Objective: Heart failure is a clinical syndrome characterized by cardiac dysfunction due to structural abnormalities of the myocardium that results in the inability of the heart to eject sufficient volume of the blood to the circulation. HF is a major cause of morbidity and mortality in the world. The ECM is a highly dynamic structure controlling the proper functioning of heart muscle. ECM remodelling with enhanced deposition of collagen due to hemodynamic overload impairs cardiomyocyte adhesion and electrical coupling that contributes to cardiac mechanical dysfunction and arrhythmias. Design and method: We aimed to explore ECM and connexin-43 (Cx43) signalling pathways in hemodynamically overloaded rat heart as well as the possible implication of Ang(1-7) to prevent/attenuate adverse myocardial remodelling. Males, 8 weeks-old, normotensive Hannover Spraque-Dawley rats (HSD), hypertensive Ren-2 transgenic rats (TGR) and Ang(1-7)-transgenic rats (TGR(A1-7)) underwent aortocaval fistula (ACF) to produce volume overload. 5 weeks later, were right and left ventricular tissue samples used for biometric, biochemical, and proteomic analyzes. Results: Cardiac hypertrophy in response to volume overload was significantly less pronounced in TGR(A1-7) comparing to HSD rats. Moreover, a marker of fibrosis, hydroxyproline, was increased in both ventricles of volume overloaded TGR while reduced in Ang(1-7) right heart ventricle. Protein level and activity of MMP-2 were reduced in both ventricles of volume overloaded TGR/TGR(A1-7) compared to HSD. SMAD2/3 protein levels were decreased in the right ventricle of TGR(A1-7) compared to HSD/TGR in response to volume overload. In parallel, Cx43 and pCx43 implicated in electrical coupling were increased in TGR(A1-7) versus HSD/TGR. PKC↑ and PKC↗ showed a decrease in the TGR group compared to HSD and a decrease in the TGR(A1-7) ACF group compared to HSD and TGR after ACF in both chambers. Conclusions: It can be concluded that Ang(1-7) exhibits cardioprotective and anti-fibrotic potential in condition of cardiac volume overload. If we look at the results comprehensively, it can be concluded that Ang(1-7) exhibits cardioprotective and antifibrotic potential in condition of cardiac volume overload. Supported by grants VEGA 2/0006/23, APVV 21-0410.