Simple Summary: One of the oldest anticancer drugs in use is doxorubicin. It associates with cardiotoxicity that until now remains unresolved. Diastolic cardiac dysfunction can be the first sign, and develops at the very low doses. We asked whether short-term changes were preceded by even earlier effects. We tested functional and molecular changes in cardiac fibroblasts after very brief exposure to doxorubicin. Cardiac fibroblasts increased energy metabolism and differentiated into myofibroblast, expressing markers of cell activation and phenotype change. These results demonstrate that cardiac fibroblasts are effector cells that swiftly react to systemically administered drug. From the very first contact with doxorubicin, they can actively contribute to the initial phase of pathological myocardial remodeling. Thus, it is conceivable to foresee these cells as a target to counteract the cardiotoxicity of doxorubicin at the very early stage. The application of doxorubicin (DOX) is hampered by cardiotoxicity, with diastolic dysfunction as the earliest manifestation. Fibrosis leads to impaired relaxation, but the mechanisms that operate shortly after DOX exposure are not clear. We asked whether the activation of cardiac fibroblasts (CFs) anticipates myocardial dysfunction and evaluated the effects of DOX on CF metabolism. CFs were isolated from the hearts of rats after the first injection of DOX. In another experiment, CFs were exposed to DOX in vitro. Cell phenotype and metabolism were determined. Early effects of DOX consisted of diastolic dysfunction and unchanged ejection fraction. Markers of pro-fibrotic remodeling and evidence of CF transformation were present immediately after treatment completion. Oxygen consumption rate and extracellular acidification revealed an increased metabolic activity of CFs and a switch to glycolytic energy production. These effects were consistent in CFs isolated from the hearts of DOX-treated animals and in naïve CFs exposed to DOX in vitro. The metabolic switch was paralleled with the phenotype change of CFs that upregulated markers of myofibroblast differentiation and the activation of pro-fibrotic signaling. In conclusion, the metabolic switch and activation of CFs anticipate DOX-induced damage and represent a novel target in the early phase of anthracycline cardiomyopathy. [ABSTRACT FROM AUTHOR]