Different from the human heart, zebrafish are capable of fully regenerating its damaged heart after ventricular resection, cryoinjury or genetic ablation of cardiomyocytes, but the underlying mechanisms are not completely understood. In this study, we report that Itga4, a subunit of integrin heterodimer, is necessary for adult zebrafish heart regeneration. Both mRNA and protein of Itga4 were dynamically expressed during heart regeneration. Homozygous Itga4 mutants showed deficiency in the recruitment of leucocytes and angiogenesis in the injury site, and therefore had severe cardiac fibrosis and compromised myocardial regeneration. Itga4 was found to mainly colocalize with Lcp1-positive and coronin1a-EGFP-positive inflammatory cells, suggesting Itga4 regulates heart regeneration mainly through its function in leucocytes. Taken together, our study has obtained novel insights of Itga4-mediated leukocyte signaling into zebrafish heart regeneration.