ADRENERGIC CONTROL OF HEART & MUSCLE PHOSPHOFRUCTOKINASE

肾上腺素能控制心脏

• 批准号: 3357492
• 负责人: TAG E. MANSOUR
• 金额: $ 12.58万
• 依托单位: STANFORD UNIVERSITY
• 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 1994-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3357492
• 关键词: 6 phosphofructokinase; adrenergic agents; antiadrenergic agents; cytoplasm; enzyme mechanism; epinephrine; glycolysis; guinea pigs; hormone regulation /control mechanism; hypoxia; isoproterenol; laboratory rabbit; muscle cells; muscle pharmacology; myocardial ischemia /hypoxia; myocardium; myofibrils; phosphorylation; protein kinase; radiotracer; striated muscles; tissue /cell culture

Phosphofructokinase (PFK) is a rate-limiting enzyme that regulates the glycolytic flux in the heart and in skeletal muscle. Because of the critical role it plays during anoxia, information about its modulation contributes to a better understanding of many cardiovascular and muscular abnormalities such as myocardial ischemia and muscle hypoxia. The impetus for the present proposal is our discovery that epinephrine and isoproteronol increase PFK activity in perfused heart, isolated rat diaphragm, and skeletal of intact rabbits. Both agents are known to stimulate glycolysis in these organs. This laboratory has been studying the different regulatory mechanisms of PFK for the past 25 years. In spite of the expensive knowledge gained, the molecular details of the in vivo action of adrenergic agents on PFK to study the molecular and pharmacological events involved in activation of PFK by adrenergic agents. We intend to use three biological systems that offer different experimental advantages: (1) Myotubes derived from a myogenic cell line are more amenable to experimental manipulation (e.g., labeling with 32P) that intact animals and isolated organs; (2) Perfused guinea pig hearts are used to demonstrate activation of cardiac PFK by adrenergic agents; (3) Skeletal muscles from rabbits are used for studying activation of muscle PFK. The effects of adrenergic agents on the following regulatory parameters of PFK in these systems will be studied. First, we will study the effects of adrenergic agents on phosphorylation sites in PFK. These sites will be compared with phosphorylation sites in pure PFK following its phosphorylation by known protein kinases. The possibility that PFK is phosphorylated by as yet unknown protein kinases will be examined. Secondly, we will study the effects of adrenergic agents on PFK activity using both conventional assay methods that favor enzyme deaggregation and newly reported assay methods that favor enzyme aggregation. Third, we will determine whether adrenergic agents affect the levels of fru-2,6-P2, the most potent activator of PFK, and the activity of the kinase that catalyzes its synthesis, fru-6-P,2-P kinase. Finally, we will study whether adrenergic agents influence the translocation of PFK from cytosol to the particulate fraction (myofibril-rich) of heart and of muscle. Through the use of specific alpha and beta adrenergic blockers we intend to ascertain the pharmacological nature of activation of this enzyme by epinephrine and its congeners.

磷酸果糖激酶（PFK）是一种限制酶， 调节心脏和骨骼肌中的糖酵解通量。 由于它在缺氧期间扮演的关键作用，所以信息 关于它的调制有助于更好地理解 许多心血管和肌肉异常，例如 心肌缺血和肌肉缺氧。 推动 目前的建议是我们发现肾上腺素和 异丙醇醇增加灌注心脏，孤立的大鼠的PFK活性 隔膜和完整兔子的骨骼。 两种代理都是已知的 刺激这些器官中的糖酵解。 这个实验室已经 研究PFK的不同调节机制 25年。 尽管获得了昂贵的知识，但 肾上腺素能在体内作用的分子细节 PFK研究涉及的分子和药理事件 肾上腺素能剂激活PFK。 我们打算使用三个 具有不同实验优势的生物系统： （1）从肌原细胞系中得出的肌管更多 适合实验操作（例如，用32p标记） 那完整的动物和孤立的器官； （2）灌注豚鼠 心脏被用来证明通过 肾上腺素药； （3）兔子的骨骼肌用于 研究肌肉PFK的激活。 肾上腺素能的影响 PFK的以下监管参数的代理 系统将被研究。 首先，我们将研究 PFK中磷酸化位点上的肾上腺素能。 这些站点 将与纯PFK中的磷酸化位点进行比较 它通过已知蛋白激酶的磷酸化。 可能性 PFK被AS磷酸化，但未知蛋白激酶将是 检查。 其次，我们将研究肾上腺素能的影响 使用两种常规测定方法的PFK活性代理 这种有利于酶脱缘物和新报告的测定法 有利于酶聚集的方法。 第三，我们将确定 肾上腺素能量是否影响FRU-2,6-P2的水平， 最有效的PFK激活剂，以及激酶的活性 催化其合成，即FRU-6-P，2-P激酶。 最后，我们将学习 肾上腺素能是否影响PFK的易位 从细胞质到心脏的颗粒级分（肌纤维富含颗粒） 和肌肉。 通过使用特定的alpha和beta 肾上腺素阻滞剂我们打算确定药理 肾上腺素及其激活这种酶的性质 同型人。