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Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM) are a promising tool for drug testing and modelling genetic disorders. Abnormally low upstroke velocity is a current limitation. Here we investigated the use of 3D engineered heart tissue (EHT) as a culture method with greater resemblance to human heart tissue in comparison to standard technique of 2D monolayer (ML) format. I Na  was measured in ML or EHT using the standard patch-clamp technique. I Na  density was ~1.8 fold larger in EHT (−18.5 ± 1.9 pA/pF; n = 17) than in ML (−10.3 ± 1.2 pA/pF; n = 23; p < 0.001), approaching densities reported for human CM. Inactivation kinetics, voltage dependency of steady-state inactivation and activation of I Na  did not differ between EHT and ML and were similar to previously reported values for human CM. Action potential recordings with sharp microelectrodes showed similar upstroke velocities in EHT (219 ± 15 V/s, n = 13) and human left ventricle tissue (LV, 253 ± 7 V/s, n = 25). EHT showed a greater resemblance to LV in CM morphology and subcellular Na V 1.5 distribution. I Na  in hiPSC-CM showed similar biophysical properties as in human CM. The EHT format promotes I Na  density and action potential upstroke velocity of hiPSC-CM towards adult values, indicating its usefulness as a model for excitability of human cardiac tissue.

Human iPSC-derived cardiomyocytes cultured in 3D engineered heart tissue show physiological upstroke velocity and sodium current density