POLARIZED EXPRESSION OF NA+/H+ EXCHANGE ACTIVITIES IN CLONAL LLC-PK1 CELLS (CLONE4 AND PKE20) .1. BASIC CHARACTERIZATION

We have analysed the mechanisms of Na+-dependent pH(i) recovery from an acid load in LLC-PK1/Clone4 and LLC-PK1/PKE20 cells by using the intracellular pH indicator 2',7'-bis(carboxyethyl)-5,6-carboxyfluorescein acetoxymethyl ester. By analysis using single-cell microspectrofluorometry, we obtained evidence for polarized expression of Na+/H+ exchange activities with different properties in apical and basolateral cell surfaces, respectively. In Clone4 cells, Na+/H+ exchange activity is only visible on the basolateral cell surface; in PKE20 cells, Na+/H+ exchange activities with equal capacities are present on both cell surfaces. In Clone4 cells, the apparent K(m) value for Na+ is around 10 mM; in PKE20 cells it is around 20 mM and indistinguishable for the two cell poles. Ethylisopropylamiloride (EIPA) inhibition for all three activities measured in monolayer configuration is reduced by increasing Na+ concentration. Measured in the same cells, EIPA inhibition of transport of PKE20 cells is weaker for apical Na+/H+ exchange as compared to basolateral activity. In Clone4 and PKE20 cells kept in suspension, Na+/H+ exchange activities with similar properties for the two cell lines are observed. However, Na+/H+ exchange activities in cells in suspension are different from either activity measured in monolayer configuration: affinity for Na+ is higher (PKE20 cells) and inhibition by amiloride is weak and not influenced by increasing Na+ concentrations (PKE20 and Clone4 cells). It is concluded that PKE20 cells contain different Na+/H+ exchange activities on the two cell surfaces; this cell line should be a useful model to study regulatory aspect of different Na+/H+ exchange functions ("epithelial"/"housekeeping"). Furthermore, measurements on cells in suspension do not reflect the properties of Na+/H+ exchange activities of cells in epithelial configuration.