Contribution of Macrophages and Fractalkine Towards Degeneration and Repair of Cochlear Synapses

项目摘要/摘要 噪声损伤主要损害内毛细胞和外周毛细胞之间的突触连接。 螺旋神经节神经元的轴突。噪声诱导的突触病变归因于谷氨酸兴奋性毒性和 导致轴突逐渐退化，最终导致螺旋神经节神经元死亡。后果是什么？ 突触和神经元的丧失包括导致言语困难的听觉知觉功能障碍 在嘈杂的环境中进行识别和倾听。这种类型的听觉功能障碍被称为“隐性听力” 损失“，因为它不容易通过标准的听力测试来诊断。此外，没有螺旋线 神经节神经元限制了听力损失的初级治疗的效果，如人工耳蜗术和 未来的毛细胞再生策略。目前，还没有被批准的促进神经元存活的药物 或者诱导失去的听神经再生，并补充它们与幸存毛细胞的突触连接。 因此，了解突触和神经元退变的机制是非常有意义的。 再生，以发展更好的耳疗。我们最近证明了突触致病噪音 创伤足以将巨噬细胞(天然免疫细胞)招募到受损的内毛细胞-突触 区域。尽管受损的突触可以进行自发修复，但Fractalkine的破坏 信号(通过巨噬细胞上FKN受体CX3CR1的遗传缺失)损害了这种 创伤后自发的突触修复和增加螺旋神经节神经元的丢失。这些数据意味着 完整的Fractalkine信号是损伤耳蜗区突触修复和神经元存活所必需的。这里, 我们建议研究激活Fractalkine信号在预防和修复丢失中的作用 对耳蜗伤后突触和神经元存活的影响。目标1将决定FKN治疗 修复噪音、创伤或兴奋性毒性伤害后的哺乳动物小鼠耳蜗区受损的突触。 具体地说，FKN多肽将在体内突触病理性噪声损伤后注射(经鼓室) 或在谷氨酸诱导的耳蜗外植体兴奋毒性后。FKN膜的精确贡献或 我们将研究与突触修复有关的可溶型异构体。目标2将确定FKN治疗 减少噪音创伤或谷氨酸兴奋性毒性后突触的退化。我们会用FKN治疗 谷氨酸处理耳蜗体外植体或噪声损伤前的膜或可溶性异构体 活体内(经鼓室)。在目标3中，我们将消除耳蜗巨噬细胞，并检查 这种干预对突触病理性噪声损伤后突触退行性变和修复程度的影响。为 每个目标、听觉功能以及毛细胞、巨噬细胞、突触和螺旋的形态计量分析 将进行神经节神经元计数。总之，这项研究设计将有助于研究 巨噬细胞联合Fractalkine治疗耳蜗神经突触退变修复及听力恢复 并可能导致发现新的基于Fractalkine的治疗“隐性听力损失”的方法。

PROJECT SUMMARY/ABSTRACT Noise trauma can primarily damage the synaptic connections between the inner hair cells and the peripheral axons of the spiral ganglion neurons. Noise-induced synaptopathy is attributed to glutamate excitotoxicity and leads to gradual axonal degeneration and ultimately death of the spiral ganglion neurons. The consequences of loss of synapses and neurons include auditory perceptual dysfunctions leading to difficulty in speech recognition and listening in noisy environments. This type of auditory dysfunction is known as “hidden hearing loss” because it is not readily diagnosed through standard hearing tests. Moreover, absence of spiral ganglion neurons limits the performance of primary therapies for hearing loss such as cochlear implants and future hair cell regeneration strategies. Currently, there are no approved drugs that promote neuron survival or elicit regeneration of lost auditory nerves and replenish their synaptic connections with surviving hair cells. Therefore, it is of great interest to understand the mechanisms for synaptic and neuron degeneration and regeneration for the development of better ototherapeutics. We recently demonstrated that synaptopathic noise trauma is sufficient to recruit macrophages (innate-immune cells) towards the damaged inner hair cell-synaptic region. While the damaged synapses can undergo spontaneous repair however, disruption of fractalkine signaling (by genetic deletion of fractalkine (FKN) receptor CX3CR1 on macrophages) impairs such spontaneous synaptic repair and increases spiral ganglion neuron loss after trauma. These data imply that intact fractalkine signaling is necessary for synaptic repair and neuron survival in the damaged cochlea. Here, we propose to investigate the effect of activation of fractalkine signaling on prevention and repair of loss of synapses and neuron survival following cochlear trauma. Aim 1 will determine whether FKN treatment repairs damaged synapses after noise trauma or excitotoxic insult in mammalian mouse cochlea. Specifically, FKN peptide will be injected either (transtympanically) after synaptopathic noise trauma in vivo or after glutamate- induced excitotoxicity in cochlear explants. The precise contribution of FKN membrane or soluble isoforms towards synaptic repair will be examined. Aim 2 will determine whether FKN treatment reduces degeneration of synapses following noise trauma or glutamate excitotoxicity. We will treat with FKN membrane or soluble isoforms prior to glutamate treatment in ex vivo cochlear explants or prior to noise trauma in vivo (transtympanically). In Aim 3, we will eliminate cochlear macrophages and examine the influence of this intervention on the degree of synaptic degeneration and repair after synaptopathic noise trauma. For each aim, auditory function along with morphometric analyses of hair cell, macrophage, synapse and spiral ganglion neuron counts will be performed. Together, the study design will aid in investigating the effect of macrophages and fractalkine treatment on cochlear synapse degeneration and repair and hearing restoration and may lead to identification of novel fractalkine-based therapeutics for “hidden-hearing loss”.