Nerves containing vasoactive intestinal peptide (VIP) were observed in salivary glands of rat, cat and man. VIP nerves were numerous in the cat while they were moderate in number in rat and man. The measured concentrations of immunoassayable VIP were in agreement with the immunohistochemical findings. Electrical stimulation of the feline chorda lingual nerve, which stimulates salivary secretion and local blood flow, resulted in a marked elevation of VIP in the venous effluent from the submandibular gland. VIP was not measurable in saliva. Gel permeation chromatography of extracts from cat submandibular gland and from venous plasma collected before and during nervous stimulation revealed one immunoreactive peak with an elution position identical to that of highly purified porcine VIP. The finding of neuronal VIP in salivary glands, its release upon nerve stimulation and its known effect on local blood flow support the view that VIP is a neurotransmitter in the salivary glands.