A large field-of-view, single-cell-resolution two- and three-photon microscope for deep and wide imaging
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Citation
A.T. Mok, T. Wang, S. Zhao, K.E. Kolkman, D. Wu, D.G. Ouzounov, C. Seo, C. Wu, J.R. Fetcho, C. Xu. 2024. "A large field-of-view, single-cell-resolution two- and three-photon microscope for deep and wide imaging" eLight, Volume 4, Issue 1. https://doi.org/10.1186/s43593-024-00076-4
Abstract
In vivo imaging of large-scale neuronal activity plays a pivotal role in unraveling the function of the brain's circuitry. Multiphoton microscopy, a powerful tool for deep-tissue imaging, has received sustained interest in advancing its speed, field of view and imaging depth. However, to avoid thermal damage in scattering biological tissue, field of view decreases exponentially as imaging depth increases. We present a suite of innovations to optimize three-photon microscopy for large field-of-view imaging at depths unreachable by two-photon microscopy. These techniques enable us to image neuronal activities of transgenic animals expressing protein calcium sensors in a ~ 3.5-mm diameter field-of-view with single-cell resolution in the deepest cortical layer of mouse brains. We further demonstrate simultaneous large field-of-view two-photon and three-photon imaging, subcortical imaging in the mouse brain, and whole-brain imaging in adult zebrafish. The demonstrated techniques can be integrated into typical multiphoton microscopes to enlarge field of view for system-level neural circuit research.
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© The Author(s) 2024. Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.