Mesoscopic Calcium Imaging of Mouse Brain

Calcium imaging data were acquired using a two-photon random access mesoscope (2p-RAM, Thorlabs)^{11 (link)} controlled with ScanImage (Vidrio Technologies) with a laser (InSight X3, Spectra-Physics) whose excitation wavelength was tuned to 940 nm with the power of ~40 mW at the objective lens. The imaging frame rate was ~9.35 Hz and the imaging resolution was 1 $\times$ 0.4 pixel/μm with eight fields of view (FOVs) of 500 $\times$ 500 μm, which were imaged simultaneously at the depth of ~150–200 μm. The stereotaxic coordinates for these eight FOVs relative to the bregma were: ALM: 2.25 mm AP, 1.65 mm ML; M1a: 1.65 mm AP, 2.75 mm ML; M1p: 0.65 mm AP, 1.75 mm ML; S1fl: 0.25 mm AP, 2.65 mm ML; vS1: −1.15 mm AP, 3.45 mm ML; M2: 0.50 mm AP, 0.45 mm ML; RSC: −1.25 mm AP, 0.55 mm ML; PPC: −1.75 mm AP, 2.05 mm ML. The same FOVs were identified in every session and imaged longitudinally.