Colon tumors expressing high levels of beta-catenin and c-myc have been reported in male F344 rats given three short cycles of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) alternating with a high-fat (HF) diet. Using the same experimental protocol, rats were euthanized 24 h after the last dose of PhIP so as to examine early changes in colonic crypt homeostasis and beta-catenin expression, before the onset of frank tumors. PhIP/HF dosing caused a significant increase in the bromodeoxyuridine labeling index throughout the entire colon, and within the colonic crypt column cleaved caspase-3 was elevated in the basal and central zones, but reduced in the luminal region. In vehicle/HF controls, beta-catenin was immunolocalized primarily at the border between cells at the top of the crypt, whereas in rats given PhIP/HF diet there was strong cytoplasmic staining, which appeared as a gradient of increased beta-catenin extending from the base of the crypt column to the luminal region. Quantitative real-time PCR and immunoblot analyses confirmed that beta-catenin and c-myc were increased significantly in the colonic mucosa of rats given PhIP/HF diet. Collectively, these findings suggest that PhIP/HF cycling alters beta-catenin and c-myc expression in the colonic mucosa, resulting in expansion of the proliferative zone and redistribution of apoptotic cells from the lumen to the central and basal regions of the colonic crypt. Thus, during the early stages of colon carcinogenesis, alternating exposure to heterocyclic amines and a high-fat diet might facilitate molecular changes resulting in dysregulated beta-catenin and c-myc expression.