Exogenously administered cholecystokinin is a potent stimulant of pancreatic exocrine secretion and pancreatic polypeptide release. Release of cholecystokinin by amino acids and fats is strongly correlated with both pancreatic exocrine secretion and pancreatic polypeptide release. Despite this correlation, direct evidence that cholecystokinin is a physiologic mediator of these actions is not available. We have studied this problem in fasted dogs with chronic pancreatic fistulas by means of a specific cholecystokinin antagonist, proglumide, to inhibit the effect of cholecystokinin. Secretion, neurotensin (with secretin stimulation infusion), or cholecystokinin-octapeptide was infused intravenously, either with saline solution or with proglumide (300 mg/kg/hr). For endogenous release of cholecystokinin, intraduodenal infusions of phenylalanine and tryptophan or of sodium oleate were given with either intravenous saline solution or intravenous proglumide. Pancreatic secretion and release of cholecystokinin and pancreatic polypeptide were measured in plasma. Cholecystokinin-octapeptide stimulated pancreatic secretion of water and protein; both of these were significantly inhibited by proglumide. Intraduodenal amino acids and sodium oleate both caused significant release of cholecystokinin, which was not altered by proglumide; however, proglumide inhibited pancreatic secretion stimulated by intraduodenal amino acids and sodium oleate. Release of pancreatic polypeptide stimulated by amino acid and sodium oleate was also significantly inhibited by proglumide. Since proglumide appears to block actions of cholecystokinin, our results show that cholecystokinin is physiologically important for pancreatic secretion and for release of pancreatic polypeptide.