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. 1995 Jun 1;105(6):861–896. doi: 10.1085/jgp.105.6.861

The voltage-activated hydrogen ion conductance in rat alveolar epithelial cells is determined by the pH gradient

PMCID: PMC2216954  PMID: 7561747

Abstract

Voltage-activated H+ currents were studied in rat alveolar epithelial cells using tight-seal whole-cell voltage clamp recording and highly buffered, EGTA-containing solutions. Under these conditions, the tail current reversal potential, Vrev, was close to the Nernst potential, EH, varying 52 mV/U pH over four delta pH units (delta pH = pHo - pHi). This result indicates that H+ channels are extremely selective, PH/PTMA > 10(7), and that both internal and external pH, pHi, and pHo, were well controlled. The H+ current amplitude was practically constant at any fixed delta pH, in spite of up to 100-fold symmetrical changes in H+ concentration. Thus, the rate-limiting step in H+ permeation is pH independent, must be localized to the channel (entry, permeation, or exit), and is not bulk diffusion limitation. The instantaneous current- voltage relationship exhibited distinct outward rectification at symmetrical pH, suggesting asymmetry in the permeation pathway. Sigmoid activation kinetics and biexponential decay of tail currents near threshold potentials indicate that H+ channels pass through at least two closed states before opening. The steady state H+ conductance, gH, as well as activation and deactivation kinetic parameters were all shifted along the voltage axis by approximately 40 mV/U pH by changes in pHi or pHo, with the exception of the fast component of tail currents which was shifted less if at all. The threshold potential at which H+ currents were detectably activated can be described empirically as approximately 20-40(pHo-pHi) mV. If internal and external protons regulate the voltage dependence of gH gating at separate sites, then they must be equally effective. A simpler interpretation is that gating is controlled by the pH gradient, delta pH. We propose a simple general model to account for the observed delta pH dependence. Protonation at an externally accessible site stabilizes the closed channel conformation. Deprotonation of this site permits a conformational change resulting in the appearance of a protonation site, possibly the same one, which is accessible via the internal solution. Protonation of the internal site stabilizes the open conformation of the channel. In summary, within the physiological range of pH, the voltage dependence of H+ channel gating depends on delta pH and not on the absolute pH.

