Advertisement

The Journal of Membrane Biology

, Volume 121, Issue 1, pp 79–90 | Cite as

Na+ channel activity in cultured renal (A6) epithelium: Regulation by solution osmolarity

  • N. K. Wills
  • L. P. Millinoff
  • W. E. Crowe
Articles

Summary

Solution osmolarity is known to affect Na+ transport rates across tight epithelia but this variable has been relatively ignored in studies of cultured renal epithelia. Using electrophysiological methods to study A6 epithelial monolayers, we observed a marked effect of solution tonicity on amiloride-sensitive Na+ currents (Isc).Isc for tissues bathed in symmetrical hyposmotic (170 mOsm), isosmotic (200 mOsm), and hyperosmotic (230 or 290 mOsm) NaCl Ringer's solutions averaged 25±2, 9±2, 3±0.4, and 0.6±0.5 μA/cm2, respectively. Similar results were obtained following changes in the serosal tonicity; mucosal changes did not significantly affectIsc. The changes inIsc were slow and reached steady-state within 30 min. Current fluctuation analysis measurements indicated that single-channel currents and Na+ channel blocker kinetics were similar for isosmotic and hyposmotic conditions. However, the number of conducting Na+ channels was approximately threefold higher for tissues bathed in hyposmotic solutions. No channel activity was detected during hyperosmotic conditions. The results suggest that Na+ channels in A6 epithelia are highly sensitive to relatively small changes in serosal solution tonicity. Consequently, osmotic effects may partly account for the large variability in Na+ transport rates for A6 epithelia reported in the literature.

