Polarized expression of Na+/H+ exchange activities in clonal LLC-PK1 cells (Clone4 and PKE20) I. Basic characterization.
We have analysed the mechanisms of Na(+)-dependent pHi 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 Km 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').(ABSTRACT TRUNCATED AT 250 WORDS)
Casavola
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Department of Physiology, University of Züich, Switzerland.
Helmle-Kolb
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Montrose
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Murer
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H
D000584:Amiloride / Q000031:analogs & derivatives / Q000494:pharmacology; D000643:Ammonium Chloride / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002999:Clone Cells; D005452:Fluoresceins; D005456:Fluorescent Dyes; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D012964:Sodium / Q000494:pharmacology; D017923:Sodium-Hydrogen Exchangers; D013050:Spectrometry, Fluorescence
D002352:Carrier Proteins; D005452:Fluoresceins; D005456:Fluorescent Dyes; D017923:Sodium-Hydrogen Exchangers; D000643:Ammonium Chloride; D000584:Amiloride; C043829:2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein; D012964:Sodium; C039614:ethylisopropylamiloride