Na+/K+ ATPase and its functional coupling with Na+/Ca2+ exchanger in mouse embryonic stem cells during differentiation into cardiomyocytes

• Published in: Cell calcium (Edinburgh) 2005-02, Vol.37 (2), p.137-151
• Main Authors: Otsu, Keishi, Kuruma, Akinori, Yanagida, Eri, Shoji, Satoshi, Inoue, Takafumi, Hirayama, Yoshiyuki, Uematsu, Hiroshi, Hara, Yukichi, Kawano, Seiko
• Format: Article
• Language: English
• Subjects: Animals; Cardiotonic Agents - pharmacology; Cell Differentiation - physiology; Membrane Potentials - physiology; Mice; Microscopy, Fluorescence; Myocytes, Cardiac - cytology; Myocytes, Cardiac - drug effects; Myocytes, Cardiac - metabolism; Ouabain - pharmacology; Patch-Clamp Techniques; Sodium - metabolism; Sodium-Calcium Exchanger - metabolism; Sodium-Potassium-Exchanging ATPase - metabolism; Stem Cells - cytology; Stem Cells - drug effects; Stem Cells - metabolism
• ISSN: 0143-4160
• DOI: 10.1016/j.ceca.2004.08.004

Cardiomyocytes derived from mouse embryonic stem (mES) cells have been demonstrated to exhibit a time-dependent expression of ion channels and signal transduction pathways in electrophysiological studies. However, ion transporters, such as Na+/K+ ATPase (Na+ pump) or Na+/Ca2+ exchanger, which play crucial roles for cardiac function, have not been well studied in this system. In this study, we investigated the functional expression of Na+/K+ ATPase and Na+/Ca2+ exchanger in mES cells during in vitro differentiation into cardiomyocytes, as well as the functional coupling between the two transporters. By measuring [Na+]i and Na+ pump current (Ip), it was shown that an ouabain-high sensitive Na+/K+ ATPase was expressed functionally in undifferentiated mES cells and these activities increased during a time course of differentiation. Using RT-PCR, the expression of mRNA for alpha1-subunit and alpha3-subunit of the Na+/K+ ATPase could be detected in both undifferentiated mES cells and derived cardiomyocytes. In contrast alpha2-subunit mRNA could be detected only in derived cardiomyocytes but not in undifferentiated mES cells. mRNA for the Na+/Ca2+ exchanger 1 isoform (NCX1) could be detected in undifferentiated mES cells and its expression levels seemed to gradually increase throughout the differentiation accompanied by increasing its Ca2+ extrusion function. At the middle stages of differentiation (after 10-day induction), more than 75% derived cardiomyocytes exhibited [Ca2+]i oscillations by blocking of Na+/K+ ATPase, suggesting the functional coupling with Na+/Ca2+ exchanger. From these results and RT-PCR analysis, we conclude that alpha2-subunit Na+/K+ ATPase mainly contributes to establish the functional coupling with NCX1 at the middle stages of differentiation of cardiomyocytes.