Pflügers Archiv

, Volume 450, Issue 6, pp 415–428 | Cite as

NH3 and NH4+ permeability in aquaporin-expressing Xenopus oocytes

  • Lars M. Holm
  • Thomas P. Jahn
  • Anders L. B. Møller
  • Jan K. Schjoerring
  • Domenico Ferri
  • Dan A. Klaerke
  • Thomas Zeuthen
Ion Channels, Transporters

Abstract

We have shown recently, in a yeast expression system, that some aquaporins are permeable to ammonia. In the present study, we expressed the mammalian aquaporins AQP8, AQP9, AQP3, AQP1 and a plant aquaporin TIP2;1 in Xenopus oocytes to study the transport of ammonia (NH3) and ammonium (NH4+) under open-circuit and voltage-clamped conditions. TIP2;1 was tested as the wild-type and in a mutated version (tip2;1) in which the water permeability is intact. When AQP8-, AQP9-, AQP3- and TIP2;1-expressing oocytes were placed in a well-stirred bathing medium of low buffer capacity, NH3 permeability was evident from the acidification of the bathing medium; the effects observed with AQP1 and tip2;1 did not exceed that of native oocytes. AQP8, AQP9, AQP3, and TIP2;1 were permeable to larger amides, while AQP1 was not. Under voltage-clamp conditions, given sufficient NH3, AQP8, AQP9, AQP3, and TIP2;1 supported inwards currents carried by NH4+. This conductivity increased as a sigmoid function of external [NH3]: for AQP8 at a bath pH (pHe) of 6.5, the conductance was abolished, at pHe 7.4 it was half maximal and at pHe 7.8 it saturated. NH4+ influx was associated with oocyte swelling. In comparison, native oocytes as well as AQP1 and tip2;1-expressing oocytes showed small currents that were associated with small and even negative volume changes. We conclude that AQP8, AQP9, AQP3, and TIP2;1, apart from being water channels, also support significant fluxes of NH3. These aquaporins could support NH4+ transport and have physiological implications for liver and kidney function.