Key Words

A6 epithelium Na+ channel current fluctuation analysis CDPC amiloride osmolarity 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Cantiello, H.F., Patenause, C.R., Ausiello, D.A. 1989. G protein subunit, αi-3 activates a pertussis toxin-sensitive Na+ channel from the epithelial cell line, A6}.J. Biol. Chem. 264:20867–20870PubMedGoogle Scholar
  2. Crowe, W.C., Wills, N.K. 1990. Simultaneous monitoring of cell height and transepithelial Na+ transport using fluorescence microscopy.J. Am. Soc. Nephrol. 1:714Google Scholar
  3. Davis, C.W., Finn, A.L. 1987. Interactions of sodium transport, cell volume, and calcium in frog urinary bladder.J. Gen. Physiol. 89:687–702PubMedGoogle Scholar
  4. Diamond, J.M. 1982. Transcellular cross-talk between epithelial membranes.Nature 300:683–685PubMedGoogle Scholar
  5. Eaton, D.C., Ling, B.N. 1989. The effect of cholera toxin (CTX), pertussis toxin (PTX) and GTPγS on highly selective Na+ channels in A6 cells.Proc. 22nd Annual Meeting American Society of Nephrologists. (Washington, D.C.) p. 25AGoogle Scholar
  6. Fidelman, M.L., May, J.M., Biber, T.U.L., Watlington, C.O. 1982. Insulin stimulation of Na+ transport and glucose metabolism in cultured kidney cells.Am. J. Physiol. 242:C121-C123PubMedGoogle Scholar
  7. Garty, H., Asher, C. 1985. Ca++-dependent temperature-sensitive regulation of Na+ channels in tight epithelia.J. Biol. Chem. 260:8330–8335PubMedGoogle Scholar
  8. Garty, H., Asher, C., Yeger, O. 1987. Direct inhibition of epithelial Na+ channels by a pH-dependent interaction with calcium, and by other divalent ions.J. Membrane Biol. 95:151–162Google Scholar
  9. Garty, H., Yeger, O., Yanovsky, A., Asher, C. 1989. Guanosine nucleotide-dependent activation of the amiloride-blockable Na+ channel.Am. J. Physiol. 256:F965-F969PubMedGoogle Scholar
  10. Hazama, A. Okada, Y. 1988. Ca++ sensitivity of volume-regulatory K+ and Cl channels in cultured human epithelial cells.J. Physiol. 402:687–702PubMedGoogle Scholar
  11. Helman, S.I., Baxendale, L.M. 1990. Blocker related changes of channel density: Analysis of a three state model for apical Na channels of frog skin.J. Gen. Physiol. 95:647–678PubMedGoogle Scholar
  12. Heusser, H.R. 1968. Frogs and toads.In: Animal Life Encyclopedia. Vol. 5: Fishes II and Amphibia. H.C.B. Grinek, editor. pp 365–366. Kindler-Verlaz, ZurichGoogle Scholar
  13. Hoffman, E.K., Simonsen, L.O. 1989. Membrane mechanisms in volume and pH regulation in vertebrate cells.Physiol. Rev. 69:315–382PubMedGoogle Scholar
  14. Keeler, R., Wong, N.L.M. 1986. Evidence that prostaglandin E2 stimulates chloride secretion in cultured A6 renal epithelial cells.Am. J. Physiol. 250:F511-F515PubMedGoogle Scholar
  15. Krattenmacher, R., Fischer, H., Van Driessche, W., Clauss, W. 1988. Noise analysis of cAMP-stimulated Na current in frog colon.Pfluegers Arch. 412:568–573Google Scholar
  16. Lewis, S.A. 1977. A reinvestigation of the function of the mammalian urinary bladder.Am. J. Physiol. 232:F187-F295Google Scholar
  17. Lewis, S.A., deMoura, J.L.C. 1984. Apical membrane area of rabbit urinary bladder increases by fusion of intracellular vesicles: An electrophysiological study.J. Membrane Biol. 82:123–136Google Scholar
  18. Lewis, S.A., Donaldson, P. 1990. Ion channels and cell volume regulation: Chaos in an organized system.News Physiol. Sci. 5:112–119Google Scholar
  19. Marunaka, Y., Eaton, D.C. 1988. The effect of amiloride and an amiloride-analogue on single Na+ channels from a renal cell line.FASEB J. 2:A750Google Scholar
  20. Mayer, N. 1969. Adaptation deRana esculenta a des milieux varies. Etude speciale de L'excretion renale de l'eau et des electrolytes au cours des changements de milieux.Comp. Biochem. Physiol. 29:27–50PubMedGoogle Scholar
  21. Palmer, L.G., Frindt, G. 1987. Effects of cell Ca and pH on Na channels from rat cortical collecting tubule.Am. J. Physiol. 253:F333-F339PubMedGoogle Scholar
  22. Perkins, F.M., Handler, J.S. 1981. Transport properties of toad kidney epithelia in culture.Am. J. Physiol. 241:C154-C159Google Scholar
  23. Rafferty, K.A. 1969. Mass culture of amphibian cells: Methods and observations concerning stability of cell type.In: Biology of Amphibian Tumors. M. Mizell, editor. pp 52–81. Springer-Verlag, New YorkGoogle Scholar
  24. Sackin, H. 1987. Stretch-activated potassium channels in renal proximal tubule.Am. J. Physiol. 253:F1253-F1262PubMedGoogle Scholar
  25. Sariban-Sohraby, S., Burg, M.B., Turner, R.J. 1983. Apical sodium uptake in toad kidney epithelial cell line A6.Am. J. Physiol. 245:C167-C171PubMedGoogle Scholar
  26. Sariban-Sohraby, S., Burg, M., Wiesmann, W.P., Chiang, P.K., Johnson, J.P. 1984. Methylation increases sodium transport into A6 apical membrane vesicles: Possible mode of aldosterone action.Science 225:745–746PubMedGoogle Scholar
  27. Sariban-Sohraby, S., Sorscher, E.J., Brenner, B.M., Benos, D.J. 1988. Phosphorylation of a single subunit of the epithelial Na+ channel protein following vasopressin treatment of A6 cells.J. Biol. Chem. 263:13875–13879PubMedGoogle Scholar
  28. Schultz, S.G. 1981. Homocellular regulating mechanisms in sodium-transporting epithelia: Avoidance of extinction by “flush-through.”Am. J. Physiol. 241:F579-F598Google Scholar
  29. Thomas, S.R., Mintz, E. 1987. Time-dependent apical membrane K+ and Na+ selectivity in cultured kidney cells.Am. J. Physiol. 253:C1-C6PubMedGoogle Scholar
  30. Ussing, H.H. 1965. Relationship between osmotic reactions and active sodium transport in frog skin epithelium.Acta Physiol. Scand. 63:141–155PubMedGoogle Scholar
  31. Van Driessche, W., Lindemann, B. 1978. Low noise amplification of voltage and current fluctuations arising in epithelia.Rev. Sci. Instrum. 49:52–57Google Scholar
  32. Van Driessche, W., Zeiske, W. 1980. Spontaneous fluctuations of potassium channels in the apical membrane of frog skin.J. Physiol. 299:1–16PubMedGoogle Scholar
  33. Wade, J.B., Stetson, D.L., Lewis, S.A. 1981. ADH action: Evidence for a membrane shuttle action.Ann. NY Acad. Sci. 372:106–117PubMedGoogle Scholar
  34. Watson, P.A. 1989. Accumulation of cAMP and calcium in S49 mouse lymphoma cells following hyposmotic swelling.J. Biol. Chem. 264:14735–14740PubMedGoogle Scholar
  35. Welsh, M.J. 1988. Defective regulation of ion transport in cystic fibrosis airway epithelium.In: Cellular and Molecular Basis of Cystic Fibrosis. G. Mastella and P.M. Quinton, editors, pp. 321–344. San Francisco Press. San FranciscoGoogle Scholar
  36. Wills, N.K., Alles, W.P., Sandle, G.I., Binder, H.J. 1984. Apical membrane properties and amiloride binding kinetics of the human descending colon.Am. J. Physiol. 247:G749-G757Google Scholar
  37. Wills, N.K., Lewis, S.A., Eaton, D.C. 1979. Active and passive properties of rabbit descending colon: A microelectrode and nystatin study.J. Membrane Biol. 45:81–108Google Scholar
  38. Wills, N.K., Millinoff, L.P. 1990. Amiloride-sensitive Na+ transport across culutured renal (A6) epithelium: Evidence for large currents and high Na:K selectivity.Pfluegers Arch. 416:481–492Google Scholar
  39. Wills, N.K., Zweifach, A. 1987. Recent advances in the characterization of epithelial ionic channels.Biochim. Biophys. Acta 906:1–31PubMedGoogle Scholar
  40. Wong, S.M.E., DeBell, M.C., Chase, H.S. 1990. Cell swelling increases intracellular free [Ca] in cultured toad bladder cells.Am. J. Physiol. 258:F292-F296PubMedGoogle Scholar

Copyright information

© Springer-Verlag New York Inc. 1991

Authors and Affiliations

  • N. K. Wills
    • 1
  • L. P. Millinoff
    • 1
  • W. E. Crowe
    • 1
  1. 1.Department of Physiology and BiophysicsUniversity of Texas Medical BranchGalveston

Personalised recommendations