Extended field-of-view two-photon imaging in the deep brain of freely behaving mice using lightweight microfabricated corrective optics

Investigating neural circuits underlying complex behavior requires high-resolution imaging of large neuronal populations during naturalistic conditions. Miniaturized two-photon microscopes (MINI2Ps) enable deep brain imaging in freely moving animals when coupled with gradient index (GRIN)-based microendoscopes. However, intrinsic optical aberrations in GRIN-based microendoscopes degrade spatial resolution and limit the effective field-of-view (FOV). Here, we address these limitations using lightweight microfabricated optics to correct GRIN aberrations during deep brain MINI2P imaging. The aberration corrected system demonstrated improved and more homogeneous axial resolution across the FOV, resulting in more than fourfold increase in effective FOV. We applied the corrected system to the ventral posteromedial thalamus in freely moving mice and performed optical mapping of head direction (HD) tuned cells. These measurements revealed the absence of spatial clustering of HD preference, suggesting spatially sparse HD coding in sensory thalamus. Combining the MINI2P with aberration corrected GRIN-based microendoscopes is a powerful approach for large-scale and high-resolution functional imaging of deep brain circuits during naturalistic behavior.