Etiology of distinct membrane excitability in pre- and posthearing auditory neurons relies on activity of Cl− channel TMEM16A
Etiology of distinct membrane excitability in pre- and posthearing auditory neurons relies on activity of Cl− channel TMEM16A
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DOI：
10.1073/pnas.1414741112
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发表时间：
2015-02
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通讯作者：
Xiao-Dong Zhang;Jeong‐Han Lee;P. Lv;Wei Chun Chen;Hyo Jeong Kim;D. Wei;Wenying Wang;Choong-Ryoul Sihn;K. Doyle;J. Rock;N. Chiamvimonvat;E. Yamoah
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作者：
Xiao-Dong Zhang;Jeong‐Han Lee;P. Lv;Wei Chun Chen;Hyo Jeong Kim;D. Wei;Wenying Wang;Choong-Ryoul Sihn;K. Doyle;J. Rock;N. Chiamvimonvat;E. Yamoah
意义毛细胞和螺旋神经节神经元（SGNs）的发育过程是如何从听前特征向听后特征转变的，这是听觉神经科学研究的主要问题之一。几十年来，人们认为SGN中的自发动作电位源自自发放电的听前毛细胞。在这里，我们证明发育中的SGN使用Ca 2+激活的Cl-电导来去极化静息膜电位并使神经元处于超兴奋的预听状态。此外，SGN在发育过程中经历细胞内Cl−稳态的全局变化，以改变其编码特性。我们的研究结果解决了SGN自发活动的内源性起源，超越了神经元特异性现象，并可能为系统神经科学中Cl−调节机制的研究打开闸门。毛细胞和螺旋神经节神经元的膜特性发生了巨大的变化，这是听觉首次出现的发育预演。虽然开发HC过渡到成熟阶段的潜在机制是详细了解，SGN的成熟从超兴奋性prehearing静态posthearing神经元具有广泛的动态范围是未知的。在这里，我们使用药理学方法，笼状Ca 2+光解和短杆菌肽贴片记录证明，听前SGN使用Ca 2+激活的Cl−电导来降低静息膜电位，并使神经元处于超兴奋状态。耳蜗制备物的免疫染色揭示了SGN中Ca 2+激活的Cl−通道跨膜成员16 A（TMEM 16 A）的身份和表达。此外，TMEM 16 A的无效缺失减少了SGN中Ca 2+激活的Cl−电流和动作电位放电。为了确定Cl−离子和TMEM 16 A是否参与SGN烧前和烧后特征之间的转变，我们测量了SGN中的细胞内Cl−浓度[Cl−]i。令人惊讶的是，听前小鼠SGN中的[Cl−]i为1.90 mM，显著高于听后神经元的1.20 mM，表明Cl−通道在发育中的神经元中的作用发生了明显改变。[Cl−]i的转换源于细胞内Cl−调节机制发展的延迟表达。由于Cl−通道是唯一一个具有反转电位的活性离子选择性电导，其位于SGN动作电位的动态范围内，因此[Cl−]i的发育变化以及Cl−（ECl）的平衡电位将听觉前表型转变为听觉后表型。
Significance One of the major issues in auditory neuroscience is the mechanism by which the developing hair cells and spiral ganglion neurons (SGNs) transition from prehearing characteristics to posthearing features. For decades it was thought that spontaneous action potentials in SGNs emanate from spontaneously firing prehearing hair cells. Here, we demonstrate that developing SGNs use Ca2+-activated Cl− conductance to depolarize the resting membrane potential and to prime the neurons in a hyperexcitable prehearing state. Moreover, SGNs undergo global changes in intracellular Cl− homeostasis to alter their coding properties during development. Our findings address the endogenous origin of spontaneous activity in SGNs, transcend auditory-neuron-specific phenomena, and could open the flood gate for investigation on the mechanisms of Cl− regulation in systems neuroscience. The developmental rehearsal for the debut of hearing is marked by massive changes in the membrane properties of hair cells (HCs) and spiral ganglion neurons (SGNs). Whereas the underlying mechanisms for the developing HC transition to mature stage are understood in detail, the maturation of SGNs from hyperexcitable prehearing to quiescent posthearing neurons with broad dynamic range is unknown. Here, we demonstrated using pharmacological approaches, caged-Ca2+ photolysis, and gramicidin patch recordings that the prehearing SGN uses Ca2+-activated Cl− conductance to depolarize the resting membrane potential and to prime the neurons in a hyperexcitable state. Immunostaining of the cochlea preparation revealed the identity and expression of the Ca2+-activated Cl− channel transmembrane member 16A (TMEM16A) in SGNs. Moreover, null deletion of TMEM16A reduced the Ca2+-activated Cl− currents and action potential firing in SGNs. To determine whether Cl− ions and TMEM16A are involved in the transition between pre- and posthearing features of SGNs we measured the intracellular Cl− concentration [Cl−]i in SGNs. Surprisingly, [Cl−]i in SGNs from prehearing mice was ∼90 mM, which was significantly higher than posthearing neurons, ∼20 mM, demonstrating discernible altered roles of Cl− channels in the developing neuron. The switch in [Cl−]i stems from delayed expression of the development of intracellular Cl− regulating mechanisms. Because the Cl− channel is the only active ion-selective conductance with a reversal potential that lies within the dynamic range of SGN action potentials, developmental alteration of [Cl−]i, and hence the equilibrium potential for Cl− (ECl), transforms pre- to posthearing phenotype.