Ultrarapid Delayed Rectifier K+ Channelopathies in Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes

Abstract

Atrial fibrillation (AF) is the most common cardiac arrhythmia. About 5–15% of AF patients have a mutation in a cardiac gene, including mutations in KCNA5 , encoding the K v 1.5 α-subunit of the ion channel carrying the atrial-specific ultrarapid delayed rectifier K + current (I Kur ). Both loss-of-function and gain-of-function AF-related mutations in KCNA5 are known, but their effects on action potentials (APs) of human cardiomyocytes have been poorly studied. Here, we assessed the effects of wild-type and mutant I Kur on APs of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). We found that atrial-like hiPSC-CMs, generated by a retinoic acid-based differentiation protocol, have APs with faster repolarization compared to ventricular-like hiPSC-CMs, resulting in shorter APs with a lower AP plateau. Native I Kur , measured as current sensitive to 50 μM 4-aminopyridine, was 1.88 ± 0.49 (mean ± SEM, n = 17) and 0.26 ± 0.26 pA/pF ( n = 17) in atrial- and ventricular-like hiPSC-CMs, respectively. In both atrial- and ventricular-like hiPSC-CMs, I Kur blockade had minimal effects on AP parameters. Next, we used dynamic clamp to inject various amounts of a virtual I Kur , with characteristics as in freshly isolated human atrial myocytes, into 11 atrial-like and 10 ventricular-like hiPSC-CMs, in which native I Kur was blocked. Injection of I Kur with 100% density shortened the APs, with its effect being strongest on the AP duration at 20% repolarization (APD 20 ) of atrial-like hiPSC-CMs. At I Kur densities < 100% (compared to 100%), simulating loss-of-function mutations, significant AP prolongation and raise of plateau were observed. At I Kur densities > 100%, simulating gain-of-function mutations, APD 20 was decreased in both atrial- and ventricular-like hiPSC-CMs, but only upon a strong increase in I Kur . In ventricular-like hiPSC-CMs, lowering of the plateau resulted in AP shortening. We conclude that a decrease in I Kur , mimicking loss-of-function mutations, has a stronger effect on the AP of hiPSC-CMs than an increase, mimicking gain-of-function mutations, whereas in ventricular-like hiPSC-CMs such increase results in AP shortening, causing their AP morphology to become more atrial-like. Effects of native I Kur modulation on atrial-like hiPSC-CMs are less pronounced than effects of virtual I Kur injection because I Kur density of atrial-like hiPSC-CMs is substantially smaller than that of freshly isolated human atrial myocytes.

Human induced pluripotent stem cell-derived cardiomyocytes as an electrophysiological model: Opportunities and challenges—The Hamburg perspective

“…In the seminal paper that showed the efficacy of RA to induce atrial differentiation in human embryonic stem cell (hESC)-derived CM, currents sensitive to 4-AP (50 µM) accounted to ∼6 pA/pF (pulse potential of +50 mV) ( Devalla et al, 2015 ), which is 2-fold lower than values for I Kur in human atrial CM ( Ford et al, 2013 ). A later study in hiPSC-CM showed even smaller currents (∼2 pA/pF) ( Hilderink et al, 2020 ), but they were sufficient to produce a short APD at 20% repolarization (APD 20 ; 20 ms), close to the situation in human atrium (7 ms) ( Liang et al, 2016 ; Horváth et al, 2018 ). Furthermore, as in human atrium, plateau voltage was below 0 mV, and both short APD 20 and negative plateau voltage were sensitive to 50 µM 4-AP ( Devalla et al, 2015 ).…”

PITX2 Knockout Induces Key Findings of Electrical Remodeling as Seen in Persistent Atrial Fibrillation

“…WHAT THE STUDY ADDS the large impact of atrial-selective potassium currents on repolarization. 17,[25][26][27] The present study presents a technical improvement of hiPSC atrial differentiation, which results in APs closely replicating human atrial electrophysiology. In these hiPSC-aCMs, PITX2 deficiency was associated with some characteristics known from persistent AF in patients and animal models.…”

Molecular and electrophysiological evaluation of human cardiomyocyte subtypes to facilitate generation of composite cardiac models

“…In addition, the APD was shorter in ACM compared to VCM, likely due to the presence of a large repolarizing ultrarapid delayed rectifier K + current. 9 , 42 …”

“…In addition, the APD was shorter in ACM compared to VCM, likely due to the presence of a large repolarizing ultrarapid delayed rectifier K + current. 9,42 We also evaluated the sensitivity of the various cardiomyocyte subtypes to agents that modulate heart rate. 100 nM noradrenaline elicited a positive chronotropic response in all groups, although the effect did not reach significance in VCM.…”