The Journal of Membrane Biology

, Volume 102, Issue 1, pp 35–48 | Cite as

Na+/H+ exchange in ehrlich ascites tumor cells: Activation by cytoplasmic acidification and by treatment with cupric sulphate

  • Birte Kramhøft
  • Ian H. Lambert
  • Else K. Hoffmann


Exposure of Ehrlich cells to isotonic Na+-propionate medium induces a rapid cell swelling. This treatment is likely to impose an acid load on the cells. Cell swelling is absent in K+-propionate medium but may be induced by the ionophore nigericin, which mediates K+/H+ exchange. Cell swelling in Na+-propionate medium is blocked by amiloride, but an alternative pathway is introduced by addition of the ionophore monensin, which mediates Na+/H+ exchange. Consequently, swelling of Ehrlich cells in Na+-propionate medium is due to the operation of an amiloride-sensitive, Na+-specific mechanism. It is concluded that this mechanism is a Na+/H+ exchange system, activated by cytoplasmic acidification. We have previously demonstrated that the heavy metal salt CuSO4 in micromolar concentrations inhibits regulatory volume decrease (RVD) of Ehrlich cells following hypotonic swelling. The present work shows that CuSO4 inhibits RVD as a result of a net uptake of sodium, of which the major part is sensitive to amiloride. Measurements of intracellular pH show that CuSO4 causes significant cytoplasmic alkalinization, which is abolished by amiloride. Concomitantly, CuSO4 causes an amiloride-sensitive net proton efflux from the cells. The combined results confirm that a Na+/H+ exchange system exists in Ehrlich cells and demonstrate that the heavy metal salt CuSO4 activates this Na+/H+ exchange system.

