Data for: Multiplexed miniaturized two-photon microscopy (M-MINI2Ps)

Head-mounted miniaturized two-photon microscopes are powerful tools to
record neural activity with cellular resolution deep in the mouse brain
during unrestrained, free-moving behavior. Two-photon microscopy, however,
is traditionally limited in imaging frame rate due to the necessity of
raster scanning the laser excitation spot over a large field-of-view
(FOV). Here, we present two multiplexed miniature two-photon microscopes
(M-MINI2Ps) to increase the imaging frame rate while preserving the
spatial resolution. Two different FOVs are imaged simultaneously and then
demixed temporally or computationally. We demonstrate large-scale (500×500
µm2 FOV) multiplane calcium imaging in visual cortex and prefrontal cortex
in freely moving mice during spontaneous exploration, social behavior, and
auditory stimulus. Furthermore, the increased speed of M-MINI2Ps also
enables two-photon voltage imaging at 400 Hz over a 380×150 µm2
FOV in freely moving mice. M-MINI2Ps have compact footprints and are
compatible with the open-source MINI2P. M-MINI2Ps, together with their
design principles, allow the capture of faster physiological dynamics and
population recordings over a greater volume than currently possible in
freely moving mice, and will be a powerful tool in systems neuroscience.