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. 1980 Aug;305:215–234. doi: 10.1113/jphysiol.1980.sp013359

Voltage clamp measurements of sodium channel properties in rabbit cardiac Purkinje fibres.

T J Colatsky
PMCID: PMC1282968  PMID: 6255144

Abstract

1. Voltage clamp studies of the excitatory sodium current, INa, were carried out in rabbit cardiac Purkinje fibres using th two-micro-electrode technique. Previous work has shown the rabbit Purkinje fibre to have relatively simple morphology (Sommer & Johnson, 1968) and electrical structure (Colatsky & Tsien, 1979a) compared to other cardiac preparations. 2. Non-uniformities in membrane potential were kept small by reducing the size of INa to less than 50 microA/cm2 of total membrane surface area through prepulse inactivation or removal of external sodium, Nao. Temporal resolution was improved by cooling to 10-26 degrees C. These adjustments did not greatly alter the measured properties of the sodium channel. 3. Under these conditions, sodium currents were recorded satisfying a number of criteria for adequate voltage control. Direct measurement of longitudinal non-uniformity using a second voltage electrode showed only small deviations at the time of peak current. 4. The properties of the sodium channel were examined using conventional protocols. Both peak sodium permeability, PNa, and steady-state sodium inactivation, h infinity, showed a sigmoidal dependence on membrane potential. PNa rose steeply with small depolarizations, increasing roughly e-fold per 3.2 mV, and reaching half-maximal activation at -30 +/- 2 mV. The h infinity -V curve had a midpoint of -74.9 +/- 2 mV and a reciprocal slope of 4.56 +/- 0.13 mV at temperatures of 10-19.5 degrees C, and showed a dependence on temperature, shifting to more negative potentials with cooling (approximately 3 mV/10 degrees C). Recovery of INa from inactivation in double pulse experiments followed a single exponential time course with time constants of 108-200 msec at 19 degrees C for holding potentials near -80 mV. No attempt was made to describe the activation kinetics because of uncertainties about the early time course of the current. 5. These data predict a maximum duration for INa of less than 1-2 msec and a maximum peak current density of about 500 microA/cm2 under physiological conditions, i.e. 37 degrees C and 150 mM-Nao. This current magnitude is sufficient to discharge the membrane capacitance at rates comparable to those measured experimentally (311 +/- 27 V/sec, Colatsky & Tsien, 1979a). 6. The limitations of the method are discussed. The major problem is the longitudinal cable delay which limits the speed of voltage control. This makes it difficult to separate the activation of INa from the decay of the capacity transient for potentials positive to -15 mV. 7. It is concluded that the approach described is valid for measurements of sodium currents in the potential range where action potentials are initiated, making it possible to study cardiac sodium channels in an adult mammalian preparation which is free of enzymatic treatment.

