Plasma prolactin (Prl) titers were determined by radioimmunoassay in conscious, freely moving male rats which had either had sham operations or had been pineal-ectomized. Values were determined during the day and during the night in animals on a reversed light cycle. During the day plasma Prl levels were low in both groups with small bursts and a tendency for greater elevation toward the end of the collection period at 17.00 and 18.00 h. There were only 2 significant effects of pinealectomy on these daytime values, one of which was a reduction in the elevation of Prl at 17.30 h. There were also more frequent very low values, less than 5 ng/ml, after pinealectomy. At night there was greater variation of plasma Prl in sham-operated rats and in general the animals showed a sudden elevation just prior to the time the lights were turned on. The values remained elevated for some time thereafter. After pinealectomy the elevation when the lights were turned on was slightly delayed and the responses were smaller in magnitude or even absent. There were significantly more high Prl values in the controls than in the pinealectomized animals in darkness and the area under the curve of Prl release, which was greater at night than in the daytime, was significantly lowered by pinealectomy.