PHORBOL ESTER ENHANCES EXCITATORY AMINO ACID-INDUCED DOPAMINE RELEASE FROM MESENCEPHALIC CELL-CULTURES

The hypothesis that protein kinase C activation can modulate excitatory amino acid-induced dopamine release was tested by investigating effects of phorbol esters, direct activators of protein kinase C, on dopamine release stimulated by N-methyl-D-aspartate (NMDA) and non-NMDA sub-types of excitatory amino acid agonists in fetal rat mesencephalic cell cultures. The phorbol ester, 12-O-tetradecanoyl phorbol-13-acetate (TPA), enhanced dopamine release evoked by NMDA, kainate, quisqualate and by K+ depolarization. Release in the presence of NMDA and TPA was completely abolished by the NMDA antagonist, MK-801. TPA enhancement of NMDA-stimulated dopamine release was likely due to protein kinase C activation by the phorbol ester since (1) the NMDA response was enhanced by nanomolar concentrations of TPA, (2) two phorbol esters capable of activating protein kinase C enhanced the NMDA response while an inactive phorbol ester did not, (3) staurosporine, a potent protein kinase C inhibitor, blocked TPA enhancement of the NMDA response. TPA enhancement of NMDA-stimulated dopamine release was not blocked by H8, an inhibitor with high affinity for cyclic nucleotide dependent kinases, while forskolin, a direct activator of adenylate cyclase, had no effect on NMDA-stimulated release, indicating a lack of involvement of cAMP-dependent kinase in the TPA effect. TPA enhanced NMDA-stimulated release both in the presence and absence of Mg2+, indicating that TPA enhancement was not due to reversal of a Mg2+ blockade of the NMDA receptor. Release of glycine, a co-agonist at the NMDA receptor, by TPA could not account for TPA enhancement of NMDA-stimulated dopamine release since low concentrations of glycine had no effect while higher concentrations of glycine inhibited NMDA-stimulated dopamine release. TPA enhanced NMDA-stimulated dopamine release over a range of extracellular Ca2+ concentrations, indicating that TPA may sensitize the NMDA-mediated release process to Ca2+. It is suggested that protein kinase C activation by inputs converging on dopaminergic neurons may serve to amplify excitatory amino acid-induced dopamine release.