Protein kinase C: regulation of serotonin release from rat brain cortical slices

Abstract

The effects of phorbol esters on serotonin release were examined in an attempt to investigate the role of protein kinase C in the regulation of serotonin release. Rat brain parietal cortical slices were incubated with [ 3H]5-HT in the presence of pargyline in order to label the serotonin stores. Potassium stimulated (30 s) release and spontaneous [ 3H]5-HT efflux were examined in slices during superfusion with Krebs-Ringer solution containing chlorimipramine. Repeated K + stimulations elicited reproducible responses with release ratios of approximately 1.0. Introduction of phorbol 12-myristate,13-acetate (PMA) or phorbol 12,13-dibutyrate (PDBu) 20 min prior to S 2, or S 3 resulted in dose-related increases in [ 3H]5-HT or [ 3H]NE release. PMA was slightly more potent (93% increase) than PDBu in potentiating K +-stimulated [ 3H]5-HT release. Phorbol and 4α-phorbol 12,13-didecanoate (4αPDD) which do not activate protein kinase C did not alter serotonin release. In contrast, basal [ 3H]5-HT and [ 3H]NE release were altered to a far lesser extent which was not always dose related. The response to the phorbol esters was reversible, Ca 2+-dependent and reached maximal effect after 20 min of superfusion. The putative protein kinase C inhibitor, 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H-7) inhibited K +-induced [ 3H]5-HT release significantly (11%) but did not alter basal efflux. The PMA facilitation of serotonin release was, however, markedly prevented by the enzyme inhibitor. The effect of PMA on release was found not to be directly mediated through the prejunctional serotonin autoreceptors. This is suggested by (1) the similarity in PMA-induced release potentiation observed in the presence or absence of the serotonin agonist, 5-methoxytryptamine, and (2) by the inability of the kinase inhibitor, H-7, to alter K +-stimulated release by the serotonin receptor antagonist, methiothepin. These results on serotonin and norepinephrine release and those of previous workers dealing with other transmitters point to an important role of protein kinase C in the modulation of neurotransmitter release in the central nervous system.