High-Resolution Functional Imaging of the Retina

描述（由申请人提供）：我们希望了解维持人类视力的两个重要过程；视觉循环负责通过吸收可见光而漂白的光色素的再生，每个活细胞都需要细胞代谢来提供能量。为了研究这些过程，我们需要一种方法来可视化单个细胞和测量活体眼睛中的分子动力学。所涉及的一些分子本质上是荧光的，但在单光子荧光成像的活体眼睛中是不可接近的，因为激发落在可以穿透眼睛光学的辐射范围之外。通过使用相当长的激发波长，双光子激发荧光成像有可能激发这些难以接近的荧光团，并为成像一些视网膜结构提供内在的对比。在我们最初的实验中，我们使用了自适应光学扫描光检眼镜（AOSLO）来成像活体猕猴眼锥内段的双光子荧光（Hunter et al ., 2011）。通过校正眼睛的纵向色差和物质色散，我们计划提高成像系统的效率，并开发出可靠成像眼睛中多视网膜层的结构和功能的方法。这种能力不仅提供了对正常细胞嵌合及其生化过程的深入了解，还将提高我们对影响这些视网膜生化级联反应的许多疾病的理解，如Stargardt病、黄斑变性和Leber遗传性视神经病变。

DESCRIPTION (provided by applicant): We wish to understand two important processes that sustain human vision; the visual cycle is responsible for regeneration of photopigment bleached by absorption of visible light and cellular metabolism is required in every living cell to provide energy. To study these processes, we need a method to visualize individual cells and measure molecular dynamics in the living eye. Some of the molecules involved are intrinsically fluorescent, but are inaccessible in the living eye with single- photon fluorescence imaging because the excitation falls outside the range of radiation that can penetrate the optics of the eye. By using considerably longer excitation wavelengths, two-photon excited fluorescence imaging has the potential to excite these otherwise inaccessible fluorophores and provide intrinsic contrast for imaging a number of retinal structures. In our initial experiments, we used an adaptive optics scanning light ophthalmoscope (AOSLO) to image two-photon fluorescence from cone inner segments in the living macaque eye (Hunter et al, 2011). By correcting the longitudinal chromatic aberration and material dispersion of the eye, we plan to improve the efficiency of our imaging systems and develop methodology for reliably imaging both structure and function of multiple retinal layers in the eye. Not only will this capability provide insight ito normal cell mosaics and their biochemical processes, it will also improve our understanding of many diseases that impact these retinal biochemical cascades such as Stargardt's disease, macular degeneration and Leber's hereditary optic neuropathy. PUBLIC HEALTH RELEVANCE: My new laboratory at the University of Rochester will develop two-photon fluorescence imaging into a highly efficient technique to visualize non-invasively for the first time neural cells in the back of the eye and to monitor important cellular processes in the living eye. This research could lead to improvements in the diagnosis of some retinal diseases, monitoring of their progression and establishing the efficacy of treatment strategies.