Mechanism of automaticity in cardiomyocytes derived from human induced pluripotent stem cells
Jong J Kim1, Lei Yang2, Bo Lin2
1Department of Bioengineering, University of Pittsburgh, Pittsburgh, PA 15261, USA; Department of Medicine, University of Pittsburgh, Pittsburgh, PA 15261, USA.
Insights
Human induced pluripotent stem cell-derived cardiomyocytes (hiPS-CMs) exhibit spontaneous contractions. Automaticity in hiPS-CMs is driven by a calcium (Ca2+)-clock mechanism, not the typical pacemaker current.
Area of Science:
- Cardiology
- Stem Cell Biology
- Electrophysiology
Background:
- Human induced pluripotent stem cell-derived cardiomyocytes (hiPS-CMs) offer promise for personalized medicine in cardiovascular diseases.
- The mechanism underlying the spontaneous contractions of hiPS-CMs remains unclear.
Purpose of the Study:
- To investigate the intrinsic activity and underlying mechanisms of spontaneous contractions in hiPS-CMs.
- To determine the role of ion currents and calcium handling in hiPS-CM automaticity.
Main Methods:
- Voltage-clamp electrophysiology
- Subcellular-resolution optical mapping of action potentials and intracellular calcium transients
- Pharmacological interventions targeting ion channels and exchangers
Main Results:
- hiPS-CMs showed negligible If and IK1 currents, and their spontaneous calcium transients were unaffected by If inhibition.
- Spontaneous calcium transients increased with extracellular calcium, isoproterenol, or caffeine.
- Automaticity was abolished by inhibiting ryanodine receptors, Na-Ca exchange (NCX), or increasing extracellular potassium.
- hiPS-CMs demonstrated cell-cell coupling and synchronized contractions.
Conclusions:
- Automaticity in hiPS-CMs is primarily driven by a "Ca2+-clock" mechanism.
- This mechanism involves intracellular calcium cycling via the sarcoplasmic reticulum and NCX, which triggers action potentials.
Background And Objectives:
The creation of cardiomyocytes derived from human induced pluripotent stem cells (hiPS-CMs) has spawned broad excitement borne out of the prospects to diagnose and treat cardiovascular diseases based on personalized medicine. A common feature of hiPS-CMs is their spontaneous contractions but the mechanism(s) remain uncertain.
Methods:
Intrinsic activity was investigated by the voltage-clamp technique, optical mapping of action potentials (APs) and intracellular Ca(2+) (Cai) transients (CaiT) at subcellular-resolution and pharmacological interventions.
Results:
The frequency of spontaneous CaiT (sCaiT) in monolayers of hiPS-CMs was not altered by ivabradine, an inhibitor of the pacemaker current, If despite high levels of HCN transcripts (1-4). HiPS-CMs had negligible If and IK1 (inwardly-rectifying K(+)-current) and a minimum diastolic potential of -59.1±3.3mV (n=18). APs upstrokes were preceded by a depolarizing-foot coincident with a rise of Cai. Subcellular Cai wavelets varied in amplitude, propagated and died-off; larger Cai-waves triggered cellular sCaTs and APs. SCaiTs increased in frequency with [Ca(2+)]out (0.05-to-1.8mM), isoproterenol (1μM) or caffeine (100μM) (n≥5, p<0.05). HiPS-CMs became quiescent with ryanodine receptor stabilizers (K201=2μM); tetracaine; Na-Ca exchange (NCX) inhibition (SEA0400=2μM); higher [K(+)]out (5→8mM), and thiol-reducing agents but could still be electrically stimulated to elicit CaiTs. Cell-cell coupling of hiPS-CM in monolayers was evident from connexin-43 expression and CaiT propagation. SCaiTs from an ensemble of dispersed hiPS-CMs were out-of-phase but became synchronous through the outgrowth of inter-connecting microtubules.
Conclusions:
Automaticity in hiPS-CMs originates from a Ca(2+)-clock mechanism involving Ca(2+) cycling across the sarcoplasmic reticulum linked to NCX to trigger APs.
