Protein kinase C activation inhibits receptor- evoked inositol trisphosphate formation and induction of cytosolic calcium oscillations by decreasing the affinity-state of the cholecysto- kinin receptor in pancreatic acinar cells

Digital-imaging microscopy of Fura-2-loaded pancreatic acinar cells revealed that the C-terminal octapeptide of cholecystokinin (CCKe) dose-dependently recruited 94% of freshly isolated acinar cells in terms of receptor-evoked Ca2+ mobilization. Maximal and half-maximal cell-recruitment were reached with 0.1 nM and 16.8 pM CCKe, respectively. The upstroke of the dose-recruitment curve consisted of cells displaying oscillatory changes in free cytosolic Ca2+ concentration ([Ca2+]i). After having reached its maximum, the percent­ age oscillating cells dose-dependently decreased upon further increasing of the CCKs con­ centration. Pretreatment of the acinar cells with 0.1 |j.M TPA caused a rightward shift of the dose-recruitment curve but did not change the maximal effect of CCKs on the recruitment of oscillating cells. Half-maximal recruitment was obtained with 287 pM CCKs. This observa­ tion demonstrates that high levels of protein kinase C activation do not inhibit Ca2+ oscilla­ tions at a level downstream to receptor activation. Moreover, this observation demonstrates that protein kinase C-mediated inhibition of Ca2+ oscillations evoked by submaximai CCKs concentrations occurs at the receptor level, converting it from a high-affinlty state into a low-affinity state. This conclusion is supported by the observation that TPA completely in­ hibited the recruitment of acinar cells in response to the high-affinity receptor agonist JMV180. The inhibitory action of TPA on CCKs-evoked cell-recruitment was paralleled by an inhibitory effect of the phorbol ester on the CCKs-evoked peak increase in average inositol trisphosphate concentration in a population of acinar cells. This observation indicates that low concentrations of CCKs interact with the high-affinity CCK receptor to increase [Ca2+]j through the intermediation of inositol trisphosphate. n i O i Abbreviations: CCKs, C-terminal octapeptide of cholecystokinin; [Ca ]j, cytosolic free Ca concentration; JMV-180, Boc-Tyr(SO^~)-Nle-Gly-Trp-Nle-Asp-2-phenylethylester; TPA, 12-O-tetradecanoylphorbol 13-acetate; DAG, 1,2-diacylglycerol; Ins(l A 5)P3, inositol 1,4,5-trisphosphate.