The Ever-Growing Puzzle of Asynchronous Release

Abstract

Invasion of an action potential (AP) to presynaptic terminals triggers calcium dependent vesicle fusion in a relatively short time window, about a millisecond, after the onset of the AP. This allows fast and precise information transfer from neuron to neuron by means of synaptic transmission and phasic mediator release. However, at some synapses a single AP or a short burst of APs can generate delayed or asynchronous synaptic release lasting for tens or hundreds of milliseconds. Understanding the mechanisms underlying asynchronous release (AR) is important, since AR can better recruit extrasynaptic metabotropic receptors and maintain a high level of neurotransmitter in the extracellular space for a substantially longer period of time after presynaptic activity. Over the last decade substantial work has been done to identify the presynaptic calcium sensor that may be involved in AR. Several models have been suggested which may explain the long lasting presynaptic calcium elevation a prerequisite for prolonged delayed release. However, the presynaptic mechanisms underlying asynchronous vesicle release are still not well understood. In this review article, we provide an overview of the current state of knowledge on the molecular components involved in delayed vesicle fusion and in the maintenance of sufficient calcium concentration to trigger AR. In addition, we discuss possible alternative models that may explain intraterminal calcium dynamics underlying AR.

Keywords: calcium; calcium extrusion; presynaptic; synaptic release; synaptotagmins.

Figure 1

Asynchronous release (AR) is temporally separated from action potential (AP) generated calcium concentration microdomains. (A) Example traces of responses recorded from a pair of connected cells, a hippocampal presynaptic cholecystokinin (CCK)⁺ basket cell (black) and postsynaptic CA1 pyramidal neuron (red, three subsequently recorded traces). Five APs (50 Hz) trigger synchronized phasic IPSC [labeled with (^{^})] and delayed responses that can be observed both during the AP train [yellow window; labeled with (*)] and after termination of presynaptic stimulation (green window). (B) Schematic drawing of the presynaptic calcium concentration dynamics after a single AP. Opening of the voltage gated calcium channel (VGCC) causes formation of a calcium concentration microdomain—a short-lasting local elevation of [Ca²⁺]_i sufficient to trigger phasic release (left panel). After closure of the VGCC, [Ca²⁺]_i radially diffuses and equilibrates within the terminal and then further declines due to binding to endogenous buffers and extrusion (right panel). (C) Schematic drawing of vesicle fusion driven by an AP evoked Ca²⁺ micro/nano-domain (upper panel). Note that high synchrony arises from the low affinity of the Ca²⁺ sensor (Syt_LA) and tight spatial coupling of the Ca²⁺ source and Ca²⁺ sensor. The lower panel shows delayed vesicle fusion mediated by residual [Ca²⁺]_i remaining in terminals several milliseconds after the last AP. In this case recruitment of high affinity synaptotagmins (Syt_HA) is necessary, but vesicles can be spatially separated from VGCC, since release is triggered by bulk [Ca²⁺]_i. (D) Schematic representation of the [Ca²⁺]_i time course at the release site (blue) after AP. Dotted lines show time windows for synchronous (SR) and AR components of release that are probably mediated by synaptotagmins with different Ca²⁺ affinities.

Figure 2

Sources of calcium for AR generation. (A) Proposed sources of calcium for AR generation: (i) ATP-gated ionotropic P2X2 receptors; calcium- dependent release from intracellular calcium stores; and (ii) calcium-dependent prolongation of calcium entry through VGCC. (B) Schematic representation of a hypothetical [Ca²⁺]_i time course at the release site during SR and AR. Conventional [Ca²⁺]_i elevation due to the flux through VGCC during AP (blue) combines with Ca²⁺ entry via an additional AP activated Ca²⁺ source (green). Dotted lines show time windows for SR and AR.

Figure 3

Possible role of the presynaptic calcium extrusion pumps in AR generation. (A) Schematic drawing of presynaptic sequence of Na⁺ and Ca²⁺ fluxes triggered by a single AP: (i) Na⁺ entry trough voltage-gated sodium channels (VGSCs); (ii) Ca²⁺ entry trough VGCC; (iii) Fast Ca²⁺ extrusion via sodium/calcium exchanger (NCX); and (iv) Final clearance of the presynaptic Ca²⁺ by plasma membrane calcium-ATPase (PMCA). (B) Massive elevation of intra-terminal Na⁺ concentration during the high frequency train of APs can strongly reduce the NCX extrusion rate (a), or at extreme elevation of [Na⁺]_i, reverse the direction of Na⁺ and Ca²⁺ fluxes (b) through NCX prolonging the time course of the presynaptic calcium clearance, especially in terminals with reduced function of PMCA. (C) Schematic representation of intraterminal [Ca²⁺]_i (blue) and [Na⁺]_i (red) time courses after a single AP. (D) Schematic representation of the [Ca²⁺]_i time course (blue) after burst APs leading to the massive elevation [Na⁺]_i (red) when NCX is the only extrusion pump. Note, that direct simultaneous measurements of intraterminal [Na⁺]i and [Ca²⁺]_i are not technically possible at the moment. However, modeling studies suggest that in the case of PMCA absence the temporal dynamics of [Ca²⁺]_i and [Na⁺]_i are be tightly coupled.