Abstract Body: Cardiovascular diseases are the leading causes of death in the world, among which myocardial infarction (MI) is the most fatal form. Following MI, the death of cardiomyocytes induces cardiac remodeling, which is mainly mediated by cardiac fibroblasts but also involves other cardiac cell types. A deeper understanding of the dynamics of individual cell types among different stages after myocardial infarction may help identify novel therapeutic strategies. Here, through single-nucleus multiomic (snRNAseq and snATACseq) analysis of uninjured mouse hearts and mouse hearts at different time points (2 h, 1 d, 3 d, 7 d, and 4 w) after MI, we identified multiple cardiac cell types, including cardiomyocyte, fibroblast, epicardial cell, endothelial cell, mural cell, and immune cell, and their dynamic gene expression and chromatin accessibility after MI. Features specific to the acute and chronic responses of different cell types to MI were identified. Cell type-specific single-nucleus gene regulatory networks (snGRNs) constructed through integrated analysis of snRNAseq and snATACseq data revealed transcription factors possibly responsible for the gene expression changes in individual cell types. The cardiac fibroblast GRN was further enriched using bulk RNAseq and ATACseq data collected at more time points (uninjured, and 2 h, 12 h, 1 d, 3 d, 7 d, 2 w, and 4 w after MI) and non-coding RNA transcriptomics. Promoter and enhancer activities and chromatin interactions in cardiac fibroblasts were further verified by CUT&Tag targeting corresponding histone markers and Hi-C, respectively. The refined cardiac fibroblast GRN identified multiple key transcription factors that likely govern cardiac fibroblast activation and differentiation after MI.