Using light-sensitive channels to restore vision for blinding retinal diseases

描述(申请人提供)：在视网膜退行性疾病中，光感受器细胞的严重丢失通常会导致完全失明。目前，盲人还没有可用的治疗方法或治愈方法。目前正在考虑的恢复视杆细胞和视锥细胞变性后视力的前瞻性策略包括移植正常的光感受器或祖细胞，以及通过视网膜植入物直接电刺激存活的视网膜神经元。我们正在探索另一种恢复退化视网膜的光敏性的策略：在存活的二级或三级视网膜神经元中表达光敏膜通道。这种做法似乎是可行的。首先，最近已经克隆了直接光门微生物类型视紫红质，通道视紫红质。我们的初步结果表明，在活体正常和视网膜营养不良动物的视网膜神经元中，通道视紫红质-2(ChR2)具有功能性表达，并且能够用光去极化表达ChR2的视网膜神经元膜电位。其次，在盲人患者和动物模型中的研究表明，视网膜的二级和三级神经元部分地保存在病变的视网膜中。此外，还有可能通过基于病毒的基因治疗将外源基因导入视网膜神经元。这种方法的一个潜在优势是，它不涉及将组织或装置引入视网膜，因此可以避免免疫反应和生物相容性并发症。这项提议的目标是通过使用啮齿动物模型来解决对这一战略的可行性和成功至关重要的问题。该建议的具体目的是：1)检测ChR2在体内视网膜神经元中表达的持久性和生物相容性，并探索ChR2在某些功能亚群的视网膜内神经元中的靶向表达；2)表征表达视网膜内神经元的光诱发电流和电压响应；3)检测光感受器变性后存活的视网膜内神经元的生理特性；4)表征ChR2表达后变性视网膜的光反应特性。这些拟议的研究可以为进一步的工作奠定必要的基础，从而产生潜在的治疗失明疾病的方法，以及适用于视网膜研究的电力技术。

DESCRIPTION (provided by applicant): The severe loss of photoreceptor cells in retinal degenerative diseases often leads to total blindness. At present, there is no available treatment or cure to the blind. Prospective strategies currently being considered for restoration of vision following rod and cone degeneration include transplantation of normal photoreceptor or progenitor cells and direct electrical stimulation of the surviving retinal neurons via retinal implants. We are exploring another strategy to restore light sensitivity to the degenerate retinas: the expression of light-sensitive membrane channels in surviving second- or third-order retinal neurons. This approach is plausible. First, directly light-gated microbial-type rhodopsins, channelrhodopsins, have been recently cloned. Our preliminary results have shown the ability of functional expression of channerhodopsin-2 (ChR2) in retinal neurons in normal and retinal dystrophic animals in vivo and the capability to depolarize the membrane potential of ChR2-expressing retinal neurons by light. Second, studies in blind patients and in animal models suggest that the second- and third-order retinal neurons remain, in part, preserved in the diseased retinas. Furthermore, it is possible that foreign genes can be introduced into retinal neurons by viral-based gene therapy. A potential advantage of this approach is that it does not involve the introduction of tissues or devices into the retina and, thus, may avoid the immune reactions and biocompatibility complications. The objective of this proposal is to address questions that are fundamental to the feasibility and success of this strategy by using rodent models. The specific aims of this proposal are: 1) To examine the longevity and biocompatibility of the expression of ChR2 in retinal neurons in vivo and explore the targeting expression of ChR2 in certain functional sub-populations of inner retinal neurons; 2) To characterize the light-evoked current and voltage responses of ChR2 expressing inner retinal neurons; 3) To examine the physiological properties of the surviving inner retinal neurons after photoreceptor degeneration; 4) To characterize the light response properties in the degenerate retinas following the expression of ChR2. These proposed studies could establish the necessary foundation for further work leading to a potential treatment for blinding disorders as well as power techniques with applicability in retinal research.