The effect of vasoactive intestinal polypeptide (VIP) and acetylcholine on secretion of epidermal growth factor (EGF) from the rat salivary glands was investigated. VIP in doses of 3 X 10(-10) to 3 X 10(-8) mol/kg per h stimulated secretion of saliva and total output of EGF dose-dependently. Acetylcholine also stimulated salivation and output of EGF. VIP in a dose of 3 X 10(-11) to 3 X 10(-10) mol/kg per h enhanced the stimulatory effect of acetylcholine, but this effect disappeared when the dose of VIP was increased. Adrenalectomy decreased acetylcholine stimulated total output of EGF by approximately 50%, but only by 20% when acetylcholine plus VIP was administered. EGF was localized to the convoluted granular tubules in the submandibular gland, whereas EGF could not be detected in the remaining salivary glands. The results suggest that VIP and acetylcholine cooperate in the control of exocrine secretion from the rat salivary glands. The effect of acetylcholine, however, seems to be partly dependent on circulating catecholamines.