Ca v 2.2 channels sustain vesicle recruitment at a mature glutamatergic synapse.

The composition of voltage-gated Ca 2+ channel (Ca v ) subtypes that gate action potential (AP)-evoked release changes during development of mammalian CNS synapses. Ca v 2.2 and Ca v 2.3 lose their function in gating evoked release during postnatal synapse maturation. In mature boutons, Ca v 2.1 currents provide the almost exclusive trigger for evoked release and Ca v 2.3 currents are required for the induction of presynaptic long term potentiation. However, the functional significance of Ca v 2.2 remained elusive in mature boutons, although they remain present at active zones and continue contributing significantly to presynaptic Ca 2+ influx. Here, we addressed the functional significance of Ca v 2.2 and Ca v 2.3 at mature parallel-fiber (PF) to Purkinje-neuron synapses of mice of either sex. These synapses are known to exhibit the corresponding developmental Ca v subtype changes in gating release. We addressed two hypotheses, namely that Ca v 2.2 and Ca v 2.3 are involved in triggering spontaneous glutamate release and that they are engaged in vesicle recruitment during repetitive evoked release. We found that spontaneous miniature release is Ca 2+ -dependent. However, experiments with Ca v subtype-specific blockers excluded spontaneous opening of Ca v s as Ca 2+ source for spontaneous glutamate release. Thus, neither Ca v 2.2 nor Ca v 2.3 control spontaneous release from PF boutons. Furthermore, vesicle recruitment during brief bursts of APs was also independent of Ca 2+ influx through Ca v 2.2 and Ca v 2.3. However, Ca v 2.2 but not Ca v 2.3 currents significantly boosted vesicle recruitment during sustained high-frequency synaptic transmission. Thus, in mature PF boutons Ca v 2.2 channels are specifically required to sustain synaptic transmission during prolonged neuronal activity. Significance statement: At young CNS synapses action potential evoked release is gated via three subtypes of voltage gated Ca 2+ channels, Ca v 2.1, Ca v 2.2 and Ca v 2.3. During postnatal maturation, Ca v 2.2 and Ca v 2.3 lose their function in gating evoked release, such that at mature synapses Ca v 2.1 provide the almost exclusive source for triggering evoked release. Ca v 2.3 currents are required for the induction of presynaptic long term potentiation. However, the function of the still abundant Ca v 2.2 in mature boutons remained largely elusive. Here, we studied mature cerebellar parallel-fiber synapses and found that Ca v 2.2 do not control spontaneous release. However, Ca 2+ influx through Ca v 2.2 significantly boosted vesicle recruitment during trains of action potentials. Thus, Ca v 2.2 in mature parallel-fiber boutons participate in sustaining synaptic transmission during prolonged activity.