Advertisement

The Journal of Membrane Biology

, Volume 91, Issue 1, pp 33–41 | Cite as

Stoichiometry and ion affinities of the Na−K−Cl cotransport system in the intestine of the winter flounder (Pseudopleuronectes americanus)

  • S. M. O'Grady
  • M. W. Musch
  • M. Field
Articles

Summary

Na−K−Cl cotransport stoichiometry and affinities for Na, K and Cl were determined in flounder intestine. Measurement of simultaneous NaCl and RbCl influxes resulted in ratios of 2.2 for Cl/Na and 1.8 for Cl/Rb. The effect of Na and Rb on Rb influx showed first order kinetics withK1/2 values of 5 and 4.5mm and Hill coefficients of 0.9 and 1.2, respectively. The effect of Cl on rubidium influx showed a sigmoidal relationship withK1/2 of 20mm and a Hill coefficient of 2.0. The effects of variations in Na and Cl concentration on short-circuit current (Isc) were also determined. TheK1/2 for Na was 7mm with a Hill coefficient of 0.9 and theK1/2 for Cl was 46mm with a Hill coefficient of 1.9. Based on the simultaneous influx measurements, a cotransport stoichiometry of 1Na∶1K∶2Cl is concluded. The Hill coefficients for Cl suggest a high degree of cooperativity between Cl binding sites. Measurements of the ratio of net Na and Cl transepithelial fluxes under short-circuit conditions (using a low Na Ringer solution to minimize the passive Na flux) indicate that the Cl/Na flux ratio is approximately 2∶1. Therefore Na recycling from serosa to mucosa does not significantly contribute to theIsc. Addition of serosal ouabain (100 μm) inhibited Rb influx, indicating that Na−K−Cl cotransport is inhibited by ouabain. This finding suggests that a feedback mechanism exists between the Na−K-ATPase on the basolateral membrane and the apical Na−K−2Cl cotransporter.

