Low concentration of ouabain elevates intracellular free calcium in guinea pig ventricular myocytes: The possible signal transduction pathway / 第二军医大学学报

ABSTRACT

Objective: To study the effect of low concentration of ouabain MUM on intracellular calcium concentration ([Ca2+]i) in guinea pig ventricular myocytes and to understand whether low concentration of OUA can increase [Ca2+]i through Na+, K+-ATPase channel. Methods: The guinea pig ventricular myocytes were obtained by enzymatic digestion and the [Ca2+]i fluorescent density of individual myocytes was observed under confocal laser scanning microscope. The isolated ventricular myocytes were then incubated with different concentrations of ouabain(10-9, 10-8, 10-7, 10-6 mol · L-1). The sediment was subjected to Western blot analysis to assess the phosphorylation of Src by OUA. Results: In normal Tyrode' s solution and Ca2+-free Tyrode's solution, OUA (1 × 10-9 -1 × 10-6) mol · L-1 elevated [Ca2+]i in a concentration-dependent manner, with the elevation in normal Tyrode's solution more obvious (P < 0.05). Genistein (GST) (1, 10, 50, and 100 μmol - L-1) abolished the OUA-induced increases of [Ca2+]i in a concentration-dependent manner. There were two tyrosine-phosphorylated bands, with the molecular weights being 120 000 and 70 000. Compared with control group, the densities of the 2 bands in all OUA groups were significantly higher (P<0.05) and GST could obviously inhibit the elevating effect of OUA. Conclusion: Low concentration of OUA may promote opening of Ca2+ channel and release of intracellular Ca2+, and subsequently elevate intracellular free calcuim through phosphorylation of tyrosine and activition of OUA/Na+, k+-ATPase/Src signal transduction pathway.