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Selected References

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  1. Adrian R. H., Almers W. Membrane capacity measurements on frog skeletal muscle in media of low ion content. J Physiol. 1974 Mar;237(3):573–605. doi: 10.1113/jphysiol.1974.sp010499. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Adrian R. H., Chandler W. K., Hodgkin A. L. Voltage clamp experiments in striated muscle fibres. J Physiol. 1970 Jul;208(3):607–644. doi: 10.1113/jphysiol.1970.sp009139. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Adrian R. H., Marshall M. W. Sodium currents in mammalian muscle. J Physiol. 1977 Jun;268(1):223–250. doi: 10.1113/jphysiol.1977.sp011855. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Attwell D., Cohen I., Eisner D., Ohba M., Ojeda C. The steady state TTX-sensitive ("window") sodium current in cardiac Purkinje fibres. Pflugers Arch. 1979 Mar 16;379(2):137–142. doi: 10.1007/BF00586939. [DOI] [PubMed] [Google Scholar]
  5. Attwell D., Cohen I. The voltage clamp of multicellular preparations. Prog Biophys Mol Biol. 1977;31(3):201–245. doi: 10.1016/0079-6107(78)90009-3. [DOI] [PubMed] [Google Scholar]
  6. Baer M., Best P. M., Reuter H. Voltage-dependent action of tetrodotoxin in mammalian cardiac muscle. Nature. 1976 Sep 23;263(5575):344–345. doi: 10.1038/263344a0. [DOI] [PubMed] [Google Scholar]
  7. Baumgarten C. M., Isenberg G., McDonald T. F., Ten Eick R. E. Depletion and accumulation of potassium in the extracellular clefts of cardiac Purkinje fibers during voltage clamp hyperpolarization and depolarization: experiments in sodium-free bathing media. J Gen Physiol. 1977 Aug;70(2):149–169. doi: 10.1085/jgp.70.2.149. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Beeler G. W., Jr, Reuter H. Voltage clamp experiments on ventricular myocarial fibres. J Physiol. 1970 Mar;207(1):165–190. doi: 10.1113/jphysiol.1970.sp009055. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Beeler G. W., McGuigan J. A. Voltage clamping of multicellular myocardial preparations: capabilities and limitations of existing methods. Prog Biophys Mol Biol. 1978;34(3):219–254. doi: 10.1016/0079-6107(79)90019-1. [DOI] [PubMed] [Google Scholar]
  10. CORABOEUF E., WEIDMANN S. Temperature effects on the electrical activity of Purkinje fibres. Helv Physiol Pharmacol Acta. 1954;12(1):32–41. [PubMed] [Google Scholar]
  11. Campbell D. T., Hille B. Kinetic and pharmacological properties of the sodium channel of frog skeletal muscle. J Gen Physiol. 1976 Mar;67(3):309–323. doi: 10.1085/jgp.67.3.309. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Chandler W. K., Meves H. Rate constants associated with changes in sodium conductance in axons perfused with sodium fluoride. J Physiol. 1970 Dec;211(3):679–705. doi: 10.1113/jphysiol.1970.sp009299. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Chandler W. K., Meves H. Voltage clamp experiments on internally perfused giant axons. J Physiol. 1965 Oct;180(4):788–820. doi: 10.1113/jphysiol.1965.sp007732. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Chiu S. Y., Mrose H. E., Ritchie J. M. Anomalous temperature dependence of the sodium conductance in rabbit nerve compared with frog nerve. Nature. 1979 May 24;279(5711):327–328. doi: 10.1038/279327a0. [DOI] [PubMed] [Google Scholar]
  15. Colatsky J. J., Tsien R. W. Sodium channels in rabbit cardiac Purkinje fibres. Nature. 1979 Mar 15;278(5701):265–268. doi: 10.1038/278265a0. [DOI] [PubMed] [Google Scholar]
  16. Colatsky T. J., Tsien R. W. Electrical properties associated with wide intercellular clefts in rabbit Purkinje fibres. J Physiol. 1979 May;290(2):227–252. doi: 10.1113/jphysiol.1979.sp012769. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Connor J., Barr L., Jakobsson E. Electrical characteristics of frog atrial trabeculae in the double sucrose gap. Biophys J. 1975 Oct;15(10):1047–1067. doi: 10.1016/S0006-3495(75)85882-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Coraboeuf E., Deroubaix E., Coulombe A. Effect of tetrodotoxin on action potentials of the conducting system in the dog heart. Am J Physiol. 1979 Apr;236(4):H561–H567. doi: 10.1152/ajpheart.1979.236.4.H561. [DOI] [PubMed] [Google Scholar]
  19. DECK K. A., KERN R., TRAUTWEIN W. VOLTAGE CLAMP TECHNIQUE IN MAMMALIAN CARDIAC FIBRES. Pflugers Arch Gesamte Physiol Menschen Tiere. 1964 Jun 9;280:50–62. doi: 10.1007/BF00412615. [DOI] [PubMed] [Google Scholar]
  20. DECK K. A., TRAUTWEIN W. IONIC CURRENTS IN CARDIAC EXCITATION. Pflugers Arch Gesamte Physiol Menschen Tiere. 1964 Jun 9;280:63–80. doi: 10.1007/BF00412616. [DOI] [PubMed] [Google Scholar]
