Leak current, even with gigaohm seals, can cause misinterpretation of stem-cell derived cardiomyocyte action potential recordings

Abstract

Human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) have become an essential tool to study arrhythmia mechanisms. Much of the foundational work on these cells, and the computational models built from the resultant data, has overlooked the contribution of seal-leak current on the immature and heterogeneous phenotype that has come to define these cells. Here, we use in silico and in vitro studies to demonstrate how seal-leak current depolarises action potentials (APs), substantially affecting their morphology, even with seal resistances (Rseal) above 1 G{Omega}. We show that compensation of this leak current is difficult due to challenges with recording accurate measures of Rseal during an experiment. Using simulation, we show that Rseal measures: 1) change during an experiment, invalidating the use of pre-rupture values, and 2) are polluted by the presence of transmembrane currents at every voltage. Finally, we posit the background sodium current in baseline iPSC-CM models imitates the effects of seal-leak current and is increased to a level that masks the effects of seal-leak current on iPSC-CMs. Based on these findings, we make three recommendations to improve iPSC-CM AP data acquisition, interpretation, and model-building. Taking these recommendations into account will improve our understanding of iPSC-CM physiology and the descriptive ability of models built from such data. Key pointsO_LIHuman induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) are an essential tool in the study of cardiac arrhythmia mechanisms. C_LIO_LITheir immature and heterogeneous action potential phenotype complicates the interpretation of experimental data, and has slowed their acceptance in industry and academia. C_LIO_LIWe suggest that a leak current caused by an imperfect pipette-membrane seal during single-cell patch-clamp experiments is partly responsible for inducing this phenotype. C_LIO_LIUsing in vitro experiments and computational modelling, we show that this seal-leak current affects iPSC-CM AP morphology, even under ideal experimental conditions. C_LIO_LIBased on these findings, we make recommendations that should be considered when interpreting, analysing and fitting iPSC-CM data. C_LI