Comparing volumetric reconstruction algorithms for light field imaging of high signal-to-noise ratio neuronal calcium transients
SignificanceLight-field microscopy (LFM) enables fast, light-efficient, volumetric imaging of neuronal activity with calcium indicators. Calcium transients differ in temporal signal-to-noise ratio (tSNR) and spatial confinement when extracted from volumes reconstructed by different algorithms. AimWe evaluated the capabilities and limitations of two light-field reconstruction algorithms for calcium fluorescence imaging. ApproachWe acquired light-field image series from neurons either bulk-labeled or filled intracellularly with the red-emitting calcium dye CaSiR-1 in acute mouse brain slices. We compared the tSNR and spatial confinement of calcium signals extracted from volumes reconstructed with synthetic refocusing and Richardson-Lucy 3D deconvolution with and without total variation regularization. ResultsBoth synthetic refocusing and Richardson-Lucy deconvolution resolved calcium signals from single cells and neuronal dendrites in three dimensions. Increasing deconvolution iteration number improved spatial confinement but reduced tSNR compared to synthetic refocusing. Volumetric light-field imaging did not decrease calcium signal tSNR compared to interleaved, widefield image series acquired in matched planes. ConclusionsLFM enables high-volume rate, volumetric imaging of calcium transients in single cells (bulk-labeled), somata and dendrites (intracellular loaded). The trade-offs identified for tSNR, spatial confinement, and computational cost indicate which of synthetic refocusing or deconvolution can better realize the scientific requirements of future LFM calcium imaging applications.