  21. DRAPER M. H., WEIDMANN S. Cardiac resting and action potentials recorded with an intracellular electrode. J Physiol. 1951 Sep;115(1):74–94. doi: 10.1113/jphysiol.1951.sp004653. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. DiFrancesco D., McNaughton P. A. The effects of calcium on outward membrane currents in the cardiac Purkinje fibre. J Physiol. 1979 Apr;289:347–373. doi: 10.1113/jphysiol.1979.sp012741. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Dudel J., Peper K., Rüdel R., Trautwein W. Excitatory membrane current in heart muscle (Purkinje fibers). Pflugers Arch Gesamte Physiol Menschen Tiere. 1966;292(3):255–273. doi: 10.1007/BF00362740. [DOI] [PubMed] [Google Scholar]
  24. Dudel J., Peper K., Rüdel R., Trautwein W. The effect of tetrodotoxin on the membrane current in cardiac muscle (Purkinje fibers). Pflugers Arch Gesamte Physiol Menschen Tiere. 1967;295(3):213–226. doi: 10.1007/BF01844101. [DOI] [PubMed] [Google Scholar]
  25. Dudel J., Rüdel R. Voltage and time dependence of excitatory sodium current in cooled sheep Purkinje fibres. Pflugers Arch. 1970;315(2):136–158. doi: 10.1007/BF00586657. [DOI] [PubMed] [Google Scholar]
  26. Ellis D. The effects of external cations and ouabain on the intracellular sodium activity of sheep heart Purkinje fibres. J Physiol. 1977 Dec;273(1):211–240. doi: 10.1113/jphysiol.1977.sp012090. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Fabiato A., Fabiato F. Calcium release from the sarcoplasmic reticulum. Circ Res. 1977 Feb;40(2):119–129. doi: 10.1161/01.res.40.2.119. [DOI] [PubMed] [Google Scholar]
  28. Fozzard H. A., Beeler G. W., Jr The voltage clamp and cardiac electrophysiology. Circ Res. 1975 Oct;37(4):403–413. doi: 10.1161/01.res.37.4.403. [DOI] [PubMed] [Google Scholar]
  29. Fozzard H. A. Membrane capacity of the cardiac Purkinje fibre. J Physiol. 1966 Jan;182(2):255–267. doi: 10.1113/jphysiol.1966.sp007823. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Gadsby D. C., Cranefield P. F. Two levels of resting potential in cardiac Purkinje fibers. J Gen Physiol. 1977 Dec;70(6):725–746. doi: 10.1085/jgp.70.6.725. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Gettes L. S., Reuter H. Slow recovery from inactivation of inward currents in mammalian myocardial fibres. J Physiol. 1974 Aug;240(3):703–724. doi: 10.1113/jphysiol.1974.sp010630. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Goldman D. E. POTENTIAL, IMPEDANCE, AND RECTIFICATION IN MEMBRANES. J Gen Physiol. 1943 Sep 20;27(1):37–60. doi: 10.1085/jgp.27.1.37. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. HODGKIN A. L., HUXLEY A. F., KATZ B. Measurement of current-voltage relations in the membrane of the giant axon of Loligo. J Physiol. 1952 Apr;116(4):424–448. doi: 10.1113/jphysiol.1952.sp004716. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. HODGKIN A. L., KATZ B. The effect of sodium ions on the electrical activity of giant axon of the squid. J Physiol. 1949 Mar 1;108(1):37–77. doi: 10.1113/jphysiol.1949.sp004310. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Haas H. G., Kern R., Einwächter H. M., Tarr M. Kinetics of Na inactivation in frog atria. Pflugers Arch. 1971;323(2):141–157. doi: 10.1007/BF00586445. [DOI] [PubMed] [Google Scholar]
  36. Harrington L., Johnson E. A. Voltage clamp of cardiac muscle in a double sucrose gap. A feasibility study. Biophys J. 1973 Jul;13(7):626–647. doi: 10.1016/S0006-3495(73)86013-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Iijima T., Pappano A. J. Ontogenetic increase of the maximal rate of rise of the chick embryonic heart action potential. Relationship to voltage, time, and tetrodotoxin. Circ Res. 1979 Mar;44(3):358–367. doi: 10.1161/01.res.44.3.358. [DOI] [PubMed] [Google Scholar]
  38. Johnson E. A., Lieberman M. Heart: excitation and contraction. Annu Rev Physiol. 1971;33:479–532. doi: 10.1146/annurev.ph.33.030171.002403. [DOI] [PubMed] [Google Scholar]
  39. Johnson E. A., Sommer J. R. A strand of cardiac muscle. Its ultrastructure and the electrophysiological implications of its geometry. J Cell Biol. 1967 Apr;33(1):103–129. doi: 10.1083/jcb.33.1.103. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Kass R. S., Siegelbaum S. A., Tsien R. W. Three-micro-electrode voltage clamp experiments in calf cardiac Purkinje fibres: is slow inward current adequately measured? J Physiol. 1979 May;290(2):201–225. doi: 10.1113/jphysiol.1979.sp012768. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Kass R. S., Tsien R. W. Multiple effects of calcium antagonists on plateau currents in cardiac Purkinje fibers. J Gen Physiol. 1975 Aug;66(2):169–192. doi: 10.1085/jgp.66.2.169. