Agents influencing the output of prostaglandin F 2α (PGF 2α) from non-pregnant endometrium were investigated by in vitro incubation, for 5 and 19h, using mid-cycle (day 7), end of cycle (day 15), or ovariectomised guinea-pigs.Estradiol 17-β (l0μq/ml) stimulated PGF 2∝ output 24h after incubation with endometrium (p<0.05). This stimulation was greater at mid-cycle.Progesterone(50ng/ml) inhibited output of PGF 2α (p<0.05) in mid-cycle, end of cycle and ovariectomised guinea-pig cultures.Oxytocin (l × 10 −5 u/ml) stimulated theoutput of PGF 2α at the end of the cycle, but not at mid-cycle. However, in the presence of estradiol 17-β (lOμg/ml), oxytocin stimulation of mid-cycle PGF 2α output was observed. The calcium ionophore A23187 (5μg/ml) stimulated PGF 2α synthesis from mid-cycle and end-of-cycle endometrium, and this stimulation resembled that caused by arachidonic acid (lOOμg/ml), suggesting an action via substrate mobilisation. Co-culture of endometrium and myometrium did not influence endometrial PGF 2α or myometrial 6-oxo-PGF 2 output. It is suggested that the steroid hormones act as coarse modulators of endometrial PGF 2α output, but more rapid changes may be achieved by oxytocin and agents that mobilise substrate supply, possibly via calcium ion fluctuations.