Skip to main content
PMC home page
The Journal of General Physiology logoLink to The Journal of General Physiology
. 1990 Jul 1;96(1):167–193. doi: 10.1085/jgp.96.1.167

Anion-coupled Na efflux mediated by the human red blood cell Na/K pump

PMCID: PMC2228984  PMID: 2212979

Abstract

The red cell Na/K pump is known to continue to extrude Na when both Na and K are removed from the external medium. Because this ouabain- sensitive flux occurs in the absence of an exchangeable cation, it is referred to as uncoupled Na efflux. This flux is also known to be inhibited by 5 mM Nao but to a lesser extent than that inhibitable by ouabain. Uncoupled Na efflux via the Na/K pump therefore can be divided into a Nao-sensitive and Nao-insensitive component. We used DIDS- treated, SO4-equilibrated human red blood cells suspended in HEPES- buffered (pHo 7.4) MgSO4 or (Tris)2SO4, in which we measured 22Na efflux, 35SO4 efflux, and changes in the membrane potential with the fluorescent dye, diS-C3 (5). A principal finding is that uncoupled Na efflux occurs electroneurally, in contrast to the pump's normal electrogenic operation when exchanging Nai for Ko. This electroneutral uncoupled efflux of Na was found to be balanced by an efflux of cellular anions. (We were unable to detect any ouabain-sensitive uptake of protons, measured in an unbuffered medium at pH 7.4 with a Radiometer pH-STAT.) The Nao-sensitive efflux of Nai was found to be 1.95 +/- 0.10 times the Nao-sensitive efflux of (SO4)i, indicating that the stoichiometry of this cotransport is two Na+ per SO4=, accounting for 60-80% of the electroneutral Na efflux. The remainder portion, that is, the ouabain-sensitive Nao-insensitive component, has been identified as PO4-coupled Na transport and is the subject of a separate paper. That uncoupled Na efflux occurs as a cotransport with anions is supported by the result, obtained with resealed ghosts, that when internal and external SO4 was substituted by the impermeant anion, tartrate i,o, the efflux of Na was inhibited 60-80%. This inhibition could be relieved by the inclusion, before DIDS treatment, of 5 mM Cli,o. Addition of 10 mM Ko to tartrate i,o ghosts, with or without Cli,o, resulted in full activation of Na/K exchange and the pump's electrogenicity. Although it can be concluded that Na efflux in the uncoupled mode occurs by means of a cotransport with cellular anions, the molecular basis for this change in the internal charge structure of the pump and its change in ion selectivity is at present unknown.

