SENSING PH AT THE GASTRIC SURFACE

胃必须在胃上皮细胞分泌的盐酸中存活下来。这一挑战因化合物(如非类固醇抗炎药)或生物体(幽门螺杆菌)的频繁存在而加剧，这些化合物或生物体破坏了胃对酸的屏障，当屏障被打破时会导致溃疡。碱性粘膜旁凝胶层是胃防御的第一道防线：起到保护胃上皮细胞免受胃腔酸性物质反向扩散的作用。我们开发了一种独特的方法，可以在体内非侵入性地测量胃表面的pH值。使用共聚焦显微镜，我们已经成像了粘膜旁的碱性层，当管腔pH改变到在喂养的胃中发现的值时，它转化为酸性层。我们的结果提出了一种新的胃表面pH调节模型，其中包括控制表面pH的转运体的实质性变化。我们的目标是确定调节胃的管腔pH感觉和表面pH调节的元件。主要使用大鼠或小鼠胃的活体共聚焦显微镜，我们的第一个目标将定义胃感觉和响应管腔pH的基本要求。我们将a)确定从碱到酸分泌所需的管腔pH变化的时间和位置，以及b)测试管腔营养素和缓冲液在表面pH调节中的作用。在第二个目标中，我们将质疑哪些分子是腔pH刺激的转导分子，该刺激介导胃酸和碱分泌的综合调节。我们将重点介绍生长抑素、前列腺素E_2和含有降钙素基因相关肽的辣椒素敏感传入神经的作用。生长抑素的作用将使用针对生长抑素2型受体(Sst2)的拮抗剂和激动剂进行分析，同时对Sst2基因敲除小鼠进行平行研究。通过对COX-1和COX-2基因敲除小鼠的平行研究，将使用已知的环氧合酶(COX)亚型的选择性抑制剂来探讨前列腺素合成的作用。辣椒素敏感的传入神经将使用香草素受体激动剂、CGRP受体激动剂和拮抗剂进行分析，同时对化学去传入动物进行平行研究。第三个目标是问胃粘膜损伤如何扰乱表面的pH调节。我们将通过双光子显微镜在胃上皮细胞中产生微观损伤。使用这种新的胃局灶性损伤模型，我们将实时跟踪表面pH调节和组织修复过程的中断。我们将询问阿司匹林，临床粘膜损伤的主要原因，是否通过破坏正常和COX基因敲除小鼠的管腔pH感觉和表面pH调节来削弱胃屏障。我们将确定阿司匹林是否影响双光子显微镜造成的局灶性病变的修复。结果将把对胃防御的理解与对酸和碱分泌的调节结合起来。

The stomach must survive HCl secreted by the gastric epithelium. This challenge is heightened by the frequent presence of compounds (e.g. non-steroidal anti-inflammatory drugs) or organisms (Helicobacter pylori) which compromise the gastric barrier to acid, leading to ulcers when the barrier is breached. An alkaline juxtamucosal gel layer is the first line of gastric defense: acting to protect the gastric epithelium from back-diffusion of acid from the lumen. We have developed a unique approach to non-invasively measure pH at the gastric surface in vivo. Using confocal microscopy, we have imaged a juxtamucosal alkaline layer, which converts to an acid layer when luminal pH is changed to values found in the fed stomach. Our results suggest a new model of gastric surface pH regulation, which includes a substantial change in the transporters that control surface pH. Our objective is to define the elements regulating luminal pH sensing and surface pH regulation in the stomach. Using primarily in vivo confocal microscopy of rat or mouse stomach, our first aim will define fundamental requirements for the stomach to sense and respond to luminal pH. We will a) define the timing and location of luminal pH change required for conversion from alkali to acid secretion, and b) test the role of luminal nutrients and buffers in surface pH regulation. In the second aim, we will question which molecules are transducers of the luminal pH stimulus that mediate the integrated regulation of both gastric acid and alkali secretion. We will focus on the role of somatostatin, PGE2, and capsaicin-sensitive afferent nerves containing CGRP. The role of somatostatin will be analyzed using antagonists and agonists selective for the somatostatin type 2 receptor (SST2) with parallel studies of SST2-knockout mice. The role of prostaglandin synthesis will be approached using selective inhibitors of known cycloxygenase (COX) isoforms, with parallel studies of COX-1 and COX-2 knockout mice. Capsaicin-sensitive afferents will be analyzed using vanilloid receptor agonists, CGRP receptor agonists and antagonists, with parallel studies of chemically deafferented animals. The third aim will ask how gastric mucosal damage disrupts surface pH regulation. We will generate microscopic lesions in the gastric epithelium by two-photon microscopy. Using this new model of focal gastric damage, we will follow the disruptions in surface pH regulation and the tissue repair process in real time. We will ask if aspirin, a major cause of clinical mucosal damage, weakens the gastric barrier by disruption of luminal pH sensing and surface pH regulation in normal and COX-knockout mice. We will determine if aspirin affects the repair of focal lesions created by two-photon microscopy. Results will integrate understanding of gastric defense with regulation of acid and alkali secretion.