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High-resolution two-photon transcranial imaging of brain using direct wavefront sensing

Congping Chen,Congping Chen,Congping Chen,Zhongya Qin,Zhongya Qin,Zhongya Qin,Sicong He,Sicong He,Sicong He,Shaojun Liu,Shaojun Liu,Shun-Fat Lau,Shun-Fat Lau,Shun-Fat Lau,Wanjie Wu,Wanjie Wu,Wanjie Wu,Dan Zhu,Dan Zhu,Nancy Y. Ip,Nancy Y. Ip,Nancy Y. Ip,Jianan Y. Qu,Jianan Y. Qu,Jianan Y. Qu

Imaging of the brain in its native state at high spatial resolution poses major challenges to visualization techniques. Two-photon microscopy integrated with the thinned-skull or optical clearing skull technique provides a minimally invasive tool for in vivo imaging of the cortex of mice without activating immune response and inducing brain injury. However, the imaging contrast and spatial resolution are severely compromised by the optical heterogeneity of the skull, limiting the imaging depth to the superficial layer. In this work, an optimized configuration of an adaptive optics two-photon microscope system and an improved wavefront sensing algorithm are proposed for accurate correction for the aberrations induced by the skull window and brain tissue. Using this system, we achieved subcellular resolution transcranial imaging of layer 5 pyramidal neurons up to 700 μm below pia in living mice. In addition, we investigated microglia–plaque interaction in living brain of Alzheimer’s disease and demonstrated high-precision laser dendrotomy and single-spine ablation.