Abstract
Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM) provide a unique opportunity to study human heart physiology and pharmacology and repair injured hearts. The suitability of hiPSC-CM critically depends on how closely they share physiological properties of human adult cardiomyocytes (CM). Here we investigated whether a 3D engineered heart tissue (EHT) culture format favors maturation and addressed the L-type Ca2+-current (ICa,L) as a readout. The results were compared with hiPSC-CM cultured in conventional monolayer (ML) and to our previous data from human adult atrial and ventricular CM obtained when identical patch-clamp protocols were used. HiPSC-CM were two- to three-fold smaller than adult CM, independently of culture format [capacitance ML 45 ± 1 pF (n = 289), EHT 45 ± 1 pF (n = 460), atrial CM 87 ± 3 pF (n = 196), ventricular CM 126 ± 8 pF (n = 50)]. Only 88% of ML cells showed ICa, but all EHT. Basal ICa density was 10 ± 1 pA/pF (n = 207) for ML and 12 ± 1 pA/pF (n = 361) for EHT and was larger than in adult CM [7 ± 1 pA/pF (p < 0.05, n = 196) for atrial CM and 6 ± 1 pA/pF (p < 0.05, n = 47) for ventricular CM]. However, ML and EHT showed robust T-type Ca2+-currents (ICa,T). While (−)-Bay K 8644, that activates ICa,L directly, increased ICa,Lto the same extent in ML and EHT, β1- and β2-adrenoceptor effects were marginal in ML, but of same size as (−)-Bay K 8644 in EHT. The opposite was true for serotonin receptors. Sensitivity to β1 and β2-adrenoceptor stimulation was the same in EHT as in adult CM (−logEC50: 5.9 and 6.1 for norepinephrine (NE) and epinephrine (Epi), respectively), but very low concentrations of Rp-8-Br-cAMPS were sufficient to suppress effects (−logEC50: 5.3 and 5.3 respectively for NE and Epi). Taken together, hiPSC-CM express ICa,L at the same density as human adult CM, but, in contrast, possess robust ICa,T. Increased effects of catecholamines in EHT suggest more efficient maturation.
Highlights
The L-type Ca2+-current (ICa,L) is central for cardiac electrophysiology
When we plotted individual cell capacitance as an electrophysiological correlate of cell size in hiPSC-CM and human adult CM, we found substantial overlap between hiPSCCM and adult CM, but mean values in hiPSC-CM were twoto three-fold smaller than human atrial and ventricular CM, respectively (Figure 1)
Despite the fact that human adult CM were clearly larger than hiPSC-CM, both human adult and hiPSC-CM showed a positive correlation between cell size and ICa
Summary
The L-type Ca2+-current (ICa,L) is central for cardiac electrophysiology It contributes to the shape of the cardiac action potential and its regulation plays an important role in cardiac excitability and contractility (Tsien, 1983). HiPSC-CM display an immature cardiac phenotype, and current efforts are directed toward means to unfold the full potential of these cells by increasing their maturity (Yang et al, 2014). One such strategy could be culture in engineered heart tissue (EHT) under conditions in which hiPSC-CM form a 3-dimensional network and perform auxotonic contractile work against elastic silicone posts (Schaaf et al, 2011). We directly compared the biophysics and regulation of Ca2+-currents in hiPSC-CM cultured either in standard monolayer format (ML) or as EHT and compared the data to our previous data on human adult CM obtained under identical patch-clamp protocols
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