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The Journal of Membrane Biology

, Volume 97, Issue 2, pp 137–159 | Cite as

Ion permeation through single channels activated by acetylcholine in denervated toad sartorius skeletal muscle fibers: Effects of alkali cations

  • Nino Quartararo
  • Peter H. Barry
  • Peter W. Gage
Articles

Summary

The gigaohm seal technique was used to study ion permeation through acetylcholine-activated channels in cell-attached patches of the extrajunctional membrane of chronically denervated, enzyme-treated cells from the sartorius muscle of the toadBufo marinus. The most frequently occurring channel type (>95% of channel openings), provisionally classified as ‘extrajunctional,’ had a chord conductance of approximately 25 pS under normal conditions (−70 mV, 11°C, Normal Toad Ringer's). The less frequently observed channel type (<5% of channel openings), classified as a ‘junctional’ type, had a conductance of 35 pS under the same conditions, and a similar null potential. In many patches, a small percentage (usually <2%) of openings of the extrajunctional channel displayed a lower conductance state. The shape of theI–V curves obtained for the extrajunctional channel dependend on the predominant extracellular cation. For Cs and K, theI–V curves were essentially linear over the voltage range +50 to −150 mV across the patch, suggesting that the potential independent component of the energy profile within the channel was symmetrical. For Li, theI–V curve was very nonlinear, displaying a significant sublinearity at hyperpolarized potentials. Both an electrodiffusion and a symmetrical uniform four-barrier, three-site rate-theory model provided reasonable fits to the data, whereas symmetrical two-barrier, single-site rate-theory models did not. For the alkali cations examined, the relative permeability sequence wasPCs>PK>PNa>PLi—a “proportional” selectivity sequence. This was different from the single channel conductance sequence which was found to beγK>γCs>γNa>γLi implying that ions do not move independently through the channel. The relative binding constant sequence for the channel sites was found to be a “polarizability” sequence, i.e.,KLi>KCs>KNa>KK There was an inverse relationship for the cations examined. Under conditions when the single-channel conductance was relatively high, the conductance at depolarized potentials was lower than that predicted by both electrodiffusion and rate theory models, suggesting that there was a rate-limiting access step for ions, from the intracellular compartment into the channel.

Key Words

ions ion permeation ion selectivity channels single channels ACh channels alkali cations gigaohm seal technique patch clamp skeletal muscle electrodiffusion rate theory 

