目的 观察低浓度(1×10-8 mol·L-1)哇巴因(OUA)对大鼠心肌细胞的收缩作用,并研究低浓度哇巴因发挥正性肌力作用有关的钠钾泵(NKA)信号转导通路。方法 在急性分离的大鼠心室肌细胞上,利用全细胞膜片钳技术记录钠钾泵电流(Ip),并观察低浓度哇巴因对Ip作用的影响。利用细胞动缘探测系统测定心肌细胞的长度及收缩力的大小。①观察并比较1×10-8~1×10-3 mol·L-1浓度哇巴因增强大鼠心室肌细胞收缩力的作用。②分别用信号转导抑制剂PP2(1 μmol·L-1)、NAC(100 μmol·L-1)及PD98059(50 μmol·L-1)与1×10-8 mol·L-1哇巴因共同灌流,记录信号转导抑制剂对1×10-8 mol·L-1哇巴因增强大鼠心室肌细胞收缩力的影响。结果 1×10-8~1×10-3 mol·L-1的哇巴因均能增加大鼠心室肌细胞收缩力(P<0.01);1 μmol·L-1PP2+1×10-8 mol·L-1哇巴因组与1×10-8 mol·L-1哇巴因组相比心肌细胞收缩力减小(P<0.05),100 μmol·L-1 NAC+1×10-8 mol·L-1哇巴因组与1×10-8 mol·L-1哇巴因组相比心肌细胞收缩力也明显减小(P<0.01); 而50 μmol·L-1 PD98059+1×10-8 mol·L-1哇巴因与1×10-8 mol·L-1哇巴因组相比信息及细胞收缩力没有显著性差异(P>0.05)。结论 1×10-8~1×10-3 mol·L-1的哇巴因均能增加大鼠心室肌细胞收缩力,且具有浓度依赖性。低浓度哇巴因的正性肌力作用与离子转运功能无关而与钠钾泵的信号转导功能有关,其中Src/ROS信号途径参与调节了1×10-8 mol·L-1的哇巴因的正性肌力作用。
Abstract
OBJECTIVE To observe whether low concentration (1×10-8 mol·L-1) of ouabain (OUA)can increase the contractility in rat cardiocytes and investigate the Na/K pump signal transduction pathways related to positive inotropic action following the low concentration of OUA. METHODS On enzymatic isolation of rats ventricular myocytes, the Na+/K+ pump current (Ip) was by whole-cell patch-clamp, in order to observe the low concentration of OUA on Ip. The contraction of a single myocyte was assessed by a video-based motion edge-detection system. ①To detect and compare the potentiations of 1×10-8-1×10-3 mol·L-1 OUA on the contractility in rat cardiocytes. ②The cardiocytes were pre-treated with PP2(1 μmol·L-1), NAC(100 μmol·L-1), PD98059(50 μmol·L-1)for 5 min, and the effects of the signals transduction inhibitors on the positive inotropic effect of 1×10-8 mol·L-1 OUA was recorded. RESULTS The 1×10-8-1×10-3 mol·L-1 OUA increased the contractility of rat cardiocytes (P<0.01). Compared 1 μmol·L-1 PP2 +1×10-8 mol·L-1 OUA group with 1×10-8 mol·L-1 OUA group, the contraction amplitude decreased (P<0.05), and compared 100 μmol·L-1 NAC +1×10-8 mol·L-1 OUA group with 1×10-8 mol·L-1 OUA group, the contraction amplitude also decreased (P<0.01), no significant difference in contraction amplitudes between 50 μmol·L-1 PD98059+1×10-8 mol·L-1 OUA group and 1×10-8 mol·L-1 OUA group (P>0.05). CONCLUSION The 1×10-8-1×10-3 mol·L-1 OUA could increase the contraction amplitude of cardiocytes in rats in concentration-dependent manner. Positive inotropic effect of OUA in low concentration is related to Na/K pump signal transduction. Multiple signal pathways regulate the positive inotropic effect of 1×10-8 mol·L-1 of OUA, including the Src/ROS signal pathway.
