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Pflügers Archiv

, Volume 419, Issue 6, pp 632–638 | Cite as

Amiloride blockage of Na+ channels in amphibian epithelia does not require external Ca2+

  • Luc Desmedt
  • Jeannine Simaels
  • Willy Van Driessche
Transport Processes, Metabolism and Endocrinology; Kidney, Gastrointestinal Tract, and Exocrine Glands

Abstract

Noise analysis was used to study the influence of external Ca2+ on the blockage of Na+ transport by amiloride. Experiments were done using frog skin (Rana temporaria and Rana catesbeiana), toad urinary bladder (Bufo marinus) and epithelia of A6 cells. In non-depolarized skins and bladders, removal of Ca2+ from the mucosal bath diminished markedly the inhibitory effect of amiloride. Ca2+ depletion also gave rise to the appearance of an additional noise component related to cation movement through the poorly selective cation channel in the apical membrane [Aelvoet I, Erlij D, Van Driessche W (1988) J Physiol (Lond) 398:555–574; Van Driessche W, Desmedt L, Simaels J (1991) Pflügers Arch 418:193–203]. The amplitude of this Ca2+-blockable noise component was elevated by amiloride and markedly exceeded the amiloride-induced Lorentzian noise levels as recorded in the presence of Ca2+. On the other hand, in K+-depolarized skins and bladders as well as in non-depolarized epithelia of A6 cells, the Ca2+-blockable noise was absent or of much smaller amplitude. Depolarization of frog skin and toad urinary bladder apparently inactivated the poorly selective channels, whereas in A6 cells they were not observed. Under these conditions the typical amiloride-induced blocker noise could also be analysed in the absence of Ca2+ and demonstrated that the on and off rates for amiloride binding were not significantly altered by external Ca2+. We conclude that (a) external Ca2+ per se does not affect the inhibitory potency of amiloride, and (b) that the observed differences between frog skin, toad urinary bladder and A6 cells originate from the presence or absence of a poorly selective cation channel rather than from a different amiloride receptor structure.

Key words

Frog skin Toad urinary bladder A6 epithelia Amiloride Noise analysis Calcium 

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References

  1. 1.
    Aelvoet I, Erlij D, Van Driessche W (1988) Activation and blockage of a calcium-sensitive cation-selective pathway in the apical membrane of toad bladder. J Physiol (Lond) 398:555–574Google Scholar
  2. 2.
    Benos DJ (1982) Amiloride: a molecular probe of Na+ transport in tissues and cells. Am J Physiol 242:C131-C145Google Scholar
  3. 3.
    Benos DJ, Mandel LJ, Balaban RS (1979) On the mechanism of the amiloride — Na+ entry site interaction in anuran skin epithelia. J Gen Physiol 73:307–326Google Scholar
  4. 4.
    Cuthbert AW, Shum WK (1974) Binding of amiloride to Na+ channels in frog skin. Mol Pharmacol 10:880–891Google Scholar
  5. 5.
    Cuthbert AW, Wong PYD (1972) The role of Ca2+ ions in the interaction of amiloride with membrane receptors. Mol Pharmacol 8:222–229Google Scholar
  6. 6.
    Cuthbert AW, Wong PYD (1974) Interactions of Na+ channels in transporting epithelia: a two-state model. Mol Pharmacol 10:892–903Google Scholar
  7. 7.
    Fabiato A, Fabiato F (1979) Calculator programs for computing the composition of solutions containing multiple metals and ligands used for experiments in skinned muscle cells. J Physiol (Lond) 75:463–505Google Scholar
  8. 8.
    Granitzer M, Nagel W (1990) Dual effect of barium on basolateral membrane conductance of frog skin. Pflügers Arch 417:207–212Google Scholar
  9. 9.
    Harrison SM, Bers DM (1989) Correction of proton and Ca association constants of EGTA for temperature and ionic strength. Am J Physiol 256:C1250-C1256Google Scholar
  10. 10.
    Helman SI, Kizer NL (1990) Apical Na+ ion channels of tight epithelia as viewed from the perspective of noise analysis. In: Bronner F (ed) Current topics in membranes and transport. Channels and noise in epithelial tissues, vol 37, Academic Press, New York, pp 117–155Google Scholar
  11. 11.
    Helman SI, Cox TC, Van Driessche W (1983) Hormonal control of apical membrane Na+ transport in epithelia. J Gen Physiol 82:201–220Google Scholar
  12. 12.
    Hess P, Tsien RW (1984) Mechanism of ion permeation through calcium channels. Nature 309:453–456Google Scholar
  13. 13.
    Hoshiko T, Van Driessche W (1986) Effect of Na+ on amiloride and triamterene induced current fluctuations in isolated frog skin. J Gen Physiol 87:425–442Google Scholar
  14. 14.
    Martell AE, Smith RM (1974) Critical stability constants, vol 1. Amino acids. Plenum, New YorkGoogle Scholar
  15. 15.
    Preston AS, Muller J, Handler JS (1988) Dexamethasone accelerates differentiation of A6 epithelia and increases response to vasopressin. Am J Physiol 255:C661-C666Google Scholar
  16. 16.
    Rabito CA, Rotunno CA, Cereijido M (1978) Amiloride and calcium effect on the outer barrier of the frog skin. J Membr Biol 42:169–187Google Scholar
  17. 17.
    Tang J, Abramcheck FJ, Van Driessche W, Helman SI (1985) Electrophysiology and noise analysis of K+ depolarized epithelia of frog skin. Am J Physiol 249:C421-C429Google Scholar
  18. 18.
    Van Driessche W, Erlij D (1983) Noise analysis of inward and outward Na+ currents across the apical border of ouabain treated frog skin. Pflügers Arch 398:179–188Google Scholar
  19. 19.
    Van Driessche W, Lindemann B (1979) Concentration dependence of currents through single sodium-selective pores in frog skin. Nature 282:519–520Google Scholar
  20. 20.
    Van Driessche W, Zeiske W (1985) Ca2+ sensitive, spontaneously fluctuating, cation channels in the apical membrane of the adult frog skin epithelium. Pflügers Arch 405:250–259Google Scholar
  21. 21.
    Van Driessche W, Aelvoet I, Erlij D (1987) Oxytocin and cAMP stimulate monovalent cation movements through a Ca2+-sensitive, amiloride-insensitive channel in the apical membrane of toad urinary bladder. Proc Natl Acad Sci USA 84:313–317Google Scholar
  22. 22.
    Van Driessche W, Desmedt L, Simaels J (1991) Blockage of Na+ currents through poorly-selective cation channels in the apical membrane of frog skin and toad urinary bladder. Pflügers Arch 418:193–203Google Scholar
  23. 23.
    Wills NK, Millinoff LP (1990) Amiloride-sensitive Na+ transport across cultured renal (A6) epithelium: evidence for large currents and high Na: K selectivity. Pflügers Arch 416:481–492Google Scholar

Copyright information

© Springer-Verlag 1991

Authors and Affiliations

  • Luc Desmedt
    • 1
  • Jeannine Simaels
    • 1
  • Willy Van Driessche
    • 1
  1. 1.Laboratory for PhysiologyKULeuvenLeuvenBelgium

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