRL binding by its receptors (from Ka = 1.66 +/- 0.2 x 10(9) M(-1) for control MA-10 cell membranes to Ka 3.03 +/- 0.6 x 10(8) M(-1) for membranes incubated with 8 mM GTP). 2. Prior exposure of MA-10 cells to PRL (10 pg/ml) caused a significant reduction in the ability of a 44-kDa membrane protein to undergo PT-induced [32P]ADP-ribosylation. These results demonstrate that MA-10 Leydig cells possess highly specific and biologically functional PRL receptors mediating direct and dose-dependent biphasic effects of PRL on hCG-induced progesterone secretion. These cells thus offer a suitable model to study the mechanism(s) of PRL action and signal transduction of its receptor on a physiologically relevant differentiated function.
Weiss-Messer
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Department of Pharmacology, The Bruce Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa, Israel.
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Barkey
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D000818:Animals; D002116:Calcitonin / Q000494:pharmacology; D002417:Cattle; D002455:Cell Division / Q000187:drug effects; D006063:Chorionic Gonadotropin / Q000378:metabolism / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D019204:GTP-Binding Proteins / Q000502:physiology; D006801:Humans; D007985:Leydig Cells / Q000187:drug effects* / Q000378:metabolism*; D008297:Male; D051379:Mice; D011374:Progesterone / Q000378:metabolism; D011388:Prolactin / Q000494:pharmacology*; D012756:Sheep; D015398:Signal Transduction; D013256:Steroids / Q000096:biosynthesis*; D013997:Time Factors; D014407:Tumor Cells, Cultured
D006063:Chorionic Gonadotropin; D013256:Steroids; D011374:Progesterone; D011388:Prolactin; D002116:Calcitonin; D019204:GTP-Binding Proteins