TWO ISOFORMS OF NA,K-ATPASE IN THE HEART

这个项目的总体目标是阐明 生理学、药理学意义和生化 Na~+，K~+-ATPase两种分子形式在心肌细胞中的调控 雪貂的心。我们已经证明了两种分子形式的 雪貂心肌细胞中存在Na+，K+-ATPase 丰度相等；高(阿尔法(+))和低(阿尔法) 具有形式的分子对应于高亲和力和低亲和力 哇巴因结合部位。同工酶的抑制 哇巴因所致双相正性肌力作用提示 两个阿尔法亚基都是药理受体 心脏糖苷类药物。虽然Na+，K+-ATPase同工酶 广泛分布于不同的细胞类型，生理上 心脏钠泵存在两种异构体的意义 细胞是未知的。雪貂之心提供了一个独特的机会 这些研究。 在拟议的研究中，将进行以下实验： 1)这两种异构体可能的生理意义 将检查NA泵。阿尔法(+)和阿尔法的假设 不同形式的钠泵以不同的速度工作，而胰岛素 优先调节钠泵的α(+)形式将是 测试过。胰岛素已被证明可以刺激阿尔法(+)形式的 对脂肪细胞的钠泵不影响α-型 打气筒。2)α(+)形式的相对丰度在 新生儿和出生后早期发育。假设 这些变化与药理学的变化密切相关。 糖苷和低亲和力“新生儿”的作用 在早期发育的不同物种中是常见的 测试过。3)假设人与人的亲和力差异 糖苷的阿尔法亚基是由于 磷酸酶的周转率和/或稳定性 检查过了。4)心肌肥厚改变假说 同工酶谱与心脏的糖苷敏感性 将会受到考验。5)甲状腺激素调节广泛的 包括肌球蛋白表达在内的生理功能 同工酶和Na+，K+-ATPase单位总数。因此， 甲状腺激素调节血管紧张素转换酶表达的假说 将测试两种不同亚型的α亚基。 这些研究的结果将表明，相对活动或 Na+，K+-ATPase两种异构体的相对丰度低于 激素控制或受心肌肥厚影响，以及 这些变化与对 洋地黄总苷。

The overall objective of this project is to elucidate the physiological, and pharmacological significance and biochemical regulation of the two molecular forms of Na+,K+ -ATPase in the ferret heart. We have demonstrated that two molecular forms of Na+,K+-ATPase exist in ferret myocardial cells in approximately equal abundance; the high (alpha(alpha(+)) and the low (alpha) molecular with forms correspond to the high and low affinity ouabain binding sites, respectively. Inhibition of the isoenzymes by ouabain caused biphasic positive inotropic effects indicating that both of the alpha subunits are the pharmacological receptors for the cardiac glycosides. Although isoenzymes of Na+,K+-ATPase are widely distributed in different cell types, the physiological significance of having two isoforms of the Na pump in the cardiac cells is unknown. Ferret heart offers a unique opportunity for these studies. In the proposed study, the following experiments will be performed: 1) Possible physiological significance of the two isoforms of the Na pump will be examined. The hypothesis that alpha(+) and alpha forms of the Na pump operate at different rates and that insulin preferentially regulates the alpha(+) form of the Na pump will be tested. Insulin has been shown to stimulate the alpha(+) form of the Na pump of fat cells with no effect on the alpha form of the pump. 2) Relative abundance of the alpha(+) form increases during neonatal and early postnatal development. The hypothesis that these changes are closely related to changes in pharmacological effects of the glycoside and that a low affinity "neonatal" form is common among different species during early development will be tested. 3) The hypothesis that differences in affinity of the alpha subunits for the glycoside results from differences in turnover rate and/or stability of the phosphoenzymes will be examined. 4) The hypothesis that myocardial hypertrophy alters isoenzyme profiles and therefore glycoside sensitivity of the heart will be tested. 5) Thyroid hormone regulates a wide range of physiological functions including the expression of myosin isoenzymes and the total number of Na+,K+-ATPase units. Therefore, the hypothesis that thyroid hormone regulates the expression of the two isoforms of alpha subunits will be tested. Results of these studies will indicate if the relative activity or the relative abundance of two isoforms of Na+,K+-ATPase is under hormonal control or is affected by myocardial hypertrophy, and these changes are associated with alterations in sensitivity to the digitalis glycosides.