title: Na+/Ca2+ exchanger is a determinant of excitation-contraction coupling in human embryonic stem cell-derived ventricular cardiomyocytes.
abstract: In adult cardiomyocytes (CMs), the Na(+)/Ca(2+) exchanger (NCX) is a well-defined determinant of Ca(2+) homeostasis. Developmentally, global NCX knockout in mice leads to abnormal myofibrillar organization, electrical defects, and early embryonic death. Little is known about the expression and function of NCX in human heart development. Self-renewable, pluripotent human embryonic stem cells (hESCs) can serve as an excellent experimental model. However, hESC-derived CMs are highly heterogeneous. A stably lentivirus-transduced hESC line (MLC2v-dsRed) was generated to express dsRed under the transcriptional control of the ventricular-restricted myosin light chain-2v (MLC2v) promoter. Electrophysiologically, dsRed+ cells differentiated from MLC2vdsRed hESCs displayed ventricular action potentials (AP), exclusively. Neither atrial nor pacemaker APs were observed. While I(Ca-L), I(f), and I(Kr) were robustly expressed, I(Ks) and I(K1) were absent in dsRed+ ventricular hESCCMs. Upon differentiation (7+40 to +90 days), the basal [Ca(2+)](i), Ca(2+) transient amplitude, maximum upstroke, and decay velocities significantly increased (P < 0.05). The I(Ca-L) antagonizer nifedipine (1 microM) decreased the Ca(2+) transient amplitude (to approximately 30%) and slowed the kinetics (by approximately 2-fold), but Ca(2+) transients could still be elicited even after complete ICa-L blockade, suggesting the presence of additional Ca(2+) influx(es). Indeed, Ni(2+)-sensitive INCX could be recorded in 7+40- and +90-day dsRed+ hESC-CMs, and its densities increased from -1.2 +/- 0.6 pA/pF at -120 mV and 3.6 +/- 1.0 pA/pF at 60 mV by 6- and 2-folds, respectively. With higher [Ca(2+)](i), 7+90-day ventricular hESC-CMs spontaneously but irregularly fired transients upon a single stimulus under an external Na(+)-free condition; however, without extracellular Na(+), nifedipine could completely inhibit Ca(2+) transients. We conclude that I(NCX) is functionally expressed in developing ventricular hESC-CMs and contributes to their excitation-contraction coupling.
authors.affiliation: Human Embryonic Stem Cell Consortium, University of California, Davis, California, USA.
authors.forename: Ji-Dong
authors.initials: JD
authors.lastname: Fu
authors.forename: Peng
authors.initials: P
authors.lastname: Jiang
authors.forename: Stephanie
authors.initials: S
authors.lastname: Rushing
authors.forename: Jing
authors.initials: J
authors.lastname: Liu
authors.forename: Nipavan
authors.initials: N
authors.lastname: Chiamvimonvat
authors.forename: Ronald A
authors.initials: RA
authors.lastname: Li
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism; D020746:Calcium Channels, L-Type / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D056966:Excitation Contraction Coupling / Q000187:drug effects; D006352:Heart Ventricles / Q000166:cytology*; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects; D008164:Luminescent Proteins / Q000378:metabolism; D051379:Mice; D032383:Myocytes, Cardiac / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D009543:Nifedipine / Q000494:pharmacology; D010641:Phenotype; D019831:Sodium-Calcium Exchanger / Q000378:metabolism*; D014161:Transduction, Genetic
chemical_list: D020746:Calcium Channels, L-Type; D008164:Luminescent Proteins; D019831:Sodium-Calcium Exchanger; C413663:fluorescent protein 583; D009543:Nifedipine; D002118:Calcium