Ca2+ influx through carbachol-activated non-selective cation channels in guinea-pig gastric myocytes

1. Ca2+ microfluorometry (100 mu M K-5 fura-2) and the voltage-clamp technique were combined to study the effect of carbachol (CCh, 50 mu M) in inducing currents (I-CCh) through nonselective cation channels (NSCCCh) and increments in global cytosolic Ca2+ concentration (Delta[Ca2+](c)). 2. In Na+-containing bath solution, I-CCh fell from an initial phasic to a subsequent small (5 %) tonic component; Delta[Ca2+](c) fell to zero. Tonic I-CCh and [Ca2+](c) became prominent after substitution of extracellular 1.40 mM Na+ by 140 mM Cs+. Tonic I-CCh and Delta[Ca2+], were insensitive to intracellular heparin (3 mg ml(-1)) and ryanodine (4 mu M), i.e, they did not depend on Ca2+ release from sarcoplasmic reticulum (SR). 3. Single channel currents of NSCCCh could be resolved in whole-cell recordings. Substitution of Na+ by Cs+ increased NSCCCh activity by one order of magnitude and slope conductance from 22 to 30 pS. Extracellular quinidine (3 mu M) reversibly blocked the NSCCCh activity. 4. Both tonic I-CCh and tonic Delta[Ca2+](c) (a) followed a similar time course of activation, desensitization and facilitation, (b) were reversibly blocked by 3 mu M quinidine, and (c) persisted upon block of SR Ca2+ release. 5. A Ca2+ fractional current of tonic I-CCh (f(Ca)) of 0.009 was calculated by comparing the ratio Delta[Ca2+](c) (corrected for simultaneous Ca2+ redistribution) over I-CCh with depolarization-induced *Delta[Ca2+](c) (Delta[Ca2+](c) calculated from I-Ca induced by a 400 ms depolarization from -60 to 0 mV at 2 mM [Ca2+](o), 145 mM [Cs+](o)) over I-Ca. f(Ca) was 0.023 at [Ca2+](o) = 4 mM. 6. With 110 mM extracellular CaCl2 and 1.45 mM intracellular CsCl, I-CCh reversed at +19.5 mV suggesting a permeability ratio P-Ca/P-Cs of 2.8. 7. We conclude that Ca2+ influx through NSCCCh under physiological [Ca2+](o) could induce Delta[Ca2+](c). The f(Ca) was, however, much smaller than the one calculated from the reversal potential.