Using immunohistochemistry we have investigated the presence and cellular distribution of the 31-kDa subunit of vacuolar-type H+-ATPase (V-ATPase) in secretory endpieces and the duct system of rat major salivary glands. In all three salivary glands studied the 31-kDa subunit of V-ATPase was not expressed in secretory endpieces. In rat parotid gland V-ATPase was luminally located in main excretory and striated duct cells. In contrast, both rat submandibular and sublingual glands showed a diffuse intracellular V-ATPase distribution. The differences in V-ATPase immunolocalization in rat salivary glands probably reflect the structural heterogeneity of the different glands. The data also suggest that the duct systems of major salivary glands may modify the H+ and HCO3- concentration of the final saliva in different ways.