Ca²⁺ current facilitation determines short‐term facilitation at inhibitory synapses between cerebellar Purkinje cells

Abstract

Key pointsr Short-term facilitation takes place at GABAergic synapses between cerebellar Purkinje cells (PCs).r By directly patch clamp recording from a PC axon terminal, we studied the mechanism of short-term facilitation.r We show that the Ca 2+ currents elicited by high-frequency action potentials were augmented in a [Ca 2+ ] i -dependent manner.r The facilitation of synaptic transmission showed 4-5th power dependence on the Ca 2+ current facilitation, and was abolished when the Ca 2+ current amplitude was adjusted to be identical.r Short-term facilitation of Ca 2+ currents predominantly mediates short-term facilitation at synapses between PCs.Abstract Short-term synaptic facilitation is critical for information processing of neuronal circuits. Several Ca 2+ -dependent positive regulations of transmitter release have been suggested as candidate mechanisms underlying facilitation. However, the small sizes of presynaptic terminals have hindered the biophysical study of short-term facilitation. In the present study, by directly recording from the axon terminal of a rat cerebellar Purkinje cell (PC) in culture, we demonstrate a crucial role of [Ca 2+ ] i -dependent facilitation of Ca 2+ currents in short-term facilitation at inhibitory PC-PC synapses. Voltage clamp recording was performed from a PC axon terminal visualized by enhanced green fluorescent protein, and the Ca 2+ currents elicited by the voltage command consisting of action potential waveforms were recorded. The amplitude of presynaptic Ca 2+ current was augmented upon high-frequency paired-pulse stimulation in a [Ca 2+ ] i -dependent manner, leading to paired-pulse facilitation of Ca 2+ currents. Paired recordings from a presynaptic PC axon terminal and a postsynaptic PC soma demonstrated that the paired-pulse facilitation of inhibitory synaptic transmission between PCs showed 4-5th power dependence on that of Ca 2+ currents, and was completely abolished when the Ca 2+ current amplitude was adjusted to be identical. Thus, short-term facilitation of Ca 2+ currents predominantly mediates short-term synaptic facilitation at synapses between PCs.

The Mechanisms and Functions of Synaptic Facilitation

“…They found that the onset of facilitation is rapid (τ on =2 ms) and following a return to resting calcium levels enhancement is short-lived (τ off =31 ms). The time course of paired-pulse facilitation of calcium current had a similar time course at the calyx of Held (Cuttle et al, 1998) and in Purkinje cell presynaptic boutons (Diaz-Rojas et al, 2015) (Figure 5C). These experiments were performed at room temperature, so it is likely that the onset and decay of facilitation of calcium entry is even more rapid at physiological temperatures.…”

“…High-frequency (40 Hz) stimulation does not lead to spike broadening in presynaptic boutons of cerebellar interneurons (Pouzat and Hestrin, 1997; Rowan et al, 2014). Synapses made between cultured Purkinje cells facilitate, but waveform changes are not responsible for facilitation (Diaz-Rojas et al, 2015). …”

“…For action potential driven release at the calyx of Held synaptic facilitation more than doubled synaptic strength, and the facilitation of calcium channels accounted for less than half of the synaptic facilitation (Muller et al, 2008). At synapses between cultured Purkinje cells, there is a 16 % enhancement of calcium current and a doubling of synaptic strength for stimuli separated by 10 ms. Calcium current enhancement is sufficient to account for synaptic enhancement, provided release is steeply dependent on calcium influx (4.5 power, (Diaz-Rojas et al, 2015)). In cultured superior cervical ganglion neurons that express P-type calcium channels, both calcium entry and synaptic strength show facilitation, but quantitative comparisons were complicated because calcium currents were evoked by 5 ms voltage steps and synaptic responses were evoked by action potentials (Yan et al, 2014).…”

Axonal GABAA receptors depolarize presynaptic terminals and facilitate transmitter release in cerebellar Purkinje cells

“…A ; Díaz‐Rojas et al . ; Kawaguchi & Sakaba, ). Cultured PCs were labelled with EGFP by an AAV vector serotype 2, which shows a relatively specific transfection of PCs, and the fluorescence was used to precisely position a patch pipette with a small tip at the selected axonal varicosity.…”

“…Electrophysiological experiments on cultured PCs were performed similarly to previous studies (Díaz‐Rojas et al . ). Whole‐cell patch clamp recording from a cultured PC was performed with an amplifier (EPC10, HEKA, Germany) in a Hepes‐buffered extracellular solution containing (in m m ): 145 NaCl, 5 KCl, 2 CaCl 2 , 1 MgCl 2 , 10 Hepes, and 10 glucose (pH 7.3, 300 mOsm (kg H 2 O) −1 ).…”