The effect of protein kinase C (PKC) stimulation on Ca2+ channels was studied in frog sympathetic neurons. 12,13-Phorbol dibutyrate (PDBu) consistently augmented Ca2+ channel currents in whole-cell recordings. This enhancement was blocked by staurosporine and PKC(19-31), but not produced by 4 alpha-phorbol 12,13-didecanoate, indicating that PDBu acts via PKC. Both N- and L-type currents, as isolated pharmacologically, were increased. PKC enhancement was independent of the extent of G protein activation, indicating that it was not caused by removal of tonic G protein inhibition. In unitary recordings PDBu produced dramatic increases in single N- and L-type channel activity by sharply decreasing closed time intervals between adjacent openings, but did not alter the unitary current size or mean open time. This up-modulation by PKC may constitute a positive feedback mechanism in the regulation of neuronal Ca2+ channel activity.