Although ketamine increases pulmonary vascular resistance of patients, an occasional decrease of resistance in animals and humans has been reported. In addition, ketamine has a direct relaxant effect on isolated smooth muscle. The effects of ketamine on the main pulmonary artery rings isolated from the guinea pig were studied to elucidate the underlying mechanism of the reported relaxant effect of this anesthetic on smooth muscle. Ketamine (10–250 μg/mL) caused a concentration-dependent shift to the right of CaCl2 concentration-effect curves on artery rings, suggesting an interference with Ca2+ metabolism. In Ca2+-free buffer, ketamine (10–250 μg/mL) did not affect the magnitude of epinephrine-induced contractions but inhibited dose-dependent BaCl2-induced contractions. These observations suggest that ketamine inhibits transmembrane Ca2+ influx but does not affect its release from intracellular stores or its binding to intracellular receptor sites on the contractile system. Ketamine (25–500 μg/mL) also caused equipotent concentration-dependent relaxation of epinephrine-induced contractions in the absence and the presence of monensin, a Na+-ionophore that dissipates the Na+ gradient across the cell membrane, and in Na+-free, sucrose-substituted buffer. Ketamine (25–500 μg/mL) also relaxed ouabain-induced contractions to the baseline, an effect that was significantly attenuated in the presence of ruthenium red, a Ca2+ adenosine triphosphatase (ATPase) inhibitor. The relaxant effect of ketamine (250–750 μg/mL) of epinephrine-induced contraction also was attenuated in the presence of 0.1 mM lanthanum chloride (La3+), an inhibitor of adenosine 5′-triphosphate (ATP)-dependent Ca2+ extrusion, and completely inhibited in the presence of 10 mM La3+. These data suggest that ketamine-induced relaxation is attributed mainly to the stimulation of Ca2+-ATPase and that Na+-dependent, Ca2+ efflux participates only minimally to this relaxation. We, therefore, conclude that ketamine has two effects on Ca2+ transport in isolated pulmonary artery smooth muscle: 1) it inhibits Ca2+ influx, and 2) it stimulates Ca2+-ATPase to cause efflux.