Keywords

Ammonia Ammonium Aquaporins Conduction Oocytes Mitochondria Liver Kidney Plant 

References

  1. 1.
    Ackerman MJ, Wickman KD, Clapham DE (1994) Hypotonicity activates a native chloride current in Xenopus oocytes. J Gen Physiol 103:153–179CrossRefPubMedGoogle Scholar
  2. 2.
    Agre P, King LS, Yasui M, Guggino WB, Ottersen OP, Fujiyoshi Y, Engel A, Nielsen S (2002) Aquaporin water channels—from atomic structure to clinical medicine. J Physiol (Lond) 542:3–16CrossRefGoogle Scholar
  3. 3.
    Bakouh N, Benjelloun F, Hulin P, Brouillard F, Edelman A, Chèrif-Zahar B, Planelles G (2004) NH3 is involved in the NH4+ transport induced by the functional expression of the human Rh C glycoprotein. J Biol Biochem 279:15975–15983CrossRefGoogle Scholar
  4. 4.
    Boldt M, Burckhardt G, Burckhardt BC (2003) NH4+ conductance in Xenopus laevis oocytes. III. Effect of NH3. Pflugers Arch 446:652–657PubMedGoogle Scholar
  5. 5.
    Burckhardt B-C, Frömter E (1992) Pathways of NH3/NH4+ permeation across Xenopus laevis oocyte cell membrane. Pflugers Arch 420:83–86CrossRefPubMedGoogle Scholar
  6. 6.
    Burckhardt B-C, Burckhardt G (1997) NH4+ conductance in Xenopus laevis oocytes. Pflugers Arch 434:306–312CrossRefPubMedGoogle Scholar
  7. 7.
    Calamita G, Mazzone A, Bizzoca A, Cavalier A, Cassano G, Thomas D, Svelto M (2001) Expression and immunolocalization of the aquaporin-8 water channel in rat gastrointestinal tract. Eur J Cell Biol 80:711–719PubMedGoogle Scholar
  8. 8.
    Carbrey JM, Gorelick-Feldman DA, Kozono D, Praetorius J, Nielsen S, Agre P (2003) Aquaglyceroporin AQP9: solute permeation and metabolic control of expression in liver. Proc Natl Acad Sci USA 100:2945–2950CrossRefPubMedGoogle Scholar
  9. 9.
    Cougnon M, Bouyer P, Hulin P, Anagnostopoulos T, Planelles G (1996) Further investigation of ionic diffusive properties and of NH4+ pathways in Xenopus laevis oocyte cell membrane. Pflugers Arch 431:658–667CrossRefPubMedGoogle Scholar
  10. 10.
    Elkjær M-L, Nejsum LN, Gresz V, Kwon T-H, Jensen UB, Frøkiær J, Nielsen S (2001) Immunolocalization of aquaporin-8 in rat kidney, gastrointestinal tract, testis, and airways. Am J Physiol 281:F1047–F1057Google Scholar
  11. 11.
    Engel A, Stahlberg H (2002) Aquaglyceroporins: channel proteins with a conserved core, multiple functions, and variable surfaces. Int Rev Cytol 215:75–104PubMedGoogle Scholar
  12. 12.
    Ferri D, Mazzone A, Liquori GE, Cassano G, Svelto M, Calamita G (2003) Ontogeny, distribution, and possible functional implications of an unusual aquaporin, AQP8, in mouse liver. Hepatology 38:947–957CrossRefPubMedGoogle Scholar
  13. 13.
    Finkelstein A (1987) Water movement through lipid bilayers, pores and plasma membranes. Wiley-Interscience, New YorkGoogle Scholar
  14. 14.
    Garcia F, Kierbel A, Larocca MC, Gradilone SA, Splinter P, LaRusso NF, Marinelli RA (2001) The water channel aquaporin-8 is mainly intracellular in rat hepatocytes, and its plasma membrane insertion is stimulated by cyclic AMP. J Biol Chem 276:12147–12152CrossRefPubMedGoogle Scholar
  15. 15.
    Häussinger D (1996) Physiological functions of the liver. In: Greger R, Windhorst U (eds) Comprehensive human physiology, Vol. 2. Springer, Berlin Heidelberg New York, pp 1369–1391Google Scholar
  16. 16.
    Häussinger D (1996) Zonal metabolism in the liver. In: Greger R, Windhorst U (eds) Comprehensive human physiology, Vol 2. Springer, Berlin Heidelberg New York, pp 1393–1402Google Scholar
  17. 17.
    Hill EA (1994) Osmotic flow in membrane pores of molecular size. J Membr Biol 137:197–203PubMedGoogle Scholar
