3D functional changes in the gap junctional signaling during the age related neurodegeneration in the mammalian inner ear Organ of Corti

听力损失是全球主要的健康问题之一。超过50%的75岁以上的人患有永久性听力损失。目前，还没有一种治疗剂可以预防或减缓听力损失的进展。听力损失的主要特征是对听觉内耳的损害，包括负责听力过程的各种细胞类型的丧失，如螺旋神经节神经元、感觉毛细胞和血管纹。然而，在听力损失发生之前，在特定的细胞类型中观察到了功能变化。这些在听力开始前就表现出病理变化的细胞在功能上与缝隙连接相结合。缝隙连接是细胞间的连接，直接连接两个细胞的细胞质，并允许各种分子、离子和电脉冲直接通过细胞之间的调节门。重要的是，缝隙连接功能的改变是人类遗传性耳聋的主要原因。探索缝隙连接细胞间通讯在听力过程中的作用依赖于对表达系统和不成熟的内耳培养的研究，使用破坏天然细胞组成的侵入性技术(如将染料注入细胞以观察通过缝隙连接的细胞间染料的传播)，或使用对细胞造成损害的技术(如用于可视化注入的染料的非常高强度的激光)。在这项研究中，我们专注于成熟的天然组织，使用新的非侵入性技术来研究缝隙连接偶联，允许保存细胞和内源性代谢物和第二信使，它们是缝隙连接信号的重要调节器。因此，这将是第一次探索年龄相关性听力损失和缝隙连接信号变化之间的直接联系的研究。这项初步研究的主要目的是了解缝隙连接细胞间通讯的功能变化，这些变化有助于耳蜗神经退行性变导致听力损失。我们将用小鼠作为人类耳蜗的模型。已有研究表明，老鼠和人类的耳蜗会以相似的方式发挥作用，并分享共同的潜在感觉处理机制。小鼠是最常见的动物模型，用于研究听力过程和任何可能的听力恢复。缝隙连接耦合的研究将在急性分离的听觉内耳中进行，使用新的高分辨率光学技术和细胞渗透染料来研究缝隙连接耦合的动力学。我们还将使用已知的影响缝隙连接耦合和功能的药物调节剂，在完整的系统中评估这些调节剂如何影响细胞之间的电信号，以及它们在神经退化过程中的变化，例如在年龄相关性听力损失中。这个项目的结果可能会为理解听力损失的过程提供新的方向和新的机会。此外，从这些试验性研究中获得的知识将为耳蜗的工作原理提供新的见解，并提供未来的研究方向，这最终可能有助于为大多数听力受损社区开发未来的治疗方法，而目前的听力损失治疗方法几乎没有什么好处。

Hearing loss is one of the major health concerns worldwide. Over 50% of individuals older than 75 suffer from permanent hearing loss. Currently, there is no therapeutic agent that can prevent or reduce the progression of hearing loss. The main hallmark of hearing loss is the damage to the auditory inner ear, including loss of various cell types responsible for the process of hearing, such as spiral ganglion neurons, sensory hair cells and stria vascularis. However, functional alterations are observed in the specific cell types before the onset of hearing loss. These cells which show pathological changes before the onset of hearing are functionally coupled with gap junctions. Gap junctions are intercellular connections that directly connect the cytoplasm of two cells, and allow various molecules, ions and electrical impulses to directly pass through a regulated gate between cells. Importantly, alterations in gap junction function is the major cause of hereditary deafness in humans. Probing the role gap junctional intercellular communication in the process of hearing has relied on studies in expression systems and immature inner ear cultures, using invasive techniques which disrupt native cellular composition (such as injecting dyes into the cell to observe the propagation of dye between cells through gap junctions), or using techniques which cause the damage to the cells (such as very high intensity lasers which are used to visualize the dyes injected). In this study, we focus on mature native tissues, to study gap junctional coupling using novel, non-invasive techniques, allowing for preservation of cellular and endogenous metabolites and second messengers, which are established and important modulators of gap junctional signalling. Thus, this would be the first study to explore a direct link between the age related hearing loss and the alterations of gap junction signaling. The main objectives of this pilot study is to understand the functional alterations in the gap junction intercellular communication that contribute to neurodegeneration of the cochlea leading to hearing loss. We will use mice as models for the human cochlea. It has been shown that the mouse and human cochleae function in similar ways and share common underlying sensory processing mechanisms. Mice are most common animal model used for investigation of hearing processes and any possible hearing restoration. The studies of gap junctional coupling will be performed in acutely isolated auditory inner ear using novel high resolution optical techniques and cell permeable dyes which can defuse through gap junctions between cells to study the kinetics of gap junctional coupling. We will also use pharmacological modulators known to affect the gap junctional coupling and function to asses in intact system how these affect the electrical signalling between cells, and how they change in the process of neurodegeneration, such as in age related hearing loss. The results from this project may provide new directions and fresh opportunities to understand the process of hearing loss. Moreover, knowledge gained from these pilot studies would provide novel insights into the workings of cochlea and provide future research directions, which could ultimately contribute to development of future therapeutic treatments for the majority of the hearing impaired community that benefits very little from current treatments for hearing loss.