Imaging the neuronal activity throughout the brain with high temporal and spatial resolution is an important step in understanding how the brain works. Two-photon laser scanning microscopy with fluorescent calcium indicators has enabled this type of experiments in vivo. Most of these microscopes acquire images serially, with a single laser beam, limiting the overall imaging speed. To overcome this limit, multiple beamlets can be used to image in parallel multiple regions. Here, we demonstrate a novel scheme of a two-photon laser-scanning microscope that can simultaneously record neuronal activity at multiple planes of the sample with a single photomultiplier tube. A spatial light modulator is used to generate the designated multiple beamlets, and a constrained non-negative matrix factorization algorithm is used to demix the signals from multiple scanned planes. We simultaneously record neuronal activity of multiple layers of a mouse cortex at 10 fps in vivo. This novel imaging scheme provides a powerful tool for mapping the brain activity.
Weijian Yang, Jae-eun K Miller, Luis Carrillo-Reid, Eftychios Pnevmatikakis, Liam Paninski, Darcy S. Peterka, and Rafael Yuste, "Two-photon multiplane imaging of neural circuits
(Conference Presentation)," Proc. SPIE 9690, Clinical and Translational Neurophotonics; Neural Imaging and Sensing; and Optogenetics and Optical Manipulation, 969016 (Presented at SPIE BiOS: February 15, 2016; Published: 26 April 2016); https://doi.org/10.1117/12.2219843.4848636561001.
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