The effect of protein kinase C (PKC) on amylase discharge from streptolysin-O-permeabilized rat pancreatic acini was investigated. Addition of phorbol 12-myristate 13-acetate (PMA) to permeabilized cells potentiated Ca2+-stimulated release, but had no effect on discharge at non-stimulatory Ca2+ concentrations. PMA markedly shifted the Ca2+-concentration-dependence of amylase discharge to the left, by enhancing the time over which the permeabilized cells release. This effect was inhibited by both staurosporine and PKC-19-31-amide peptide inhibitor, indicating that the effect of PMA was due to its action on PKC. Staurosporine also partially inhibited amylase release at the optimal concentration of Ca2+; this effect was not replicated by the more specific PKC-19-3 1-amide peptide inhibitor and may be due to an effect on another second-messenger system. PKC appears to be an important modulator of release in pancreatic acini, but its activation is not an absolute requirement for Ca2+-dependent amylase discharge.