  18. 18.
    Holm LM, Klaerke DA, Zeuthen T (2004) Aquaporin 6 is permeable to glycerol and urea. Pflügers Arch 448:181–186CrossRefPubMedGoogle Scholar
  19. 19.
    Huebert RC, Splinter PL, Garcia F, Marinelli RA, LaRusso NF (2002) Expression and localization of aquaporin water channels in rat hepatocytes. J Biol Chem 277:22710–22717CrossRefPubMedGoogle Scholar
  20. 20.
    Ishibashi K, Kuwahara M, Gu Y, Tanaka Y, Marumo F, Sasaki S (1998) Cloning and functional expression of a new aquaporin (AQP9) abundantly expressed in the peripheral leukocytes permeable to water and urea, but not to glycerol. Biochem Biophys Res Commun 244:268–274CrossRefPubMedGoogle Scholar
  21. 21.
    Jahn TP, Møller ALB, Zeuthen T, Holm LM, Klaerke DA, Mohsin B, Kühlbrandt W, Schjoerring JK (2004) Aquaporin homologues in plants and mammals transport ammonia. FEBS Lett 574:31–36CrossRefPubMedGoogle Scholar
  22. 22.
    Kedem O, Katchalsky A (1961) A physical interpretation of the phenomenological coefficients of membrane permeability. J Gen Physiol 45:143–179CrossRefPubMedGoogle Scholar
  23. 23.
    Khademi S, O’Connell III J, Remis J, Robles-Colmenares Y, Miercke LJW, Stroud RM (2004) Mechanism of ammonia transport by Amt/MEP/Rh: structure of AmtB at 1.35 Å. Science 305:1587–1594CrossRefPubMedGoogle Scholar
  24. 24.
    Knepper MA, Packer R, Good DW (1989) Ammonium transport in the kidney. Physiol Rev 69:179–249PubMedGoogle Scholar
  25. 25.
    Koyama Y, Yamamoto T, Kondo D, Funaki H, Yaoita E, Kawasaki K, Sato N, Hatekeyama K, Kihara I (1997) Molecular cloning of a new aquaporin from rat pancreas and liver. J Biol Chem 272:30329–30333CrossRefPubMedGoogle Scholar
  26. 26.
    Ludewig U (2004) Electroneutral ammonium transport by basolateral rhesus B glycoprotein. J Physiol (Lond) 559:751–759Google Scholar
  27. 27.
    Ma T, Song Y, Yang B, Gillespie A, Carlson EJ, Epstein CJ, Verkman AS (2000) Nephrogenic diabetes insipidus in mice lacking aquaporin-3 water channels. Proc Natl Acad Sci USA 97:4386–4391CrossRefPubMedGoogle Scholar
  28. 28.
    Ma T, Yang B, Verkman AS (1997) Cloning of a novel water and urea-permeable aquaporin from mouse expressed strongly in colon, placenta, liver and heart. Biochem Biophys Res Commun 240:324–328CrossRefPubMedGoogle Scholar
  29. 29.
    Meinild A-K, Klaerke DA, Loo DDF, Wright EM, Zeuthen T (1998) The human Na+ /glucose cotransporter is a molecular water pump. J Physiol (Lond) 508:15–21Google Scholar
  30. 30.
    Meinild A-K, Klaerke DA, Zeuthen T (1998) Bidirectional water fluxes and specificity for small hydrophilic molecules in aquaporins 0 to 5. J Biol Chem 273:32446–32451CrossRefPubMedGoogle Scholar
  31. 31.
    Murata K, Mitsouka K, Hirai T, Walz T, Agre P, Heymann JB, Engel A, Fujiyoshi Y (2000) Structural determinants of water permeation through aquaporin-1. Nature 407:599–605CrossRefPubMedGoogle Scholar
  32. 32.
    Nakhoul NL, Hamm LL (2004) Non-erythroid Rh glycoproteins: a putative new family of mammalian ammonium transporters. Pflugers Arch 447:807–812CrossRefPubMedGoogle Scholar
  33. 33.
    Nakhoul NL, Hering-Smith KS, Abdulnour-Nakhoul SM, Hamm LL (2001) Transport of NH3/NH4+ in oocytes expressing aquaporin-1. Am J Physiol 281:F255–F263Google Scholar
  34. 34.
    Nicholls DG, Ferguson SJ (1995) Bioenergetics 2 edn. Academic Press, LondonGoogle Scholar
  35. 35.
    Nielsen S, Frøkier J, Marples D, Kwon T-H, Agre P, Knepper MA (2002) Aquaporins in the kidney: from molecules to medicine. Physiol Rev 82:205–244PubMedGoogle Scholar
