Cell-targeted Gold Nanoparticles for Photo-excitation fo Retinal Ganglion Cells

用于视网膜神经节细胞光激发的细胞靶向金纳米颗粒

• 批准号: 9999837
• 负责人: FRANCISCO J BEZANILLA
• 金额: $ 1.96万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-04-01 至 2020-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9999837
• 关键词: Cell; targeted; Gold; Nanoparticles; Photo

ABSTRACT Retinal ganglion cells (RGCs) convey visual signals to the brain in the form of action potentials, the initiation and axonal propagation of which depend on the activation of RGC voltage-gated sodium channels (NaVs). In photoreceptor degenerative diseases such as age-related macular degeneration (AMD), inner retinal neurons including RGCs in many cases remain intact and capable of generating action potential responses. RGCs thus represent a logical target for approaches aimed at restoring vision in advanced-stage AMD and related retinal diseases, by bypassing the nonfunctioning rod and cone photoreceptors and establishing direct RGC responsiveness to light. In a recent study of dorsal root ganglion cells (a non-retinal neuron widely studied as a model action-potential-generating cell type) and in hippocampal slice preparations, we have shown that gold nanoparticles (AuNPs) conjugated with a cell-targeting biomolecule enable robust light-induced NaV activation and resulting action potential generation. Essential features of the cell-targeted AuNP technique are: (i) functionalization of the light-absorbing AuNPs to localize them at or near the NaVs; (ii) upon the AuNP’s plasmon absorption of a millisecond/submillisecond light flash, AuNP radiation of the light energy as a nondamaging pulse of heat that creates a localized, transient, depolarizing capacitive current across the plasma membrane; and (iii) resulting activation, i.e., channel opening, of neighboring NaVs and thus action potential initiation by this depolarization. In this application we propose exploratory research to apply this AuNP approach to RGCs in the living eye of the rat. The project’s goal is to establish AuNP treatment conditions that achieve robust AuNP-mediated RGC photo-responsiveness in vivo. In rats for which rod and cone photoreceptor signaling to RGCs has been suppressed pharmacologically, we will intra-vitreally deliver AuNP conjugates designed for binding to the immediate vicinity of the RGC NaVs. Following treatment with the AuNP conjugates, we will employ in vivo recording of electroretinographic (ERG) signals associated with RGC activity, and of visual evoked potentials (VEPs), to analyze properties of RGC electrophysiological responses to AuNP photo-excitation. The research will involve variation of the size/structure of the AuNP, of the AuNP- conjugated RGC-targeting component, and of the duration and energy [(intensity) x (duration)] of AuNP- excitatory flashes. Accompanying the in vivo experiments will be all-optical stimulation/recording of AuNP- mediated action potentials in isolated rat retina, and histological analysis of retinas treated in vivo and in vitro with AuNPs. Results of the in vitro retina experiments will guide the selection of in vivo treatment conditions to be systematically investigated and facilitate interpretation of the in vivo data. Leading the research will be David R. Pepperberg, PhD (Univ. of Illinois at Chicago) and Francisco Bezanilla, PhD (Univ. of Chicago).

摘要 视网膜神经节细胞（RGC）以动作电位的形式将视觉信号传递到大脑， 其轴突传播依赖于RGC电压门控钠通道（NaVs）的激活。在 光感受器变性疾病，例如年龄相关性黄斑变性（AMD）、视网膜内神经元 包括RGC在许多情况下保持完整并能够产生动作电位反应。因此，RGC 代表旨在恢复晚期AMD和相关视网膜病变患者视力的方法的逻辑目标。 疾病，通过绕过无功能的杆和锥光感受器和建立直接RGC 对光的反应。在最近的一项背根神经节细胞（一种被广泛研究的非视网膜神经元， 模型动作电位产生细胞类型）和海马切片制备，我们已经表明，金 与细胞靶向生物分子缀合的纳米颗粒（AuNPs）能够实现稳健的光诱导NaV激活 以及由此产生的动作电位。细胞靶向AuNP技术的基本特征是： 光吸收AuNP的官能化以将它们定位在NaV处或附近;（ii）在AuNP的 毫秒/亚毫秒光闪光的等离子体激元吸收，光能量的AuNP辐射， 非破坏性的热脉冲，产生一个局部的，瞬态的，去极化电容电流通过 质膜;和（iii）产生的活化，即，通道开放，相邻NaV，从而作用 通过这种去极化的潜在启动。在本申请中，我们提出了探索性研究，以应用这种AuNP 在大鼠的活体眼睛中接近RGC。该项目的目标是建立AuNP处理条件， 在体内实现稳健的AuNP介导的RGC光响应性。在视杆细胞和视锥细胞 视网膜神经节细胞的光感受器信号传导已经被抑制，我们将在玻璃体内递送AuNP。 设计用于结合RGC NaV的紧邻的缀合物。在用AuNP处理之后， 结合物，我们将采用与RGC相关的视网膜电图（ERG）信号的体内记录 活动和视觉诱发电位（VEP），以分析RGC电生理反应的特性 到AuNP光激发。该研究将涉及AuNP的尺寸/结构的变化，AuNP的尺寸/结构的变化， 结合的RGC靶向组分的持续时间和能量[（强度）X（持续时间）]，以及结合的RGC靶向组分的持续时间和能量[（强度）X（持续时间）]。 兴奋的闪光伴随体内实验的将是AuNP的全光学刺激/记录， 在离体大鼠视网膜中介导的动作电位，以及在体内和体外处理的视网膜的组织学分析 关于AuNPs体外视网膜实验的结果将指导体内治疗条件的选择， 进行系统研究，并促进体内数据的解释。领导这项研究的将是 大卫河Pepperberg博士（伊利诺伊大学芝加哥分校）和弗朗西斯科Bezanilla博士（芝加哥大学）。