Although the right ventricular (RV) myocardial performance index (MPI) usually is increased in the presence of RV dysfunction and pressure overload, debate continues over the correlation between the RV MPI and functional derangement in patients with RV pressure-overload congenital heart disease (CHD). To address this controversy, this study took serial measurements of the RV MPI in addition to invasive RV hemodynamic measurements during the acute stage of mild to severe pressure overload. Right ventricle pressure overload was induced by partial pulmonary arterial banding (PAB) in 3-week-old rats. The rats were divided into two groups: mild pulmonary stenosis (PS) group (20-40 % stenosis; n = 20) and severe PS group (40-70 % stenosis; n = 28). Sham-treated animals (sham group; n = 30) underwent the same surgical procedure without PAB. Pressure-overload RV hypertrophy was documented by weighing the heart, by evaluating echocardiograms, and by evaluating cardiac hypertrophy-associated gene expression. The RV MPI was checked 1, 2, 3, 5, and 8 weeks after PAB. The MPI was calculated as the sum of the isovolumic contraction time and the isovolumic relaxation time (IRT) divided by the ejection time. The RV MPI of the mild PS group did not differ significantly from that of the sham group. The RV MPI of the severe PS group, however, was lower than that of the sham group (0.27 ± 0.01 vs 0.29 ± 0.01) 2 to 8 weeks after PAB: 0.19 ± 0.01 at 2 weeks (P < 0.001), 0.16 ± 0.01 at 3 weeks (P < 0.001), 0.20 ± 0.01 at 5 weeks (P = 0.021), and 0.18 ± 0.01 at 8 weeks (P < 0.001) after PAB. The decreased RV MPI was associated with decreased IRT and increased ejection time. RV hypertrophy contributes to the decrease in the RV MPI in the severe pressure-overload condition.