On the simultaneous electrophysiological measurements of neurotransmitter release and perineural calcium currents from frog motor nerve endings.

Ca2+ currents from the perineural region of motor nerve endings were measured together with evoked acetylcholine (ACh) release (i.e., end-plate potentials EPPs) in frog skeletal muscle in an attempt to define experimental conditions in which simultaneous measurements of both phenomena were feasible. In a solution containing low Ca2+ (0.9 mM), high Mg2+ (10 mM) and modest concentrations of K+ channel blockers (250 microM tetraethylammonium, 100 microM 3,4,-diaminopyridine), reliable measurements of perineural Ca2+ currents were possible. For convenience, this solution will be termed 'Ca2+ current' Ringer. The mean number of ACh quanta released in Ca2+ current Ringer was near the midpoint of the relationship between extracellular [Ca2+] and evoked ACh release observed previously in normal Ringer solutions. Consequently, ACh release in response to low-frequency motor nerve stimulation (0.05 Hz) was well maintained, allowing simultaneous measurements of Ca2+ currents and evoked ACh release to be made. Ca2+ currents and EPPs measured simultaneously in Ca2+ current Ringer were increased or decreased in parallel by increasing or decreasing the extracellular Ca2+ concentrations. Ca2+ channel blockers (Cd2+, 500 microM; omega-conotoxin, 3 microM) eliminated both EPPs and the Ca2+ component of the perineural current. NaF (10 mM), which stimulates ACh release, produced parallel increases in EPPs and perineural Ca2+ currents. NG-cyclohexyladenosine (CHA), an A1 adenosine receptor agonist, inhibits ACh release without effects on perineural currents. The results suggest that the concurrent electrophysiological recording of Ca2+ currents and ACh release in Ca2+ current Ringer is a reliable experimental approach for determining whether drugs or disease states affect ACh release by acting on Ca2+ channels in the presynaptic membrane.