A genetically encoded ratiometric indicator for chloride: Capturing chloride transients in cultured hippocampal neurons
A genetically encoded ratiometric indicator for chloride: Capturing chloride transients in cultured hippocampal neurons
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DOI：
10.1016/s0896-6273(00)00056-8
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发表时间：
2000-09-01
期刊：
影响因子：
16.2
通讯作者：
Augustine, GJ
中科院分区：
文献类型：
--
作者：
Kuner, T;Augustine, GJ
通过将氯离子敏感的黄色荧光蛋白与氯离子不敏感的青色荧光蛋白融合，构建了一种新型的氯离子光学指示剂。这些荧光团的FRET依赖性发射的比率与Cl-的浓度成比例地变化，并且用于测量培养的海马神经元中的细胞内氯离子浓度（[Cl-]（i））。[CI-]（i）在神经元发育期间降低，与GABA能突触成熟期间从兴奋到抑制的转变一致。GABA（A）受体的局灶性激活引起[CI-]（i）的巨大变化，这可能是GABA依赖性突触传递的使用依赖性抑制的基础。GABA诱导的体细胞[CI-]（i）的变化扩散到树突中，表明[CI-]（i）可以指示突触活动的位置。这种遗传编码的指示剂将允许从高通量药物筛选到直接记录体内突触CI信号的新方法。
We constructed a novel optical indicator for chloride ions by fusing the chloride-sensitive yellow fluorescent protein with the chloride-insensitive cyan fluorescent protein. The ratio of FRET-dependent emission of these fluorophores varied in proportion to the concentration of CI- and was used to measure intracellular chloride concentration ([CI-](i)) in cultured hippocampal neurons. [CI-](i) decreased during neuronal development, consistent with the shift from excitation to inhibition during maturation of GABAergic synapses. Focal activation of GABA(A) receptors caused large changes in [CI-](i) that could underlie use-dependent depression of GABA-dependent synaptic transmission. GABA-induced changes in somatic [CI-](i) spread into dendrites, suggesting that [CI-](i) can signal the location of synaptic activity. This genetically encoded indicator will permit new approaches ranging from high-throughput drug screening to direct recordings of synaptic CI- signals in vivo.