Key Words

Ehrlich ascites tumor cells pH regulation Na+/H+ exchange volume regulation hypotonic shock cupric sulphate cytoplasmic acidification phorbol ester TPA 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Aronson, P.S., Boron, W.F. (editors). 1986. Na+−H+ exchange, intracellular pH, and cell function.Curr. Topics Membr. Transport 26:3–305Google Scholar
  2. Benos, D.J. 1982. Amiloride: A molecular probe of sodium transport in tissues and cells.Am. J. Physiol. 242:C131-C145Google Scholar
  3. Berridge, M.J. 1984. Inositol triphosphate and diacylglycerol as second messengers.Biochem. J. 220:345–360Google Scholar
  4. Bowen, J.W., Levinson, C. 1984. H+ transport and the regulation of intracellular pH in Ehrlich ascites tumor cells.J. Membrane Biol. 79:7–18Google Scholar
  5. Cala, P.M. 1985. Volume regulation byAmphiuma red blood cells: Characteristics of volume-sensitive K+/H+ and Na+/H+ exchange.Mol. Physiol. 8:199–214Google Scholar
  6. Cala, P.M. 1986. Volume-sensitive alkali metal-H+ transport inAmphiuma red blood cells.Curr. Topics Membr. Transp. 26:79–99.Google Scholar
  7. Doppler, W., Maly, K., Grunicke, H. 1986. Role of Na+/H+ antiport in the regulation of the internal pH of Ehrlich ascites tumor cells in culture.J. Membrane Biol. 91:147–155Google Scholar
  8. Florence, T.M., Batley, G.E. 1977. Determination of the chemical forms of trace metals in natural waters with special reference to copper, lead, cadmium and zinc.Talanta 24:151–158Google Scholar
  9. Frelin, C., Barbry, P., Green, R.D., Jean, T., Vigne, P., Lazdunski, M. 1986. The Na+/H+ antiport of eukaryotic cells: Relationships between the kinetic properties of the system and its physiological function.Biochimie 68:1279–1285Google Scholar
  10. Geck, P., Pfeiffer, B. 1985. Na++K++2Cl cotransport in animal cells—its role in volume regulation.Ann. N.Y. Acad. Sci. 456:166–182Google Scholar
  11. Gillies, R.J., Ogino, T., Shulman, R.G., Ward, D.C. 1982.31P-nuclear magnetic resonance evidence for the regulation of intracellular pH by Ehrlich ascites tumor cells.J. Cell Biol. 95:24–28Google Scholar
  12. Grinstein, S., Clarke, C.A., Rothstein, A. 1983. Activation of Na+/H+ exchange in lymphocytes by osmotically induced volume changes and by cytoplasmic acidification.J. Gen. Physiol. 82:619–638Google Scholar
  13. Grinstein, S., Cohen, S., Goetz, J.D., Rothstein, A. 1985a. Na+/H+ exchange in volume regulation and cytoplasmic pH homeostasis in lymphocytes.Fed. Proc. 44:2508–2512Google Scholar
  14. Grinstein, S., Cohen, S., Goetz, J.D., Rothstein, A., Gelfand, E.W. 1985b. Characterization of the activation of Na+/H+ exchange in lymphocytes by phorbol esters: Change in cytoplasmic pH dependence of the antiport.Proc. Natl. Acad. Sci. USA 82:1429–1433Google Scholar
  15. Grinstein, S., Cohen, S., Rothstein, A. 1984a. Cytoplasmic pH regulation in thymic lymphocytes by an amiloride-sensitive Na+/H+ antiport.J. Gen. Physiol. 83:341–370Google Scholar
  16. Grinstein, S., Furuya, W. 1986. Characterization of the amiloride-sensitive Na+−H+ antiport of human neutrophils.Am. J. Physiol. 250:C283-C291Google Scholar
  17. Grinstein, S., Goetz, J.D., Furuya, W., Rothstein, A., Gelfand, E.W. 1984b. Amiloride-sensitive Na+−H+ exchange in platelets and leukocytes: Detection by electronic cell sizing.Am. J. Physiol. 247:C293-C298Google Scholar
  18. Grinstein, S., Rothstein, A. 1986. Mechanisms of regulation of the Na+/H+ exchanger.J. Membrane Biol. 90:1–12Google Scholar
  19. Grinstein, S., Rothstein, A., Sarkadi, B., Gelfand, E.W. 1984c. Responses of lymphocytes to anisotonic media: Volume-regulating behavior.Am. J. Physiol. 246:C204-C215Google Scholar
  20. Heinz, A., Sachs, G., Schafer, J.A. 1981. Evidence for activation of an active electrogenic proton pump in Ehrlich ascites tumor cells during glycolysis.J. Membrane Biol. 61:143–153Google Scholar
  21. Hendil, K.B., Hoffmann, E.K. 1974. Cell volume regulation in Ehrlich ascites tumor cells.J. Cell. Physiol. 84:115–126Google Scholar
  22. Hoffmann, E.K. 1978. Regulation of cell volume by selective changes in the leak permeabilities of Ehrlich ascites tumor cells.In: Osmotic and volume regulation. C.B. Jørgensen and E. Skadhauge, editors. pp. 377–412. Munksgaard, CopenhagenGoogle Scholar
  23. Hoffmann, E.K. 1987. Volume Regulation in Cultured cells.In: Curr. Top. Membr. Transp. 30:125–179Google Scholar
  24. Hoffmann, E.K., Hendil, K.B. 1976. The role of amino acids and taurine in isosmotic intracellular regulation in Ehrlich ascites mouse tumour cells.J. Comp. Physiol. 18:279–286Google Scholar
  25. Hoffmann, E.K., Lambert, I.H. 1983. Amino acid transport and cell volume regulation in Ehrlich ascites tumour cells.J. Physiol. (London) 338:613–625Google Scholar