Key Words

cotransport ion transport stoichiometry intestinal transport winter flounder 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Aiton, J.F., Chipperfield, A.R., Lamb, J.F., Ogden, P., Simmons, N.L. 1981. Occurrence of passive furosemide-sensitive transmembrane potassium transport in cultured cells.Biochim. Biophys. Acta 646:389–398Google Scholar
  2. 2.
    Degnan, K.J., Karnaky, K.J., Zadunaisky, J.A. 1977. Active chloride transport in the in vitro opercular skin of a teleost (Fundulus heteroclitus), a gill like epithelium rich in chloride cells.J. Physiol. (London) 271:155–191Google Scholar
  3. 3.
    Duffey, M.F., Thompson, S.M., Frizzell, R.A., Schultz, S.G. 1979. Intracellular chloride activities and active chloride absorption in the intestinal epithelium of the winter flounder.J. Membrane Biol. 50:331–341Google Scholar
  4. 4.
    Dunham, P.B., Stewart, G.W., Ellory, J.C. 1980. Chloride activated passive potassium transport in human erythrocytes.Proc. Natl. Acad. Sci. USA 77:1711–1715Google Scholar
  5. 5.
    Ellory, J.C., Stewart, G.W. 1982. The human erythrocyte Cl-dependent Na−K cotransport system as a possible model for studying the action of loop diuretics.Br. J. Pharmacol. 75:183–188Google Scholar
  6. 6.
    Evans, D.H., Mallery, C.H., Kravitz, L. 1973. Sodium extrusion by a fish acclimated to sea water; physiological and biochemical description of a Na for K exchange system.J. Exp. Biol. 58:627–636Google Scholar
  7. 7.
    Field, M., Karnaky, K.J., Smith, P.L., Bolton, J.E., Kinter, W.B. 1978. Ion transport across the isolated intestinal mucosa of the winter flounder (Pseudopleuronectes americanus).J. Membrane Biol. 41:265–293Google Scholar
  8. 8.
    Foskett, J.K., Scheffey, C. 1982. The chloride cell: Definitive identification as the salt-secretory cell in teleosts.Science 215:164–166Google Scholar
  9. 9.
    Frizzell, R.A., Halm, D.R., Musch, M.W., Stewart, C.P., Field, M. 1984. Potassium transport by flounder intestinal mucosa.Am. J. Physiol. 246:F946-F951Google Scholar
  10. 10.
    Frizzell, R.A., Smith, P.L., Vosburgh, E., Field, M. 1979. Coupled sodium-chloride influx across brush border of flounder intestine.J. Membrane Biol. 46:27–39Google Scholar
  11. 11.
    Geck, P., Pietvgyk, C., BurckHardt, B.C., Pfeiffer, B., Heinz, E. 1980. Electrically silent cotransport of Na,K and Cl in Ehrlich cells.Biochim. Biophys. Acta 600:432–447Google Scholar
  12. 12.
    Greger, R. 1981. Chloride reabsorption in the rabbit cortical thick ascending limb of the hoop of Henle.Pfluegers Arch. 390:38–43Google Scholar
  13. 13.
    Gregor, R. 1981. Coupled transport of Na and Cl in the thick ascending limb of Henle's loop of rabbit nephron.Scand. Audiol. Suppl. 14:1–15Google Scholar
  14. 14.
    Greger, R., Schlatter, E. 1981. Presence of luminal K, a prerequisite for active NaCl transport in cortical thick ascending limb of Henle's loop in rabbit kidney.Pfluegers Arch. 392:92–94Google Scholar
  15. 15.
    Greger, R., Schlatter, E. 1983. Cellular mechanisms of the action of loop diuretics on the thick ascending limb of Henle's loop.Klin. Wochenschr. 61:1019–1027Google Scholar
  16. 16.
    Greger, R., Schlatter, E. 1984. Mechanism of NaCl secretion in the rectal gland of spiny dogfish (Squalus acanthias) I.Pfluegers Arch. 402:63–75Google Scholar
  17. 17.
    Hall, A.C., Ellory, J.C. 1985. Measurement and stoichiometry of bumetanide-sensitive (2Na∶1K∶3Cl) cotransport in ferret red cells.J. Membrane Biol. 85:205–213Google Scholar
  18. 18.
    Halm, D.R., Krasny, E.J., Frizzell, R.A. 1985. Electrophysiology of flounder intestinal mucosa: Conductance properties of the cellular and paracellular pathways.J. Gen. Physiol. (in press) Google Scholar
  19. 19.
    Hannafin, J., Kinne-Saffran, E., Friedman, D., Kinne, R. 1983. Presence of a sodium-potassium chloride cotransport system in the rectal gland ofSqualus acanthias.J. Membrane Biol. 75:73–83Google Scholar
  20. 20.
    Hass, M., Schmidt, W.F., McManus, T.J. 1982. Catecholamine-stimulated ion transport in duck red cells. Gradient effects in electrically neutral [Na+K+2Cl] cotransport.J. Gen. Physiol. 80:125–147Google Scholar
  21. 21.
    Hickman, C.P. 1968. Ingestion, intestinal absorption and elimination of sea water and salts in the southern flounder,Paralichthys lethostigma.Can. J. Zool. 46:457–466Google Scholar
  22. 22.
    Karnaky, K.J. 1980. Ion secreting epithelia: Chloride cells in the head region ofFundulus heteroclitus.Am. J. Physiol. 238:R185-R198Google Scholar
  23. 23.
    Koenig, B., Ricapito, S., Kinne, R. 1983. Chloride transport in the thick ascending limb of Henle's loop: Potassium dependence and stoichiometry of the NaCl cotransport system in plasma membrane vesicles.Pfluegers Arch. 399:173–179Google Scholar
  24. 24.
    Krasny, F.J., Frizzell, R.A. 1984. Intestinal ion transport in marine teleosts.In: Chloride Transport Coupling in Biological Membranes and Epithelia. G.A. Gerencer, editor. pp. 205–220. Elsevier, New YorkGoogle Scholar
  25. 25.
    McRoberts, J.A., Erlinger, S., Rindler, M.J., Saier, M.H. 1982. Furosemide sensitive salt transport in Madin-Darby canine kidney cell line. Evidence for the cotransport of Na,K and Cl.J. Biol. Chem. 257:2260–2266Google Scholar
  26. 26.
    Musch, M.W., Orellana, S.A., Kimberg, L.S., Field, M., Halm, D.R., Krasny, E.J., Frizzell, R.A. 1982. Na−K−Cl cotransport in the intestine of a Marine teleost.Nature (London) 300:351–353Google Scholar
  27. 27.
    Oberleithner, H., Guggino, W., Glekisch, G. 1983. Effect of furosemide on luminal sodium, chloride and potassium transport in the early distal tubule ofAmphuima kidney.Pfluegers Arch. 396:27–33Google Scholar
  28. 28.
    Owen, N.E., Prastein, M.L. 1975. Na/K/Cl cotransport in cultured human fibroblasts.J. Biol. Chem. 260:1445–1451Google Scholar
  29. 29.
    Palfrey, H.C., Alper, S.L., Greengard, P. 1980. Protein phosphorylation and the regulation of a cation cotransport.J. Exp. Biol. 89:103–115Google Scholar
  30. 30.
    Rao, M.C., Nash, N.T., Field, M. 1984. Differing effects of cGMP and cAMP on ion transport in winter flounder intestine.Am. J. Physiol. 246:C167-C177Google Scholar
  31. 31.
    Russell, J.M. 1979. Chloride and sodium influx: A coupled uptake mechanism in the squid giant axon.J. Gen. Physiol. 73:801–818Google Scholar
  32. 32.
    Russell, J.M. 1983. Cation coupled chloride influx in squid axon: Role of potassium and stoichiometry of the transport process.J. Gen. Physiol. 81:909–925Google Scholar
  33. 33.
    Sleet, R.B., Weber, L.S. 1982. The rate and manner of sea water ingestion by a marine teleost and corresponding sea water modification by the gut.Comp. Biochem. Physiol. 72A:469–475Google Scholar
  34. 34.
    Thompson, S.M., Dawson, D.C. 1978. Sodium uptake across the apical border of the isolated turtle colon: Confirmation of the two-barrier model.J. Membrane Biol. 42:357–374Google Scholar

Copyright information

© Springer-Verlag New York Inc. 1986

Authors and Affiliations

  • S. M. O'Grady
    • 1
    • 2
  • M. W. Musch
    • 1
    • 2
  • M. Field
    • 1
    • 2
  1. 1.Department of Medicine and Department of PhysiologyColumbia University, College of Physicians and SurgeonsNew York
  2. 2.Mount Desert Island Biological LaboratorySalsbury Cove

Personalised recommendations