Electrical stimulation promotes cardiomyocyte differentiation and cardiac development

Electrical signals play critical roles in early heart development by regulating cardiomyocyte differentiation, proliferation, and energy metabolism. However, the mechanisms by which electrical stimulation (ES) influences early cardiac development remain incompletely understood. Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) and zebrafish embryos were employed as complementary in vitro and in vivo models. hiPSC-CMs were subjected to continuous ES from the cardiac mesoderm stage (day 4) to day 10 of differentiation. Cellular analyses included immunofluorescence, calcium imaging, proliferation assays, ATP quantification, and qRT-PCR. Zebrafish embryos were subjected to continuous ES for 10 h and 24 h post-fertilization (hpf), respectively. Subsequently, at 36, 48, and 72 hpf, the heart rate, contractile function, proliferation of cardiac cells, and vascular development were assessed. RNA sequencing was conducted on hiPSC-CMs and zebrafish embryos to assess transcriptomic changes, followed by bioinformatic pathway enrichment analysis. Functional studies involved siRNA mediated GATA4 knockdown in hiPSC-CMs, and in zebrafish embryos treated with a gata4 inhibitor, to systematically evaluate the effects of gata4 on cardiac function. ES significantly enhanced hiPSC-CMs differentiation, calcium handling, sarcomere maturation, proliferation, and energy metabolism, accompanied by upregulation of RYR2 and oxidative phosphorylation genes. In zebrafish embryos, ES accelerated body growth, yolk absorption, heart rate, cardiac contractility, and intersegmental vessel development. Transcriptomic analysis revealed upregulation of genes associated with sarcomere structure, calcium signaling, and energy metabolism, consistent with increased cardiac function and vascular development. Further investigation demonstrated that ES upregulated the expression of GATA4 ; GATA4 knockdown reduces hiPSC-CMs differentiation, while its inhibitor 3i 1000 abolishes ES enhanced zebrafish heart contractility. These findings indicate that GATA4 plays a significant role in the regulation of cardiac differentiation and heart development by ES. ES promotes early cardiac cell differentiation and cardiovascular development by enhancing energy metabolism, associated with increased GATA4 related transcriptional activity. These findings offer mechanistic insights into bioelectrical cardiac development, laying a foundation for regenerative therapies for congenital and early-stage heart diseases.