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Kimura J. E., Meves H. The effect of temperature on the asymmetrical charge movement in squid giant axons. J Physiol. 1979 Apr;289:479–500. doi: 10.1113/jphysiol.1979.sp012748. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Kootsey J. M., Johnson E. A. Voltage clamp of cardiac muscle. A theoretical analysis of early currents in the single sucrose gap. Biophys J. 1972 Nov;12(11):1496–1508. doi: 10.1016/S0006-3495(72)86177-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Lederer W. J., Tsien R. W. Transient inward current underlying arrhythmogenic effects of cardiotonic steroids in Purkinje fibres. J Physiol. 1976 Dec;263(2):73–100. doi: 10.1113/jphysiol.1976.sp011622. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Lee K. S., Weeks T. A., Kao R. L., Akaike N., Brown A. M. Sodium current in single heart muscle cells. Nature. 1979 Mar 15;278(5701):269–271. doi: 10.1038/278269a0. [DOI] [PubMed] [Google Scholar]
  46. McAllister R. E., Noble D. The time and voltage dependence of the slow outward current in cardiac Purkinje fibres. J Physiol. 1966 Oct;186(3):632–662. doi: 10.1113/jphysiol.1966.sp008060. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. McAllister R. E., Noble D., Tsien R. W. Reconstruction of the electrical activity of cardiac Purkinje fibres. J Physiol. 1975 Sep;251(1):1–59. doi: 10.1113/jphysiol.1975.sp011080. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Miura D. S., Hoffman B. F., Rosen M. R. The effect of extracellular potassium on the intracellular potassium ion activity and transmembrane potentials of beating canine cardiac Purkinje fibers. J Gen Physiol. 1977 Apr;69(4):463–474. doi: 10.1085/jgp.69.4.463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Moore L. E. Effect of temperature and calcium ions on rate constants of myelinated nerve. Am J Physiol. 1971 Jul;221(1):131–137. doi: 10.1152/ajplegacy.1971.221.1.131. [DOI] [PubMed] [Google Scholar]
  50. Nathan R. D., DeHaan R. L. Voltage clamp analysis of embryonic heart cell aggregates. J Gen Physiol. 1979 Feb;73(2):175–198. doi: 10.1085/jgp.73.2.175. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Ramón F., Anderson N., Joyner R. W., Moore J. W. Axon voltage-clamp simulations. A multicellular preparation. Biophys J. 1975 Jan;15(1):55–69. doi: 10.1016/S0006-3495(75)85791-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Reuter H., Scholz H. A study of the ion selectivity and the kinetic properties of the calcium dependent slow inward current in mammalian cardiac muscle. J Physiol. 1977 Jan;264(1):17–47. doi: 10.1113/jphysiol.1977.sp011656. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Reuter H., Seitz N. The dependence of calcium efflux from cardiac muscle on temperature and external ion composition. J Physiol. 1968 Mar;195(2):451–470. doi: 10.1113/jphysiol.1968.sp008467. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Schneider M. F., Chandler W. K. Effects of membrane potential on the capacitance of skeletal muscle fibers. J Gen Physiol. 1976 Feb;67(2):125–163. doi: 10.1085/jgp.67.2.125. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Schoenberg M., Dominguez G., Fozzard H. A. Effect of diameter on membrane capacity and conductance of sheep cardiac Purkinje fibers. J Gen Physiol. 1975 Apr;65(4):441–458. doi: 10.1085/jgp.65.4.441. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Schoenberg M., Fozzard H. A. The influence of intercellular clefts on the electrical properties of sheep cardiac Purkinje fibers. Biophys J. 1979 Feb;25(2 Pt 1):217–234. doi: 10.1016/s0006-3495(79)85287-x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Sommer J. R., Johnson E. A. Cardiac muscle. A comparative study of Purkinje fibers and ventricular fibers. J Cell Biol. 1968 Mar;36(3):497–526. doi: 10.1083/jcb.36.3.497. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Tarr M., Trank J. W. An assessment of the double sucrose-gap voltage clamp technique as applied to frog atrial muscle. Biophys J. 1974 Sep;14(9):627–643. doi: 10.1016/S0006-3495(74)85940-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Tsien R. W. Mode of action of chronotropic agents in cardiac Purkinje fibers. Does epinephrine act by directly modifying the external surface charge? J Gen Physiol. 1974 Sep;64(3):320–342. doi: 10.1085/jgp.64.3.320. [DOI] [PMC free article] [PubMed] [Google Scholar]
  60. Vitek M., Trautwein W. Slow inward current and action potential in cardiac Purkinje fibres. The effect of Mn plus,plus-ions. Pflugers Arch. 1971;323(3):204–218. doi: 10.1007/BF00586384. [DOI] [PubMed] [Google Scholar]

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