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References

  1. Adams, D.J., Dwyer, T.M., Hille, B. 1980. The permeability of end-plate channels to monovalent and divalent metal cations.J. Gen. Physiol. 75:493–510Google Scholar
  2. Adams, D.J., Nonner, W., Dwyer, T.M., Hille, B. 1981. Block of end-plate channels by permeant cations in frog skeletal muscles.J. Gen. Physiol. 78:593–615Google Scholar
  3. Andersen, O.S. 1983. Ion movement through Gramicidin A channels. Studies on the diffusion-controlled association step.Biophys. J. 41:147–165Google Scholar
  4. Anderson, C.R., Stevens, C.F. 1973. Voltage-clamp analysis of acetylcholine produced end-plate current fluctuations at the neuromuscular junction.J. Physiol. (London) 235:655–691Google Scholar
  5. Auerbach, A., Sachs, F. 1984. Patch clamp studies of single ionic channels.Annu. Rev. Biophys. Bioeng. 13:269–302Google Scholar
  6. Ayer, R.K., Jr., De Haan, R.L., Fischmeister, R. 1983. Measurement of membrane patch andseal resistance with two patch electrodes in chick embryo cardiac cells.J. Physiol. (London) 345:29PGoogle Scholar
  7. Barford, N.C. 1967. Experimental Measurements: Precision, Error and Truth. Addison-Wesley, LondonGoogle Scholar
  8. Barry, P.H., Diamond, J.M. 1970. Junction potentials, electrode standard potentials, and other problems in interpreting electrical properties of membranes.J. Membrane Biol. 3:93–122Google Scholar
  9. Barry, P.H., Gage, P.W. 1984. Ionic selectivity of channels at the end-plate.In: Ion Channels: Molecular and Physiological Aspects. W.D. Stein, editor. Ch. 1, pp. 1–51. Academic, New YorkGoogle Scholar
  10. Barry, P.H., Gage, P.W., Van Helden, D.F. 1979. Cation permeation at the amphibian motor end-plate.J. Membrane Biol. 45:245–276Google Scholar
  11. Camardo, J.S., Siegelbaum, S.A. 1983. Single channel analysis inAplysia neurones. A specific K+ channel is modulated by serotonin and cyclic AMP.In: Single Channel Recording. B. Sakmann and E. Neher, editors. Ch. 21, pp. 409–423. Plenum, New YorkGoogle Scholar
  12. Corey, D.P., Stevens, C.F. 1983. Science and technology of patch-recording electrodes.In: Single-Channel Recording. B. Sakmann and E. Neher, editors. Ch. 3 pp. 53–68. Plenum, New YorkGoogle Scholar
  13. Coronado, R., Rosenberg, R.L., Miller, C. 1980. Ionic selectivity, saturation and block in a K+-selective channel from sarcoplasmic reticulum.J. Gen. Physiol. 76:425–446Google Scholar
  14. Dulhunty, A.F., Gage, P.W. 1973. Electrical properties of toad sartorius muscle fibers in summer and winter.J. Physiol. (London) 230:619–641Google Scholar
  15. Dwyer, T.M. 1986. Guanidine block of single channel currents activated by acetylcholine.J. Gen. Physiol. 88:635–650Google Scholar
  16. Dwyer, T.M., Adams, D.J., Hille, B. 1980. The permeability of end-plate channels to organic cations in frog muscle.J. Gen. Physiol. 75:469–492Google Scholar
  17. Dwyer, T.M., Farley, J.M. 1984. Permeability properties of chick myotube acetylcholine-activated channels.Biophys. J. 45:529–539Google Scholar
  18. Eisenman, G., Horn, R. 1983. Ionic selectivity revisited: The role of kinetic and equilibrium processes in ion permeation through channels.J. Membrane Biol. 76:197–225Google Scholar
  19. Eyring, H. 1935. The activated complex in chemical reactions.J. Chem. Phys. 3:107–115Google Scholar
  20. Fenwick, E.M., Marty, A., Neher, E. 1982. A patch clamp study of bovine chromaffin cells and of their sensitivity to acetylcholine.J. Physiol. (London) 331:577–597Google Scholar
  21. Gage, P.W., Hamill, O.P. 1980. Lifetime and conductance of acetylcholine-activated channels in normal and denervated toad sartorius muscle.J. Physiol. (London) 298:525–538Google Scholar
  22. Gage, P.W., Van Helden, D.F. 1979. Effects of permeant monovalent cations on end-plate channels.J. Physiol. (london) 288:509–528Google Scholar
  23. Gagne, S., Plamondon, R. 1983. Tip potential of open tip glass microelectrodes: Theoretical and experimental studies.Can. J. Physiol. Pharmacol. 61:857–869Google Scholar
  24. Gardener, P., Ogden, D.C., Colquhoun, D. 1984. Conductances of single ion channels opened by nicotinic agonists are indistinguishable.Nature (London) 309:160–162Google Scholar
  25. Glasstone, S., Laidler, K.J., Eyring, H. 1941. The Theory of Rate Processes. McGraw-Hill, New YorkGoogle Scholar
  26. Goldman, D. 1943. Potential, impedance and rectification in membranes.J. Gen. Physiol. 27:37–60Google Scholar
  27. Hamill, O.P., 1983. Membrane ion channels.In: Topics in Molecular Pharmacology. A.S.V. Burgen and G.C.K. Roberts, editors. pp. 181–205. Elsevier Science Publishers, AmsterdamGoogle Scholar
  28. Hamill, O.P., Marty, A., Neher, E., Sakmann, B., Sigworth, F.J. 1981. Improved patch-clamp techniques for high resolution current recording from cells and cell free membrane patches.Pfluegers Arch. 391:85–100Google Scholar
  29. Hamill, O.P., Sakmann, B. 1981. Multiple conductance states of single acetylcholine receptor channels in embryonic muscle cells.Nature (London) 294:462–464Google Scholar
  30. Hille, B. 1975a. Ionic selectivity, saturation and block in sodium channels. A four-barrier model.J. Gen. Physiol. 66:535–560Google Scholar
