Role of PIEZO Channels in Bladder Function and Dysfunction

PIEZO 通道在膀胱功能和功能障碍中的作用

基本信息

批准号：
9815767
负责人：
Gerard L Apodaca
金额：
$ 65.09万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-08-01 至 2024-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9815767
关键词：
Affect Afferent Neurons Biological Assay Bladder Bladder Dysfunction Bowman&apos s space Cations Cell membrane Cells Communication Cyclophosphamide Cystitis Defect Epithelial Esthesia Event Exocytosis Functional disorder Goals Guns Hyperreflexia Image Ion Transport Knockout Mice Lead Lower urinary tract Mechanics Mediator of activation protein Membrane Membrane Protein Traffic Molecular Conformation Monitor Nature Neuraxis Overactive Bladder Pathway interactions Pelvis Physiological Play Process Reflex action Role Sensory Signal Transduction Stimulus Stratified Epithelium Stratum Basale Stretching Surface Testing Time Tissues Urodynamics Urothelial Cell Urothelium Viral afferent nerve allodynia gain of function gain of function mutation insight loss of function mechanotransduction multidisciplinary mutant novel patch clamp response tool

Affect Afferent Neurons Biological Assay Bladder Bladder Dysfunction Bowman&apos s space Cations Cell membrane Cells Communication Cyclophosphamide Cystitis Defect Epithelial Esthesia Event Exocytosis Functional disorder Goals Guns Hyperreflexia Image Ion Transport Knockout Mice Lead Lower urinary tract Mechanics Mediator of activation protein Membrane Membrane Protein Traffic Molecular Conformation Monitor Nature Neuraxis Overactive Bladder Pathway interactions Pelvis Physiological Play Process Reflex action Role Sensory Signal Transduction Stimulus Stratified Epithelium Stratum Basale Stretching Surface Testing Time Tissues Urodynamics Urothelial Cell Urothelium Viral afferent nerve allodynia gain of function gain of function mutation insight loss of function mechanotransduction multidisciplinary mutant novel patch clamp response tool

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项目摘要

Abstract: During filling and voiding, the degree of tension in the bladder wall must be sensed and then relayed to the central nervous system, otherwise bladder dysfunction ensues. While there is a general understanding of the role of sensory neurons (i.e., wall mechanoceptors) in these events, we have fewer insights into how non-neuronal tissues contribute to tension sensation and transduction in the bladder. Our studies focus on the urothelium, which forms the direct interface between the bladder wall and the urinary space. This tissue responds to changes in tension by modulating its ion transport, membrane traffic, and release of mediators, which are hypothesized to alter bladder function, in part via a local urothelial:afferent reflex. However, we still have limited insights into how tension in the plasma membrane of urothelial cells is sensed, what is the nature of the downstream pathways that are activated in response to stretch, or how do these events contribute to bladder function and dysfunction. We hypothesize that urothelial-expressed PIEZO channels act as mechanosensors that in response to bladder filling promote Ca2+ entry, mediator release, and signaling to afferent nerve processes, promoting normal bladder function. We further hypothesize that dysregulation of PIEZO-dependent mechanotransduction in the urothelium will lead to bladder dysfunction. In Specific Aim 1, we will determine if PIEZO channels act as bona fide mechanosensors by demonstrating the following: (i) that functional PIEZO channels are expressed at the surface of urothelial cells; (ii) that urothelial expressed PIEZO channels respond to physiologically relevant stimuli (i.e., bladder filling); (iii) that PIEZO channels are required for mechanically regulated events including membrane traffic and mediator release; (iv) and that expression of loss-of-function or gain-of-function PIEZO mutants will lead to altered urothelial responses. In Specific Aim 2, we seek to understand how PIEZO channels promote urothelial:afferent signaling. Using novel tools, including ex vivo bladder imaging, we will explore the mechanisms by which PIEZO channels stimulate increases in intracellular Ca2+ ([Ca2+]i). Because PIEZO channels rapidly inactivate, there is likely to be a mechanism to amplify the original signal. Thus, we will define whether Ca2+-induced Ca2+ release or membrane depolarization act downstream of PIEZO channels to increase [Ca2+]i. We will also determine if the PIEZO-triggered rise in [Ca2+]i is critical for stimulating exocytosis and mediator release in the urothelium, and if urothelial PIEZO channels modulate urothelial:afferent signaling. In Specific Aim 3, we will begin to explore whether urothelial PIEZO channels contribute to bladder function and/or dysfunction. Specifically, we will determine whether loss of PIEZO expression or function results in bladder underactivity and if PIEZO channels with gain-of-function mutations lead to bladder overactivity. Finally, we will determine whether urothelial-expressed PIEZO channels contribute to the bladder hyperreflexia and pelvic allodynia associated with cyclophosphamide-induced cystitis.

摘要：在填充和空隙期间，必须感知膀胱壁中的张力程度，然后传递到中枢神经系统，否则膀胱功能障碍随之而来。虽然有一般了解感官神经元（即壁式受体）在这些事件中的作用，我们的作用更少对非神经组织如何促进膀胱的张力和转导的洞察力。我们的研究的重点是尿路上皮，该尿皮细胞形成膀胱壁与尿中的直接接口空间。该组织通过调节其离子传输，膜流量和介质的释放，该介体被认为可以改变膀胱功能，部分通过局部尿路上皮：传入反射。但是，我们仍然对尿路上皮细胞质膜的张力如何是有限的见解感觉到，下游途径的性质是什么，响应拉伸或如何激活这些事件导致膀胱功能和功能障碍。我们假设尿路上皮表达的压电渠道充当机械传感器，以响应膀胱填充促进CA2+进入，调解员释放和信号传导到传入神经过程，促进正常的膀胱功能。我们进一步假设尿皮上皮中的压电依赖性机械转导的失调将导致膀胱功能障碍。在具体目标1，我们将通过证明该电压通道是否充当真正的机械传感器以下：（i）功能性压电通道在尿路细胞的表面表达；（ii）尿路上皮表达的压电通道对生理上相关的刺激有反应（即膀胱填充）；（iii）那个压电机械调节事件（包括膜流量和介质释放）所需的渠道是必需的；（iv）并且表达功能丧失或功能获得的压电突变体将导致尿路上皮改变回答。在特定目标2中，我们试图了解压电频道如何促进尿路上皮：传播信号。使用新的工具，包括离体膀胱成像，我们将探讨该机制压电通道刺激细胞内Ca2+（[Ca2+] i）的增加。因为压电通道迅速失活，所以可能会有一种机制来扩大原始信号。因此，我们将定义CA2+诱导的Ca2+ 压电通道下游的释放或膜去极化会增加[Ca2+] i。我们也会确定[Ca2+] I中对压电触发的上升是否对于刺激胞吐作用和介体释放至关重要尿路上皮，如果尿路上皮压电通道调节尿路上皮：传入信号传导。在特定的目标3中，我们将开始探索尿路上皮压电通道是否有助于膀胱功能和/或功能障碍。具体而言，我们将确定压电表达或功能的丧失是否导致膀胱不足如果具有功能收益突变的压电通道导致膀胱过度活动过度。最后，我们将确定尿路上皮表达的压电通道是否有助于膀胱高反射和骨盆异常与环磷酰胺诱导的膀胱炎有关。