Protein kinase C-␦ (PKC-␦) is involved in growth, differentiation, tumor suppression, and regulation of other cellular processes. PKC-␦ activation causes translocation, tyrosine phosphorylation, and serine-threonine kinase activity. However, little is known about the ability of G protein-coupled receptors to activate these processes or the mediators involved. In the present study, we explored the ability of the neurotransmitter/hormone, CCK, to stimulate these changes in PKC-␦ and explored the mechanisms. In rat pancreatic acini under basal conditions, PKC-␦ is almost exclusively located in cytosol. CCK and TPA stimulated a rapid PKC-␦ translocation to membrane and nuclear fractions, which was transient with CCK. CCK stimulated rapid tyrosine phosphorylation of PKC-␦ and increased kinase activity. Using tyrosine kinase (B44) and a tyrosine phosphatase inhibitor (orthovanadate), changes in both CCK-and TPA-stimulated PKC-␦ tyrosine phosphorylation were shown to correlate with changes in its kinase activity but not translocation. Both PKC-␦ tyrosine phosphorylation and activation occur exclusively in particulate fractions. The Src kinase inhibitors, SU6656 and PP2, but not the inactive related compound, PP3, inhibited CCK-and TPA-stimulated PKC-␦ tyrosine phosphorylation and activation. In contrast, PP2 also had a lesser effect on CCK-but not TPA-stimulated PKC-␦ translocation. CCK stimulated the association of Src kinases with PKC-␦, demonstrated by co-immunoprecipitation. These results demonstrate that CCK A receptor activation results in rapid translocation, tyrosine phosphorylation, and activation of PKC-␦. Stimulation of PKC-␦ translocation precedes tyrosine phosphorylation, which is essential for activation to occur. Activation of Src kinases is essential for the tyrosine phosphorylation and kinase activation to occur and plays a partial role in translocation.