Asynchronous Ca2+ current conducted by voltage-gated Ca2+ (CaV)-2.1 and CaV2.2 channels and its implications for asynchronous neurotransmitter release

We have identified an asynchronously activated Ca2+ current through voltage-gated Ca2+ (Ca-V)-2.1 and Ca(V)2.2 channels, which conduct P/Q- and N-type Ca2+ currents that initiate neurotransmitter release. In nonneuronal cells expressing Ca(V)2.1 or CaV2.2 channels and in hippocampal neurons, prolonged Ca2+ entry activates a Ca2+ current, I-Async, which is observed on repolarization and decays slowly with a half-time of 150-300 ms. I-Async is not observed after L-type Ca2+ currents of similar size conducted by Ca(V)1.2 channels. I-Async is Ca2+-selective, and it is unaffected by changes in Na+, K+, Cl-, or H+ or by inhibitors of a broad range of ion channels. During trains of repetitive depolarizations, I-Async increases in a pulse-wise manner, providing Ca2+ entry that persists between depolarizations. In single-cultured hippocampal neurons, trains of depolarizations evoke excitatory postsynaptic currents that show facilitation followed by depression accompanied by asynchronous postsynaptic currents that increase steadily during the train in parallel with I-Async. I-Async is much larger for slowly inactivating Ca(V)2.1 channels containing beta(2a)-subunits than for rapidly inactivating channels containing beta(1b)-subunits. I-Async requires global rises in intracellular Ca2+, because it is blocked when Ca2+ is chelated by 10 mM EGTA in the patch pipette. Neither mutations that prevent Ca2+ binding to calmodulin nor mutations that prevent calmodulin regulation of Ca(V)2.1 block I-Async. The rise of I-Async during trains of stimuli, its decay after repolarization, its dependence on global increases of Ca2+, and its enhancement by beta(2a)-subunits all resemble asynchronous release, suggesting that I-Async is a Ca2+ source for asynchronous neurotransmission.