High-fidelity estimates of spikes and subthreshold waveforms from 1-photon voltage imaging in vivo

The ability to probe the membrane potential of multiple genetically defined neurons simultaneously would have a profound impact on neuroscience research. Genetically encoded voltage indicators are a promising tool for this purpose, and recent developments have achieved a high signal-to-noise ratio in vivo with 1-photon fluorescence imaging. However, these recordings exhibit several sources of noise and signal extraction remains a challenge. We present an improved signal extraction pipeline, spike-guided penalized matrix decomposition-nonnegative matrix factorization (SGPMD-NMF), which resolves supra- and subthreshold voltages in vivo. The method incorporates biophysical and optical constraints. We validate the pipeline with simultaneous patch-clamp and optical recordings from mouse layer 1 in vivo and with simulated and composite datasets with realistic noise. We demonstrate applications to mouse hippocampus expressing paQuasAr3-s or SomArchon1, mouse cortex expressing SomArchon1 or Voltron, and zebrafish spines expressing zArchon1.