External Ca2+ dependence of acetylcholine- and succinylcholine-induced changes in channel conductance, open time and frequency at endplates of single muscle cells of adult mice.

The involvement of external Ca2+ in channel activity of nicotinic acetylcholine (nACh) receptors was investigated in the dissociated skeletal muscle fibers of adult mice. Single cells were prepared from flexor digitorum brevis muscles by treatment with collagenase and trypsin. Acetylcholine receptor-channel currents were recorded at endplates in the cell-attached mode, using the patch-clamp technique. Opening frequency, opening pattern and channel conductance were measured at different concentrations of ACh and succinylcholine (SuCh), using a patch pipette. Bath-applied ACh (30 microM) and SuCh (3-30 microM) decreased the ACh (1 microM)-induced channel conductance (SuCh was more potent than ACh). Large concentrations of both ACh (100-1000 microM) and SuCh (30-300 microM), which were applied via a patch pipette, also decreased channel conductance in a concentration-dependent manner. Increasing the concentration of calcium [Ca2+]0 in the patch pipette from 0 to 10 mM CaCl2 reduced ACh (1 microM)- and SuCh (1 microM)-activated single channel conductances. The increase in [Ca2+]0 prolonged the mean open times by 34% for ACh and by 22% for SuCh and decreased the channel opening frequency by 50% and 60%, respectively. These results demonstrate that ACh receptor-channel properties are dependent on [Ca2+]0 and that the ACh- and SuCh-induced decrease in channel currents may be also related to intracellular concentration of Ca2+. The effect of SuCh was greater than that produced by ACh.