Morphological and functional characteristics of cells cultured from the endolymphatic sac

The endolymphatic sac is a part of the homeostasis-regulating system of the membranous labyrinth of the inner ear. Disturbances in the function of the endolymphatic sac are believed to be involved in the genesis of different inner ear disorders, such as endolymphatic hydrops and Ménière's disease. To make studies of the ion- and fluid-regulating mechanisms of the sac possible, a method to culture the tissue in vitro was developed. Epithelial cells and fibroblasts were morphologically characterised in the cell cultures with light and electron microscopy as well as immunohistochemically using antibodies against cytokeratin and vimentin. Since mesenchymal cells have been shown to express vimentin and epithelial cells cytokeratin, the antibodies against these two intermediate filament proteins were used to further confirm the morphological identification. In addition, some functional characteristics of the cultured cells from the endolymphatic sac were studied. ATP and K(+) were added to the cell cultures and changes in cytoplasmic free Ca(2+) concentration ([Ca(2+)](i)) were determined with the fura-2 method. A rapid and transient increase in [Ca(2+)](i) could be seen in both epithelial cells and fibroblasts after applying ATP (200 microM) extracellularly. However, when K(+) was added in concentrations of 50 mM and 100 mM, no changes in [Ca(2+)](i) could be seen in either the epithelial cells or the fibroblasts. The results show that the cultured endolymphatic sac cells preserve their morphological characteristics and maintain a high viability. Accordingly, this method provides a tool for further studies of ion transport mechanisms and fluid homeostasis in the endolymphatic sac.