DEVELOPMENT AND USE OF METHODS TO MEASURE CELLULAR PH

细胞 pH 值测量方法的开发和使用

• 批准号: 3292501
• 负责人: JOHN E WAMPLER
• 金额: $ 6.81万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-07-01 至 1989-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3292501
• 关键词: Dictyostelium; acid base balance; cell differentiation; cell migration; chemotaxis; cytoplasm; egg /ovum; fertilization; fluorescence spectrometry; fluorescent dye /probe; fresh water environment; growth /development; phagocytosis

Regulation of pH is essential to the life of all living cells. For instance, the pH of the cytosol must be maintained near neutrality by extrusion of acid generated as a result of metabolism. The pH of lysosomes must be acidic to increase the activity of hydrolytic enzymes. The pH of mitochondria is alkaline. Movement of protons in these organelles are required for production of ATP. Recent evidence indicates that changes in the pH in the cytosol may regulate numerous cellular activities. Changes in pH may be required for initiation of growth of quiescent, non-dividing cells. Alterations in intracellular pH may also dictate the pathway of differentiation selected by developing cells. Local changes in pH may contribute to movements of cells, such as locomotion and division. Techniques for measurement of pH have been developed that employ the pH dependence of fluorescence emitted by fluorescent probes that have been incorporated into living cells. A microspectrofluorometer can be used to continuously record the pH and its changes in local regions of single living cells. In the present proposal, the microspectrofluorometer will first be used to determine the accuracy and precision of fluorometric measurements of intracellular pH in the egg of the Medaka fish and in amoeba in the cellular slime mold, Dictyostelium discoideum. The changes in pH in the Medaka egg will be determined, localized and correlated with ongoing developmental processes. Changes in intracellular pH during development in D. discoideum will be analyzed in detail to test the hypothesis that changes in pH contribute to cell differentiation. Mechanisms or regulation of intracellular pH will be assessed by use of specific pharmacologic agents. In addition, changes in intracellular pH during cell movement in D. discoideum will be examined in order to test the hypotesis that pH is a regulator of the structure of cytoplasm, and the shape and movements of cells. Cell movements and differentiation are fundamental processes in the life of all vertebrate organisms. Elucidation of the role of pH in regulating these processes will advance our knowledge of the fundamental mechanisms that regulate cell activities, direct future studies of these phenomena in normal and pathologic states, and may eventually contribute to development of diagnostic procedures.

pH 值的调节对于所有活细胞的生命至关重要。 为了 例如，细胞质的 pH 值必须保持在中性附近 新陈代谢产生的酸的排出。 溶酶体的pH值 必须呈酸性以增加水解酶的活性。 PH值 线粒体呈碱性。 这些细胞器中质子的运动是 生产 ATP 所需的。 最近的证据表明，细胞质 p​​H 值的变化可能 调节许多细胞活动。 可能需要改变 pH 值 静止的、不分裂的细胞开始生长。 改动于 细胞内 pH 值也可能决定所选的分化途径 通过发育细胞。 pH 值的局部变化可能会导致 细胞，例如运动和分裂。 已经开发出利用 pH 值测量 pH 值的技术 荧光探针发射的荧光的依赖性 并入活细胞。 显微分光荧光计可用于 连续记录单个区域局部区域的pH值及其变化 活细胞。 在本提案中，显微分光荧光计将首先用于 确定荧光测量的准确性和精密度 青鳉鱼卵和变形虫的细胞内 pH 值 细胞粘菌，盘基网柄菌。 pH值的变化 青鳉蛋将被确定、定位并与正在进行的相关联 发展过程。 发育过程中细胞内pH值的变化 将详细分析盘状 D. 以检验以下假设： pH值的变化有助于细胞分化。 机制或监管 细胞内 pH 值将通过使用特定的药理学方法进行评估 代理。 此外，细胞运动过程中细胞内pH值的变化 将检查 D. discoideum，以检验 pH 值是一个假设 细胞质结构、形状和运动的调节者 细胞。 细胞运动和分化是生命的基本过程 所有脊椎动物。 阐明 pH 值的调节作用 这些过程将增进我们对基本机制的了解 调节细胞活动，指导这些现象的未来研究 正常和病理状态，最终可能有助于发育 诊断程序。