We provided novel evidence as follows: (1) rapid atrial pacing increased the release of plasma and atrial exosomes in canines; (2) increased exosomes mainly played a pro-fibrotic role in AF; thus, blockade of exosome release with GW4869 suppressed AF by alleviating atrial fibrosis; and (3) this pro-fibrotic effect of exosomes partly resulted from miR-21-5p enrichment in exosomes and its downstream pathway of TIMP3/TGF-β1. Here, TGFB1 is linked to atrial fibrillation.