In this study, the effects of different concentrations of protamine sulfate in the isolated rat heart papillary muscle’s parameters of contraction and contracture were studied. 20 male Wistar rats weighing between 210-260 g were used in the study. From each left ventricle, a papillary muscle of 5-6 mm in length 1-1.5 mm in width and 32 ± 4 mg in weight was isolated. With isometric transducer, in 30 °C, The parameters of contraction and contracture were measured after 0.1 Hz 0.5 ms of impulse with control (0), 20, 40 and 80 m/ml protamine and an equal proportion of heparin, protamine concentration. The concentration of papillary muscle decreased significantly (p < 0.001) and in 40, 80 mg/ml protamine concentration and half relaxation time were prolonged (p < 0.05). After a dose of 40 mg/ml protamine, the muscle contracted, the peak tension (TPT) decreased. The contraction and relaxation speed (± dF/dt) decreased with dosa. Papillary muscle showed a weak negative inotropic effect in solutions containing equal proportions of heparin and protamine.