The modulation of acetylcholine-activated current (I(ACh)) by protein kinase C (PKC) was studied in opus laevis oocytes microinjected with either mRNA extracted from C2C12 myotubes (C2C12 mRNA) or RNAs encoding murine αβγδ subunits of the nicotinic ACh receptor (nAChR). Voltage-clamped oocytes were treated for 90 sec with 12-O-tetradecanoylphorbol-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 were independent of extracellular Ca2+ concentration. Both parameters recovered to control within 20-30 min; then a slight depression of I(ACh) developed. By this time, an initial PKC down regulation was observed. At the peak of TPA-induced potentiation, dose- response relations suggested an increased binding affinity of nAChR for the neurotransmitter. 4α-phorbol 12,13-didecanoate (300 nM), a biologically inactive analogue of TPA, did not affect I(ACh), while staurosporine (5-10 μM), 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 μM), whose receptors are coupled to phosphoinositide hydrolysis. The nAChR-channel activity in cell- attached patches increased when TPA was applied to the oocytes. In 50% of the oocytes examined, a sustained depression of the single channel activity followed. We conclude that in Xenopus oocytes an endogenous PKC system regulates the function of embryonic-type muscle nAChRs.
- C2C12 myotube
- embryonic-type muscle nicotinic acetylcholine receptor
- receptor modulation
ASJC Scopus subject areas