  31. Hille, B. 1975b. Ionic selectivity of Na and K channels of nerve membranes.In: Membranes—A Series of Advances. G. Eisenman, editor. Vol. 3, Ch. 4, pp. 255–323. Marcel Dekker, New YorkGoogle Scholar
  32. Hodgkin, A.L., Katz, B. 1949. The effects of sodium ions on the electrical activity of the giant axon of the squid.J. Physiol. (London) 108:37–77Google Scholar
  33. Horn, R., Brodwick, M.S. 1980. Acetylcholine-induced current in perfused rat myoballs.J. Gen. Physiol. 75:297–321Google Scholar
  34. Horn, R., Patlak, J. 1980. Single channel currents from excised patches of muscle membranes.Proc. Natl. Acad. Sci. USA 77:6930–6934Google Scholar
  35. Horn, R., Stevens, C.F. 1980. Relation between structure and function of ion channels.Com. Mol. Cell Biophys. 1:57–68Google Scholar
  36. Labarca, P., Lindstrom, J., Montal, M. 1984. Acetylcholine receptors in planar lipid bilayers.J. Gen. Physiol. 83:473–496Google Scholar
  37. Lev, A.A., Armstrong, W. McD. 1975. Ion activities in cells.Curr. Top. Membr. Transp. 6:59–123Google Scholar
  38. Lewis, C.A. 1979. Ion concentration dependence of the reversal potential and the single channel conductance of ion channels at the frog neuromuscular junction.J. Physiol. (London) 286:417–445Google Scholar
  39. Lewis, C.A., Stevens, C.F. 1979. Mechanisms of ion permeation through channels in a post-synaptic membrane.In: Membrane Transport Processes. C.F. Stevens and R.W. Tsien, editors. pp. 133–151. Raven, New YorkGoogle Scholar
  40. Lewis, C.A., Stevens, C.F. 1983. Acetylcholine receptor channel ionic selectivity: Ions experience an aqueous environment.Proc. Natl. Acad. Sci. USA 80:6110–6113Google Scholar
  41. MacInnes, D.A. 1961. The Principles of Electrochemistry. Dover, New YorkGoogle Scholar
  42. Maeno, T., Edwards, C., Anraku, M. 1977. Permeability of end-plate membrane activated by acetylcholine to some organic cations.J. Neurobiol. 8:173–184Google Scholar
  43. Maruyama, Y., Petersen, O.H. 1982. Cholecystokinin activation of single channel currents is mediated by internal messenger in pancreatic acinar cells.Nature (London) 300:61–63Google Scholar
  44. Mullins, L.J. 1975. Ion selectivity of carriers and channels.Biophys. J. 15:921–931Google Scholar
  45. Neher, E., Sakmann, B., Steinbach, J.H. 1978. The extracellular patch clamp: A method for resolving currents through individual open channels in biological membranes.Pfluegers Arch. 375:219–230Google Scholar
  46. Purves, R.D. 1981. Microelectrode Methods for Intracellular Recording and Ionophoresis. Academic, LondonGoogle Scholar
  47. Rae, J.L., Levis, R.A. 1984. Patch voltage clamp of lens epithelial cells: Theory and practice.Mol. Physiol. 6:115–162Google Scholar
  48. Redmann, G.A., Clark, R.B., Adams, P.R. 1982. Single acetylcholine channel block in elevated sodium and lithium.Biophys. J. 37:324aGoogle Scholar
  49. Robinson, R.A., Stokes, R.H. 1965. Electrolyte Solutions (2nd Ed.) Butterworth, LondonGoogle Scholar
  50. Sakmann, B., Neher, E. 1983. Geometric parameters of pipettes and membrane patches.In: Single-Channel Recording. B. Sakmann and E. Neher, editors. Ch. 2, pp. 37–51. Plenum, New YorkGoogle Scholar
  51. Sanchez, J.A., Dani, J.A., Siemen, D., Hille, B. 1986. Slow permeation of organic cations in acetylcholine receptor channels.J. Gen. Physiol. 87:985–1001Google Scholar
  52. Siegelbaum, S.A., Camardo, J.S., Kandel, E.R. 1982. Serotonin and cyclic AMP close single K+ channels inAplysia sensory neurones.Nature (London) 299:413–417Google Scholar
  53. Sine, S.M., Steinbach, J.H. 1984a. Activation of a nicotinic acetylcholine receptor.Biophys. J. 45:175–185Google Scholar
  54. Sine, S.M., Steinbach, J.H. 1984b. Agonists block currents through acetylcholine receptor channels.Biophys. J. 46:277–284Google Scholar
  55. Takeda, K., Barry, P.H., Gage, P.W. 1980. Effects of ammonium ions on end-plate channels.J. Gen. Physiol. 75:589–613Google Scholar
  56. Takeda, K., Barry, P.H., Gage, P.W. 1982a. Effects of extracellular sodium concentrations on null potential, conductance and open time of end-plate channels.Proc. R. Soc. London B 216:225–251Google Scholar
  57. Takeda, K., Gage, P.W., Barry, P.H. 1982b. Effects of divalent cations on toad end-plate channels.J. Membrane Biol. 64:55–66Google Scholar
  58. Takeuchi, A., Takeuchi, N. 1959. Active phase of frog's end-plate potential.J. Neurophysiol. 22:395–411Google Scholar
  59. Takeuchi, A., Takeuchi, N. 1960. On the permeability of end-plate membrane during the action of transmitter.J. Physiol. (London) 154:52–67Google Scholar
  60. Van Helden, D.F., Hamill, O.P., Gage, P.W. 1977. Permeant cations alter end-plate channel characteristics.Nature (London) 269:711–713Google Scholar
  61. Woodbury, J.W. 1971. Eyring rate-theory model of the currentvoltage relationships of ion channels in excitable membranes.In: Chemical Dynamics: Papers in Honor of Henry Eyring. J.O. Hirschfelder, editor. pp. 601–617. Wiley, New YorkGoogle Scholar

Copyright information

© Springer-Verlag New York Inc 1987

Authors and Affiliations

  • Nino Quartararo
    • 1
  • Peter H. Barry
    • 1
  • Peter W. Gage
    • 1
  1. 1.School of Physiology and PharmacologyUniversity of New South WalesSydneyAustralia

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