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Selected References

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  1. Akeson M., Deamer D. W. Proton conductance by the gramicidin water wire. Model for proton conductance in the F1F0 ATPases? Biophys J. 1991 Jul;60(1):101–109. doi: 10.1016/S0006-3495(91)82034-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Andersen O. S. Ion movement through gramicidin A channels. Studies on the diffusion-controlled association step. Biophys J. 1983 Feb;41(2):147–165. doi: 10.1016/S0006-3495(83)84416-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Barish M. E., Baud C. A voltage-gated hydrogen ion current in the oocyte membrane of the axolotl, Ambystoma. J Physiol. 1984 Jul;352:243–263. doi: 10.1113/jphysiol.1984.sp015289. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bernheim L., Krause R. M., Baroffio A., Hamann M., Kaelin A., Bader C. R. A voltage-dependent proton current in cultured human skeletal muscle myotubes. J Physiol. 1993 Oct;470:313–333. doi: 10.1113/jphysiol.1993.sp019860. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Byerly L., Meech R., Moody W., Jr Rapidly activating hydrogen ion currents in perfused neurones of the snail, Lymnaea stagnalis. J Physiol. 1984 Jun;351:199–216. doi: 10.1113/jphysiol.1984.sp015241. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Byerly L., Moody W. J. Membrane currents of internally perfused neurones of the snail, Lymnaea stagnalis, at low intracellular pH. J Physiol. 1986 Jul;376:477–491. doi: 10.1113/jphysiol.1986.sp016165. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Byerly L., Suen Y. Characterization of proton currents in neurones of the snail, Lymnaea stagnalis. J Physiol. 1989 Jun;413:75–89. doi: 10.1113/jphysiol.1989.sp017642. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Chung L. A., Lear J. D., DeGrado W. F. Fluorescence studies of the secondary structure and orientation of a model ion channel peptide in phospholipid vesicles. Biochemistry. 1992 Jul 21;31(28):6608–6616. doi: 10.1021/bi00143a035. [DOI] [PubMed] [Google Scholar]
  9. Ciani S., Krasne S., Miyazaki S., Hagiwara S. A model for anomalous rectification: electrochemical-potential-dependent gating of membrane channels. J Membr Biol. 1978 Dec 15;44(2):103–134. doi: 10.1007/BF01976035. [DOI] [PubMed] [Google Scholar]
  10. DeCoursey T. E., Cherny V. V. Na(+)-H+ antiport detected through hydrogen ion currents in rat alveolar epithelial cells and human neutrophils. J Gen Physiol. 1994 May;103(5):755–785. doi: 10.1085/jgp.103.5.755. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. DeCoursey T. E., Cherny V. V. Potential, pH, and arachidonate gate hydrogen ion currents in human neutrophils. Biophys J. 1993 Oct;65(4):1590–1598. doi: 10.1016/S0006-3495(93)81198-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. DeCoursey T. E., Cherny V. V. Voltage-activated hydrogen ion currents. J Membr Biol. 1994 Sep;141(3):203–223. doi: 10.1007/BF00235130. [DOI] [PubMed] [Google Scholar]
  13. DeCoursey T. E. Hydrogen ion currents in rat alveolar epithelial cells. Biophys J. 1991 Nov;60(5):1243–1253. doi: 10.1016/S0006-3495(91)82158-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. DeCoursey T. E., Jacobs E. R., Silver M. R. Potassium currents in rat type II alveolar epithelial cells. J Physiol. 1988 Jan;395:487–505. doi: 10.1113/jphysiol.1988.sp016931. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. DeCoursey T. E. State-dependent inactivation of K+ currents in rat type II alveolar epithelial cells. J Gen Physiol. 1990 Apr;95(4):617–646. doi: 10.1085/jgp.95.4.617. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Decker E. R., Levitt D. G. Use of weak acids to determine the bulk diffusion limitation of H+ ion conductance through the gramicidin channel. Biophys J. 1988 Jan;53(1):25–32. doi: 10.1016/S0006-3495(88)83062-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Demaurex N., Grinstein S., Jaconi M., Schlegel W., Lew D. P., Krause K. H. Proton currents in human granulocytes: regulation by membrane potential and intracellular pH. J Physiol. 1993 Jul;466:329–344. [PMC free article] [PubMed] [Google Scholar]
  18. Gadsby D. C., Rakowski R. F., De Weer P. Extracellular access to the Na,K pump: pathway similar to ion channel. Science. 1993 Apr 2;260(5104):100–103. doi: 10.1126/science.7682009. [DOI] [PubMed] [Google Scholar]
