An Exclusion Zone for Ca(2+) Channels around Docked Vesicles Explains Release Control by Multiple Channels at a CNS Synapse

The spatial arrangement of Ca(2+) channels and vesicles remains unknown for most CNS synapses, despite of the crucial importance of this geometrical parameter for the Ca(2+) control of transmitter release. At a large model synapse, the calyx of Held, transmitter release is controlled by several Ca(2+) channels in a "domain overlap" mode, at least in young animals. To study the geometrical constraints of Ca(2+) channel placement in domain overlap control of release, we used stochastic MCell modelling, at active zones for which the position of docked vesicles was derived from electron microscopy (EM). We found that random placement of Ca(2+) channels was unable to produce high slope values between release and presynaptic Ca(2+) entry, a hallmark of domain overlap, and yielded excessively large release probabilities. The simple assumption that Ca(2+) channels can be located anywhere at active zones, except below a critical distance of ~ 30 nm away from docked vesicles ("exclusion zone"), rescued high slope values and low release probabilities. Alternatively, high slope values can also be obtained by placing all Ca(2+) channels into a single supercluster, which however results in significantly higher heterogeneity of release probabilities. We also show experimentally that high slope values, and the sensitivity to the slow Ca(2+) chelator EGTA-AM, are maintained with developmental maturation of the calyx synapse. Taken together, domain overlap control of release represents a highly organized active zone architecture in which Ca(2+) channels must obey a certain distance to docked vesicles. Furthermore, domain overlap can be employed by near-mature, fast-releasing synapses.