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Electrophysiological Analysis of human Pluripotent Stem Cell-derived Cardiomyocytes hPSC-CMs Using Multi-electrode Arrays MEAs
Published on: May 12, 2017
cAMP-PKA signaling modulates the automaticity of human iPSC-derived cardiomyocytes
Savyon Mazgaoker1, Ido Weiser-Bitoun1, Inbar Brosh1
1Laboratory of Bioelectric and Bioenergetic Systems, Faculty of Biomedical Engineering, Technion-Israel Institute of Technology, Haifa, Israel.
Insights
Human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) automaticity relies on coupled Ca2+ and membrane clocks. This study reveals Ca2+-cAMP-PKA signaling mediates crosstalk, crucial for normal cardiac function.
Area of Science:
- Cardiology
- Stem Cell Biology
- Molecular Physiology
Background:
- Human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) are valuable for drug screening and disease modeling.
- Understanding the physiological mechanisms of hiPSC-CMs, particularly automaticity, is crucial for their reliable use.
- Automaticity in hiPSC-CMs is governed by the interplay of the Ca2+ clock and the membrane (M) clock.
Purpose of the Study:
- To investigate the coupling mechanisms between the Ca2+ and M clocks in hiPSC-CMs.
- To test hypotheses regarding the role of local Ca2+ releases (LCRs) and cAMP/PKA signaling in regulating automaticity.
- To determine if LCR period reflects crosstalk within the coupled-clock system and if perturbing one clock affects overall automaticity.
Main Methods:
- Measurement of local and global Ca2+ transients in hiPSC-CMs.
- Pharmacological interventions targeting β-adrenergic and cholinergic receptors, PKA signaling, pacemaker current If, and sarcoplasmic reticulum Ca2+ kinetics.
- Comparison of LCR characteristics between hiPSC-CMs and rabbit sinoatrial node (SAN) cells.
Main Results:
- LCR properties directly correlate with the spontaneous beat interval of hiPSC-CMs.
- Modulating cAMP-dependent coupling (via receptor activation or PKA manipulation) altered LCR properties and hiPSC-CM automaticity.
- Disrupting clock coupling (attenuating If or SR Ca2+ kinetics) reduced beating rate and prolonged LCR period.
- hiPSC-CMs and rabbit SAN cells exhibit similar LCR characteristics at comparable beating rates.
Conclusions:
- hiPSC-CM automaticity is regulated by a coupled-clock system.
- Ca2+-cAMP-PKA signaling is the key mediator of functional crosstalk between the Ca2+ and M clocks.
- This coupled-clock system is essential for maintaining normal hiPSC-CM automaticity and cardiac function.
Abstract:
Human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) have been used to screen and characterize drugs and to reveal mechanisms underlying cardiac diseases. However, before hiPSC-CMs can be used as a reliable experimental model, the physiological mechanisms underlying their normal function should be further explored. Accordingly, a major feature of hiPSC-CMs is automaticity, which is regulated by both Ca2+ and membrane clocks. To investigate the mechanisms coupling these clocks, we tested three hypotheses: (1) normal automaticity of spontaneously beating hiPSC-CMs is regulated by local Ca2+ releases (LCRs) and cAMP/PKA-dependent coupling of Ca2+ clock to M clock; (2) the LCR period indicates the level of crosstalk within the coupled-clock system; and (3) perturbing the activity of even one clock can lead to hiPSC-CM-altered automaticity due to diminished crosstalk within the coupled-clock system. By measuring the local and global Ca2+ transients, we found that the LCRs properties are correlated with the spontaneous beat interval. Changes in cAMP-dependent coupling of the Ca2+ and M clocks, caused by a pharmacological intervention that either activates the β-adrenergic or cholinergic receptor or upregulates/downregulates PKA signaling, affected LCR properties, which in turn altered hiPSC-CMs automaticity. Clocks' uncoupling by attenuating the pacemaker current If or the sarcoplasmic reticulum Ca2+ kinetics, decreased hiPSC-CMs beating rate, and prolonged the LCR period. Finally, LCR characteristics of spontaneously beating (at comparable rates) hiPSC-CMs and rabbit SAN are similar. In conclusion, hiPSC-CM automaticity is controlled by the coupled-clock system whose function is mediated by Ca2+-cAMP-PKA signaling.
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