Objective: Left-ventricular hypertrophy and interstitial fibrosis are the main pathophysiological factors of heart failure with preserved ejection fraction. Blockade of the serotonin 5-HT 2B receptor (5-HT 2B R) has been shown to reduce cardiac hypertrophy, oxidative stress, and extracellular cell matrix activation. In this study, we evaluated the effects of the 5-HT 2B R blockade, on hemodynamic and cardiac remodeling, in spontaneously hypertensive rats (SHRs) that display a diastolic dysfunction with preserved ejection fraction. Method: Thirty-seven-week-old SHRs were randomized in four groups receiving either saline, the selective 5-HT 2B R antagonist RS-127445 (1 mg/kg per day), a calcium channel blocker nicardipine (6 mg/kg per day), or RS-127445 R nicardipine. During the 14 weeks of treatment period, cardiac function and blood pressure were monitored by echocardiography and tail-cuff. Finally, electrocardiograms and invasive hemodynamics were obtained before blood collection. Heart was analyzed for morphology and mRNA expression. A complementary study evaluated the cardiac and vascular effects of serotonin on wild-type and mice knockout for the 5-HT 2B R (Htr 2B À/À) and/or the 5-HT 2A R (Htr 2A À/À). Results: Despite the left ventricular 5-HT 2B R overexpression, 5-HT 2B R blockade by RS-127445 did not affect left ventricular hypertrophy and fibrosis in SHRs. This antagonist did not improve diastolic dysfunction, neither alone nor in combination with nicardipine, although it induced plasma brain natriuretic peptide decrease. Moreover, RS-127445 amplified subendocardial fibrosis and favored left ventricular dilatation. Finally, a subendocardial left ventricular fibrosis was induced by chronic serotonin in wild-type mice, which was increased in Htr 2B À/À animals, but prevented in Htr 2A À/À and Htr 2A/2B À/À mice, and could be explained by a contribution of the endothelial 5-HT 2B Rs to coronary vasodilatation. Conclusion: This work is the first to identify a cardioprotective function of the 5-HT 2B R in an integrated model of diastolic dysfunction with preserved ejection fraction.