After motion correction, each image frame was re-expressed in units of relative changes in fluorescence, ΔF(t)/F0 = (F(t) − F0)/F0, where F0 is the mean image obtained by averaging the entire video. Spatial filters corresponding to individual neurons were identified using Inscopix Data Processing Software (v1.0.0.2273, Inscopix, Palo Alto, CA) based on principal and independent component analyses 54 (link). Cells spatial filters were identified based on the calcium data acquired over the entire session. For each filter, all pixels were then zeroed with values <50% of that filter’s maximum intensity. To obtain time traces of calcium activity, each cell thresholded spatial filter was added to the ΔF(t)/F0 movie. As previously described 55 (link), the extracted spatial filters generally had sizes, morphologies and activity traces that were characteristic of individual neurons. Every cell included in the analyses was validated by visual inspection.
Cell registration across 10 recording sessions using Inscopix Data Processing Software (v1.0.0.2273, Inscopix, Palo Alto, CA). This corrected for potential slight translations, rotations, or focus-dependent magnification changes between sessions and yielded the location of each cell in the reference coordinate system.