Physiochemical Investigation of Taste

描述(由申请人提供)：在生活系统面临的挑战中 相遇，很少有像这样基本的要求，那就是 细胞内和细胞外的液体隔间应保持在狭窄的范围内 生理极限。有一点很重要，但到目前为止还只有部分答案， 问题是：pH敏感细胞是如何检测各种液体中的酸度的 车厢被监控了吗？更具体地说，对于味觉感受器细胞 问题变成了：检测到了什么，细胞事件是什么 导致味觉传入神经兴奋？如果是这样的话是合理的 假设味觉感受器细胞将监测潜在食物的pH值或 饮料(胞外pH值)。尽管看起来可能令人惊讶，但事实并非如此 似乎是真的。我们已经确定，对酸中毒的近端刺激 反应是细胞内的pH值。这意味着酸必须首先进入 味觉感受器细胞才能被检测到。这项提议的一个主要目的是 研究酸进入味觉感受器细胞的各种可能途径。 这些包括作为中性分子(如乙酸乙酯)在细胞膜上扩散 酸)，作为气体(例如二氧化碳)，通过氢离子的电扩散， 以及特殊的转运体(例如单羧酸转运体)。感官 细胞本身只能在很小的细胞内pH范围内发挥作用 值，所以第二个问题是确定pH调节机制 在味觉感受器细胞中的存在以及它们的功能如何随 胞内pH。很有可能是3型钠氢交换器 是味觉细胞中重要的pH调节剂。这一点将被确定并扩大范围 探测器启动。转导最终使味觉细胞去极化并 细胞内钙离子的变化是可以预料的。这些也将被探讨。我们 将利用两种基本方法来开展这些研究。我们将衡量 细胞内pH、细胞内钙和膜电位的变化 单个菌状乳头中味觉细胞的荧光成像研究 在电压钳条件下保留上皮组织的极性。这些 研究将得到鼓索录音的补充， 电压钳下的语言感受野。我们还观察到，各种 苦味物质会使细胞内的pH变得更碱性。 初步研究表明，这种碱度在 关于G蛋白阻滞剂S的国内生产总值。我们将检验这一假设 在苦味转导过程中，味觉细胞变得碱性。对于品尝者来说 碱化作用本身不是碱(如变性)的假设吗 通过钠氢交换器或其他pH的激活而发生 监管机制。我们将检验这样一种假设，即pH阻滞剂 调控机制也将阻止变性诱导的增加。 细胞内钙离子。我们已经证明鼓索对 变性是电压敏感的，表明转导涉及 电导的调制。已经确立了细胞内pH的关键作用 在酸感方面，这些涉及苦味化合物的研究将 确定细胞内pH是否也是苦味的关键中间体 转导。

DESCRIPTION (provided by applicant): Among the challenges that living systems encounter, few are as basic as the requirement that the acidity of both the intracellular and extracellular fluid compartments be maintained within narrow physiological limits. One important, and as yet only partially answered, question is: how do pH-sensing cells detect hyperacidity in the various fluid compartments being monitored? More specifically for taste receptor cells the question becomes: what is being detected and what are the cellular events resulting in excitation of the taste afferent nerves? It would be reasonable to assume that taste receptor cells would monitor the pH of potential foods or beverages (the extracellular pH). Surprising as it might seem that does not appear to be true. We have established that the proximal stimulus for sour response is the intracellular pH. This means that an acid must first enter the taste receptor cell before it can be detected. A major aim of this proposal is to research the various possible ways acids ma enter taste receptor cells. These include diffusion across cell membranes as neutral molecules (e.g. acetic acid), as gases (e.g. carbon dioxide), by electrodiffusion of hydrogen ions, and on special transporters (e.g. monocarboxylate transporters). The sensing cells themselves can only function within a narrow range of intracellular pH values, so a second issue is the determination of the pH regulatory mechanisms present in taste receptor cells and how their function may vary with intracellular pH. It is highly probable that a type 3 sodium-hydrogen exchanger is an important pH regulator in taste cells. This will be ascertained and wider probes initiated. Transduction ultimately depolarizes the taste cells and changes in intracellular calcium can be expected. These also will be probed. We will utilize two basic methods to carry out these studies. We will measure changes in intracellular pH, intracellular calcium, and membrane potential in the taste cells using fluorescent imaging method in a single fungiform papilla with epithelial tissue polarity preserved under voltage clamp conditions. These studies will be complemented by recordings from the chorda tympani with the lingual receptive field under voltage clamp. We have also observed that various bitter tasting substances cause the intracellular pH to become more alkaline. Preliminary studies show that this alkalinity is significantly reduced in the presence of GDP about S, a G-protein blocker. We will test the hypothesis that during bitter taste transduction taste cells become alkaline. For tastants that are not themselves bases (e.g. denatonium) w hypothesize that alkalinization occurs through the activation of a sodium-hydrogen exchanger or other pH regulatory mechanisms. We will test the hypothesis that blockers of pH regulatory mechanisms will also block increases in denatonium induced intracellular calcium. We have shown that the chorda tympani response to denatonium is voltage sensitive indicating that transduction involves modulation of a conductance. Having established a key role for intracellular pH in acid-sensing, these studies involving bitter-tasting compounds will determine if intracellular pH is also a key intermediate in bitter-taste transduction.