The Role of Pannexin1 in Ischemic Injury of Retinal Ganglion Cells

描述(申请人提供)：我们的长期目标是防止视网膜缺血中的神经元丢失和功能缺陷。根据我们的目标，我们已经确定了一种新的，几乎没有被探索的膜通道蛋白pAnnexin1(Panx1)，作为发现新疗法的一个有希望的来源。我们建立了Panx1作为缺血性神经元损伤的分子介质的功能，并且该通道的遗传消融可以保护视网膜神经节细胞免受 缺血性损伤。我们的中心假设是，Panx1是外部应激源的关键“汇聚中心”。我们建议的研究将比较Panx1介导的不同毒性机制的相对贡献，并确定哪些刺激或它们的组合在缺血时触发脆弱的视网膜神经节细胞该通道的病理性开放。具体目的是：1)分析Panx1通道开放引发的毒性通路对RGC损伤的相对贡献毒性；2)在实验性视网膜缺血再灌注模型中比较部分抑制与完全阻断或阻断Panx1通道的保护作用。我们将关注缺血后神经元的存活和视网膜功能的保存。意义重大。保护视网膜神经元免受缺血损伤 对于全面的治疗策略是必不可少的。了解Panx1介导的毒性通路及其对神经元损伤的贡献将永久性地改变视网膜和脑缺血的概念和治疗方法。我们将使用为该提案开发的专业知识和设计的独特工具来评估Panx1受体阻滞剂抑制或预防短暂性脑缺血中视力丧失的可行性，这是一个重要的临床问题，也是NIH NEI的研究目标。 公共卫生相关性：我们的长期目标是防止视网膜缺血中的神经元丢失和功能缺陷。我们已经确定了一种新的、以前未被探索的缺血性神经元损伤的分子介质，一种通道蛋白PAnnexin1(Panx1)。根据我们的主要发现，Panx1基因的去除抑制了缺血再灌流中神经元的丢失，我们假设在缺血中Panx1作为神经元损伤的分子介质发挥作用。我们的中心假设是，Panx1通过打开一个大的膜孔，促进视网膜的缺血性损伤和氧化应激。我们试图揭示Panx1与缺血时不同毒性通路的激活之间的因果关系。我们开发了一种独特的Panx1条件性基因敲除小鼠和新颖的Panx1特异性膜片钳协议来实现这一提议的目标。本研究的主要成果包括对Panx1介导的毒性通路及其在神经元损伤中的作用的新认识。我们还将测试Panx1阻滞剂在缓解视网膜缺血方面的治疗潜力 采用动物缺血再灌流损伤模型。

DESCRIPTION (provided by applicant): Our long-term goal is to prevent neuronal loss and functional deficits in retinal ischemia. Pursuant to our objective, we have identified a new, scantily explored membrane channel protein pannexin1 (Panx1), as a promising source for discovering new therapies. We established that Panx1 function as a molecular mediator of ischemic neuronal injury, and genetic ablation of this channel protects retinal ganglion cells from ischemic damage. Our central hypothesis states that Panx1 serves as a critical "convergence hub" for external stressors. Our proposed studies will compare relative contribution of distinct toxicity mechanisms mediated by Panx1 and identify which stimuli or their combinations trigger pathological opening of this channel in vulnerable retinal ganglion cells in ischemia. Specific Aims are designed to: 1) analyze toxicity pathways triggered by the Panx1 channel opening for their relative contribution toxicity to RGC injury~ 2) Compare protective effects of partial inhibition vs. full blockade or ablation of Panx1 channel in vivo in experimental retinal ischemia-reperfusion model. We will focus on both neuronal survival and preservation of retinal functionality following ischemia. Significance. Protecting retinal neurons from ischemic injury is essential for a comprehensive therapeutic strategy. The understanding of the Panx1-mediated toxicity pathways and their contribution to neuronal injury will permanently alter both conceptual and therapeutic approaches to retinal and brain ischemia. We will use the expertise developed and the unique tools designed for this proposal to evaluate the feasibility of Panx1 blockade for suppressing or preventing the vision loss in transient ischemia, an important clinical problem and the NIH NEI research objective. PUBLIC HEALTH RELEVANCE: Our long-term goal is to prevent neuronal loss and functional deficits in retinal ischemia. We have identified a new, previously unexplored molecular mediator of ischemic neuronal injury, a channel protein pannexin1 (Panx1). Based on our main finding demonstrating that the Panx1 gene ablation suppresses neuronal loss in ischemia-reperfusion, we hypothesize that in ischemia Panx1 function as a molecular mediator of neuronal injury. Our central hypothesis states that Panx1, through opening of a large membrane pore, facilitates ischemic injury and oxidative stress to the retina. We seek to reveal cause-and-effect relationship between Panx1 and the activation of distinct toxicity pathways in ischemia. We have developed a unique Panx1 conditional knockout mouse and novel Panx1-specific patch clamp protocol to reach the objectives of this proposal. The major outcomes of this study include the new knowledge on the Panx1-mediated toxicity pathways and their contribution to neuronal injury. We will also test a therapeutic potential of Panx1 blockade in alleviating retinal ischemic injury using animal model of ischemia-reperfusion.