AbstractThe growth factor Neuregulin‐1 (NRG‐1) regulates myocardial growth and is currently under clinical investigation as a treatment for heart failure. Here, we demonstrate in severalin vitroandin vivomodels that STAT5b mediates NRG‐1/EBBB4‐stimulated cardiomyocyte growth. Genetic and chemical disruption of the NRG‐1/ERBB4 pathway reduces STAT5b activation and transcription of STAT5b target genesIgf1,Myc, andCdkn1ain murine cardiomyocytes. Loss ofStat5balso ablates NRG‐1‐induced cardiomyocyte hypertrophy. Dynamin‐2 is shown to control the cell surface localization of ERBB4 and chemical inhibition of Dynamin‐2 downregulates STAT5b activation and cardiomyocyte hypertrophy. In zebrafish embryos, Stat5 is activated during NRG‐1‐induced hyperplastic myocardial growth, and chemical inhibition of the Nrg‐1/Erbb4 pathway or Dynamin‐2 leads to loss of myocardial growth and Stat5 activation. Moreover, CRISPR/Cas9‐mediated knockdown ofstat5bresults in reduced myocardial growth and cardiac function. Finally, the NRG‐1/ERBB4/STAT5b signaling pathway is differentially regulated at mRNA and protein levels in the myocardium of patients with pathological cardiac hypertrophy as compared to control human subjects, consistent with a role of the NRG‐1/ERBB4/STAT5b pathway in myocardial growth.