SUMMARY: Ovine prolactin injected subcutaneously for 2 days stimulated an increase in fluid uptake in vitro by everted gut sacs from male Sprague—Dawley rats. Four intestinal segments from each rat were incubated, but only two of the four segments tested showed a prolactin-stimulated increase in fluid transfer in intact rats. In adrenalectomized—nephrectomized rats all four sacs showed an increased fluid transfer after prolactin treatment. Sacs incubated for 1 h in glucose—Ringer solution containing 0·1 mg ovine prolactin/ml did not show an increased water transfer as compared with controls incubated in medium containing albumin. Dietary changes seemed to alter the sensitivity of the gut to prolactin.