Full Text

The Full Text of this article is available as a PDF (1.6 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. BAKER P. F. AN EFFLUX OF NINHYDRIN-POSITIVE MATERIAL ASSOCIATED WITH THE OPERATION OF THE NA+ PUMP IN INTACT CRAB NERVE IMMERSED IN NA+-FREE SOLUTIONS. Biochim Biophys Acta. 1964 Sep 25;88:458–460. doi: 10.1016/0926-6577(64)90208-6. [DOI] [PubMed] [Google Scholar]
  2. Beaugé L. A., Glynn I. M. Sodium ions, acting at high-affinity extracellular sites, inhibit sodium-ATPase activity of the sodium pump by slowing dephosphorylation. J Physiol. 1979 Apr;289:17–31. doi: 10.1113/jphysiol.1979.sp012722. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Becker B. F., Duhm J. Evidence for anionic cation transport of lithium, sodium and potassium across the human erythrocyte membrane induced by divalent anions. J Physiol. 1978 Sep;282:149–168. doi: 10.1113/jphysiol.1978.sp012454. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Blostein R. Sodium pump-catalyzed sodium-sodium exchange associated with ATP hydrolysis. J Biol Chem. 1983 Jul 10;258(13):7948–7953. [PubMed] [Google Scholar]
  5. Bodemann H., Passow H. Factors controlling the resealing of the membrane of human erythrocyte ghosts after hypotonic hemolysis. J Membr Biol. 1972;8(1):1–26. doi: 10.1007/BF01868092. [DOI] [PubMed] [Google Scholar]
  6. Cornelius F., Skou J. C. The sided action of Na+ on reconstituted shark Na+/K+-ATPase engaged in Na+-Na+ exchange accompanied by ATP hydrolysis. II. Transmembrane allosteric effects on Na+ affinity. Biochim Biophys Acta. 1988 Oct 6;944(2):223–232. doi: 10.1016/0005-2736(88)90435-x. [DOI] [PubMed] [Google Scholar]
  7. Cornelius F. Uncoupled Na+-efflux on reconstituted shark Na,K-ATPase is electrogenic. Biochem Biophys Res Commun. 1989 Apr 28;160(2):801–807. doi: 10.1016/0006-291x(89)92504-7. [DOI] [PubMed] [Google Scholar]
  8. De Weer P., Gadsby D. C., Rakowski R. F. Voltage dependence of the Na-K pump. Annu Rev Physiol. 1988;50:225–241. doi: 10.1146/annurev.ph.50.030188.001301. [DOI] [PubMed] [Google Scholar]
  9. Eilam Y., Stein W. D. The efflux of sodium from human red blood cells. Biochim Biophys Acta. 1973 Nov 16;323(4):606–619. doi: 10.1016/0005-2736(73)90169-7. [DOI] [PubMed] [Google Scholar]
  10. Flatman P. W., Lew V. L. The magnesium dependence of sodium-pump-mediated sodium-potassium and sodium-sodium exchange in intact human red cells. J Physiol. 1981 Jun;315:421–446. doi: 10.1113/jphysiol.1981.sp013756. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Garay R. P., Garrahan P. J. The interaction of sodium and potassium with the sodium pump in red cells. J Physiol. 1973 Jun;231(2):297–325. doi: 10.1113/jphysiol.1973.sp010234. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Garrahan P. J., Glynn I. M. Facftors affecting the relative magnitudes of the sodium:potassium and sodium:sodium exchanges catalysed by the sodium pump. J Physiol. 1967 Sep;192(1):189–216. doi: 10.1113/jphysiol.1967.sp008296. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Garrahan P. J., Glynn I. M. The behaviour of the sodium pump in red cells in the absence of external potassium. J Physiol. 1967 Sep;192(1):159–174. doi: 10.1113/jphysiol.1967.sp008294. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Garrahan P. J., Glynn I. M. The sensitivity of the sodium pump to external sodium. J Physiol. 1967 Sep;192(1):175–188. doi: 10.1113/jphysiol.1967.sp008295. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Glynn I. M., Hara Y., Richards D. E. The occlusion of sodium ions within the mammalian sodium-potassium pump: its role in sodium transport. J Physiol. 1984 Jun;351:531–547. doi: 10.1113/jphysiol.1984.sp015261. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Glynn I. M., Hoffman J. F. Nucleotide requirements for sodium-sodium exchange catalysed by the sodium pump in human red cells. J Physiol. 1971 Oct;218(1):239–256. doi: 10.1113/jphysiol.1971.sp009612. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Glynn I. M., Karlish S. J. ATP hydrolysis associated with an uncoupled sodium flux through the sodium pump: evidence for allosteric effects of intracellular ATP and extracellular sodium. J Physiol. 1976 Apr;256(2):465–496. doi: 10.1113/jphysiol.1976.sp011333. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Goldshleger R., Shahak Y., Karlish S. J. Electrogenic and electroneutral transport modes of renal Na/K ATPase reconstituted into proteoliposomes. J Membr Biol. 1990 Feb;113(2):139–154. doi: 10.1007/BF01872888. [DOI] [PubMed] [Google Scholar]
  19. HOFFMAN J. F. The active transport of sodium by ghosts of human red blood cells. J Gen Physiol. 1962 May;45:837–859. doi: 10.1085/jgp.45.5.837. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Hara Y., Nakao M. ATP-dependent proton uptake by proteoliposomes reconstituted with purified Na+,K+-ATPase. J Biol Chem. 1986 Sep 25;261(27):12655–12658. [PubMed] [Google Scholar]
