PROTEIN-KINASE C-MEDIATED DOWN-REGULATION OF VOLTAGE-DEPENDENT SODIUM-CHANNELS IN ADRENAL CHROMAFFIN CELLS

Treatment of cultured bovine adrenal chromaffin cells with 12-O-tetrad ecanoylphorbol 13-acetate (TPA), an activator of protein kinase C (PKC ), decreased [H-3]saxitoxin ([H-3]STX) binding in a concentration (IC5 0 = 19 nM)- and time (t(1/2) = 4.5 h)-dependent manner. TPA (100 nM fo r 15 h) lowered the B-max of [H-3]STX binding by 53% without altering the K-D value. Phorbol 12,13-dibutyrate (PDBu) also reduced [H-3]STX b inding, whereas 4 alpha-TPA, an inactive analogue, had no effect. The inhibitory effect of TPA was abolished when H-7 (an inhibitor of PKC), but not H-89 (an inhibitor of cyclic AMP-dependent protein kinase), w as included in the culture medium for 1 h before and during TPA treatm ent, Simultaneous treatment with TPA in combination with either actino mycin D or cycloheximide, an inhibitor of protein synthesis, nullified the effect of TPA. TPA treatment also attenuated veratridine-induced Na-22(+) influx but did not alter the affinity of veratridine for Na c hannels as well as an allosteric potentiation of veratridine-induced N a-22(+) influx by brevetoxin. These results suggest that an activation of PKC down-regulates the density of Na channels without altering the ir pharmacological features; this down-regulation is mediated via the de novo synthesis of an as yet unidentified protein(s), rather than an immediate effect of Na channel phosphorylation.