Role of acid-sensing ion channels in bladder sensory signaling

（请保持文字，不要PDF） 膀胱是内脏疼痛的常见来源，也是患者就医的最常见原因之一。尽管传入通路在正常膀胱感觉和伤害感受中具有原始作用，但我们对这些过程的分子事件的理解有限。本研究旨在了解酸敏感离子通道（ASIC）调节膀胱传入信号的机制。该提案建立在以下发现的基础上，即未处理小鼠中ASIC3的丢失降低了触发排尿所需的膀胱内压，而在化学诱导的膀胱炎的情况下，它导致伤害性通路和慢性疼痛的过度激活。这些发现令人困惑，因为ASIC可能负责将细胞外pH瞬变转换为膀胱传入神经外周末端的电信号。我们的工作假设是，ASIC控制传入流出触发尖峰适应。我们认为，Na+通过ASIC内流启动了一系列事件，导致小电导Ca 2+激活的K+（SK）通道的激活和适应。目的1中提出的实验将确定ASIC是否通过Ca2+介导的机制调节传入放电。为了测试我们的模型，我们将使用感觉神经元条件Asic3基因敲除小鼠，Ca2+成像，膜片钳分析，单单位记录，和药理学工具的集合。在目标2中，我们将评估ASIC和SK通道是否协同工作以控制持续刺激期间的膀胱传入放电。为了评估这一点，我们将使用生理工具结合遗传和药理学操作。在完成这些研究后，我们将对调节内脏传入适应的机制有一个全面的了解。

(PLEASE KEEP IN WORD, DO NOT PDF) The urinary bladder is a common source of visceral pain, one of the most frequent reasons why patients seek medical attention. Despite the primordial role that afferent pathways have in normal bladder sensation and nociception, we have limited understanding of the molecular events contributing to these processes. This proposal aims to understand the mechanisms by which acid-sensing ion channels (ASICs) regulate bladder afferent signaling. The proposal builds on the findings that the loss of ASIC3 in naïve mice reduces the intravesical pressure required to trigger micturition, while in the setting of chemically-induced cystitis it results in the hyperactivation of nociceptive pathways and chronic pain. These findings are puzzling, because ASICs are presumably responsible for the transduction of extracellular pH transients into electrical signals at the peripheral terminal of bladder afferents. Our working hypothesis is that ASICs control afferent outflow by triggering spike adaptation. We posit that Na+ influx through ASICs initiates a cascade of events that results in the activation of small conductance Ca2+-activated K+ (SK) channels and adaptation. Experiments proposed in Aim 1 will define whether ASICs regulate afferent firing through a mechanism mediated by Ca2+. To test our model, we will use sensory neuron conditional Asic3 knockout mice, Ca2+ imaging, patch-clamp analysis, single-unit recordings, and a collection of pharmacological tools. In Aim 2, we will evaluate whether ASICs and SK channels work in concert to control bladder afferent discharge during sustained stimulation. To assess this, we will use physiological tools in combination with genetic and pharmacological manipulations. Upon completion of these studies, we will have gained a thorough understanding of mechanisms that mediate the adaptation of visceral afferents.