Polarized Expression of Na+/H+ Exchange Activities in Clonal LLC-PK1 Cells (Clone4 and PKE20) I. Basic Characterization

Overview

Journal Pflugers Arch

Specialty Physiology

Date 1991 Apr 1

PMID 1649993

Citations 9

Authors

V Casavola

M H Montrose

H Murer

Affiliations

Soon will be listed here.

Abstract

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)

Citing Articles

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Na/H exchange activities in NHE1-transfected OK-cells: cell polarity and regulation.

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Identification of PTH-responsive Na/H-exchanger isoforms in a rabbit proximal tubule cell line (RKPC-2).

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References

Aronson P . Mechanisms of active H+ secretion in the proximal tubule. Am J Physiol. 1983; 245(6):F647-59. DOI: 10.1152/ajprenal.1983.245.6.F647. View

Grinstein S, Rothstein A . Mechanisms of regulation of the Na+/H+ exchanger. J Membr Biol. 1986; 90(1):1-12. DOI: 10.1007/BF01869680. View

Chan Y, Biagi B, Giebisch G . Control mechanisms of bicarbonate transport across the rat proximal convoluted tubule. Am J Physiol. 1982; 242(5):F532-43. DOI: 10.1152/ajprenal.1982.242.5.F532. View

Kurtz I . Basolateral membrane Na+/H+ antiport, Na+/base cotransport, and Na+-independent Cl-/base exchange in the rabbit S3 proximal tubule. J Clin Invest. 1989; 83(2):616-22. PMC: 303722. DOI: 10.1172/JCI113925. View

Murer H, Hopfer U, KINNE R . Sodium/proton antiport in brush-border-membrane vesicles isolated from rat small intestine and kidney. Biochem J. 1976; 154(3):597-604. PMC: 1172760. View

Boron W, Boulpaep E . Intracellular pH regulation in the renal proximal tubule of the salamander. Na-H exchange. J Gen Physiol. 1983; 81(1):29-52. PMC: 2215563. DOI: 10.1085/jgp.81.1.29. View

Murer H, Burckhardt G . Membrane transport of anions across epithelia of mammalian small intestine and kidney proximal tubule. Rev Physiol Biochem Pharmacol. 1983; 96:1-51. DOI: 10.1007/BFb0031006. View

VIGNE P, Frelin C, Cragoe Jr E, Lazdunski M . Ethylisopropyl-amiloride: a new and highly potent derivative of amiloride for the inhibition of the Na+/H+ exchange system in various cell types. Biochem Biophys Res Commun. 1983; 116(1):86-90. DOI: 10.1016/0006-291x(83)90384-4. View

Cantiello H, Scott J, Rabito C . Polarized distribution of the Na+/H+ exchange system in a renal cell line (LLC-PK1) with characteristics of proximal tubular cells. J Biol Chem. 1986; 261(7):3252-8. View

10.

Haggerty J, Agarwal N, Reilly R, Adelberg E, SLAYMAN C . Pharmacologically different Na/H antiporters on the apical and basolateral surfaces of cultured porcine kidney cells (LLC-PK1). Proc Natl Acad Sci U S A. 1988; 85(18):6797-801. PMC: 282065. DOI: 10.1073/pnas.85.18.6797. View

11.

Montrose M, Murer H . Regulation of intracellular pH by cultured opossum kidney cells. Am J Physiol. 1990; 259(1 Pt 1):C110-20. DOI: 10.1152/ajpcell.1990.259.1.C110. View

12.

Roos A, Boron W . Intracellular pH. Physiol Rev. 1981; 61(2):296-434. DOI: 10.1152/physrev.1981.61.2.296. View

13.

Gstraunthaler G . Epithelial cells in tissue culture. Ren Physiol Biochem. 1988; 11(1-2):1-42. DOI: 10.1159/000173147. View

14.

Biber J, Malmstrom K, Reshkin S, Murer H . Phosphate transport in established renal epithelial cell lines. Methods Enzymol. 1990; 191:494-505. DOI: 10.1016/0076-6879(90)91032-2. View

15.

Barros F, Dominguez P, Velasco G, Lazo P . Na+/H+ exchange is present in basolateral membranes from rabbit small intestine. Biochem Biophys Res Commun. 1986; 134(2):827-34. DOI: 10.1016/s0006-291x(86)80495-8. View

16.

Montrose M, Friedrich T, Murer H . Measurements of intracellular pH in single LLC-PK1 cells: recovery from an acid load via basolateral Na+/H+ exchange. J Membr Biol. 1987; 97(1):63-78. DOI: 10.1007/BF01869615. View

17.

Alpern R . Cell mechanisms of proximal tubule acidification. Physiol Rev. 1990; 70(1):79-114. DOI: 10.1152/physrev.1990.70.1.79. View

18.

Frelin C, VIGNE P, Ladoux A, Lazdunski M . The regulation of the intracellular pH in cells from vertebrates. Eur J Biochem. 1988; 174(1):3-14. DOI: 10.1111/j.1432-1033.1988.tb14055.x. View

19.

Montrose M, Murer H . Polarity and kinetics of Na(+)-H+ exchange in cultured opossum kidney cells. Am J Physiol. 1990; 259(1 Pt 1):C121-33. DOI: 10.1152/ajpcell.1990.259.1.C121. View

20.

Alpern R, Chambers M . Cell pH in the rat proximal convoluted tubule. Regulation by luminal and peritubular pH and sodium concentration. J Clin Invest. 1986; 78(2):502-10. PMC: 423587. DOI: 10.1172/JCI112602. View