Previous studies have demonstrated that parathyroid hormone (PTH) and human α-thrombin mobilize intracellular calcium from distinct pools in UMR 106-H5 rat osteosarcoma cells. The present studies were designed to explore the molecular basis of this differential signaling. Maximally effective concentrations of both PTH (240 nM) and thrombin (10 U/ml) produced a rapid intracellular free calcium (Ca(i)++) transient (a 2- to 3-fold increase) that was inhibited by pretreatment with the phospholipase C inhibitor 1-[6- [[17β-3-methoxyestra-1,3,5(10)-trien-17-yl]amino]hexyl-1H-pyrrole-2,5-dion e (U73,122) in a dose-dependent manner (IC50 = 3 μM). Inhibition by U73,122 was not associated with a change in PTH-stimulated adenylate cyclase activity, whereas inositol phosphate accumulation, detected only in response to thrombin, was inhibited 23 to 45%. Prior exposure of cells for 5 min with the protein kinase C activators phorbol 12-myristate 13-acetate (8-80 nM) and phorbol 12,13-dibutyrate (80 nM) weakly inhibited (≤ 30%) the peak Ca(i)++ increase in response to thrombin but completely blocked the Ca(i)++ response to PTH. In contrast, 12-myristate 13-acetate produced a 1.55-fold increase in the maximal stimulatory effect of PTH on adenylate cyclase activity. These data suggest that activation of phospholipase C is a prerequisite for both PTH- and thrombin-stimulated increases in Ca(i)++ and that protein kinase C differentially regulates the ability of these agents to raise Ca(i)++. Collectively the results support the notion that the IP3/calcium mobilizing pathways utilized by PTH and thrombin are compartmentalized. Further, the finding that phorbol esters produce opposite effects on PTH-stimulated adenylate cyclase and Ca(i)++ suggests that protein kinase C can alter the cellular response to PTH by regulating the relative strength of signaling via these pathways.
|Original language||English (US)|
|Number of pages||6|
|Journal||Journal of Pharmacology and Experimental Therapeutics|
|State||Published - 1994|
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