Quantitation and Simulation of Single Action Potential-Evoked Ca2+ Signals in CA1 Pyramida Neuron Presynaptic Terminals
Quantitation and Simulation of Single Action Potential-Evoked Ca2+ Signals in CA1 Pyramida Neuron Presynaptic Terminals
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DOI：
10.1523/eneuro.0343-19.2019
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发表时间：
2019-09-01
期刊：
影响因子：
3.4
通讯作者：
Alford, Simon
中科院分区：
文献类型：
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作者：
Hamid, Edaeni;Church, Emily;Alford, Simon
突触前钙离子可引起胞吐、内吞和突触可塑性。然而，由于荧光成像的分辨率有限，突触前分子靶点上的钙离子流量和相互作用很难量化。在不同性别的大鼠中，我们用钙离子染料作为缓冲，测量了单个精索静脉曲张突触前的钙离子浓度，并建立了钙离子弥散模型。用低亲和力染料测得动作电位可诱发钙瞬变，但变化不大(峰值789+/-39 nM，刺激后2ms内；衰减时间119-+/-10ms)。以Ca(2+)染料为缓冲液，测定内源性Ca~(2+)缓冲容量、动作电位诱发的游离[Ca~(2+)](I)和进入终末的总钙总量。这些数据约束的蒙特卡罗(Mcell)模拟钙离子进入、缓冲和清除。用模拟的钙结合蛋白28K缓冲的实验确定的钙离子通量的模拟很好地符合数据，并与随后在4ms内通过扩散在静脉曲张中聚集的钙离子进入一致。重复刺激可引起静脉曲张内游离钙离子浓度升高。然而，在纳米区域模拟，泵和缓冲去除可以忽略不计，而局部扩散占主导地位。因此，进入几十纳米范围内的钙离子没有积累。突触素(SYTL)-钙结合模型表明，即使在10微米的游离精索静脉曲张引起的钙离子，syt1必须在通道的几十纳米范围内，以确保占据其所有的钙结合部位。重复刺激引起短期突触增强，但不改变钙离子完全占据Syt1‘S C2结构域的概率，提示这种增强不是由Ca2+-Syt1相互作用介导的。我们得出结论，在融合机的时空尺度上，激活它们所需的钙离子是扩散主导的。
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 Ca 2 transients with little variation when measured with low-affinity dye (peak amplitude 789 +/- 39 nM, within 2 ms of stimulation; decay times, 119 -+/- 10 ms). Endogenous Ca2+ buffering capacity, action potential-evoked free [Ca2+ ](i), and total Ca2+ amounts entering terminals were determined using Ca(2+)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 calbindin 28K 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 synaptotagminl (sytl)-Ca2+ binding indicates that even with 10 mu 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'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.