Ca 2+ role on the effect of phorbol esters on the spontaneous quantal release of neurotransmitter at the mouse neuromuscular junction

Abstract

The effect of the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) on the release process was studied in presence of different extracellular Ca 2+ concentrations, in the mouse phrenic-diaphragm preparation. Hemidiaphragms were incubated for 2 h at room temperature in the presence or absence of TPA. TPA increased the basal frequency of miniature end plate potentials (mepp's) in a dose-dependent manner, resulting in a maximal increase of 280% at a concentration of 0.5 μM. An inverse relationship between extracellular Ca 2+ concentration and TPA effect was observed: at high extracellular concentrations of Ca 2+ the action of TPA decreased significatively, while at low Ca 2+ concentrations the effect of TPA was remarkably augmented. The highest effect of TPA was obtained when tested in a calcium-free medium. TPA also increased mepp frequency stimulated by 10 mM K +. As at basal conditions, the effect of TPA was higher at lower concentrations of extracellular calcium. The results suggest that the effect of stimulation of PKC on neurotransmitter release at the mice neuromuscular junction is not exerted at the level of calcium influx to the nerve terminal. Moreover the action of calcium and TPA seems to be superimposed. The effect of K + on neurotransmitter release could be explained not only by depolarization of the nerve terminal but by increasing the pool of activable PKC.