Abstract
In rat pancreatic acini, we previously demonstrated that depending on the agonist used, activation of cholecystokinin type A (CCK A) receptor (CCK-AR) results in the differential involvement of the cytosolic phospholipase A 2 (cPLA 2), phospholipase C ß1 (PLC ß1) and Src/protein tyrosine kinase (PTK) pathways. The high-affinity CCK-AR appears to be coupled to the Gß/cPLA 2/arachidonic acid (AA) cascade in mediating Ca 2+ oscillations. The low-affinity CCK-AR is coupled to both the Gaqα q/11/PLC ß1/inositol 1,4,5-trisphosphate (IP 3) to evoke intracellular Ca 2+ release and the Src/PTK pathway to mediate extracellular Ca 2+ influx. The objectives of this study were to provide evidence that cPLA 2 is present in pancreatic acini and to evaluate the possibility that its activation results in Ca 2+ oscillations and amylase secretion. Furthermore, we investigated the mechanism of Ca 2+ oscillations mediated by the high-affinity CCK-AR. In rat pancreatic acini, immunoprecipitation studies using an anti-cPLA 2 monoclonal antibody, demonstrated a cPLA 2 band at the location of 110 kDa. A selective inhibitor of cPLA 2, AACOCF 3 (100 μM), inhibited production of AA metabolites, Ca 2+ oscillations and amylase secretion elicited by the high-affinity CCK-AR agonist, CCKOPE (10-1000 nM). In addition, through the repetitive release of intracellular Ca 2+, CCK-OPE enhanced phosphotransferase activities of Ca 2+/calmodulin-dependent protein kinase type IV (CaMK IV), which were inhibited by AACOCF 3. The CaMK inhibitor, K252-a (1-3 μM), also abolished basal and CCK-OPE-stimulated CaMK IV activities. The CaM inhibitor, W-7 (100 μM), and K252-a inhibited Ca 2+ oscillations and amylase secretion evoked by CCK-OPE without affecting the AA formation. Therefore, it appears that Ca 2+ oscillations elicited by the high-affinity CCK-AR/Gß/ cPLA 2/AA pathway activate CaMK IV. Activated CaMK, in turn, regulates Ca 2+ oscillations through a positive feedback mechanism to mediate pancreatic exocytosis.
Published Version
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