Ji-dong Fu1, Hui-mei Yu, Rong Wang, Ji Liang, Huang-tian Yang. 1. Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences and Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China.
Abstract
AIM: To investigate the developmental regulation of intracellular Ca2+ transients, an essential event in excitation-contraction coupling, during cardiomyocyte differentiation. METHODS: Using the embryonic stem (ES) cell in vitro differentiation system and pharmacological intervention, we investigated the molecular and functional regulation of Ca2+ handling proteins on the Ca2+ transients at early, intermediate and later differentiation stages of ES cell-derived cardiomyocytes (ESCM). RESULTS: Nifedipine, a selective antagonist of L-type Ca2+ channels, totally blocked Ca2+ transients even in the condition of field-electric stimulation in ESCM at three differentiation stages. The Ca2+ transients of ESCM were also inhibited by both ryanodine [an inhibitor of ryanodine receptors (RyRs)] and 2-aminoethoxydipheylborate [2-APB, an inhibitor of inositol-1,4,5-trisphosphate receptors (IP3Rs)]. The inhibitory effect of ryanodine increased with the time of differentiation, while the effect of 2-APB decreased with the differentiation. Thapsigargin, an inhibitor of SR Ca2+-pump ATPase, inhibited Ca2+ transients equally at three differentiation stages that matched the expression profile. Na+ free solution, which inhibits Na+-Ca2+ exchanger (NCX) to extrude Ca2+ from cytosol, did not affect the amplitude of Ca2+ transients of ESCM until the latter differentiation stage, but it significantly enhanced the basal Ca2+ concentration. CONCLUSION: The Ca2+ transients in ESCM depend on both the sarcolemmal Ca2+ entry via L-type Ca2+ channels and the SR Ca2+ release from RyRs and IP3Rs even at the early differentiation stage; but NCX seems not to regulate the peak of Ca2+ transients until the latter differentiation stage.
AIM: To investigate the developmental regulation of intracellular Ca2+ transients, an essential event in excitation-contraction coupling, during cardiomyocyte differentiation. METHODS: Using the embryonic stem (ES) cell in vitro differentiation system and pharmacological intervention, we investigated the molecular and functional regulation of Ca2+ handling proteins on the Ca2+ transients at early, intermediate and later differentiation stages of ES cell-derived cardiomyocytes (ESCM). RESULTS:Nifedipine, a selective antagonist of L-type Ca2+ channels, totally blocked Ca2+ transients even in the condition of field-electric stimulation in ESCM at three differentiation stages. The Ca2+ transients of ESCM were also inhibited by both ryanodine [an inhibitor of ryanodine receptors (RyRs)] and 2-aminoethoxydipheylborate [2-APB, an inhibitor of inositol-1,4,5-trisphosphate receptors (IP3Rs)]. The inhibitory effect of ryanodine increased with the time of differentiation, while the effect of 2-APB decreased with the differentiation. Thapsigargin, an inhibitor of SR Ca2+-pump ATPase, inhibited Ca2+ transients equally at three differentiation stages that matched the expression profile. Na+ free solution, which inhibits Na+-Ca2+ exchanger (NCX) to extrude Ca2+ from cytosol, did not affect the amplitude of Ca2+ transients of ESCM until the latter differentiation stage, but it significantly enhanced the basal Ca2+ concentration. CONCLUSION: The Ca2+ transients in ESCM depend on both the sarcolemmal Ca2+ entry via L-type Ca2+ channels and the SR Ca2+ release from RyRs and IP3Rs even at the early differentiation stage; but NCX seems not to regulate the peak of Ca2+ transients until the latter differentiation stage.
Authors: Worawan Limpitikul; Nicolas Christoforou; Susan A Thompson; John D Gearhart; Leslie Tung; Elizabeth A Lipke Journal: Cardiovasc Eng Technol Date: 2010-08-06 Impact factor: 2.495