Dye-coupling in three-dimensional histoculture of rat lingual frenulum

A three-dimensional histoculture of wet stratified squamous epithelium of rat lingual frenulum was cultured on a liquid-air interface. The tissue retained its morphology for many days in culture. During this period the vast majority of the epithelial cells remained viable and exhibited dye (lucifer yellow) coupling in all living epithelial strata. Dye coupling was determined using two methods: the conventional intracellular injection method, and a new method--"cut-loading." In the cut-loading method, an incision is made in the epithelium in the presence of dye, and intracellular diffusion of dye throughout the epithelium was measured using confocal microscopy. The basolateral surface of the lingual frenulum also acted as a substrate for neuroblastoma cells to grow without exogenously added trophic factors. These neuroblastoma cells grow neurites that establish contacts with epithelial cells. This preparation can serve as a model for investigating interactions among epithelial cells and between nerves and epithelial cells.