HEPATIC UREAGENESIS--STUDIES WITH 15-N GC/MS & 13C-NMR

肝性尿失禁的生理学已经提出了一种更复杂的 代谢调节方面的问题。这项建议需要一项全面的 S调节谷氨酰胺代谢和尿素的机理评价 急性或慢性分离的肝细胞的产生 酸性或碱性大鼠。需要解决的主要问题包括： 1)TCA循环代谢在肝脏控制中的作用是什么？ 尿失禁？2)HCO3和/或PCO2的变化如何改变尿素的形成 来自谷氨酰胺？3)钾、磷和钙稳态的变化是如何改变的 尿失禁和控制部位是什么？4)激素(胰岛素， 高血糖素、儿茶酚胺)促进或抑制尿素形成的变化 酸碱动态平衡改变时会发生哪些反应？ 现场(S)的荷尔蒙作用如何？它是否与改变 细胞内pH、Ca~(2+)、PI对磷脂酰肌醇变化的影响 蛋白激酶C的周转、激活和/或Na+-H+-的调节 交易所？有待探讨的假设包括：(A)最初的 谷氨酰胺调节尿素pH的信号是一种 TCA循环代谢的改变；(B)除H+-外， 尿素调节细胞内钙离子或钙/PI比值的动态平衡变化 由于TCA循环代谢的改变而形成的；(C) 尿失禁的激素调节是通过改变 细胞内钙离子，从而调节TCA循环代谢和/或 蛋白激酶C活性；(D)或者，激素影响尿素 谷氨酰胺次级生成对Na~+-H~+-的影响 因此，细胞内pH的变化，以及通过 谷氨酰胺酶和谷氨酸脱氢酶途径，因此，氨 尿素合成的可用性。 我们将通过用15N孵育分离的肝细胞来探索这些主题 和/或13C标记底物。研究将在#年进行。 诱发急性酸碱中毒或在诱发急性酸碱中毒后 有无代谢的慢性酸中毒和碱中毒 调制器。我们将定义前体-产品关系和流量 用气相色谱-质谱仪和/或核磁共振测定。 建议的实验具有重要的科学意义和临床意义。 通过加深对肝脏谷氨酰胺代谢和尿素的了解 与H+相关的H+微扰的合成 独立于其他生理因素的。我们得到的结果应该是 提供肝脏调节机制的全景图(S) 尿失禁。

The physiology of hepatic ureagenesis has posed one of the more complex problems in metabolic regulation. this proposal entails a comprehensive evaluation of the mechanism(s) regulating glutamine metabolism and urea production in isolated hepatocytes obtained from acute or chronically acidotic or alkalotic rats. The primary questions to be addressed are: 1) What is the role of TCA-cycle metabolism in the control of hepatic ureagenesis? 2) How do changes in HCO3 and/or PCO2 alter urea formation from glutamine? 3) How do changes in K+, Pi and Ca2+ homeostasis alter ureagenesis and what is the control site? 4) Do hormones (insulin, glucagon, catecholamine) enhance or inhibit changes in urea formation which occur in response to altered acid-base homeostasis? 5) What is the site(s) of the hormonal action? Is it linked to alterations of intracellular pH, Ca2+, Pi or to changes of phosphatidylinositol turnover, activation of protein kinase C and/or modulation of the Na+-H+- exchanger? Hypotheses to be explored include: (a) that the initial signal in evoking pH modulation of urea formation from glutamine is an alteration of TCA-cycle metabolism; (b) that in addition to H+- homeostasis changes in intracellular Ca2+ or Ca2+/Pi ratio modulate urea formation secondary to alteration of TCA-cycle metabolism; (c) that the hormonal regulation of ureagenesis is mediated through changes in intracellular Ca2+ and thereby, modulation of TCA-cycle metabolism and/or protein kinase C activity; (d) alternatively, hormones affect urea formation from glutamine secondary to their effect on the Na+-H+- exchanger and hence, changes in intracellular pH, and fluxes via the glutaminase and glutamate dehydrogenase pathways, and therefore, ammonia availability for urea synthesis. We will explore these themes by incubating isolated hepatocytes with 15N and/or 13C labeled substrates. Studies will be carried out during induction of acute acidosis and alkalosis or following induction of chronic acidosis and alkalosis in the presence and absence of metabolic modulators. We will define precursor-product relationships and flux rates using GC-MS and/or NMR. The proposed experiments are of scientific as well as clinical importance by deepening our understanding of hepatic glutamine metabolism and urea synthesis in response to perturbations of H+ associated with/or independent of other physiological factors. The results we obtain should provide a panoramic view of the mechanism(s) regulating hepatic ureagenesis.