Nanomolar level of ouabain increases intracellular calcium to produce nitric oxide in rat aortic endothelial cells

被引:29
作者
Dong, XH [1 ]
Komiyama, Y [1 ]
Nishimura, N [1 ]
Masuda, M [1 ]
Takahashi, H [1 ]
机构
[1] Kansai Med Univ, Dept Clin Sci & Lab Med, Osaka 5708507, Japan
关键词
intracellular calcium; Na+/Ca2+ exchange; Na+/K+-ATPase; nitric oxide; ouabain;
D O I
10.1111/j.1440-1681.2004.03995.x
中图分类号
R9 [药学];
学科分类号
1007 ;
摘要
1. Changes in [Ca2+](i) across the cell membrane and/or the sarcoplasmic reticulum regulate endothelial nitric oxide (NO) synthase activity 2. In the present study, we investigated the effect of ouabain, a specific inhibitor of Na+/K+-ATPase, on NO release and [Ca2+](i) movements in cultured rat aortic endothelial cells (RAEC) by monitoring NO production continuously using an NO-specific real-time sensor and by measuring the change in [Ca2+](i) using a fluorescence microscopic imaging technique with high-speed wavelength switching. The t(1/2) (half-time of the decline of [Ca2+](i), to basal levels after stimulation with 10 mumol/L bradykinin) was used as an index of [Ca2+](i) extrusion. 3. A very low concentration of ouabain (10 nmol/L) did not increase the peak of NO production, but decreased the decay of NO release and, accordingly, increased integral NO production by the maximal dose-response concentration induced by bradykinin. The same dose of ouabain affected [Ca2+](i) movements across the cell membrane and/or sarcoplasmic reticulum induced by bradykinin with a time-course similar to that of NO release. Moreover, the t(1/2) was significantly increased. 4. Pretreatment of RAEC with Na+-free solution, an inhibitor of the Na+/Ca2+ exchanger, and nickel chloride hexahydrate prevented the effects induced by bradykinin and ouabain. 5. These observations using real-time recording indicate that a small amount of ouabain contributes to the bradykinin-stimulated increase of NO production through inhibition of plasma membrane Na+/K+-ATPase activity and an increase in intracellular Na+ concentrations. The membrane was then depolarized, leading to a decline in the bradykinin-stimulated increase in [Ca2+](i) by forward mode Na+/Ca2+ exchange to prolong the Ca2+ signal time.
引用
收藏
页码:276 / 283
页数:8
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