STUDIES OF CARDIAC METABOLISM BY 13C AND 31P NMR

通过 13C 和 31P NMR 研究心脏代谢

• 批准号: 3340788
• 负责人: JOHN R. WILLIAMSON
• 金额: $ 10.53万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 1983
• 资助国家: 美国
• 起止时间: 1983-07-01 至 1987-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3340788
• 关键词: Krebs' cycle; aspartate; aspartate transaminase; carbohydrate metabolism; cell free system; citrate synthase; dicarboxylate; heart metabolism; malate dehydrogenase; malates; mitochondria; myocardial ischemia /hypoxia; nuclear magnetic resonance spectroscopy; oxaloacetates; oxoglutarate dehydrogenase; perfusion; propionates; radiotracer

The major goal of this project is to take advantage of the recent advances made in the field of 31p and 13C nuclear magnetic resonances (NMR) spectroscopy to study the energetics and metabolism of perfused rat hearts under different conditions of substrate supply and work. Emphasis will be given to the use of 13C-enriched substrates to study regulation of the citric acid cycle, its interactions with the malate-asparate cycle and mechanisms for increasing or decreasing net pool sizes of citric acid cycle intermediates. 13C NMR spectra will be obtained using both intact hearts and perchloric acid extracts. The latter will also be used for enzymatic assay of metabolites to aid identification and quantitation of resonances obtained from individual 13C-labeled carbon atoms from the 13C spectra. The information provided from 13C specific labeling patterns of intermediates, together with knowledge of their total pool sizes as determined by metabolic analyses, will be used to construct a flux model of the citric acid cycle and interrelated transamination steps. The model will be used to test various hypotheses concerning regulation of flux in the citric acid cycle under different conditions of substrate supply and work output of the heart during the steady state as well as transition states when cycle flux is nonuniform. The following major specific aims will be addressed. 1. Identification of rate-controlling steps in the citric acid cycle and assessment of enzyme control strengths by inhibitor titration studies. 2. Assessment of the existence and metabolic significance of metabolite compartmention and substrate channeling. 3. Effects of varying flux rates through the malate-asparate cycle on the regulation of flux through citrate syntase, Alpha-ketoglutarate dehydrogenase and asparate aminotransferase. 4. Investigation of the mechanisms responsible for causing net changes of the tissue contents of citric acid cycle intermediates. 5. Effects of acetoacetate in the absence and presence of propionate on flux through succinate thiokinase and regulation of the citric acid cycle from Alpha-keto-glutarate to malate. 6. Investigation of citric acid cycle function in reoxygenated post-ischemic hearts.

该项目的主要目标是利用最新进展 在 31p 和 13C 核磁共振 (NMR) 领域制造 光谱学研究灌注大鼠心脏的能量学和代谢 在不同的底物供应和工作条件下。 重点将是 考虑使用富含 13C 的底物来研究 柠檬酸循环，其与苹果酸-天门冬氨酸循环的相互作用以及 增加或减少柠檬酸循环净池大小的机制 中间体。 13C NMR 谱将使用两个完整的心脏获得 和高氯酸提取物。 后者也将用于酶促 代谢物测定有助于共振的识别和定量 从 13C 光谱中单个 13C 标记的碳原子获得。 13C 特定标签模式提供的信息 中间体，以及它们的总池大小的知识 通过代谢分析确定，将用于构建通量模型 柠檬酸循环和相关的氨基转移步骤。 型号 将用于测试有关流量调节的各种假设 不同底物供应条件下的柠檬酸循环 稳态和过渡期间心脏的工作输出 表明循环通量不均匀。 主要具体目标如下 将得到解决。 1. 速率控制步骤的识别 柠檬酸循环和抑制剂对酶控制强度的评估 滴定研究。 2. 存在和代谢的评估 代谢物区室和底物通道的重要性。 3. 苹果酸-天门冬氨酸循环中不同通量速率对 通过柠檬酸合酶、α-酮戊二酸调节流量 脱氢酶和天冬氨酸转氨酶。 4. 调查 导致组织内容净变化的机制 柠檬酸循环中间体。 5. 乙酰乙酸的作用 通过琥珀酸硫激酶的通量中丙酸盐的存在和不存在以及 从α-酮-戊二酸到苹果酸的柠檬酸循环的调节。 6.复氧中柠檬酸循环功能的研究 缺血后的心脏。