  19. Gutknecht J. Proton conductance through phospholipid bilayers: water wires or weak acids? J Bioenerg Biomembr. 1987 Oct;19(5):427–442. doi: 10.1007/BF00770028. [DOI] [PubMed] [Google Scholar]
  20. Gutknecht J., Tosteson D. C. Diffusion of weak acids across lipid bilayer membranes: effects of chemical reactions in the unstirred layers. Science. 1973 Dec 21;182(4118):1258–1261. doi: 10.1126/science.182.4118.1258. [DOI] [PubMed] [Google Scholar]
  21. Hagiwara S., Takahashi K. The anomalous rectification and cation selectivity of the membrane of a starfish egg cell. J Membr Biol. 1974;18(1):61–80. doi: 10.1007/BF01870103. [DOI] [PubMed] [Google Scholar]
  22. Haines T. H. Anionic lipid headgroups as a proton-conducting pathway along the surface of membranes: a hypothesis. Proc Natl Acad Sci U S A. 1983 Jan;80(1):160–164. doi: 10.1073/pnas.80.1.160. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Hamill O. P., Marty A., Neher E., Sakmann B., Sigworth F. J. Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membrane patches. Pflugers Arch. 1981 Aug;391(2):85–100. doi: 10.1007/BF00656997. [DOI] [PubMed] [Google Scholar]
  24. Henderson L. M., Chappell J. B. The NADPH-oxidase-associated H+ channel is opened by arachidonate. Biochem J. 1992 Apr 1;283(Pt 1):171–175. doi: 10.1042/bj2830171. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Heyer R. J., Muller R. U., Finkelstein A. Inactivation of monazomycin-induced voltage-dependent conductance in thin lipid membranes. II. Inactivation produced by monazomycin transport through the membrane. J Gen Physiol. 1976 Jun;67(6):731–748. doi: 10.1085/jgp.67.6.731. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Hilgemann D. W. Channel-like function of the Na,K pump probed at microsecond resolution in giant membrane patches. Science. 1994 Mar 11;263(5152):1429–1432. doi: 10.1126/science.8128223. [DOI] [PubMed] [Google Scholar]
  27. Junge W. Protons, the thylakoid membrane, and the chloroplast ATP synthase. Ann N Y Acad Sci. 1989;574:268–286. doi: 10.1111/j.1749-6632.1989.tb25164.x. [DOI] [PubMed] [Google Scholar]
  28. Kapus A., Romanek R., Grinstein S. Arachidonic acid stimulates the plasma membrane H+ conductance of macrophages. J Biol Chem. 1994 Feb 18;269(7):4736–4745. [PubMed] [Google Scholar]
  29. Kapus A., Romanek R., Qu A. Y., Rotstein O. D., Grinstein S. A pH-sensitive and voltage-dependent proton conductance in the plasma membrane of macrophages. J Gen Physiol. 1993 Oct;102(4):729–760. doi: 10.1085/jgp.102.4.729. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Kasianowicz J., Benz R., McLaughlin S. How do protons cross the membrane-solution interface? Kinetic studies on bilayer membranes exposed to the protonophore S-13 (5-chloro-3-tert-butyl-2'-chloro-4' nitrosalicylanilide). J Membr Biol. 1987;95(1):73–89. doi: 10.1007/BF01869632. [DOI] [PubMed] [Google Scholar]
  31. Kempf C., Klausner R. D., Weinstein J. N., Van Renswoude J., Pincus M., Blumenthal R. Voltage-dependent trans-bilayer orientation of melittin. J Biol Chem. 1982 Mar 10;257(5):2469–2476. [PubMed] [Google Scholar]
  32. Krishnamoorthy G. Temperature jump as a new technique to study the kinetics of fast transport of protons across membranes. Biochemistry. 1986 Oct 21;25(21):6666–6671. doi: 10.1021/bi00369a051. [DOI] [PubMed] [Google Scholar]
  33. Mathias R. T., Cohen I. S., Oliva C. Limitations of the whole cell patch clamp technique in the control of intracellular concentrations. Biophys J. 1990 Sep;58(3):759–770. doi: 10.1016/S0006-3495(90)82418-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. McGuigan J. A., Lüthi D., Buri A. Calcium buffer solutions and how to make them: a do it yourself guide. Can J Physiol Pharmacol. 1991 Nov;69(11):1733–1749. doi: 10.1139/y91-257. [DOI] [PubMed] [Google Scholar]
  35. Meech R. W., Thomas R. C. Voltage-dependent intracellular pH in Helix aspersa neurones. J Physiol. 1987 Sep;390:433–452. doi: 10.1113/jphysiol.1987.sp016710. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Nachliel E., Gutman M. Kinetic analysis of proton transfer between reactants adsorbed to the same micelle. The effect of proximity on the rate constants. Eur J Biochem. 1984 Aug 15;143(1):83–88. doi: 10.1111/j.1432-1033.1984.tb08344.x. [DOI] [PubMed] [Google Scholar]
  37. Nagle J. F., Dilley R. A. Models of localized energy coupling. J Bioenerg Biomembr. 1986 Feb;18(1):55–64. doi: 10.1007/BF00743612. [DOI] [PubMed] [Google Scholar]
