Phorbol esters modulate the high Ca2+-stimulated accumulation of inositol phosphates in bovine parathyroid cells

We examined the effects of TPA on the high Ca2+-stimulated accumulation of inositol phosphates in bovine parathyoid cells to determine whether protein kinase C modulates phosphoinositide turnover in a fashion similar to that observed in other cell types stimulated by more classic Ca2+ mobilizing hormones. Following exposure of parathyroid cells to TPA (10−6 M) for 10 or 30 minutes, there was a time- and dose-dependent inhibition of the accumulation of inositol monophosphate (IP), inositol bisphosphate (IP2), and inositol trisphosphate (IP3) stimulated by 3 mM Ca2+. Half the maximal observed inhibition took place at 1–10 nM TPA, with 50–60% inhibition of high Ca2+-stimulated accumulation of inositol phosphates at 10−6 M TPA. The active phorbol ester, 4β-phorbol didecanoate, produced similar effects; the inactive derivative, 4α-phorbol didecanoate, was without effect. When parathyroid cells were exposed to TPA (10−6 M) for varying times and were then incubated with high (3 mM) Ca2+, inhibition of inositol phosphate accumulation was observed with 10 or 30 minutes preincubation. In contrast, preincubation of cells with TPA for 3 or 18 h markedly enhanced the high (3 mM) Ca2+-induced increase in inositol phosphates. In cells preincubated with TPA for 18 h, binding sites for [3H]phorbol dibutyrate and total protein kinase C (PKC) activity were reduced by greater than 95% and by 71%, respectively, consistent with downregulation of the enzyme. These results suggest that the high extracellular Ca2+-stimulated increase in accumulation of inositol phosphates in parathyroid cells, which has been postulated to result from a receptorlike process, can be modulated by agonists of protein kinase C in a fashion similar to that observed with more classic Ca2+ mobilizing hormones. Activators of kinase C initially inhibit the generation of inositol phosphates, presumably as a result of reduced turnover of phosphoinositides, but subsequently enhance inositol phosphate accumulation, probably because of down-regulation of protein kinase C.