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_v1.5 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 (APD₂₀) 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, APD₂₀ 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.

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