  26. Hoffmann, E.K., Lambert, I.H., Simonsen, L.O. 1986. Separate, Ca2+ activated K+ and Cl transport pathways in Ehrlich ascites tumor cells.J. Membrane Biol. 91:227–244Google Scholar
  27. Hoffmann, E.K., Simonsen, L.O., Lambert, I.H. 1984. Volume-induced increase of K+ and Cl permeabilities in Ehrlich ascites tumor cells. Role of internal Ca2+.J. Membrane Biol. 78:211–222Google Scholar
  28. Hoffmann, E.K., Simonsen, L.O., Sjøholm, C. 1979. Membrane potential, chloride exchange, and chloride conductance in Ehrlich mouse ascites tumour cells.J. Physiol. (London) 296:61–84Google Scholar
  29. Hoffmann, E.K., Sjøholm, C., Simonsen, L.O. 1983. Na+, Cl cotransport in Ehrlich ascites tumor cells activated during volume regulation (regulatory volume increase).J. Membrane Biol. 76:269–280Google Scholar
  30. Igarashi, P., Aronson, P.S. 1987. Covalent modification of the renal Na/H exchanger by N,N′-dicyclohexylcarbodiimide.J. Biol. Chem. 262:860–868Google Scholar
  31. Jennings, M.L., Douglas, S.M., McAndrew, P.E. 1986. Amiloride-sensitive sodium-hydrogen exchange in osmotically shrunken rabbit red blood cells.Am. J. Physiol. 251:C32-C40Google Scholar
  32. Koefoed-Johnsen, V., Ussing, H.H. 1973. Transport pathways in frog skin and their modification by copper.In: Secretory Mechanisms of Exocrine Glands. N.A. Thorn and O.H. Peterson, editors. pp. 411–419. Alfred Benzon Symposium VII. Munksgaard, CopenhagenGoogle Scholar
  33. Kramhøft, B., Lambert, I.H., Hoffmann, E.K. 1987a. Demonstration of Na+/H+ exchange in Ehrlich ascites tumor cells by electronic cell sizing.Acta Physiol. Scand. 129:20AGoogle Scholar
  34. Kramhøft, B., Lambert, I.H., Hoffmann, E.K. 1987b. Activation of Na+/H+ exchange in Ehrlich ascites tumor cells by cupric sulfate.Acta Physiol. Scand. 129:19AGoogle Scholar
  35. Kregenow, F.M. 1981. Osmoregulatory salt transporting mechanisms: Control of cell volume in anisotonic medium.Annu. Rev. Physiol. 43:493–505Google Scholar
  36. Lambert, I.H., Kramhøft, B., Hoffmann, E.K. 1984. Effect of copper on volume regulation in Ehrlich ascites tumour cells.Mol. Physiol. 6:83–98Google Scholar
  37. Livne, A., Hoffmann, E.K. 1988. Cytoplasmic acidification during regulatory volume decrease in Ehrlich ascites tumour cells.J. Membrane Biol. (Submitted) Google Scholar
  38. Moolenaar, W.H. 1986. Effects of growth factors on intracellular pH regulation.Annu. Rev. Physiol. 48:363–376Google Scholar
  39. Moolenaar, W.H., Tertoolen, L.G.J., Laat, S.W. de 1984. Phorbol ester and diacylglycerol mimic growth factors in raising cytoplasmic pH. Nature (London)312:371–374Google Scholar
  40. Montrose, M.H., Murer, H. 1986. Regulation of intracellular pH in LLC-PK1 cells by Na+/H+ exchange.J. Membrane Biol. 93:33–42Google Scholar
  41. Parker, J.C. 1983. Volume-responsive sodium movements in dog red blood cells.Am. J. Physiol. 244:C324-C330Google Scholar
  42. Passow, H., Rothstein, A., Clarkson, T.W. 1961. The general pharmacology of the heavy metals.Pharmacol. Rev. 13:185–223Google Scholar
  43. Riisgård, H.U. 1979. Effect of copper on volume regulation in the marine flagellateDunaliella marina.Marine Biol. 50:189–193Google Scholar
  44. Riisgård, H.U., Nørgård-Nielsen, K., Søgård-Jensen, B. 1980. Further studies on volume regulation and effect of copper in relation to pH and EDTA in the naked marine flagellateDunaliella marina.Marine Biol. 56:267–276Google Scholar
  45. Rittenhouse, H.G., Rittenhouse, J.W., Takemoto, L. 1978. Characterization of the cell coat of Ehrlich ascites tumor cells.Biochemistry 17:829–837Google Scholar
  46. Schmitt, R.C., Darwish, H.M., Cheney, J.G., Ettinger, M.J. 1983. Copper transport kinetics by isolated rat hepatocytes.Am. J. Physiol. 244:G183-G191Google Scholar
  47. Thomas, J.A., Buchsbaum, R.N., Zimniak, A., Racher, E. 1979. Intracellular pH measurements in Ehrlich ascites tumour cells utilizing spectroscopic probes generated in situ.Biochemistry 18:2210–2218Google Scholar
  48. Tiffert, T., Garcia-Sancho, J., Lew, L.V. 1984. Irreversible ATP depletion caused by low concentrations of formaldehyde and of calcium-chelator esters in intact human red cells.Biochim. Biophys. Acta 773:143–156Google Scholar
  49. Wiener, E., Dubyak, G., Scarpa, A. 1986. Na+/H+ exchange in Ehrlich ascites tumor cells. Regulation by extracellular ATP and 12-O-tetradecanoylphorbol 13-acetate.J. Biol. Chem. 261:4529–4534Google Scholar

Copyright information

© Springer-Verlag New York Inc. 1988

Authors and Affiliations

  • Birte Kramhøft
    • 1
  • Ian H. Lambert
    • 1
  • Else K. Hoffmann
    • 1
  1. 1.Institute of Biological Chemistry AAugust Krogh InstituteCopenhagen ØDenmark

Personalised recommendations