Investigations into the mouse olivocochlear system

描述（由申请人提供）：最近发现促肾上腺皮质激素释放激素（CRH）受体和尿皮质素（CRH肽家族的成员）分别在外毛细胞和橄榄耳蜗末梢中表达。虽然已经使用尿皮质素缺陷小鼠分析了urocorUn（唯一已知的在内耳中表达的能够激活CRH受体的配体）所起的作用，但对CRH受体在内耳中的发育表达模式或作用一无所知。CRH受体是G蛋白偶联受体，并刺激cAMP第二信使信号级联。我们假设CRH受体的激活可能代表了一种保护噪声性听力损失的现象。作用机制可能包括磷酸化和sK2钙激活的apamin敏感性钾通道的失活。为了进一步分析耳蜗中CRH系统的形态学方面，其在听力和保护噪声引起的听力损失中的功能作用，以及最后，为了评估与CRH受体激活相关的细胞作用机制，提出了三个具体目标。首先，我们将建立CRH受体在内耳中的发育和成人表达模式。成功完成这一特定目标的实验将建立内耳内表达CRH受体的细胞的精确身份，并在超微结构水平上识别尿皮质素免疫阳性纤维的突触后元件。其次，我们将确定CRH在听力中发挥的功能作用，以及它们是否参与保护内耳免受噪音引起的听力损失。这一目标将使用缺乏1型或2型CRH受体基因或两者都缺乏的小鼠来实现。最后，我们将使用这些小鼠来确定CRH受体基因消融后外毛细胞中cAMP诱导的靶点磷酸化是否发生改变。这一目标的成功将使我们能够识别由于CRH受体激活而磷酸化的单个蛋白质，以及它们在调节正常橄榄耳蜗突触活动中的作用。这将在功能上将尿皮质素激素/CRH受体和经典的ACh神经递质系统连接在一起，形成一个统一的模型，解释基于内耳的保护免受噪声引起的听力损失。

DESCRIPTION (provided by applicant): Corticotropin releasing hormone (CRH) receptors, and urocortin, a member of the CRH family of peptides, has recently been discovered expressed in outer hair cells and olivocochlear terminals, respectively. While analysis of the role played by urocorUn, the only known ligand expressed in the inner ear capable of activating the CRH receptors, has been analyzed using urocortin deficient mice, nothing is known of the developmental expression pattern or role of the CRH receptors in the inner ear. The CRH receptors are G protein coupled receptors, and stimulate the cAMP second messenger-signaling cascade. We hypothesize that activation of the CRH receptors may represent one phenomenon underlying protection from noise induced hearing loss. The mechanisms of action may include phosphorylation and inactivation of the sK2 calcium-activated apamin-sensitive potassium channel. In order to further analyze the morphological aspects of the CRH system in the cochlea, its functional role in hearing and protection from noise induced hearing loss, and finally, to assess the cellular mechanisms of action associated with activation of the CRH receptors, three specific aims are proposed. First, we will establish the developmental and adult expression pattern of the CRH receptors in the inner ear. Successful completion of the experiments of this specific aim will establish the precise identity of the cells within the inner ear that express the CRH receptors, and identify the postsynaptic elements of the urocortin immunopositive fibers at the ultra structural level. Second, we will establish the functional roles CRH plays in hearing, and whether they participate in protection of the inner ear from noise induced hearing loss. This aim will be accomplished using mice that lack the gene for either the type 1 or the type 2 CRH receptor, or that lack both. Finally, we will use these mice to establish whether there are alterations in cAMP induced phosphorylation of targets in the outer hair cells following CRH receptors gene ablation. Success in this aim will allow us to identify individual proteins phosphorylated due to activation of the CRH receptors, as well as their role in modulating normal olivocochlear synaptic activity. This will functionally link the urocortin hormone/CRH receptor and classical ACh neurotransmitter systems together in a unified model explaining inner ear based protection from noise induced hearing loss.