Video rate volumetric Ca2+ imaging across cortical layersusing Seeded Iterative Demixing (SID) microscopy

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Light-field microscopy (LFM) is a scalable approach for volumetric Ca- imaging with the highest volumetric acquisition rates (up to 100Hz). While this has enabled high-speed whole-brain Ca imaging in small semi-transparent specimen, tissue scattering has limited its application in the rodent brain. Here we introduce Seeded Iterative Demixing (SID), a computational source extraction technique that extends LFM to the scattering mammalian cortex. Using GCaMP expressing mice we demonstrate SIDs ability to capture neuronal dynamics in vivo within a volume of 900x900x260um located as deep as 380um in the mouse cortex and hippocampus at 30Hz volume rate while faithfully discriminating signals from neurons as close as 20um, at three orders of magnitude reduced computational cost. The simplicity and scalability of LFM, coupled with the performance of SID opens up a range of new applications including closed loop experiments and is expected to propel its wide dissemination within the neuroscience community.