Differentiation of proton-pumping activity in cultured renal inner medullary collecting duct cells

Abstract

Cultured inner medullary collecting duct (IMCD) cells have been shown to secrete protons (H+) by two mechanisms: an N-ethylmaleimide- and dicyclohexyl-carbodiimide-sensitive electrogenic H(+)-ATPase or H+ pump, and an amiloride-sensitive, secondary active Na+H+ exchanger. These cells also express Cl-/HCO3- exchange and carbonic anhydrase activity in common with other renal epithelial cells involved in acid-base transport. Video fluorescence microscopy of individual cells using 2',7'-biscarboxyethyl-5(6)-carboxyfluorescein has demonstrated that adjacent-cultured IMCD cells show substantial functional intercellular heterogeneity. The development of H(+)-pumping activity is associated with high-baseline intracellular pH and peanut agglutinin (PNA) affinity, and loss of mitotic activity and of Na+/H+ exchange. The H(+)-pumping activity may be further enhanced by removal of fetal calf serum for 6-54 h or by selecting cells with high PNA affinity. IMCD cells in their most differentiated state form domes, which consistently showed the highest rates of H(+)-pumping activity, as well as high affinity for peanut lectin. When IMCD were plated at low density, domes developed relatively late (2-4 weeks), at which time cells located in the center of nests of contiguously growing cells were quiescent and showed H(+)-pumping activity but no Na+/H+ exchange. On the other hand, dense plating was associated with early development of domes (end of 1st week), at which time adjacent cells showed a high mitotic activity and Na+/H+ exchange, but no H(+)-pumping activity. We speculate that differentiation of IMCD cells results in the development of cell polarity.(ABSTRACT TRUNCATED AT 250 WORDS)