Sims Ruth R, Bendifallah Imane, Grimm Christiane, Mohamed-Lafirdeen Aysha, Lu Xiaoyu, St-Pierre François, Papagiakoumou Eirini, Emiliani Valentina
Institut de la Vision, Sorbonne Université, INSERM, CNRS, F-75012 Paris, France.
Systems, Synthetic, and Physical Biology Program, Rice University, Houston, TX, USA.
Res Sq. 2023 Jan 24:rs.3.rs-2412371. doi: 10.21203/rs.3.rs-2412371/v1.
Parallel light-sculpting methods have been used to perform scanless two-photon photostimulation of multiple neurons simultaneously during all-optical neurophysiology experiments. We demonstrate that scanless two-photon excitation also enables high-resolution, high-contrast, voltage imaging by efficiently exciting fluorescence in a large fraction of the cellular soma. We present a thorough characterisation of scanless two-photon voltage imaging using existing parallel approaches and lasers with different repetition rates. We demonstrate voltage recordings of high frequency spike trains and sub-threshold depolarizations in intact brain tissue from neurons expressing the soma-targeted genetically encoded voltage indicator JEDI-2P-kv. Using a low repetition-rate laser, we perform recordings from up to ten neurons simultaneously. Finally, by co-expressing JEDI-2P-kv and the channelrhodopsin ChroME-ST in neurons of hippocampal organotypic slices, we perform single-beam, simultaneous, two-photon voltage imaging and photostimulation. This enables in-situ validation of the precise number and timing of light evoked action potentials and will pave the way for rapid and scalable identification of functional brain connections in intact neural circuits.
在全光学神经生理学实验中，平行光雕刻方法已被用于同时对多个神经元进行无扫描双光子光刺激。我们证明，无扫描双光子激发还能通过有效激发大部分细胞体中的荧光来实现高分辨率、高对比度的电压成像。我们使用现有的并行方法和不同重复率的激光对无扫描双光子电压成像进行了全面表征。我们展示了在表达靶向细胞体的基因编码电压指示剂JEDI-2P-kv的完整脑组织中高频尖峰序列和阈下去极化的电压记录。使用低重复率激光，我们同时对多达十个神经元进行记录。最后，通过在海马器官型切片的神经元中共表达JEDI-2P-kv和通道视紫红质ChroME-ST，我们进行了单光束、同步双光子电压成像和光刺激。这使得能够原位验证光诱发动作电位的精确数量和时间，并将为在完整神经回路中快速、可扩展地识别功能性脑连接铺平道路。
