Spontaneous activity in the developing cochlea

发育中的耳蜗的自发活动

• 批准号: 7535210
• 负责人: DWIGHT E BERGLES
• 金额: $ 34.85万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-12-01 至 2010-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7535210
• 关键词: ATP Receptors; Acoustic Nerve; Action Potentials; Acute; Afferent Neurons; Anions; Auditory; Automobile Driving; Biological Preservation; Brain; Brain Stem; Cell Communication; Cell Nucleus; Cells; Characteristics; Cochlea; Connexins; Dendrites; Development; Event; Exhibits; Frequencies; Gap Junctions; Generations; Growth; Hair Cells; Hearing; Human; Image; Inner Hair Cells; Inner Supporting Cell; Ions; Lead; Maintenance; Maps; Measures; Mediating; Membrane; Membrane Potentials; Monitor; Mutation; Nerve; Nerve Fibers; Neurons; Optics; Organ of Corti; Outer Hair Cells; Pattern; Permeability; Physiological; Play; Property; Proteins; Purinergic P2 Receptors; Rattus; Relative (related person); Resistance; Rest; Rodent; Role; Sensory; Signal Transduction; Stimulus; Supporting Cell; Synapses; Testing; Time; Tinnitus; Tissues; Whole-Cell Recordings; afferent nerve; auditory pathway; caged IP3; deafness; extracellular; fluorescence imaging; human GJB2 protein; inhibitor/antagonist; photolysis; response; sensory system; sound; spiral ganglion

DESCRIPTION (provided by applicant): Spontaneous activity in developing sensory systems has been shown to be important for the growth and survival of projection neurons as well as the refinement and stabilization of sensory maps in the brain. In the developing cochlea, bursts of action potentials occur in afferent spiral ganglion neurons prior to the onset of hearing, activity that has been traced to inner hair cells (IHCs). Although IHCs are capable of generating Ca2+ action potentials during this period, the depolarizing stimulus required to initiate these events has not been identified. Whole-cell recordings from IHCs and supporting cells located adjacent in IHCs in ex vivo cochleas from young rodents revealed the presence of spontaneous inward currents that were capable of inducing large depolarizations. This activity was coincident with changes in the optical properties of the tissue when visualized using IR/DIC imaging, indicating that these events can be monitored non- invasively. Spontaneous electrical and optical activity was blocked by P2 purinergic receptor antagonists and gap junction inhibitors, suggesting that ATP and gap junctions/hemichannels are involved in initiating these events. Remarkably, this activity is no longer observed after the onset of hearing. This discovery of spontaneous purinergic signaling in the developing organ of Corti raises many new questions about the mechanisms responsible for producing this activity, the role that this ATP-mediated signaling plays in driving afferent firing, and the cause of the disappearance of the activity after hearing onset. We hypothesize that these ATP driven depolarizations of IHCs are responsible for initiating activity in developing auditory pathways. The preservation of this activity in both acute and cultured cochleas in which appropriate cell- cell interactions are maintained provides us with an unprecedented opportunity to understand the mechanisms responsible for these robust phenomena. We propose to use IR/DIC and confocal fluorescence imaging, photolysis, and both whole cell and extracellular recording to investigate the mechanisms underlying spontaneous activity in supporting cells and hair cells in the developing organ of Corti. These studies will evaluate the specific hypothesis that spontaneous oscillations in [Ca2+]i within supporting cells triggers both inward currents and the release of ATP that depolarizes IHCs.Relevance The studies outlined in this proposal seek to understand the mechanisms responsible for initiating spontaneous activity in supporting cells, hair cells, and afferent dendrites in the developing cochlea. This activity has been shown to have a profound influence on survival of target neurons in brainstem nuclei, the physiological properties of these auditory neurons, and the pattern of synaptic connectivity in these regions. Most congenital forms of deafness result from mutations in connexin 26, a gap junction protein highly expressed by cochelar supporting cells. As our preliminary results suggest that connexin hemichannels may play a role in ATP release from supporting cells, the studies outlined here may help explain how these mutations lead to deafness. Furthermore, these studies may reveal one mechanism by which activity can be induced in afferent nerves in the absence of sound, which may have direct relevance to human conditions such as tinnitus.

描述（由申请人提供）：已证明感觉系统发育中的自发活动对于投射神经元的生长和存活以及大脑中感觉图的细化和稳定非常重要。在发育中的耳蜗中，在出现听觉之前，传入螺旋神经节神经元中会出现动作电位的爆发，这种活动已被追踪到内毛细胞（IHC）。尽管 IHC 能够在此期间产生 Ca2+ 动作电位，但启动这些事件所需的去极化刺激尚未确定。来自年轻啮齿动物离体耳蜗中 IHC 和 IHC 邻近支持细胞的全细胞记录揭示了自发内向电流的存在，能够诱导大量去极化。当使用 IR/DIC 成像可视化时，这种活动与组织光学特性的变化一致，表明可以非侵入性地监测这些事件。自发的电和光活动被 P2 嘌呤能受体拮抗剂和间隙连接抑制剂阻断，表明 ATP 和间隙连接/半通道参与启动这些事件。值得注意的是，在出现听力后就不再观察到这种活动。正在发育的柯蒂氏器官中发现自发的嘌呤能信号，引发了许多新的问题：产生这种活动的机制、这种 ATP 介导的信号在驱动传入放电中所起的作用，以及听力后活动消失的原因。发病。我们假设这些 ATP 驱动的 IHC 去极化负责启动发育听觉通路的活动。在急性和培养的耳蜗中保留这种活性，并维持适当的细胞-细胞相互作用，为我们提供了前所未有的机会来了解造成这些强大现象的机制。我们建议使用 IR/DIC 和共焦荧光成像、光解以及全细胞和细胞外记录来研究发育中的 Corti 器官中支持细胞和毛细胞自发活动的机制。这些研究将评估支持细胞内 [Ca2+]i 自发振荡触发内向电流和去极化 IHC 的 ATP 释放的具体假设。 相关性 本提案中概述的研究旨在了解在支持细胞中启动自发活动的机制发育中的耳蜗中的细胞、毛细胞和传入树突。这种活动已被证明对脑干核中目标神经元的存活、这些听觉神经元的生理特性以及这些区域的突触连接模式具有深远的影响。大多数先天性耳聋是由连接蛋白 26 突变引起的，连接蛋白 26 是一种由耳蜗支持细胞高度表达的间隙连接蛋白。由于我们的初步结果表明连接蛋白半通道可能在支持细胞释放 ATP 中发挥作用，因此此处概述的研究可能有助于解释这些突变如何导致耳聋。此外，这些研究可能揭示了一种在没有声音的情况下诱导传入神经活动的机制，这可能与耳鸣等人类状况直接相关。