Differentiated function in cultured epithelia derived from thick ascending limbs

Abstract

Medullary thick ascending limbs were dissected from rabbit kidney and cultured on thin collagen membranes. The collagen membranes were supported by a special device that limited the exposed surface area to a disk 1 mm in diameter. The cells usually proliferated to form a confluent epithelial monolayer covering the collagen membrane within 8-10 days. Transepithelial voltage was measured between bridges immersed in the solutions bathing the bottom of the collagen membrane and the apical (upper) surface of the epithelium. The voltage typically was oriented positive toward the apical surface and persisted for a few weeks. Furosemide but not amiloride reduced the voltage reversibly. Both the orientation of the voltage and its reduction by furosemide are characteristic of thick ascending limbs. Therefore, the observed voltage supports the identification of the epithelial cells in culture and is evidence of differentiated function.