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通过双光子激发荧光显微镜比较用于监测哺乳动物大脑动作电位的基因编码钙指示剂。

Comparison of genetically encoded calcium indicators for monitoring action potentials in mammalian brain by two-photon excitation fluorescence microscopy.

机构信息

Dalhousie University , Department of Physiology and Biophysics, Halifax, Nova Scotia B3H 4R2, Canada.

Saitama University , Brain Science Institute, Saitama 338-8570, Japan.

出版信息

Neurophotonics. 2015 Apr;2(2):021014. doi: 10.1117/1.NPh.2.2.021014. Epub 2015 Apr 30.

Abstract

Imaging calcium transients associated with neuronal activity has yielded important insights into neural physiology. Genetically encoded calcium indicators (GECIs) offer conspicuous potential advantages for this purpose, including exquisite targeting. While the catalogue of available GECIs is steadily growing, many newly developed sensors that appear promising in vitro or in model cells appear to be less useful when expressed in mammalian neurons. We have, therefore, evaluated the performance of GECIs from two of the most promising families of sensors, G-CaMPs [Nat. Biotechnol.2(2), 137-141 (2001)] and GECOs [Science2(2), 1888-1891 (2011)], for monitoring action potentials in rat brain. Specifically, we used two-photon excitation fluorescence microscopy to compare calcium transients detected by G-CaMP3; GCaMP6f; G-CaMP7; Green-GECO1.0, 1.1 and 1.2; Blue-GECO; Red-GECO; Rex-GECO0.9; Rex-GECO1; Carmine-GECO; Orange-GECO; and Yellow-GECO1s. After optimizing excitation wavelengths, we monitored fluorescence signals associated with increasing numbers of action potentials evoked by current injection in CA1 pyramidal neurons in rat organotypic hippocampal slices. Some GECIs, particularly Green-GECO1.2, GCaMP6f, and G-CaMP7, were able to detect single action potentials with high reliability. By virtue of greatest sensitivity and fast kinetics, G-CaMP7 may be the best currently available GECI for monitoring calcium transients in mammalian neurons.

摘要

对与神经元活动相关的钙瞬变进行成像,为神经生理学研究提供了重要的见解。基因编码的钙指示剂(GECIs)在这方面具有明显的优势,包括精确的靶向性。虽然可用的 GECI 目录在不断增加,但许多在体外或模型细胞中表现出前景的新开发传感器,在表达于哺乳动物神经元时似乎用处不大。因此,我们评估了两种最有前途的传感器家族(G-CaMPs [Nat. Biotechnol.2(2), 137-141 (2001)] 和 GECOs [Science2(2), 1888-1891 (2011))的 GECIs 在大鼠大脑中监测动作电位的性能。具体来说,我们使用双光子激发荧光显微镜比较了 G-CaMP3、GCaMP6f、G-CaMP7、Green-GECO1.0、1.1 和 1.2、Blue-GECO、Red-GECO、Rex-GECO0.9、Rex-GECO1、Carmine-GECO、Orange-GECO 和 Yellow-GECO1s 检测到的钙瞬变。在优化激发波长后,我们监测了电流注入诱发 CA1 锥体神经元中增加的动作电位数量相关的荧光信号。一些 GECIs,特别是 Green-GECO1.2、GCaMP6f 和 G-CaMP7,能够以高可靠性检测单个动作电位。由于最大的灵敏度和快速动力学,G-CaMP7 可能是目前监测哺乳动物神经元钙瞬变的最佳 GECI。

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