A new strategy for vision restoration based on melanopsin transduction mechanisms

基于黑视蛋白转导机制的视力恢复新策略

• 批准号: 10502717
• 负责人: Zheng Jiang
• 金额: $ 40万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10502717
• 关键词: Animals; Blindness; Brain; Cell Death; Cells; Cyclic Nucleotides; Data; Death Rate; Dominant-Negative Mutation; Effectiveness; FOS gene; Family; Future; G-Protein-Coupled Receptors; Image; Inflammatory Response; Ion Channel; Kinetics; Knowledge; Light; Measures; Mediating; Modality; Modification; Monitor; Mus; Names; PDE4A4; Pathway interactions; Pattern; Phase; Photophobia; Photoreceptors; Photosensitivity; Phototransduction; Proteins; Retina; Retinal Diseases; Retinal Ganglion Cells; Reverse Transcriptase Polymerase Chain Reaction; Signal Pathway; Signal Transduction; Signaling Molecule; Testing; Therapeutic; Vertebrate Photoreceptors; Viral; Vision; Visual; Water; Work; activity marker; base; behavior test; cyclic-nucleotide gated ion channels; density; gene therapy; immunocytochemistry; improved; innovation; knowledge base; light intensity; melanopsin; member; mouse model; multi-electrode arrays; mutant; novel; optogenetics; overexpression; patch clamp; photoactivation; photoreceptor degeneration; relating to nervous system; response; restoration; retinal neuron; sight restoration; temporal measurement; theories; treatment strategy; trend; vision aid

PROJECT SUMMARY A pressing challenge in the treatment of retinal diseases is to restore vision in the retina with irreversible photoreceptor degeneration. In theory, even in the absence of rods and cones, such diseased retina should still be able to sense light through intrinsically photosensitive retinal ganglion cells (ipRGCs) using a photopigment named melanopsin. However, the kinetics of melanopsin-mediated light response is slow, thus giving only very poor temporal resolution and therefore largely limiting its ability to provide image-forming information. For the same reason, a vision restoration approach previously proposed – which made use of virally expressed melanopsin to endow light sensitivity to conventional retinal ganglion cells (RGCs) – is also limited by the slow response kinetics of melanopsin. Our recent study has firmly established that melanopsin-mediated phototransduction is rate-limited by its downstream components and that it can be accelerated by virally expressed ion channels with faster kinetics. We propose to develop a novel vision restoration strategy by accelerating melanopsin signaling via manipulating downstream transduction components in photoreceptor degenerated mouse models. We have two major aims. In Aim 1, we shall focus on enhancing the light response of endogenous melanopsin in M4- and M5-subtypes of ipRGCs because they have been shown to project to image-forming brain centers, allowing them to contribute to the image-forming vision. We have discovered that M4-cells respond to melanopsin photoactivation by elevating intracellular levels of cyclic nucleotides (cNMP) and subsequent opening of cNMP-sensitive HCN channels. Our new preliminary data suggest that M5 cells also use the HCN pathway. We have further shown that the response of the HCN-dependent signaling pathway can be sped up by introducing a member of the cyclic nucleotide-gated channel family, CNGA2, resulting in faster response kinetics and larger amplitudes, proving the concept of our strategy. In Aim 2, we shall restore light response in conventional retinal ganglion cells using virally expressed melanopsin together with faster signaling molecules. Our multielectrode array results show that simultaneously expressing melanopsin and CNGA2 in cRGCs provides higher light sensitivities and faster rising phases than expressing melanopsin alone. Using water-based vision-guided maze tests, we have further found that exogenous melanopsin and CNGA2 not only restore light sensitivity but also confer pattern vision in photoreceptor-degenerated animals. Together, these exciting preliminary observations raise the prospect of using CNG channels and/or modifications to other phototransduction components of melanopsin as a therapeutic modality to restore vision following photoreceptor degeneration. Innovation. All optogenetic approaches proposed thus far focus on finding a better light-sensing protein with higher expression, better light sensitivity, and faster response kinetics. We are the first to exploit downstream components and novel signaling molecules of melanopsin for vision restoration purposes. This represents a conceptual innovation and an out-of-the-box strategy for vision restoration.

项目摘要 视网膜疾病治疗中的一个紧迫挑战是在具有不可逆损伤的视网膜中恢复视力。 光感受器变性理论上，即使没有视杆细胞和视锥细胞，这种病变的视网膜仍然应该 能够通过使用一种光敏色素的内在光敏视网膜神经节细胞（ipRGC）来感知光 叫做黑视素然而，黑视素介导的光响应的动力学是缓慢的，因此仅给出非常小的光响应。 时间分辨率差，因此极大地限制了其提供成像信息的能力。为 出于同样的原因，先前提出的一种视力恢复方法--利用病毒表达的 黑视素赋予常规视网膜神经节细胞（RGC）光敏感性-也受到视网膜神经节细胞（RGCs）的缓慢生长的限制。 黑视素的反应动力学。我们最近的研究已经确定，黑视素介导的 光转导受其下游组分的速率限制，并且可以通过病毒加速 表达的离子通道具有更快的动力学。我们建议开发一种新的视力恢复策略， 通过操纵感光细胞中的下游转导元件加速黑视蛋白信号传导 退化的小鼠模型。我们有两个主要目标。在目标1中，我们将专注于增强光响应 内源性黑视素在ipRGC的M4-和M5-亚型中的表达，因为它们已被证明投射到 图像形成的大脑中心，使他们有助于图像形成的视觉。我们发现 M4-细胞通过升高细胞内环核苷酸（cNMP）水平来响应黑视素光活化， 随后打开cNMP敏感的HCN通道。我们新的初步数据表明，M5细胞也使用 HCN途径。我们进一步表明，HCN依赖性信号通路的反应可以是 通过引入环核苷酸门控通道家族的成员CNGA 2来加速， 响应动力学和更大的振幅，证明了我们的策略的概念。在目标2中，我们将恢复光明 使用病毒表达的黑视蛋白以及更快的信号传导在常规视网膜神经节细胞中的反应 分子。我们的多电极阵列结果表明，在细胞中同时表达黑视素和CNGA 2， 与单独表达黑视蛋白相比，cRGC提供更高的光敏感性和更快的上升期。使用 基于水的视觉引导迷宫测试，我们进一步发现，外源性黑视蛋白和CNGA 2不仅 恢复光敏感性，但也赋予光感受器退化的动物图案视觉。所有这些 令人兴奋的初步观察提出了使用CNG通道和/或对其他通道的修改的前景。 黑视蛋白的光转导成分作为光感受器损伤后恢复视力的治疗方式 退化创新到目前为止，所有提出的光遗传学方法都集中在寻找更好的光传感器。 蛋白质具有更高的表达、更好的光敏感性和更快的响应动力学。我们是第一个 用于视力恢复目的的黑视蛋白的下游组分和新的信号分子。这 代表了一种概念创新和一种开箱即用的视力恢复策略。