Developmental changes in calcium domain at the presynaptic terminal of the central synapse

突触前神经末梢的Ca结构域决定突触传递的速度和保真度。在大鼠听性脑干Held萼，终末内注射EGTA可在出生后第7天听力开始前使EPSC振幅衰减（P <0.05）。但这种抑制作用几乎可以忽略不计后，听力发作（P14），这意味着钙结构域是从微域到纳米域发育转化。为了观察神经末梢中的Ca结构域，我采用了局部共聚焦点检测方法和低亲和力Ca^2+指示剂俄勒冈州绿色BAPTA/5 N，以评估P7和P14末梢中单个动作电位（AP）诱发的Ca^2+梯度的时空分布。在P7时，几乎所有共焦点沿着突触表面都观察到Ca瞬变，但在P14时观察到的频率较低，幅度变小。电压钳实验表明，突触前动作电位的缩短和钙通道密度的降低都有助于这种减少。为了研究Ca结构域发育变化的两种机制，我同时进行了突触前和突触后的电生理记录。用肽类Ca^<2+>通道阻断剂减少P7末端功能性Ca通道的数量可显著降低末端内注射EGTA对EPSC的抑制作用。相反，在P13-15突触，使用TEA延长AP持续时间增加EGTA对EPSC的作用。我们的结论是，发育过程中Ca^2+通道密度和AP时程的降低都通过开放通道密度的降低，促进了Held突触萼Ca结构域的发育转化。

The Ca domain in presynaptic nerve terminal can determine speed and fidelity of synaptic transmission. At the calyx of Held, in the rat auditory brainstem, intraterminal injection of EGTA attenuates EPSC amplitude before hearing onset postnatal day (P)7). But this inhibitory effect is almost negligible after hearing onset (P14), implying that Ca domain is developmentally transformed from microdomain into nanodomain. To visualize Ca domain in the nerve terminal, I employed a localized confocal spot detection method and a low affinity Ca^<2+> indicator Oregon Green BAPTA/5N, to assess the spatiotemporal profile of single action potential (AP)-evoked Ca^<2+> gradients in P7 and P14 terminals. Ca transient were observed in almost every confocal spot locations along synaptic surface at P7, but they were less frequently observed and their amplitude became smaller at P14. Voltage-clamp experiments revealed that both shortening of presynaptic action potential and decrease in Ca channel density contribute to this reduction in Catransients. To investigate these two mechanisms underlie developmental changes in Ca domain, I performed simultaneous pre^- and postsynaptic electrophysiological recordings. Reducing the number of functional Ca channels in P7 terminals with peptide Ca^<2+> channel blockers markedly reduced the inhibitory effect of intraterminal EGTA injection on EPSCs. Conversely, at P13-15 synapses, prolongation of AP duration using TEA increased the effect of EGTA on EPSCs. We conclude that both the developmental decreases in Ca^<2+> channel density and AP duration contribute to developmental transformation of Ca domain at the calyx of Held synapse, through a decrease in the density of open channels.