INHIBITION OF N-TYPE CA2-SY5Y) CELLS BY MUSCARINE VIA STIMULATION OF M(3) RECEPTORS( CHANNEL CURRENTS IN HUMAN NEUROBLASTOMA (SH)

The effects of muscarine on whole-cell Ca2+ channel currents in SH-SY5 Y cells were studied using conventional and perforated-patch-clamp tec hniques, with 10 mM Ba?2+ as charge carrier. Muscarine (10-300 mu M) c aused concentration-dependent inhibitions of Ca2+ channel currents whi ch were only reversible when perforated-patch recordings were used. In hibition of currents was associated with slowing of activation kinetic s in approximately 50% of cells. In the presence of 5 mu M nifedipine, muscarine was still able to inhibit currents, but after pre-exposure of cells to 1 mu M omega-conotoxin GVIA the inhibitory effects of musc arine were almost completely lost. In the presence of 100 mu M muscari ne, Bay K 8644 (5 mu M) was still able to enhance current amplitudes. Pre-treatment of cells with pertussis toxin (250 ng/ml for 16-24 hr) o r inclusion of 1 mM GDP-beta-S in the patch-pipette prevented the inhi bitory actions of muscarine. Hexahydrosiladifenidol (0.1-1 mu M) antag onized the actions of muscarine (calculated pA(2) 7.1) but the presenc e of 10 mu M pirenzipine or 0.1 mu M methoctramine in the bath solutio n did not alter the degree of current inhibition caused by 100 mu M mu scarine. In summary, these results indicate that muscarine in SH-SY5Y cells causes inhibition of N-type Ca2+ channels via a M(3) receptor co upled to a pertussis toxin-sensitive G-protein.