Type II Afferents and Cochlear Damage

II 型传入神经和耳蜗损伤

• 批准号: 10164751
• 负责人: ELISABETH GLOWATZKI
• 金额: $ 39.25万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-06-16 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10164751
• 关键词: Acoustic Trauma; Action Potentials; Acute; Age; Anatomy; Antibiotics; Antibodies; Biological Markers; C Fiber; Calcium; Cochlea; Cre driver; Cues; DRD2 gene; Dose; Drug Delivery Systems; Electron Microscopy; Electrophysiology (science); Enterobacteria phage P1 Cre recombinase; Fiber; Future; G-Protein-Coupled Receptors; Genetic; Genetic Models; Glutamates; Goals; Hearing; Human; Hyperactivity; Hyperacusis; Hyperalgesia; Hypersensitivity; Injections; Label; Labyrinth; Ligands; Membrane; Methods; Microtomy; Modeling; Molecular; Mus; Neurobiology; Nociception; Nociceptors; Outer Hair Cells; Pattern; Perception; Pharmacology; Physiological; Property; Proteins; Protocols documentation; Rattus; Reporter; Role; Site; Specificity; Structure; Synapses; Testing; Tissues; Toxin; Transgenic Model; Trauma; Tyrosine 3-Monooxygenase; Validation; Work; animal tissue; base; cell injury; designer receptors exclusively activated by designer drugs; experimental study; extracellular; hearing impairment; light microscopy; mouse model; nerve supply; novel; noxacusis; optogenetics; ototoxicity; post-trauma; postsynaptic; prevent; programs; promoter; response; round window; sound; therapeutic target; tool; voltage

This proposal aims to test the hypothesis that type II afferents serve as cochlear nociceptors. Taking cues from the human complaint of hyperacusis after hearing loss, we will examine the structure and function of type II afferents in normal and post‐trauma cochleas. The working hypothesis is that painful hyperacusis, noxacusis, includes hyperactivity of type II afferents, by analogy to hyperalgesia of somatic nociceptive C‐fibers. Thus we will examine type II structure and function in normal and post‐trauma cochleas of rats and mice. In parallel we will investigate the properties of surviving outer hair cells in post‐ trauma cochleas. Our methods include: ex vivo electrophysiology, light and electron microscopy, utilization of optogenetic and chemogenetic tools,and validation and quantification of mouse models in which type II specific bio‐markers are expressed. A necessary first step is to extend our ex vivo experimental approach to older cochleas so that changes wrought by acoustic trauma can be compared to the normal condition. We will compare damaging sound, ototoxic antibiotics and genetically encoded biotoxins to produce experimentally tractable effects on tissue for ex vivo experiments. The properties and synaptic connections of type II afferents and outer hair cells will be examined in the excised cochlear tissue of these animals. We will continue to explore type II specific genetic mouse models. Genetically‐encoded reporter proteins, voltage‐ and calcium‐sensitive indicators, biotoxins, and opto‐ and chemo‐genetic modulators have become highly informative tools in neurobiology generally and for the inner ear specifically. Our ongoing work has characterized one mouse line, tyrosine hydroxylase promoter driven Cre‐recombinase expression. Three other candidate type II specific Cre lines will be validated and quantified. With such transgenic models it becomes possible to study innervation patterns by expression of fluorescent reporter proteins, and to activate, eliminate, or modulate type II activity for anatomical and physiological studies. Cre‐dependent expression of genetically‐modified G‐protein‐coupled receptors (DREADDS) will provide mice in which type II activity can be increased or decreased by injection of a novel synthetic ligand, depending on the specific construct. Varying combinations of systemic and round window drug delivery will be employed to increase the specificity of experimental manipulations. The over‐arching goal of this program of experiments is to complete the description of type II afferents, a still‐unresolved component of cochlear innervation. The working hypothesis is that these serve as cochlear nociceptors. If correct these are a likely neurobiological substrate for noxacusis (painful hyperacusis). By defining the basic cellular and molecular mechanisms of type II function and plasticity, future therapeutic targets can be identified to ameliorate or prevent noxacusis.

该提案旨在检验以下假设：II型传入是人工耳蜗伤害感受器。 听取听力丧失后，从人类对Hypersis的抱怨中获取线索，我们将检查结构 II型传入的功能以及在正常和创伤后耳蜗中的功能。工作的假设是 痛苦的超源，Noxacusis，包括II型传入的多动症 体细胞伤害性C纤维。我们将在正常和创伤后检查II型结构和功能 大鼠和小鼠的耳蜗。同时，我们将研究生存后毛细胞的特性 创伤耳蜗。我们的方法包括：离体电生理学，光和电子显微镜，利用率 光遗传学和化学遗传学工具，以及II型小鼠模型的验证和定量 表达了特定的生物标记。 一个必要的第一步是将我们的离体实验方法扩展到较旧的耳蜗，以便 通过声学创伤的变化可以与正常情况进行比较。我们将比较 破坏性声音，耳毒性抗生素和遗传编码的生物毒素以实验性产生 对组织实验的组织的易作用。 II型的属性和突触连接 将在这些动物的出色耳蜗组织中检查传入和外毛细胞。我们将 继续探索II型特异性遗传小鼠模型。遗传编码的记者蛋白， 电压和钙敏感指标，生物毒素和光遗传调节剂具有 普遍和内耳都成为神经生物学的高度信息工具。我们的 正在进行的工作表征了一条小鼠系，酪氨酸羟化酶启动子驱动的CRE聚合酶 表达。其他三个候选II型特异性CRE线将得到验证和量化。这样 转基因模型可以通过表达荧光来研究神经支配模式 报告蛋白，并激活，消除或调节解剖学和生理的II型活动 研究。 CRE依赖性的表达一般修饰的G蛋白偶联受体（Dreadds）将会 提供小鼠，可以通过注射新型合成可以增加或减少II型活动 配体，取决于特定构造。全身和圆形窗户药物的不同组合 交付将用于提高实验操作的特异性。 该实验程序的整个目标是完成II型的描述 传统，是人工耳蜗神经的仍未解决的组成部分。工作假设是这些 充当耳蜗伤害感受器。如果正确，则可能是NoxaCusis的神经生物学底物 （痛苦的超源）。通过定义II型功能的基本细胞和分子机制以及 可以确定可塑性，未来的治疗靶标可以改善或预防Noxacusis。