  38. Nagle J. F., Morowitz H. J. Molecular mechanisms for proton transport in membranes. Proc Natl Acad Sci U S A. 1978 Jan;75(1):298–302. doi: 10.1073/pnas.75.1.298. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Nagle J. F. Theory of passive proton conductance in lipid bilayers. J Bioenerg Biomembr. 1987 Oct;19(5):413–426. doi: 10.1007/BF00770027. [DOI] [PubMed] [Google Scholar]
  40. Nagle J. F., Tristram-Nagle S. Hydrogen bonded chain mechanisms for proton conduction and proton pumping. J Membr Biol. 1983;74(1):1–14. doi: 10.1007/BF01870590. [DOI] [PubMed] [Google Scholar]
  41. Nunogaki K., Kasai M. The H+/OH- flux localizes around the channel mouth in buffered solution. J Theor Biol. 1988 Oct 7;134(3):403–415. doi: 10.1016/s0022-5193(88)80070-5. [DOI] [PubMed] [Google Scholar]
  42. Pennefather P. S., DeCoursey T. E. A scheme to account for the effects of Rb+ and K+ on inward rectifier K channels of bovine artery endothelial cells. J Gen Physiol. 1994 Apr;103(4):549–581. doi: 10.1085/jgp.103.4.549. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Pennefather P., Oliva C., Mulrine N. Origin of the potassium and voltage dependence of the cardiac inwardly rectifying K-current (IK1). Biophys J. 1992 Feb;61(2):448–462. doi: 10.1016/S0006-3495(92)81850-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Peskoff A., Bers D. M. Electrodiffusion of ions approaching the mouth of a conducting membrane channel. Biophys J. 1988 Jun;53(6):863–875. doi: 10.1016/S0006-3495(88)83167-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Prats M., Tocanne J. F., Teissie J. Lateral proton conduction at a lipid/water interface. Effect of lipid nature and ionic content of the aqueous phase. Eur J Biochem. 1987 Jan 15;162(2):379–385. doi: 10.1111/j.1432-1033.1987.tb10612.x. [DOI] [PubMed] [Google Scholar]
  46. Raymond L., Slatin S. L., Finkelstein A., Liu Q. R., Levinthal C. Gating of a voltage-dependent channel (colicin E1) in planar lipid bilayers: translocation of regions outside the channel-forming domain. J Membr Biol. 1986;92(3):255–268. doi: 10.1007/BF01869394. [DOI] [PubMed] [Google Scholar]
  47. Saigusa A., Matsuda H. Outward currents through the inwardly rectifying potassium channel of guinea-pig ventricular cells. Jpn J Physiol. 1988;38(1):77–91. doi: 10.2170/jjphysiol.38.77. [DOI] [PubMed] [Google Scholar]
  48. Scheiner S., Hillenbrand E. A. Modification of pK values caused by change in H-bond geometry. Proc Natl Acad Sci U S A. 1985 May;82(9):2741–2745. doi: 10.1073/pnas.82.9.2741. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Schulten Z., Schulten K. A model for the resistance of the proton channel formed by the proteolipid of ATPase. Eur Biophys J. 1985;11(3):149–155. doi: 10.1007/BF00257393. [DOI] [PubMed] [Google Scholar]
  50. Siczkowski M., Davies J. E., Ng L. L. Activity and density of the Na+/H+ antiporter in normal and transformed human lymphocytes and fibroblasts. Am J Physiol. 1994 Sep;267(3 Pt 1):C745–C752. doi: 10.1152/ajpcell.1994.267.3.C745. [DOI] [PubMed] [Google Scholar]
  51. Silver M. R., DeCoursey T. E. Intrinsic gating of inward rectifier in bovine pulmonary artery endothelial cells in the presence or absence of internal Mg2+. J Gen Physiol. 1990 Jul;96(1):109–133. doi: 10.1085/jgp.96.1.109. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Stoeckenius W., Lozier R. H., Bogomolni R. A. Bacteriorhodopsin and the purple membrane of halobacteria. Biochim Biophys Acta. 1979 Mar 14;505(3-4):215–278. doi: 10.1016/0304-4173(79)90006-5. [DOI] [PubMed] [Google Scholar]
  53. Thomas R. C., Meech R. W. Hydrogen ion currents and intracellular pH in depolarized voltage-clamped snail neurones. Nature. 1982 Oct 28;299(5886):826–828. doi: 10.1038/299826a0. [DOI] [PubMed] [Google Scholar]
  54. Wagner R., Apley E. C., Hanke W. Single channel H+ currents through reconstituted chloroplast ATP synthase CF0-CF1. EMBO J. 1989 Oct;8(10):2827–2834. doi: 10.1002/j.1460-2075.1989.tb08429.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Woolley G. A., Wallace B. A. Model ion channels: gramicidin and alamethicin. J Membr Biol. 1992 Aug;129(2):109–136. doi: 10.1007/BF00219508. [DOI] [PubMed] [Google Scholar]
  56. Zagotta W. N., Hoshi T., Dittman J., Aldrich R. W. Shaker potassium channel gating. II: Transitions in the activation pathway. J Gen Physiol. 1994 Feb;103(2):279–319. doi: 10.1085/jgp.103.2.279. [DOI] [PMC free article] [PubMed] [Google Scholar]

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