VASOTOCIN RECEPTORS

加压素受体

• 批准号: 3233666
• 负责人: PATRICK EGGENA
• 金额: $ 16.47万
• 依托单位: MOUNT SINAI SCHOOL OF MEDICINE OF CUNY
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-04-01 至 1991-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3233666
• 关键词: Anura; adenylate cyclase; affinity chromatography; affinity labeling; biological fluid transport; cyclic AMP; hormone metabolism; hormone receptor; membrane permeability; protein metabolism; radiotracer; tritium; urea; urinary bladder epithelium; vasopressins; vasotocin

The antidiuretic hormone, vasopressin (AVP), plays an important role in the regulation of plasma volume and osmolality by triggering the reabsorption of water, urea, and sodium from the forming uring in the kidney. The isolated urinary bladder of the toad, Bufo marinus, has been used extensively to study the cellular mechanism of action of AVP (or the amphibian antidiuretic hormone, vasotocin (AVT), which is similar to AVP). It is not known whether different receptors and (or) different target cells are involved in the three transport functions of the hormone. We plan to approach this problem by isolating and purifying AVT receptors from isolated toad bladders by Percoll- and sucrose- density gradient centrifugation, affinity chromatography with a biotinyl derivative of AVT linked to an avidin column, and photoaffinity labeling of receptors with radio-active AVT analogues followed by SDS-PAGE. Both agonistic and antagonistic radio-labeled photoaffinity analogues will be synthesized for this purpose. After AVT receptor proteins have been purified from toad bladder, antibodies will be raised in rabbits to these receptors, and the action of antireceptor antibodies to inhibit binding of radio-labeled hormone to one or more classes of receptors in membranes will be compared with their effect on adenylate cyclase activity and on water, urea, and sodium transport across the bladder epithelium. AVT analogues containing fluorescein or rhodamine moieties in positions 7 (agonists) or 4 (antagonists) will be synthesized and their localization in frozen tissue slices of toad bladder studied by quantitative fluorescence microscopy. Displacement of fluorescent AVT by different receptor antibodies will be used as a way of matching different cell types with different transport functions. Similarly, a second antibody with a fluorescent label will be used to localize the distribution of anti-receptor antibodies. It is hoped that these studies will shed light on the question of how a single hormone can differentially regulate three independent transport functions in the same target organ.

抗利尿激素、加压素 (AVP) 在 血浆容量和渗透压的调节作用 触发水、尿素和钠的重吸收 在肾脏中形成尿。 离体膀胱 蟾蜍（Bufo marinus）已被广泛用于研究 AVP（或两栖动物）的细胞作用机制 抗利尿激素、催产素（AVT），与 AVP 类似）。 目前尚不清楚是否不同的受体和（或）不同的 靶细胞参与三种运输功能 激素。 我们计划通过分离和纯化 AVT 来解决这个问题 Percoll-和蔗糖-从分离的蟾蜍膀胱中提取受体 密度梯度离心、亲和层析 连接到抗生物素蛋白柱的 AVT 的生物素衍生物，以及 用放射性 AVT 光亲和标记受体 类似物，然后进行 SDS-PAGE。 既具有对抗性又 拮抗性放射性标记的光亲和类似物将 为此目的合成的。 从蟾蜍中纯化 AVT 受体蛋白后 膀胱，兔子体内会产生针对这些受体的抗体， 以及抗受体抗体抑制结合的作用 对一类或多类受体进行放射性标记的激素 将比较膜对腺苷酸的影响 环化酶活性以及水、尿素和钠转运 膀胱上皮。 含有荧光素或罗丹明部分的 AVT 类似物 位置 7（激动剂）或 4（拮抗剂）将被合成并 研究人员研究了它们在蟾蜍膀胱冷冻组织切片中的定位 定量荧光显微镜。 位移 不同受体抗体的荧光 AVT 将用作 将不同细胞类型与不同运输相匹配的方法 功能。 同样，带有荧光标记的第二抗体 将用于定位抗受体的分布 抗体。 希望这些研究能够阐明这个问题 单一激素如何差异化调节三种激素 在同一靶器官中具有独立的运输功能。