关键词
钠钾ATP酶 /
心室肌细胞 /
哇巴因 /
正性肌力 /
活性氧
{{custom_keyword}} /
Key words
Na+-K+-ATPase /
ventricular myocyte /
ouabain /
positive inotropic effect /
ROS
{{custom_keyword}} /
中图分类号：
R965
{{custom_clc.code}}
({{custom_clc.text}})
{{custom_sec.title}}
{{custom_sec.title}}
{{custom_sec.content}}
参考文献
[1] BERS D M. Calcium cycling and signaling in cardiac myocytes. Annu Rev Physiol, 2008, 70:23-49.
[2] GAO J, WYMORE R S, WANG Y, et al. Isoform-specific stimulation of cardiac Na/K pumps by nanomolar concentrations of glycosides. J Gen Physiol, 2002, 119(4): 297-312.
[3] SWIFT F, TOVSRUD N, ENGER U H, et al. The Na+/K+-ATPase alpha2-isoform regulates cardiac contractility in rat cardiomyocytes. Cardiovasc Res, 2007, 75(1): 109-117.
[4] SU S W, XU Y F, MEI H S, et al. Effects of strophanthidin on intracellular calcium concentration in ventricular myocytes of guinea pig. Acta Pharm Sin(药学学报),2008, 43(3):259-266.
[5] SU S W, WANG Y L, LI J X, et al. Relationship between cardiotonic effects and inhibition on cardiac sarcolemmal Na+,K+-ATPase of strophanthidin at low concentration. Acta Pharmacol Sin(中国药理学报), 2003, 24(11): 1103-1107.
[6] YIN J X, WANG Y L, LI Q, et al. Effects of low concentration of dihydroouabain on intracellular calcium in guinea pig ventricular myocytes.Acta Physiol Sin(生理学报),2002, 54(5): 385-389.
[7] NOBLE D. Mechanism of action of therapeutic levels of cardiac glycosides. Cardiovasc Res, 1980, 14(9):495-514.
[8] DOSTANIC-LARSON I, LORENZ J N, VAN HUYSSE J W, et al. Physiological role of the alpha1- and alpha2-isoforms of the Na+-K+-ATPase and biological significance of their cardiac glycoside binding site. Am J Physiol Regul Integr Comp Physiol, 2006, 290(3): 524-528.
[9] KATZ A, LIFSHITZ Y, BAB-DINITZ E, et al. Selectivity of digitalis glycosides for isoforms of human Na,K-ATPase. J Biol Chem, 2010, 285(25): 19582-19592.
[10] ADAMS R J, SCHWARTZ A, GRUPP G, et al. High-affinity ouabain binding site and low-dose inotropic effect in rat myocardium. Nature, 1982, 296(5853): 167-169.
[11] XIONG C, LI J X, GUO H C, et al. The H1-H2 domain of the α1 isoform of Na+-K+-ATPase is involved in ouabain toxicity in rat ventricular myocytes. Toxicol Appl Pharmacol, 2012,262(1): 32-42.
[12] SU S W, WANG U L, ZHANG Y J. Effect of low concentrations of strophanthidin on isolated guinea pig failure hearts.Chin Pharmcol Bull(中国药理学通报), 2003, 19(2):167-171.
[13] ZHANG L, ZHANG Z, GUO H, et al. Na+/K+-ATPase mediated signal transduction and Na+/K+-ATPase regulation. Fundam Clin Pharmacol, 2008, 22(6): 615-621.
[14] XIE Z. Ouabain interaction with cardiac Na/K-ATPase reveals that the enzyme can act as a pump and as a signal transducer. Cell Mol Biol, 2001, 47 (2): 383-390.
{{custom_fnGroup.title_cn}}
脚注
{{custom_fn.content}}
基金
国家自然科学基金资助项目(81302773);河北省自然科学基金项目(H2014206319)
{{custom_fund}}
PDF(938 KB)
