胞质NADH/NAD⁺水平对通透化大鼠心室肌细胞肌浆网Ca²⁺释放的影响
Effects of cytosolic NADH/NAD(+) levels on sarcoplasmic reticulum Ca(2+) release in permeabilized rat ventricular myocytes.
作者信息
Zima Aleksey V, Copello Julio A, Blatter Lothar A
机构信息
Department of Physiology, Stritch School of Medicine, Loyola University Chicago, 2160 South First Avenue, Maywood, IL 60153, USA.
出版信息
J Physiol. 2004 Mar 16;555(Pt 3):727-41. doi: 10.1113/jphysiol.2003.055848. Epub 2004 Jan 14.
In the heart ischaemic conditions induce metabolic changes known to have profound effects on Ca(2+) signalling during excitation-contraction coupling. Ischaemia also affects the redox state of the cell. However, the role of cytosolic redox couples, such as the NADH/NAD(+) redox system, for the regulation of Ca(2+) homeostasis has remained elusive. We studied the effects of NADH and NAD(+) on sarcoplasmic reticulum (SR) Ca(2+) release in permeabilized rat ventricular myocytes as well as on Ca(2+) uptake by SR microsomes and ryanodine receptor (RyR) single channel activity. Exposure of permeabilized myocytes to NADH (2 mm; Ca(2+)= 100nm) decreased the frequency and the amplitude of spontaneous Ca(2+) sparks by 62% and 24%, respectively. This inhibitory effect was reversed by NAD(+) (2 mm) and did not depend on mitochondrial function. The inhibition of Ca(2+) sparks by NADH was associated with a 52% decrease in SR Ca(2+) load. Some of the effects observed with NADH may involve the generation of superoxide anion (O(2)(-).) as they were attenuated to just a transient decrease of Ca(2+) spark frequency by superoxide dismutase (SOD). O(2)(-). generated in situ from the xanthine/xanthine oxidase reaction caused a slowly developing decrease of Ca(2+) spark frequency and SR Ca(2+) load by 44% and 32%, respectively. Furthermore, in studies with cardiac SR microsomes NADH slowed the rate of ATP-dependent Ca(2+) uptake by 39%. This effect also appeared to depend on O(2)(-). formation. Single channel recordings from RyRs incorporated into lipid bilayers revealed that NADH (2 mm) inhibited the activity of RyR channels by 84%. However, NADH inhibition of RyR activity was O(2)(-).-independent. In summary, an increase of the cytoplasmic NADH/NAD(+) ratio depresses SR Ca(2+) release in ventricular cardiomyocytes. The effect appears to be mediated by direct NADH inhibition of RyR channel activity and by indirect NADH inhibition (O(2)(-). mediated) of SR Ca(2+)-ATPase activity with a subsequent decrease in SR Ca(2+) content.
在心脏缺血条件下会引发代谢变化，已知这些变化在兴奋 - 收缩偶联过程中对Ca(2+)信号传导有深远影响。缺血还会影响细胞的氧化还原状态。然而，胞质氧化还原对，如NADH/NAD(+)氧化还原系统，在调节Ca(2+)稳态中的作用仍不清楚。我们研究了NADH和NAD(+)对通透化大鼠心室肌细胞肌浆网（SR）Ca(2+)释放的影响，以及对SR微粒体Ca(2+)摄取和兰尼碱受体（RyR）单通道活性的影响。将通透化的心肌细胞暴露于NADH（2 mM；[Ca(2+)]cyt = 100 nM）后，自发Ca(2+)火花的频率和幅度分别降低了62%和24%。NAD(+)（2 mM）可逆转这种抑制作用，且该作用不依赖于线粒体功能。NADH对Ca(2+)火花的抑制作用与SR Ca(2+)负荷降低52%有关。NADH观察到的一些效应可能涉及超氧阴离子（O(2)(-).）的产生，因为超氧化物歧化酶（SOD）将其减弱为Ca(2+)火花频率仅短暂下降。由黄嘌呤/黄嘌呤氧化酶反应原位产生的O(2)(-).分别导致Ca(2+)火花频率和SR Ca(2+)负荷缓慢下降44%和32%。此外，在对心脏SR微粒体的研究中，NADH使ATP依赖性Ca(2+)摄取速率减慢39%。这种效应似乎也依赖于O(2)(-).的形成。整合到脂质双层中的RyR单通道记录显示，NADH（2 mM）使RyR通道活性抑制84%。然而，NADH对RyR活性的抑制不依赖于O(2)(-).。总之，细胞质NADH/NAD(+)比值的增加会抑制心室心肌细胞中SR Ca(2+)的释放。这种效应似乎是由NADH直接抑制RyR通道活性以及NADH间接抑制（O(2)(-).介导）SR Ca(2+)-ATPase活性，随后SR Ca(2+)含量降低所介导的。
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