The biosynthesis of polyamines, a group of growth-related amines, varied in the rat pituitary gland with the different stages of the estrous cycle. The activity of ornithine decarboxylase (ODC), which catalyzes the rate-limiting step in the biosynthesis of polyamines, was greatly increased in the pituitary gland during the evening of proestrus. Ovariectomy resulted in a disappearance of this cyclic change in polyamine synthesis, whereas estrogen treatment of the ovariectomized rats gave rise to daily afternoon peaks in pituitary ODC activity. Like the prolactin cells, pituitary polyamine biosynthesis appeared to be regulated by dopamine. The induction of ODC activity in the pituitary gland during proestrus or after estrogen treatment was almost completely repressed by bromocriptine, a dopaminergic compound. The dopamine antagonist haloperidol, on the other hand, was a potent inducer of pituitary ODC activity. Neither pituitary DNA synthesis nor prolactin secretion, which both are induced by estrogen, were affected by inhibition of polyamine synthesis, suggesting a biological function of the pituitary polyamines unrelated to these events.