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Anterior pituitary glands from female rats were dispersed enzymically in the absence of dopamine. Dispersed cells (10⁶–10⁷) were layered onto columns containing Bio-Gel P-2 and were then perifused for 3 h with Dulbecco's Modified Eagle's Medium. The prolactin content of the perifusate and cell homogenates was determined by radioimmunoassay. Prolactin secretion during the third hour of perifusion increased as the loading of cells increased. However, the increase was not linear, and when secretion rate per 10⁶ cells was calculated it was found that increased loading decreased the rate, which fell to a plateau of 1·3 ± 0·1 (S.E.M.) ng/min per 10⁶ cells at a loading of about 8 x 10⁶ cells from 3·8 ± 0·1 ng/min per 10⁶ cells for a loading of 10⁶ cells. This cell-density dependence of the rate of prolactin secretion in the perifusion system may be due to intercellular contact since the isolation of the tissue removes the influence of hypothalamic factors, while localized build up of prolactin (potentially causing direct autoregulation on the lactotroph) seems unlikely because of the continuous flow of medium.