Video rate volumetric Ca²⁺ imaging across cortex using seeded iterative demixing (SID) microscopy.

Abstract

Light-field microscopy (LFM) is a scalable approach for volumetric Ca²⁺ imaging with high volumetric acquisition rates (up to 100 Hz). Although the technology has enabled whole-brain Ca²⁺ imaging in semi-transparent specimens, tissue scattering has limited its application in the rodent brain. We introduce seeded iterative demixing (SID), a computational source-extraction technique that extends LFM to the mammalian cortex. SID can capture neuronal dynamics in vivo within a volume of 900 × 900 × 260 μm located as deep as 380 μm in the mouse cortex or hippocampus at a 30-Hz volume rate while discriminating signals from neurons as close as 20 μm apart, at a computational cost three orders of magnitude less than that of frame-by-frame image reconstruction. We expect that the simplicity and scalability of LFM, coupled with the performance of SID, will open up a range of applications including closed-loop experiments.