  21. Hara Y., Nakao M. Sodium ion discharge from pig kidney Na+, K+-ATPase Na+-dependency of the E1P-E2P equilibrium in the absence of KCl. J Biochem. 1981 Oct;90(4):923–931. doi: 10.1093/oxfordjournals.jbchem.a133580. [DOI] [PubMed] [Google Scholar]
  22. Hoffman J. F., Kaplan J. H., Callahan T. J. The Na:K pump in red cells is electrogenic. Fed Proc. 1979 Oct;38(11):2440–2441. [PubMed] [Google Scholar]
  23. Hoffman J. F., Laris P. C. Determination of membrane potentials in human and Amphiuma red blood cells by means of fluorescent probe. J Physiol. 1974 Jun;239(3):519–552. doi: 10.1113/jphysiol.1974.sp010581. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Karlish S. J., Glynn I. M. An uncoupled efflux of sodium ions from human red cells, probably associated with Na-dependent ATPase activity. Ann N Y Acad Sci. 1974;242(0):461–470. doi: 10.1111/j.1749-6632.1974.tb19110.x. [DOI] [PubMed] [Google Scholar]
  25. Karlish S. J., Yates D. W., Glynn I. M. Conformational transitions between Na+-bound and K+-bound forms of (Na+ + K+)-ATPase, studied with formycin nucleotides. Biochim Biophys Acta. 1978 Jul 7;525(1):252–264. doi: 10.1016/0005-2744(78)90219-x. [DOI] [PubMed] [Google Scholar]
  26. Kennedy B. G., Lunn G., Hoffman J. F. Effects of altering the ATP/ADP ratio on pump-mediated Na/K and Na/Na exchanges in resealed human red blood cell ghosts. J Gen Physiol. 1986 Jan;87(1):47–72. doi: 10.1085/jgp.87.1.47. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Knauf P. A., Fuhrmann G. F., Rothstein S., Rothstein A. The relationship between anion exchange and net anion flow across the human red blood cell membrane. J Gen Physiol. 1977 Mar;69(3):363–386. doi: 10.1085/jgp.69.3.363. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Lee K. H., Blostein R. Red cell sodium fluxes catalysed by the sodium pump in the absence of K+ and ADP. Nature. 1980 May 29;285(5763):338–339. doi: 10.1038/285338a0. [DOI] [PubMed] [Google Scholar]
  29. Lew V. L., Hardy M. A., Jr, Ellory J. C. The uncoupled extrusion of Na+ through the Na+ pump. Biochim Biophys Acta. 1973 Oct 11;323(2):251–266. doi: 10.1016/0005-2736(73)90149-1. [DOI] [PubMed] [Google Scholar]
  30. Marin R., Hoffman J. F. Two different types of ATP-dependent anion coupled Na transport are mediated by the human red blood cell and Na/K pump. Prog Clin Biol Res. 1988;268A:539–544. [PubMed] [Google Scholar]
  31. POST R. L., JOLLY P. C. The linkage of sodium, potassium, and ammonium active transport across the human erythrocyte membrane. Biochim Biophys Acta. 1957 Jul;25(1):118–128. doi: 10.1016/0006-3002(57)90426-2. [DOI] [PubMed] [Google Scholar]
  32. Pedemonte C. H. Kinetic mechanism of inhibition of the Na+-pump and some of its partial reactions by external Na+ (Na+o). J Theor Biol. 1988 Sep 17;134(2):165–182. doi: 10.1016/s0022-5193(88)80200-5. [DOI] [PubMed] [Google Scholar]
  33. Polvani C., Blostein R. Protons as substitutes for sodium and potassium in the sodium pump reaction. J Biol Chem. 1988 Nov 15;263(32):16757–16763. [PubMed] [Google Scholar]
  34. Reed P. W. Effects of divalent cation ionophore A23187 on potassium permeability of rat erythrocytes. J Biol Chem. 1976 Jun 10;251(11):3489–3494. [PubMed] [Google Scholar]
  35. Sachs J. R. Sodium movements in the human red blood cell. J Gen Physiol. 1970 Sep;56(3):322–341. doi: 10.1085/jgp.56.3.322. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Sachs J. R. The order of addition of sodium and release of potassium at the inside of the sodium pump of the human red cell. J Physiol. 1986 Dec;381:149–168. doi: 10.1113/jphysiol.1986.sp016319. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Sims P. J., Waggoner A. S., Wang C. H., Hoffman J. F. Studies on the mechanism by which cyanine dyes measure membrane potential in red blood cells and phosphatidylcholine vesicles. Biochemistry. 1974 Jul 30;13(16):3315–3330. doi: 10.1021/bi00713a022. [DOI] [PubMed] [Google Scholar]
  38. Yingst D. R., Hoffman J. F. Ca-induced K transport in human red blood cell ghosts containing arsenazo III. Transmembrane interactions of Na, K, and Ca and the relationship to the functioning Na-K pump. J Gen Physiol. 1984 Jan;83(1):19–45. doi: 10.1085/jgp.83.1.19. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Yingst D. R., Hoffman J. F. Changes of intracellular Ca++ as measured by arsenazo III in relation to the K permeability of human erythrocyte ghosts. Biophys J. 1978 Sep;23(3):463–471. doi: 10.1016/S0006-3495(78)85462-9. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from The Journal of General Physiology are provided here courtesy of The Rockefeller University Press

RESOURCES