Molecular events involved in acute myocardial infarction (AMI) still remain unclear. A rat AMI model and cardiomyocytes cultured in vitro were used to mimic hypoxic conditions, and the profiles of histone methylation-related gene expression were explored. The demethylase Kdm6a expression was significantly upregulated in the rat AMI model and in hypoxia induction. The apoptosis rate of cardiomyocytes was significantly exacerbated when Kdm6a was knocked down. The expression of the Na + /Ca 2+ exchanger (Ncx) was significantly upregulated in cardiomyocytes under hypoxia. Knockdown of Kdm6a downregulated the Ncx expression via enhancing H3K27me3 modification on Ncx gene promoter, and attenuated the intracellular calcium influx ability in cardiomyocytes as a consequence. Kdm6a regulates Ncx expression through reducing the H3K27me3 level on the Ncx promoter or enhancer. This finding provides a basis for further study of Kdm6a as a new regulator for AMI development. © 2019, Springer Science+Business Media, LLC, part of Springer Nature.