The permeability of skin and oral mucosa to various compounds has been measured but the actual pathways substances take in traversing the epithelia have not been identified. In this study, radiolabelled cholesterol, ethanol or water were placed on the surface of porcine skin, keratinized gingiva, or nonkeratinized floor of mouth mucosa, and incubated at 37 degrees C for 2 h. The tissue was then snap-frozen, and sectioned in a cryostat, picked up on precoated slides and exposed at -20 degrees C for 40 days for light microscopic autoradiography. Some tissues were freeze-dried and directly embedded in low viscosity resin and prepared for electron microscopic autoradiography. Examination of autoradiographs revealed silver grains over, or adjacent to, intercellular spaces. Counts of the grains over the extra- and intracellular compartments were made in random light and electron microscope fields. For all compounds and tissue regions, there were significantly more (p less than 0.05) grains over the intercellular spaces than over the cells. The results indicate that the intercellular compartment is the predominant route for compounds moving across the superficial barrier layer of epidermis and oral epithelia. The nature of the intercellular material is, thus, a primary determinant of epithelial permeability.