Rats received a daily injection of 2, 6, or 12 mg kg-1 bethanechol for 7 or 14 days. Pancreatic weight and content of protein, amylase, and chymotrypsinogen were increased in a dose and time dependent fashion by bethanechol. After 14 days of treatment with the highest dose, pancreatic weight and total pancreatic DNA content increased to 1.18 times control (P less than 0.05) while protein increased to 1.25, amylase to 1.85, and chymotrypsinogen to 1.72 times control. There was a slight increase in oxyntic gland area weight in bethanechol-treated rats, but content of DNA, protein, and pepsinogen were not changed. Bethanechol had no effect on duodenal weight or content of DNA, protein, or maltase. Acute administration of 12 mg kg-1 bethanechol increased circulating gastrin levels for at least 4 h in unanesthetized rats. We conclude that long-term treatment with a cholinergic agonist produces pancreatic hyperplasia and increases protein and enzyme content. Under the conditions of this study, no effects were seen on oxyntic gland area of stomach or duodenum.