Retinal dehydrogenase type 1 (RALDH1) is involved in the biosynthesis of retinoic acid (RA), a modulator of gene expression and cell differentiation. RALDH1 mRNA transcripts are present in the stomach and small intestine, and their expression is regulated by vitamin A status. In situ hybridization demonstrated RALDH1 mRNA expression in epithelial cells of the stomach, small intestine, and large intestine. Strong hybridization was also seen in the lamina propria of small intestinal mucosa and the smooth muscle layer of the small and large intestines. Immunocytochemical localization revealed RALDH1 staining in parietal cells of the stomach and prismatic cells of the small and large intestines. The presence of RALDH1 protein was also detectable within supportive glial cells around neuronal fibers throughout the muscular layers of the stomach as well as the small and large intestines. These data suggest an important role for RALDH1 in generating RA needed for the differentiation of specific epithelial cells in the stomach and intestines.