Assessment of Retinal Function in Health and Disease From Mouse To Human

评估从小鼠到人类的健康和疾病中的视网膜功能

• 批准号: 9535533
• 负责人: Frans Vinberg
• 金额: $ 23.64万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-01 至 2020-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9535533
• 关键词: Affect; Age related macular degeneration; Animal Model; Animal Testing; Area; Argon; Blindness; Burn injury; Cell physiology; Cells; Cessation of life; Clinical Trials; Collaborations; Cone; Data; Developed Countries; Developing Countries; Devices; Diabetes Mellitus; Diabetic Retinopathy; Disease; Disease model; Dissection; Electrophysiology (science); Electroretinography; Environment; Euthanasia; Eye; Eye diseases; Family suidae; Functional disorder; Genetic; Glaucoma; Health; Human; Individual; Lasers; Lateral; Light; Link; Macular degeneration; Measures; Mediating; Methodology; Methods; Molecular; Muller' s cell; Mus; Mutation; Neuroglia; Neurons; Noise; Opsin; Peripheral; Pharmaceutical Preparations; Pharmacology; Photoreceptors; Positioning Attribute; Primates; Protocols documentation; Retina; Retinal; Retinal Diseases; Retinitis Pigmentosa; Rhodopsin; Rodent; Safety; Shapes; Signal Transduction; Streptozocin; Techniques; Time; Vertebrate Photoreceptors; Vision; Wild Type Mouse; base; cell type; design; experimental study; fovea centralis; ganglion cell; in vivo; injured; macula; mouse model; multi-electrode arrays; novel; patch clamp; public health relevance; response; scale up; screening

 DESCRIPTION (provided by applicant): Leading causes of blindness in developed countries include macular degeneration, glaucoma and diabetic retinopathy which are all manifested as dysfunction and degeneration of specific cell types in the retina. The molecular mechanisms and pathophysiology of these diseases are not well understood. Widely used techniques to study the pathophysiology of retinal diseases include In Vivo electroretinogram (ERG) and single cell electrophysiology. In Vivo ERG can assess the overall health state of the retina but is limited in providing quantitative information about the cellular and molecular origin of the functional deficits in a diseased retina. Single cell recordings, on the other hand, provide quantitative data from individual cells but are challenging, offer only limited recording time and are not easily scaled up to assess the function and signaling across the whole retina. The objective of this proposal is to advance techniques and methodology to dissect the function of photoreceptor, bipolar and Müller-glial cells in the intact mouse, primate, pig and ultimately human retinas. I wll first develop methodology to quantitatively assess the intrinsic functional "state" of these cells y using Ex Vivo ERG from isolated wild-type mouse retinas (K99). Ex Vivo ERG provides information about the average functional state of all cells across the lateral axis of the retina. However, some of the retinal diseases affect retina only locally (e.g. DR) or target primarily ganglion cells (e.g. glaucoma) that will not necessarily be observable in the Ex Vivo ERG signal. One objective of this proposal is to develop a novel device combining Ex Vivo ERG and multi-electrode array (MEA) methods. This device will be used to assess the local function of photoreceptors, bipolar, Müller-glial and ganglion cells across the whole wild-type mouse retinas (K99). Ex Vivo ERG (K99) will be applied to determine how the function of rod and cone bipolar, Müller-glial and ganglion cells are affected in the mouse models of retinitis pigmentosa (RP, P23H rhodopsin mutation) and diabetic retinopathy (DR, Streptozotocin-induced diabetes) and MEA-ERG device will be used to assess the local function of the retina that has been focally injured by laser (K99 and R00). These experiments will advance the understanding of pathophysiology of these diseases known to affect primarily outer and inner retina, respectively. The methodology developed for mouse retinas will be used to establish protocols to dissect the function of photoreceptors, bipolar and Müller-glial cells in primate, pig, and ultimately human donor retinas. I will first develop the recording protocols to obtain viable responses from primate retinas dissected from eye balls enucleated immediately following the euthanasia (K99). Then, in collaboration with Dr. Hanneken, we will determine the acceptable time frame between death and enucleation by using pig eyes (R00). Finally, based on the primate and pig experiments we will design and conduct recordings to assess function of photoreceptor, bipolar and Müller glia cells in a macula and peripheral regions of the human retina (R00).

 描述（申请人提供）：发达国家失明的主要原因包括黄斑变性、青光眼和糖尿病性视网膜病变，这些疾病均表现为视网膜中特定细胞类型的功能障碍和变性。这些疾病的分子机制和病理生理学尚不清楚。研究视网膜疾病病理生理学的广泛使用的技术包括体内视网膜电图（ERG）和单细胞体内电生理学，可以评估整体。视网膜的健康状况，但在提供有关患病视网膜功能缺陷的细胞和分子起源的定量信息方面受到限制，另一方面，单细胞记录提供定量数据。 该提案的目的是推进技术和方法来剖析光感受器、双极和穆勒的功能。 -完整小鼠、灵长类动物、猪以及最终人类视网膜中的神经胶质细胞，我将首先使用来自分离的野生型小鼠的体外 ERG 来定量评估这些细胞的内在功能“状态”。视网膜（K99）。体外 ERG 提供有关视网膜横轴上所有细胞的平均功能状态的信息。然而，一些视网膜疾病仅影响视网膜局部（例如 DR）或主要针对神经节细胞（例如青光眼）。该提案的一个目标是开发一种结合了 Ex Vivo ERG 和多电极阵列 (MEA) 的新型设备。该装置将用于评估整个野生型小鼠视网膜 (K99) 的光感受器、双极细胞、穆勒神经胶质细胞和神经节细胞的局部功能，以确定如何应用离体 ERG (K99) 的功能。在色素性视网膜炎（RP、P23H 视紫红质突变）和糖尿病小鼠模型中，视杆细胞和视锥细胞双极细胞、米勒神经胶质细胞和神经节细胞受到影响视网膜病变（DR，链脲佐菌素诱导的糖尿病）和 MEA-ERG 装置将用于评估激光（K99 和 R00）局部损伤的视网膜的局部功能。这些实验将增进对这些已知疾病的病理生理学的理解。分别主要影响外视网膜和内视网膜。为小鼠视网膜开发的方法将用于建立解剖光感受器、双极细胞和米勒神经胶质细胞功能的方案。我将首先在灵长类动物、猪以及最终人类捐赠者的视网膜中开发记录协议以获得灵长类动物的可行反应。 安乐死后立即从眼球上切下视网膜（K99） 然后，我们将与 Hanneken 博士合作，使用猪眼确定死亡和摘除之间可接受的时间范围（R00）。我们将设计并进行记录实验，以评估黄斑和人类视网膜周边区域的感光细胞、双极细胞和米勒神经胶质细胞的功能（R00）。