Although histone deacetylases (HDACs) was are the promising target for the initiation and development of diabetic cardiomyopathy (DCM), which isoform of them plays the key role remains unclear. The present study was designed to determine whether DCM could be prevented by selective inhibition of HDAC3 and the underlying mechanism. Male type 1 diabetic OVE26 and age-matched wild-type mice were given the selective HDAC3 inhibitor (HDAC3i) RGFP966 or vehicle for 3 months before the cardiac function was examined with Echo. Results indicated that HDAC3i treatment improved cardiac function in the diabetes group. HDAC3 activity was significantly increased in the heart of diabetic mice, which was blocked by the treatment of HDAC3i. Oxidative stress as a major cause of DCM is also inhibited by HDAC3i. Mechanistically cardiac miR-200a, which targets and destabilizes kelch-like ECH-associated protein 1 (KEAP1) mRNA, was significantly up-regulated and KEAP1 expression was markedly inhibited by HDAC3i in the diabetic mouse. Immunoprecipitation shows that the binding of KEAP1 in the diabetic heart was decreased by the treatment of HDAC3i when we pull down nuclear factor-E2-related factor 2 (NRF2). Meanwhile, the nuclear localization of NRF2 and its downstream anti-oxidative stress genes, NADPH quinone oxidoreductase 1 (NQO1), heme oxygenase-1 (HO-1), were markedly up-regulated in the HDAC3i treated diabetic mouse group. These results suggest that HDAC3i prevents DCM likely via miR-200a-mediated degradation of KEAP1 and consequently activation of the Nrf2-regulated antioxidant pathway.