In vivo fundus imaging offers non-invasive access to neuron structures and biochemical processes in the retina. However, optical aberrations of the eye degrade the imaging resolution and prevent visualization of subcellular retinal structures. We developed an adaptive optics two-photon excitation fluorescence microscopy (AO-TPEFM) system to correct ocular aberrations based on a nonlinear fluorescent guide star and achieved subcellular resolution for in vivo fluorescence imaging of the mouse retina. With accurate wavefront sensing and rapid aberration correction, AO-TPEFM permits structural and functional imaging of the mouse retina with submicron resolution. Specifically, simultaneous functional calcium imaging of neuronal somas and dendrites was demonstrated. Moreover, the time-lapse morphological alteration and dynamics of microglia were characterized in a mouse model of retinal disorder. In addition, precise laser axotomy was achieved, and degeneration of retinal nerve fibres was studied. This high-resolution AO-TPEFM is a promising tool for non-invasive retinal imaging and can facilitate the understanding of a variety of eye diseases as well as neurodegenerative disorders in the central nervous system.

中文翻译：

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自适应光学双光子显微镜可以在体内实现近衍射极限和功能性视网膜成像。

体内眼底成像可无创地进入视网膜的神经元结构和生化过程。但是，眼睛的光学像差会降低成像分辨率并阻止亚细胞视网膜结构的可视化。我们开发了一种自适应光学两光子激发荧光显微镜（AO-TPEFM）系统，以校正基于非线性荧光导星的眼像差，并实现了小鼠视网膜体内荧光成像的亚细胞分辨率。借助精确的波前感测和快速的像差校正，AO-TPEFM可以对小鼠视网膜进行亚微米分辨率的结构和功能成像。具体而言，证实了神经元体和树突的同时功能性钙成像。此外，在视网膜疾病的小鼠模型中表征了小胶质细胞的时态形态变化和动态。此外，实现了精确的激光轴切术，并研究了视网膜神经纤维的变性。这种高分辨率的AO-TPEFM是用于非侵入性视网膜成像的有前途的工具，可以促进对多种眼部疾病以及中枢神经系统神经退行性疾病的理解。