PROTEIN-KINASE-C MODULATES EXOGENOUS ACETYLCHOLINE CURRENT IN XENOPUSOOCYTES

The modulation of acetylcholine-activated current (I-ACh) by protein k inase C (PKC) was studied in Xenopus laevis oocytes microinjected with either mRNA extracted from C2C12 myotubes (C2C12 mRNA) or RNAs encodi ng murine alpha beta gamma delta subunits of the nicotinic ACh recepto r (nAChR), Voltage-clamped oocytes were treated for 90 sec with 12-O-t etradecanoylphorbol-13-acetate (TPA, 300 nM), a potent PKC activator. Transient increase in the amplitude and acceleration in the decay of I -ACh were invariably observed within minutes of TPA application, and w ere independent of extracellular Ca2+ concentration. Both parameters r ecovered to control within 20-30 min; then a slight depression of I-AC h developed, By this time, an initial PKC down regulation was observed . At the peak of TPA-induced potentiation, dose-response relations sug gested an increased binding affinity of nAChR for the neurotransmitter . 4 alpha-phorbol 12,13-didecanoate (300 nM), a biologically inactive analogue of TPA, did not affect I-ACh, while staurosporine (5-10 mu M) , a potent inhibitor of PKC activity, suppressed the action of TPA on I-ACh, In oocytes co-injected with C2C12 mRNA and with rat brain mRNA, I-ACh was potentiated by 5-hydroxytryptamine (10 mu M), whose recepto rs are coupled to phosphoinositide hydrolysis. The nAChR-channel activ ity in cell-attached patches increased when TPA was applied to the ooc ytes, In 50% of the oocytes examined, a sustained depression of the si ngle channel activity followed. We conclude that in Xenopus oocytes an endogenous PKC system regulates the function of embryonic-type muscle nAChRs. (C) 1995 Wiley-Liss, Inc.