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Application of ionizing radiation to adult rat major salivary glands tested tenets of the bicellular reserve cell hypothesis for the induction of salivary gland tumors, namely, that stem cells preferentially located to luminal cells of the intercalated duct and basal cells of the excretory duct in normal salivary glands. The effect of a single, low dose (3000 cGy) of x-radiation administered to the parotid and submandibular glands was followed with the use of immunocytochemistry and an antibody to the cell cycle-related protein proliferating cell nuclear antigen to detect the kinetics and localization of cycling cells up to 15 days postirradiation. Maximal responses occurred in acinar cells (12.6-fold increase) of submandibular glands on day 7 postirradiation. Similar but less dramatic concurrent increases in proliferating cells were evident in intercalated (3.4-fold) and striated (2.2-fold) duct cells, but little response was seen in basal or luminal cells of submandibular gland excretory ducts. A limited but maximal proliferative response again occurred on day 7 in the parotid gland. Neither in the steady state nor irradiated submandibular gland was there evidence of specific stem ("reserve") cells associated with the intercalated or excretory ducts. It appears unnecessary to invoke stem cells in a model of cellular proliferation in salivary glands. Therefore current concepts of salivary gland tumorigenesis require modification because all cell types, including acinar cells, are at risk in the carcinogenic process.