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

项目总结/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联系 螺旋神经节神经元的轴突。噪声诱导的突触病归因于谷氨酸兴奋性毒性， 导致螺旋神经节神经元的逐渐轴突变性和最终死亡。后果 突触和神经元的丧失包括听觉感知功能障碍，导致言语困难 在嘈杂的环境中识别和倾听。这种类型的听觉功能障碍被称为“隐性听力 因为它不容易通过标准的听力测试来诊断。此外，没有螺旋 神经节神经元限制了用于听力损失的主要疗法如耳蜗植入物的性能， 未来毛细胞再生策略。目前，还没有批准的药物可以促进神经元存活 或者引发失去的听觉神经的再生，并用存活的毛细胞补充它们的突触连接。 因此，了解突触和神经元变性的机制以及 为了开发更好的耳疗法。我们最近证明了突触病变噪音 创伤足以招募巨噬细胞（先天免疫细胞）向受损的内毛细胞-突触 地区虽然受损的突触可以进行自发修复，但是， 信号传导（通过巨噬细胞上的Fractalkine（FKN）受体CX 3CR 1的基因缺失）损害这种 自发突触修复和增加创伤后螺旋神经节神经元损失。这些数据表明， 完整的Fractalkine信号传导对于受损耳蜗中的突触修复和神经元存活是必需的。在这里， 我们建议研究fractalkine信号传导的激活对预防和修复损失的影响， 突触和神经元存活率的变化。目标1将确定FKN治疗是否 修复哺乳动物小鼠耳蜗中噪声创伤或兴奋性毒性损伤后受损的突触。 具体地，FKN肽将在体内突触病变性噪声创伤后（经鼓膜）注射， 或在耳蜗外植体中谷氨酸诱导的兴奋性毒性之后。FKN膜的精确贡献或 将检查朝向突触修复的可溶性同种型。目标2将确定FKN治疗是否 减少噪音创伤或谷氨酸兴奋毒性后突触的退化。我们将用FKN治疗 在离体耳蜗外植体中谷氨酸盐处理之前或在噪声创伤之前， 体内（经鼓膜）。在目标3中，我们将消除耳蜗巨噬细胞，并检查 这种干预对突触损伤性噪声损伤后突触变性和修复程度的影响。为 每个目标，听觉功能沿着以及毛细胞，巨噬细胞，突触和螺旋的形态计量学分析 将进行神经节神经元计数。总之，研究设计将有助于调查 巨噬细胞和Fractalkine治疗耳蜗突触变性和修复以及听力恢复 并且可能导致鉴定用于“隐性听力损失”的基于Fractalkine的新疗法。

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”.