Monolayer cultures of epithelial cells grown from explants of fetal rat small intestine can differentiate into columnar as well as goblet cells in tissue culture (Kondo et al., Exp cell res 153 (1984) 12) . In this study we have cultured tissue from various levels of the fetal rat gastrointestinal tract in order to characterize those cell types that can be cultured from these tissues and to determine the growth potential of these cells using this culture system. Explants of esophagus and forestomach tissue yielded monolayer outgrowths of squamous epithelial cells which grew as closely apposed polygonal cells capable of developing cornified envelopes. Explants of the glandular portion of the stomach yielded outgrowths of densely packed cells which were more pleiomorphic and which did not desquamate or form cornified envelopes. Explants of small intestine and colon yielded outgrowths of epithelial cells, some of which differentiated into goblet cells, while others developed a 'columnar' cell morphology. The following differences between squamous- and non-squamous cultures were observed.1.Squamous epithelium showed self-sustained growth, while growth of non-squamous epithelial cells became self-limiting.2.Cytochalasin B (CB) (5 μg/ml for 1 h) induced contraction of the whole cell sheet of non-squamous cells, but of only individual squamous cells.3.Squamous cell outgrowth was observed from tissues derived from fetuses of all ages (13-day fetuses through to 3-week-old rats); whereas non-squamous epithelial outgrowth was poor when rats older than the 21st gestational day were studied. These results indicate that cultures established in this manner developed the general characteristics of the cells of the organ from which they were derived, even when undifferentiated fetal tissue was used. The growth conditions needed for columnar epithelium are more stringent than those needed for squamous tissues. This technique opens the way for further characterization of growth requirements of gastrointestinal columnar epithelium.