The effects of protein kinase-C agonists on parathyroid hormone release and intracellular free Ca2+ in bovine parathyroid cells.
High extracellular Ca2+ stimulates the accumulation of inositol trisphosphate and diacylglycerol in parathyroid cells and suppresses PTH release. Since diacylglycerol is an endogenous activator of protein kinase-C, these observations would suggest that activation of protein kinase-C is associated with inhibition of PTH release. However, phorbol esters, which stimulate protein kinase-C activity, have been reported to enhance PTH release. To clarify the role of protein kinase-C in the regulation of PTH secretion, we studied the responses of parathyroid cells to phorbol myristate acetate (PMA), bryostatin-1, and 1,2-dioctanoylglycerol (diC8). PMA and bryostatin-1 translocated protein kinase-C activity from the soluble to particulate fractions of cell homogenates. Phosphotransferase activity in the particulate fractions increased from 21 +/- 4% to 93 +/- 6% of the total activity after 10 min of exposure to PMA (10(-6) M) and from 21 +/- 2% to 69 +/- 2% after 5 min of exposure to bryostatin-1 (10(-7) M). These three structurally different agonists of protein kinase-C also altered the typical secretory response to Ca2+ in parathyroid cells. At 2.0 mM extracellular Ca2+, PMA (10(-6) M) bryostatin-1 (10(-7) M), and 1,2-dioctanoylglycerol (3 x 10(-4) M) blunted the suppressive effects of high Ca2+ on secretion, thus stimulating PTH release 252 +/- 45%, 122 +/- 20%, and 485 +/- 95% over control levels, respectively. However, at low extracellular Ca2+, these agents inhibited maximal PTH release. Since changes in the intracellular free Ca2+ concentration ([Ca2+]i) may be important in the control of PTH release, we investigated whether protein kinase-C agonists changed the relationship between extracellular Ca2+ and PTH release by affecting [Ca2+]i. In PMA-treated cells, the intracellular Ca2+ response to raising extracellular Ca2+ from 0.5 to 1.5 and 2.0 mM was reduced to 50 +/- 1% and 63 +/- 3% of that in control cells, respectively (P less than 0.005; n = 7-11). Specifically, PMA preincubation reduced the initial intracellular Ca2+ transient with raising extracellular Ca2+ from 0.5 to 2.0 mM and with adding 4.0 mM Sr2+. The sustained phase response to high Ca2+, but not to Sr2+, was also attenuated after incubation with PMA. We conclude that protein kinase-C agonists suppress PTH release at low extracellular Ca2+ and enhance PTH release at high extracellular Ca2+. The effects on secretion at high extracellular Ca2+ may be related to the ability of protein kinase-C agonists to change the sensitivity of [Ca2+]i to high extracellular Ca2+ in these cells.
Membreño
L
L
Endocrine Research Unit, San Francisco Veterans Administration Medical Center, California 94121.
Chen
T H
TH
Woodley
S
S
Gagucas
R
R
Shoback
D
D
D000818:Animals; D054713:Bryostatins; D002118:Calcium / Q000378:metabolism*; D002417:Cattle; D004075:Diglycerides / Q000494:pharmacology*; D005989:Glycerides / Q000494:pharmacology*; D066298:In Vitro Techniques; D007424:Intracellular Fluid / Q000378:metabolism; D007700:Kinetics; D007783:Lactones / Q000494:pharmacology*; D018942:Macrolides; D010280:Parathyroid Glands / Q000187:drug effects / Q000378:metabolism*; D010281:Parathyroid Hormone / Q000378:metabolism*; D010770:Phosphotransferases / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D054713:Bryostatins; D004075:Diglycerides; D005989:Glycerides; D007783:Lactones; D018942:Macrolides; D010281:Parathyroid Hormone; C044295:1,2-dioctanoylglycerol; C046785:bryostatin 1; D010770:Phosphotransferases; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium