Relative Distribution of Ca Channels at the Crayfish Inhibitory Neuromuscular Junction

Allana, Tariq N. and Jen-Wei Lin. Relative distribution of Ca channels at the crayfish inhibitory neuromuscular junction. J Neurophysiol 92: 1491–1500, 2004. First published May 12, 2004; 10.1152/ jn.00287.2004. We investigated the Ca channel-synaptic vesicle topography at the inhibitor of the crayfish (Procambarus Clarkii) neuromuscular junction (NMJ) by analyzing the effect of different modes of Ca channel block on transmitter release. Initial identification of Ca channels revealed the presence of two classes, P and non-P-type with P-type channels governing 70% of the total Ca influx. The remaining Ca influx was completely blocked by Cd but not by saturating concentrations of -conotoxins MVIIC and GVIA, or nifedipine and SNX-482. To examine the relative spatial distribution of Ca channels with respect to synaptic vesicles, we compared changes in inhibitory postsynaptic current amplitude and synaptic delay resulting from different spatial profiles of [Ca ]i around release sites. Specifically, addition of either [Mg ]o, which decreases single-channel current, or -Aga IVA, which completely blocks P-type channels, prolonged synaptic delay by a similar amount when Ca influx block was 40%. Because non-P-type channels are able to compensate for blocked P-type channels, it suggests that these channels overlap considerably in their distribution. However, when Ca influx was blocked by 50%, -Aga IVA increased delay significantly more than Mg , suggesting that P-type channels are located closer than non-P-type channels to synaptic vesicles. This distribution of Ca channels was further supported by the observations that non-P-type channels are unable to trigger release in physiological saline and EGTA preferentially prolongs synaptic delay dominated by non-P-type channels when transmitter release is evoked with broad action potentials. We therefore conclude that although non-P-type channels do not directly trigger release under physiological conditions, their distribution partially overlaps with P-type channels.