  36. 36.
    Portincasa P, Moschetta A, Mazzone A, Palasciano G, Svelto M, Calamita G (2003) Water handling and aquaporins in bile formation: recent advances and research trends. J Hepatol 39:864–874CrossRefPubMedGoogle Scholar
  37. 37.
    Preston GM, Carroll TP, Guggino WB, Agre P (1992) Appearance of water channels in Xenopus oocytes expressing red cell CHIP28 protein. Science 256:385–389PubMedGoogle Scholar
  38. 38.
    Sasaki S, Ishibashi K, Marumo F (1998) Aquaporin-2 and -3: representatives of two subgroups of the aquaporin family colocalized in the kidney collecting duct. Annu Rev Physiol 60:199–220CrossRefPubMedGoogle Scholar
  39. 39.
    Sasaki S, Ishibashi K, Nagai T, Marumo F (1992) Regulation mechanisms of intracellular pH of Xenopus laevis oocyte. Biochim Biophys Acta 1137:45–51CrossRefPubMedGoogle Scholar
  40. 40.
    Tsukaguchi H, Shayakul C, Berfer UV, Mackenzie B, Devidas S, Guggino WB, VanHoek AN, Hediger MA (1998) Molecular characterization of a broad selectivity neutral solute channel. J Biol Biochem 273:24737–24743CrossRefGoogle Scholar
  41. 41.
    Tsukaguchi H, Weremowicz S, Morton CC, Hediger MA (1999) Functional and molecular characterization of the human neutral solute channel aquaporin-9. Am J Physiol 277:F685–F696PubMedGoogle Scholar
  42. 42.
    Vaughan-Jones RD, Peercy BE, Keener JP, Spitzer KW (2002) Intrinsic H+ ion mobility in the rabbit ventricular myocyte. J Physiol (Lond) 541:139–158CrossRefGoogle Scholar
  43. 43.
    Verkman AS, Yang B, Song Y, Manley GT, Ma T (2000) Role of water channels in fluid transport studied by phenotype analysis of aquaporin knockout mice. Exp Physiol 85S:233S–241SCrossRefGoogle Scholar
  44. 44.
    Zampighi GA, Kreman M, Boorer KJ, Loo DDF, Bezanilla F, Chandy G, Hall JE, Wright EM (1995) A method for determining the unitary functional capacity of cloned channels and transporters expressed in Xenopus laevis oocytes. J Membr Biol 148:65–78PubMedGoogle Scholar
  45. 45.
    Zeuthen T (1980) How to make and use double-barreled ion-selective microelectrodes. Curr Top Membr Trans 13:31–47Google Scholar
  46. 46.
    Zeuthen T, Hamann S, la Cour M (1996) Cotransport of H+, lactate and H2O by membrane proteins in retinal pigment epithelium of bullfrog. J Physiol (Lond) 497:3–17Google Scholar
  47. 47.
    Zeuthen T, Klaerke DA (1999) Transport of water and glycerol in aquaporin 3 is gated by H+. J Biol Chem 274:21631–21636CrossRefPubMedGoogle Scholar
  48. 48.
    Zeuthen T, Meinild A-K, Klaerke DA, Loo DDF, Wright EM, Belhage B, Litman T (1997) Water transport by the Na+ /glucose cotransporter under isotonic conditions. Biol Cell 89:307–312CrossRefPubMedGoogle Scholar
  49. 49.
    Zeuthen T, Zeuthen E, Klaerke DA (2002) Mobility of ions, sugar, and water in the cytoplasm of Xenopus oocytes expressing Na+-coupled sugar transporters (SGLT1). J Physiol (Lond) 542:71–87CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2005

Authors and Affiliations

  • Lars M. Holm
    • 1
  • Thomas P. Jahn
    • 2
  • Anders L. B. Møller
    • 2
  • Jan K. Schjoerring
    • 2
  • Domenico Ferri
    • 3
  • Dan A. Klaerke
    • 1
  • Thomas Zeuthen
    • 1
  1. 1.Nordic Centre for Water Imbalance Related Disorders. Department of Medical PhysiologyPanum Institute, University of CopenhagenDenmark
  2. 2.Plant Nutrition Laboratory, Department of Agricultural SciencesRoyal Veterinary and Agricultural UniversityDenmark
  3. 3.Department of Zoology, Laboratory of Histology and Comparative AnatomyUniversity of BariItaly

Personalised recommendations