The effect of the rate of intravenous infusion of protamihe on the acute hemodynamic and pulmonary effects of heparin neutralization was investigated in six adult sheep surgically instrumented for chronic studies. Bovine lung heparin at a dose of 200 IU/kg was injected intravenously over 10 sec, 5 min before the start of protamine administration. On separate experimental days, each sheep received protamine at the same dose of 2 mg/kg, but it was infused over four different time periods: 3 s, 30 s, 300 s, or 30 min. At an additional session, protamine was administered over 3 s without prior heparinization to assess the effect of protamine alone. The sequence of the sessions was randomized and performed blindly. Injecting protamine in unheparinized sheep produced no change in any of the measured variables. In contrast, when protamine was injected over 3 s in heparinized sheep, it induced a transient and significant (P< 0.001) pulmonary hypertension (from 17.2 ± 1.5 to 45.6 ± 2.4 mmHg at 1 min) with an increased pulmonary (five-fold) and systemic (2.5-fold) vascular resistance; a decrease of cardiac output (from 3.85 ± 0.43 to 1.93 ± 0.291/min) without change in left atrial pressure (from 5.3 ± 1.3 to 6.0 ± 1.7 mmHg;P= NS); a significant (P< 0.001) increase of plasma thromboxane B2(TxB2) concentrations (from 349 ± 131 to 974 ± 218 pg/ml); leukopenia (76 ± 4% of baseline white blood cell counts); and hypoxemia (Pao2decreased from 81 ± 3 to 63 ± 4 mmHg at 2 min). Administering the same amount of protamine after heparin at a slower infusion rate significantly attenuated and delayed all components of the adverse response to protamine. This attenuation occurred in an infusion rate-dependent fashion, so that when protamine was infused over 30 min, no significant changes in any of the measured variables were noted. The time course of plasma heparin concentrations following protamine indicated that chemical heparin was completely neutralized over the time period of protamine infusion. These results demonstrate that therateof generation of heparin–protamine complexes (as detected by changes of plasma concentrations of chemical heparin) during iv protamine infusion started 5 min after heparin administration is a factor involved in the generation of sufficient mediators required to initiate a characteristic physiologic response in sheep, including systemic and pulmonary vasoconstriction, TxB2generation, and leukopenia. Infusing a neutralizing dose of protamine over 30 min avoids these adverse reactions in sheep.