45Ca was used to investigate the effect of oxytocin on calcium movements in uterine smooth muscle from estrogenized rats. Upon exposure to oxytocin the influx of 45Ca in myometrial strips increased rapidly. After 2 min of incubation with oxytocin (1 microM) there was a 30% increase over the control. Prolonged stimulation with oxytocin caused no further accumulation of 45Ca. The amount of Ca gained following oxytocin stimulation could be removed by washing with a lanthanum solution. The rate of 45Ca efflux from preloaded tissues was substantially increased by oxytocin. In the absence of external Ca, the stimulation of 45Ca efflux by oxytocin was much reduced. The data indicate that oxytocin-induced contraction of the myometrium is largely dependent on the influx of extracellular calcium. Oxytocin appears to be capable of releasing small amounts of calcium from intracellular storage sites. The data also provide evidence against the inhibition by oxytocin of the calcium extrusion pump.