Dynamic Factors for Transmitter Release at Small Presynaptic Boutons Revealed by Direct Patch-Clamp Recordings

Abstract

Small size of an axon and presynaptic structures have hindered direct functional analysis of axonal signaling and transmitter release at presynaptic boutons in the central nervous system. However, recent technical advances in subcellular patch-clamp recordings and in fluorescent imagings are shedding light on the dynamic nature of axonal and presynaptic mechanisms. Here I summarize the functional design of an axon and presynaptic boutons, such as diversity and activity-dependent changes of action potential (AP) waveforms, Ca²⁺ influx, and kinetics of transmitter release, revealed by the technical tour de force of direct patch-clamp recordings and the leading-edge fluorescent imagings. I highlight the critical factors for dynamic modulation of transmitter release and presynaptic short-term plasticity.

Keywords: action potential; axon; patch-clamp; presynaptic terminal; short-term plasticity; transmitter release.

FIGURE 1

Presynaptic transmitter releases studied by direct patch clamp recordings. (A,B) Patch-clamp recordings from a EGFP-labeled axon varicosity of a cerebellar PC (A) or of GC (B), adapted from Kawaguchi and Sakaba (2015, 2017), respectively, with permission from Elsevier. Top, images of EGFP-labeled axon terminals of each neuron. Middle, presynaptic Ca²⁺ currents (I_Ca) upon different durations of depolarization pulses to 0 mV. Bottom, presynaptic Cm increase triggered by Ca²⁺ currents indicated in the middle. Average Cm increases plotted against duration of depolarization pulses are also shown.

FIGURE 2

Kinetics of APs, Ca²⁺ influx, and transmitter release in a PC and GC bouton. Peak-scaled representative waveforms of APs, presynaptic Ca²⁺ currents (I_Ca), and transmitter release rate obtained by simultaneous recordings from a representative pair of presynaptic bouton (A, PC; B, GC) and postsynaptic cell. Spontaneous APs (blue), recorded from an EGFP-labeled axon varicosity of a PC (A, peak of +30 mV) and of a GC (B, peak of +37 mV), were used as voltage commands for stimulation to the voltage-clamped bouton, and presynaptic I_Ca (red, average of 5 traces from a pair) and postsynaptic currents were simultaneously recorded. Time courses of transmitter release rate (black, average of 5 traces recorded from a pair) were calculated by the deconvolution of the postsynaptic responses evoked by the presynaptic I_Ca, based on the miniature postsynaptic responses. Presynaptic recordings from a bouton of PC axon and that of GC axon had similar recording conditions, precluding the possibility of the distinct kinetics being due to the quality of patch-clamp recordings. All traces are adapted from Kawaguchi and Sakaba (2015, 2017) with permission from Elsevier.