EXCITAION-CONTRACTION COUPLINGS DURING IN VITRO DIFFERENTIATION INTO CARDIOMYOCYTES FROM MOUSE EMBRYONIC STEM CELLS

Abstract

Mouse embryonic stem (mES) cells have abilities to differentiate into various types of the cells, including cardiomyocytes. In this study we examined Ca²⁺ signaling pathways in mES cells for excitation-contraction (E-C) coupling during the differentiation into cardiomyocytes. To identify the cardiac precursor cells, we used the mES cells transfected with the vector containing GFP driven by mouse cardiac α-actin promoter. Caffeine effects on [Ca²⁺]_i were observed in GFP-positive precursor cells cultured for longer than 5 days without leukemia inhibitory factor and these effects increased during the differentiation. A depolarization induced elevation of [Ca²⁺]_i was observed in 10%, 13%, 25% and 46% of 4-day, 5-7 day, 12-14 day and 24-27 day differentiated cells, respectively. In whole cell patch clamp experiments, T-type but not L-type Ca²⁺ currents could be recorded in the precursor cells at 12-day stages. In 19-day or beating cells, L-type Ca²⁺ currents were observed. The expressions of mRNAs for RyR2 and T-type Ca²⁺ channel were detected from 4-day differentiated cells, but mRNAs for L-type Ca²⁺ channel could be detected from 7-day differentiated cells. The expression levels of these genes increased during the differentiation. From these results we conclude that main Ca²⁺ entry pathways switch from T-type Ca²⁺ channel to L-type one during the differentiation, which might contribute to E-C coupling. [Jpn J Physiol 54 Suppl:S103 (2004)]