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The Journal of Neuroscience, September 1, 2000, 20(17):6326-6332

Analysis of Presynaptic Ca²⁺ Influx and Transmitter Release Kinetics during Facilitation at the Inhibitor of the Crayfish Neuromuscular Junction

Andrey Vyshedskiy, Tariq Allana, and Jen-Wei Lin

Department of Biology, Boston University, Boston, Massachusetts 02215

The inhibitory synapse of the crayfish neuromuscular junction was used to examine mechanisms underlying the F2 component of synaptic facilitation. Because previous studies have shown accelerated transmitter release during facilitation, we examined whether an activity-dependent plasticity in I_Ca could underlie this acceleration. We established that fluorescent transients generated by Magnesium Green can resolve small differences in presynaptic Ca²⁺ influx that correlate with changes in IPSC waveform. However, there was no change in Ca²⁺ transients associated with the accelerated release. Analyzing the initial rise of IPSC and the duration of the presynaptic spike yielded a depolarization-release coupling plot that captures the impact of spike waveform on the initial rate of release. We conclude that accelerated release during F2 facilitation cannot be attributed to plasticity of I_Ca or modulation of spike waveform. Kinetic analysis showed a reduction in synaptic delay during facilitation only when broad action potentials were used. In unfacilitated release, synaptic delay increased as spike duration lengthened. We propose that small single Ca²⁺ channel currents during the plateau phase of broad action potentials raise local Ca²⁺ concentration only enough to fill a high-affinity site. Occupation of this site in itself, or events downstream, would convert a vesicle from control to facilitated state. If the conversion were a slow process, it could explain the changes in synaptic delay reported here. This hypothesis can also account for a number of observations related to Ca²⁺ cooperativity and synaptic facilitation.

Key words: synaptic delay; synapse; crayfish inhibitor; neuromuscular junction; facilitation; calcium indicator

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Copyright © 2000 Society for Neuroscience  0270-6474/00/20176326-07$05.00/0

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