In vitro functional imaging in brain slices using fast voltage-sensitive dye imaging combined with whole-cell patch recording
In vitro functional imaging in brain slices using fast voltage-sensitive dye imaging combined with whole-cell patch recording
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DOI：
10.1038/nprot.2007.539
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发表时间：
2008-01-01
期刊：
影响因子：
14.8
通讯作者：
Coulter, Douglas A.
中科院分区：
文献类型：
--
作者：
Carlson, Greg C.;Coulter, Douglas A.
在许多大脑区域，回路连接被分成特定的层或肾小球。这些解剖学离散区域的功能成像对于表征电路特性特别有用。电压敏感染料（VSD）成像直接分析神经元活动的时空动态，包括相关神经元的功能连接。在空间隔离的结构中，VSD 成像可以定义生理学和连接性如何相互作用，并可以识别神经和精神疾病模型中的功能异常。在以下协议中，我们描述了基于快速电荷耦合器件 (CCD) 的 VSD 成像与同时全细胞贴片记录相结合所需的体外切片制备、落射荧光设置和分析。添加单细胞记录可验证成像结果，并可以揭示单细胞活动与 VSD 成像群体反应之间的关系；在同步激活的神经元中，全细胞记录的 V-m 变化可以准确地代表驱动 VSD 反应的群体 Vm 变化。因此，VSD 成像和全细胞贴片相结合的方法提供了从单细胞电生理学到复杂局部电路响应的实验分辨率。
In many brain areas, circuit connectivity is segregated into specific lamina or glomerula. Functional imaging in these anatomically discrete areas is particularly useful in characterizing circuit properties. Voltage-sensitive dye (VSD) imaging directly assays the spatiotemporal dynamics of neuronal activity, including the functional connectivity of the neurons involved. In spatially segregated structures, VSD imaging can define how physiology and connectivity interact, and can identify functional abnormalities in models of neurological and psychiatric disorders. In the following protocol, we describe the in vitro slice preparation, epifluorescence setup and analyses necessary for fast charge-coupled device (CCD)-based VSD imaging combined with simultaneous whole-cell patch recording. The addition of single-cell recordings validates imaging results, and can reveal the relationship between single-cell activity and the VSD-imaged population response; in synchronously activated neurons, this change in whole-cell recorded V-m can accurately represent population Vm changes driving the VSD responses. Thus, the combined VSD imaging and whole-cell patch approach provides experimental resolution spanning single-cell electrophysiology to complex local circuit responses.