REGULATION OF MORPHOGENESIS BY IONS, CAMP AND INOSITIDES

离子、CAMP 和肌醇酯对形态发生的调节

• 批准号: 3469481
• 负责人: WILLIAM B BUSA
• 金额: $ 8.34万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1987
• 资助国家: 美国
• 起止时间: 1987-05-01 至 1992-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3469481
• 关键词: Xenopus; acid base balance; adenylate cyclase; calcium; congenital disorders; cyclic AMP; enzyme mechanism; fluorescent dye /probe; histogenesis; image processing; ion exchange chromatography; lithium; microelectrodes; microinjections; phosphatidylcholines; protein kinase; radioimmunoassay; teratogens

Pattern formation during development is widely thought to involve specification of cells' positions in the embryo by gradients of informational molecules collectively known as morphogens. This is in keeping with much embryological data, but little success has been made in identifying such morphogens. In some lower organisms cyclic AMP, intracellular free calcium, or pH have been implicated as diffusible morphogens, but the involvement of these agents in vertebrate pattern formation has not been carefully investigated. Teratogens are often employed in the study of pattern formation, in particular the lithium ion, but its pathological effects on whole embryos are difficult to interpret. The recent demonstration that microinjection of Li+ into a specific blastomere of the early frog embryo induces formation of a second complete head now provides a powerful tool for such studies. Li+ has two well-established biochemical effects: inhibition of the phosphatidylinositol cycle (PI cycle) and of adenylate cyclase. Since the former's products, inositol trisphosphate and diacylglycerol, are involved in intracellular free calcium and pH regulation and the latter enzyme produces cyclic AMP, these agents may be involved in vertebrate pattern formation. The long-term goal of this study is to employ the teratogen, Li+, to test the involvement of pH, free Ca2+, cAMP, and inositol phosphates in pattern formation. Specific aims are to: (1) determine the distribution of PI cycle intermediates, cAMP levels, and intracellular free Ca2+ and pH values in normal frog embryos; (2) determine the effects of Li+ microinjection on these distributions and attempt to correlate these with head formation; (3) determine whether other inhibitors of the PI cycle and cAMP action mimic lithium's head-inducing ability; and (4) test whether direct manipulations of these distributions can mimic or prevent lithium's effects. Methodologies employed include measurement of intracellular free Ca2+ and pH with fluorescent probes and digital image analysis, chromatographic assay of PI cycle intermediates, radioimmunoassay of cAMP, microinjection, and microelectrodes. Health relatedness: These studies may contribute to our understanding of the mechanisms of teratogens such as Li+ (which is widely employed clinically), and of some birth defects of the neural tube and head, such as anencephaly. They may also fundamentally contribute to our understanding of vertebrate pattern formation.

人们普遍认为开发过程中的模式形成涉及 通过 信息分子统称为形态剂。 这是 与大量的胚胎学数据保持一致，但是在 识别这种形态剂。 在某些较低的生物体中，循环放大器， 细胞内游离钙或pH已被认为是可扩散的 形态剂，但这些药物在脊椎动物模式中的参与 尚未仔细研究组。 特拉致元通常用于模式形成的研究， 特别是锂离子，但其病理对整个胚胎的影响 很难解释。 最近的显微注射的演示 li+进入早期青蛙胚胎的特定胚胎诱导 现在，第二个完整头的形成为这种情况提供了强大的工具 研究。 Li+具有两个完善的生化效应：抑制 磷脂酰肌醇循环（PI循环）和腺苷酸环化酶。 自从 前者的产品，肌醇三磷酸和二酰基甘油，是 参与细胞内游离钙和pH调节以及后者 酶产生环状AMP，这些药物可能参与脊椎动物 图案形成。 这项研究的长期目标是采用 Teratogen，Li+，以测试pH，游离Ca2+，CAMP和 图案形成中的肌醇磷酸盐。 具体目的是：（1）确定PI周期的分布 中间人，营地水平和细胞内的无CA2+和pH值 正常的青蛙胚胎； （2）确定Li+微注射对 这些分布并试图将它们与头部形成相关联； （3） 确定PI周期和CAMP动作的其他抑制剂是否模仿 锂的头部诱导能力； （4）测试是否直接操纵 这些分布可以模仿或预防锂的作用。 采用的方法包括测量细胞内游离Ca2+和 带有荧光探针和数字图像分析，色谱法的pH PI周期中间体的测定 和微电极。 健康相关性：这些研究可能有助于我们对 Teratogens（例如Li+）的机制（已广泛使用 在临床上），以及神经管和头部的某些先天缺陷，例如 脑脓肿。 他们也可能从根本上有助于我们的理解 脊椎动物模式的形成。