Optogenetic strategies for high-efficiency all-optical interrogation using blue-light-sensitive opsins.
Optogenetic strategies for high-efficiency all-optical interrogation using blue-light-sensitive opsins.
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DOI：
10.7554/elife.63359
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发表时间：
2021-05-25
期刊：
影响因子：
7.7
通讯作者：
Fellin T
中科院分区：
文献类型：
--
作者：
Forli A;Pisoni M;Printz Y;Yizhar O;Fellin T
以高空间分辨率成像和操纵大脑网络的全光学方法是研究神经元集合如何驱动行为的基础。使用双光子全息技术刺激神经元系综需要高灵敏度的致动器，以避免光损伤和加热。此外，双光子可激发视蛋白应该对用于成像的波长的光不敏感。为了实现这一目标，我们开发了一种新的大电导蓝光敏感视蛋白CoChR（stCoChR）的胞体靶向变体。在小鼠体内皮层中，我们将stCoChR的全息双光子刺激与在视蛋白吸收峰处调谐的放大激光和红移指示剂jRCaMP1a的双光子成像相结合。与先前表征的蓝光敏感的体细胞靶向视蛋白在体内相比，stCoChR允许神经元刺激，平均功率低10倍以上，并且没有光谱串扰。stCoChR，调谐放大激光刺激，和红移功能指标的组合有望成为一个强大的工具，在完整的大脑神经网络的大规模审讯。
All-optical methods for imaging and manipulating brain networks with high spatial resolution are fundamental to study how neuronal ensembles drive behavior. Stimulation of neuronal ensembles using two-photon holographic techniques requires high-sensitivity actuators to avoid photodamage and heating. Moreover, two-photon-excitable opsins should be insensitive to light at wavelengths used for imaging. To achieve this goal, we developed a novel soma-targeted variant of the large-conductance blue-light-sensitive opsin CoChR (stCoChR). In the mouse cortex in vivo, we combined holographic two-photon stimulation of stCoChR with an amplified laser tuned at the opsin absorption peak and two-photon imaging of the red-shifted indicator jRCaMP1a. Compared to previously characterized blue-light-sensitive soma-targeted opsins in vivo, stCoChR allowed neuronal stimulation with more than 10-fold lower average power and no spectral crosstalk. The combination of stCoChR, tuned amplified laser stimulation, and red-shifted functional indicators promises to be a powerful tool for large-scale interrogation of neural networks in the intact brain.