Prolactin signaling in porcine adrenocortical cells: involvement of protein kinases.
Prolactin (PRL) was found to have a stimulatory effect on adrenal steroidogenesis in vivo and in vitro in several species including pigs. PRL signal transduction pathways, however, in adrenocortical cells are poorly recognized. Therefore, the goal of this paper is to ascertain the involvement of protein kinase C (PKC) and tyrosine kinases in PRL signaling in porcine adrenal cortex. Adrenals were harvested from locally slaughtered mature gilts. Cortical cells were dispersed by sequential treatment with collagenase. The cells were seeded into 24-well culture plates at a density of 3 x 10(5)/mL. Cells were incubated with or without PRL (500 ng/mL), ACTH (5 nM--a positive control), tyrosine kinase inhibitor--genistein (1; 2.5 or 5 microM), PKC inhibitor--sphingosine (20-1000 nM) and PKC activators--diacylglycerol (DiC8; 10-100 microM) and phorbol ester (PMA; 1-1000 nM). All incubations were performed for 8 h (95% air and 5% CO(2), 37 degrees C). PRL and ACTH (P < 0.05) increased cortisol and androstenedione (A(4)) secretion. DiC8 and PMA mimicked the stimulatory effect of PRL. Sphingosine (P < 0.05) suppressed basal and PRL-stimulated steroid secretion. Genistein inhibited (P < 0.05) PRL-stimulated cortisol secretion and enhanced (P < 0.05) basal and PRL-stimulated A(4) secretion. Moreover, PKC activation was assessed by measuring the specific association of [3H]phorbol dibutyrate ([3H]PDBu) with adrenocortical cells after treatment with PRL or ionomycin (a positive control). PRL (within 2-3 min) and ionomycin (within 2-5 min) increased (P < 0.05) specific binding of [3H]PDBu to the porcine adrenocortical cells. In addition, PRL did not augment the cortisol and A(4) secretion by PKC-deficient adrenocortical cells. In conclusion, presented results support the hypothesis that PKC and tyrosine kinases are involved in PRL signaling in adrenocortical cells in pigs. Moreover, activation of PKC is associated with the increased secretion of cortisol and A(4).
Kaminska
B
B
Department of Animal Physiology, University of Warmia and Mazury in Olsztyn, Oczapowskiego 5, 10-718 Olsztyn, Poland. tkam@uwm.edu.pl
Ciereszko
R E
RE
Opalka
M
M
Dusza
L
L
D000302:Adrenal Cortex / Q000166:cytology / Q000187:drug effects / Q000201:enzymology / Q000502:physiology*; D000324:Adrenocorticotropic Hormone / Q000378:metabolism / Q000502:physiology; D000735:Androstenedione / Q000378:metabolism; D000818:Animals; D004075:Diglycerides / Q000494:pharmacology; D020536:Enzyme Activators / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D019833:Genistein / Q000494:pharmacology; D006854:Hydrocortisone / Q000378:metabolism; D015759:Ionomycin / Q000494:pharmacology; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D011388:Prolactin / Q000494:pharmacology* / Q000502:physiology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology; D013110:Sphingosine / Q000494:pharmacology; D013552:Swine / Q000378:metabolism / Q000502:physiology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D004075:Diglycerides; D020536:Enzyme Activators; D004791:Enzyme Inhibitors; C044295:1,2-dioctanoylglycerol; D015240:Phorbol 12,13-Dibutyrate; D000735:Androstenedione; D015759:Ionomycin; D000324:Adrenocorticotropic Hormone; D011388:Prolactin; D019833:Genistein; D011505:Protein-Tyrosine Kinases; D011493:Protein Kinase C; D013110:Sphingosine; D013755:Tetradecanoylphorbol Acetate; D006854:Hydrocortisone