Local control of excitation-contraction coupling in human embryonic stem cell-derived cardiomyocytes.
We investigated the mechanisms of excitation-contraction (EC) coupling in human embryonic stem cell-derived cardiomyocytes (hESC-CMs) and fetal ventricular myocytes (hFVMs) using patch-clamp electrophysiology and confocal microscopy. We tested the hypothesis that Ca(2+) influx via voltage-gated L-type Ca(2+) channels activates Ca(2+) release from the sarcoplasmic reticulum (SR) via a local control mechanism in hESC-CMs and hFVMs. Field-stimulated, whole-cell [Ca(2+)](i) transients in hESC-CMs required Ca(2+) entry through L-type Ca(2+) channels, as evidenced by the elimination of such transients by either removal of extracellular Ca(2+) or treatment with diltiazem, an L-type channel inhibitor. Ca(2+) release from the SR also contributes to the [Ca(2+)](i) transient in these cells, as evidenced by studies with drugs interfering with either SR Ca(2+) release (i.e. ryanodine and caffeine) or reuptake (i.e. thapsigargin and cyclopiazonic acid). As in adult ventricular myocytes, membrane depolarization evoked large L-type Ca(2+) currents (I(Ca)) and corresponding whole-cell [Ca(2+)](i) transients in hESC-CMs and hFVMs, and the amplitude of both I(Ca) and the [Ca(2+)](i) transients were finely graded by the magnitude of the depolarization. hESC-CMs exhibit a decreasing EC coupling gain with depolarization to more positive test potentials, "tail" [Ca(2+)](i) transients upon repolarization from extremely positive test potentials, and co-localized ryanodine and sarcolemmal L-type Ca(2+) channels, all findings that are consistent with the local control hypothesis. Finally, we recorded Ca(2+) sparks in hESC-CMs and hFVMs. Collectively, these data support a model in which tight, local control of SR Ca(2+) release by the I(Ca) during EC coupling develops early in human cardiomyocytes.
Zhu
Wei-Zhong
WZ
Department of Pathology, University of Washington, Seattle, Washington, United States of America.
Santana
Luis F
LF
Laflamme
Michael A
MA
D002118:Calcium / Q000378:metabolism; D020746:Calcium Channels, L-Type / Q000378:metabolism*; D002478:Cells, Cultured; D004594:Electrophysiology; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000502:physiology; D006801:Humans; D008564:Membrane Potentials; D009200:Myocardial Contraction; D032383:Myocytes, Cardiac / Q000166:cytology / Q000502:physiology*; D012519:Sarcoplasmic Reticulum / Q000378:metabolism
D020746:Calcium Channels, L-Type; D002118:Calcium