@article {HamidENEURO.0343-19.2019, author = {Edaeni Hamid and Emily Church and Simon Alford}, title = {Quantitation and Simulation of Single Action Potential-Evoked Ca2+ Signals in CA1 Pyramidal Neuron Presynaptic Terminals}, volume = {6}, number = {5}, elocation-id = {ENEURO.0343-19.2019}, year = {2019}, doi = {10.1523/ENEURO.0343-19.2019}, publisher = {Society for Neuroscience}, abstract = {Presynaptic Ca2+ evokes exocytosis, endocytosis, and synaptic plasticity. However, Ca2+ flux and interactions at presynaptic molecular targets are difficult to quantify because fluorescence imaging has limited resolution. In rats of either sex, we measured single varicosity presynaptic Ca2+ using Ca2+ dyes as buffers, and constructed models of Ca2+ dispersal. Action potentials evoked Ca2+ transients with little variation when measured with low-affinity dye (peak amplitude 789 {\textpm} 39 nM, within 2 ms of stimulation; decay times, 119 {\textpm} 10 ms). Endogenous Ca2+ buffering capacity, action potential-evoked free [Ca2+]i, and total Ca2+ amounts entering terminals were determined using Ca2+ dyes as buffers. These data constrained Monte Carlo (MCell) simulations of Ca2+ entry, buffering, and removal. Simulations of experimentally-determined Ca2+ fluxes, buffered by simulated calbindin28K well fit data, and were consistent with clustered Ca2+ entry followed within 4 ms by diffusion throughout the varicosity. Repetitive stimulation caused free varicosity Ca2+ to sum. However, simulated in nanometer domains, its removal by pumps and buffering was negligible, while local diffusion dominated. Thus, Ca2+ within tens of nanometers of entry, did not accumulate. A model of synaptotagmin1 (syt1)-Ca2+ binding indicates that even with 10 {\textmu}M free varicosity evoked Ca2+, syt1 must be within tens of nanometers of channels to ensure occupation of all its Ca2+-binding sites. Repetitive stimulation, evoking short-term synaptic enhancement, does not modify probabilities of Ca2+ fully occupying syt1{\textquoteright}s C2 domains, suggesting that enhancement is not mediated by Ca2+-syt1 interactions. We conclude that at spatiotemporal scales of fusion machines, Ca2+ necessary for their activation is diffusion dominated.}, URL = {https://www.eneuro.org/content/6/5/ENEURO.0343-19.2019}, eprint = {https://www.eneuro.org/content/6/5/ENEURO.0343-19.2019.full.pdf}, journal = {eNeuro} }