Reduced endogenous Ca²⁺ buffering speeds active zone Ca²⁺ signaling

Fast synchronous neurotransmitter release at the presynaptic active zone is triggered by local Ca²⁺ signals, which are confined in their spatiotemporal extent by endogenous Ca²⁺ buffers. However, it remains elusive how rapid and reliable Ca²⁺ signaling can be sustained during repetitive release. Here, we established quantitative two-photon Ca²⁺ imaging in cerebellar mossy fiber boutons, which fire at exceptionally high rates. We show that endogenous fixed buffers have a surprisingly low Ca²⁺-binding ratio (~15) and low affinity, whereas mobile buffers have high affinity. Experimentally constrained modeling revealed that the low endogenous buffering promotes fast clearance of Ca²⁺ from the active zone during repetitive firing. Measuring Ca²⁺ signals at different distances from active zones with ultra-high-resolution confirmed our model predictions. Our results lead to the concept that reduced Ca²⁺ buffering enables fast active zone Ca²⁺ signaling, suggesting that the strength of endogenous Ca²⁺ buffering limits the rate of synchronous synaptic transmission.