Role of voltage-gated calcium channel alpha2delta subunits for spiral ganglion neurons and afferent auditory processing

听觉通路的毛细胞和神经元使用不同的离子通道、递质受体和突触专门化，以确保超快信号具有极高的时间精确度。这也适用于电压门控钙通道(VGCC)，它是突触前钙内流和递质释放所必需的。VGCC是由形成孔洞的α1亚基、胞内β亚基和胞外α2β(A2d)亚基组成的多聚体蛋白质复合体。生物物理和药理学特性在很大程度上由十个α1亚基中的一个决定，而β和a2d亚基有助于VGCC的表面表达和微调通道门控。存在四个不同的A2D亚基，具有部分冗余和部分特定的功能。我们之前已经证明，A2d3缺陷小鼠的听觉诱发脑干反应扭曲，尽管听力阈值只有轻微提高，毛细胞的生理和形态正常。缺乏a2d3可降低培养的螺旋神经节神经元(SGN)的体细胞钙电流。听神经纤维向耳蜗核丛状细胞的突触传递受阻，听神经纤维终末大小明显减小。此外，小鼠不能在行为任务中辨别调幅音调，从而将a2d3缺陷小鼠建立为听觉处理障碍的模型。由于a2d3在SGN以外的传入听觉通路的几个核团中表达，我们将通过分析在SGN中特异缺乏a2d3的条件小鼠模型来剖析a2d3的具体作用，并结合(A)钙电流记录，(B)下丘神经元的活体记录和(C)听觉辨别学习实验。越来越多的证据表明，A2d亚基除了通过VGCC调节钙离子流量外，还发挥着额外的作用，例如对突触前和突触后的结构和功能。由于SGN也表达a2d1和a2d2的mRNA，我们将利用a2d1基因敲除和a2d2突变小鼠研究这些a2d亚基对SGN钙电流、钙通道蛋白表达和突触形态的作用。对下丘神经元的活体记录将使我们深入了解不同小鼠处理时间信息的能力。最后，行为听觉辨别学习实验将补充2d亚单位在听觉处理中的作用。

Hair cells and neurons of the auditory pathway use distinct ion channels, transmitter receptors and synapse specializations to guarantee ultrafast signalling with exquisitely high temporal precision. This also applies to voltage-gated Ca2+ channels (VGCC), which are required for presynaptic Ca2+ influx and transmitter release. VGCCs are multimeric protein complexes consisting of a pore-forming alpha1 subunit, an intracellular beta-subunit and an extracellular alpha2delta (a2d) subunit. Biophysical and pharmacological properties are largely determined by one of the ten alpha1 subunits, whereas beta and a2d subunits assist in surface expression of VGCCs and fine-tune channel gating. Four different a2d subunits exist with partially redundant and partially specific functions. We have previously shown that a2d3-deficient mice have distorted auditory evoked brainstem responses despite only mildly elevated hearing thresholds and normal physiology and morphology of hair cells. Lack of a2d3 reduced somatic Ca2+ currents of cultured spiral ganglion neurons (SGN). Synaptic transmission from auditory nerve fibers to bushy cells of the cochlear nucleus was compromised, and sizes of auditory nerve fiber terminals were markedly reduced. Moreover, the mice were unable to discriminate amplitude-modulated tones in a behavioral task, establishing the a2d3-deficient mouse as model for an auditory processing disorder. Because a2d3 is expressed beyond SGN in several nuclei of the afferent auditory pathway, we will dissect the specific roles of a2d3 by analysing a conditional mouse model that specifically lacks a2d3 in SGN with (a) Ca2+ current recordings, (b) in-vivo recordings of neurons in the inferior colliculus and (c) auditory discrimination learning experiments. Evidence is accumulating that a2d subunits play additional roles than merely modulating Ca2+ flux through VGCCs, e.g. for the structure and function of pre- and postsynapses. Because SGN express mRNA for a2d1 and a2d2 as well, we will study the roles of these a2d subunits for SGN Ca2+ currents, Ca2+ channel protein expression and synapse morphology using a2d1 knockout and a2d2 mutant mice. In vivo recordings of neurons in the inferior colliculus will give insights into the ability of the different mice to process temporal information. Finally, behavioral auditory discrimination learning experiments will complement the picture of the role of a2d subunits in auditory processing.