Opposing effects of phorbol esters on transmitter release and calcium currents at frog motor nerve endings

1. Phorbol esters activate protein kinase C (PKC) and also increase the secretion of neurotransmitter substances by an unknown mechanism. To evaluate whether the stimulatory effects of such agents on acetylcholine (ACh) secretion occur as a consequence of stimulation of Ca2+ entry, we made electrophysiological measurements of ACh secretion (i.e. endplate potentials, EPPs) and the component of the prejunctional perineural voltage change associated with nerve terminal calcium currents (perineural calcium current) at frog neuromuscular junctions. 2. In the first series of experiments, modest concentrations of K+ channel blockers were employed so that simultaneous measurements of EPP amplitudes and perineural calcium currents could be made. In these experiments, 12-O-tetradecanoylphorbol 13-acetate (TPA; 162 nM) and phorbol 12,13-dibutyrate (PDBu; 100-200 nM) each increased ACh release but simultaneously decreased the calcium component of the prejunctional perineural current. TPA and PDBu also inhibited perineural calcium currents in the presence of higher concentrations of K+ channel blockers. 3. Blockade of Ca2+ channels by Cd2+ prevented the action of PKC stimulators on perineural waveforms. 4. The inactive compound 4-alpha-phorbol 12-myristate 13-acetate (150 nM) did not affect EPP amplitudes or perineural currents. 5. The extracellular [Ca2+]-ACh release relationship was increased in maximum by PDBu without any change in the potency of Ca2+ to support evoked ACh release. 6. The results demonstrate that phorbol esters increase neurotransmitter secretion whilst simultaneously decreasing the nerve ending calcium currents that promote evoked release. The results, which suggest that the optimal control point for secretion might not be the calcium channel but rather a component of the secretory apparatus, are discussed in conjunction with the possible target sites for phorbol esters in the nerve ending.