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Patch clamp characterization of sodium channels expressed from rat brain cDNA

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Abstract

Sodium currents, I Na, were recorded from Xenopus laevis oocytes which had been injected with mRNA synthesized by in vitro transcription of the rat brain sodium channel II cDNA (Noda et al. 1986 a,b). Patch pipettes were used to apply depolarizing voltage steps and to record macroscopic sodium currents of between 50 and 750 pA from cellattached patches of the oocyte membrane. With a combination of whole-cell and patch clamp recording, the properties of the implanted sodium channels could be studied in detail. They were analyzed according to the model of Hodgkin and Huxley (1952a) assuming three activation gates. The activation of the sodium currents is characterized by an equilibrium potential of-29 mV and an apparent gating charge of 8.7 e 0. At-64 mV half of the sodium currents were inactivated. From singlechennel current recordings, an elementary sodium channel conductance of 19 pS and an average open time of 0.43 ms were obtained at-32 mV membrane potential and 16°C. The single-channel and activation properties of rat brain sodium channel II are therefore comparable to those found in peripheral nerve and skeletal muscle, but inactivation occurs at less negative potentials. This could be a specific property of the brain sodium channels and may underlie the maintained inward sodium currents reported in brain neurones (French and Gage 1985).

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References

  • Almers W, Roberts WM, Ruff RL (1984) Voltage clamp of rat and human skeletal muscle: measurements with an improved loose-patch technique. J Physiol 347:751–768

    Article  CAS  Google Scholar 

  • Baud C, Kado RT, Marcher K (1982) Sodium channels induced by depolarization of the Xenopus laevis oocyte. Proc Natl Acad Sci USA 79:3188–3192

    Article  CAS  Google Scholar 

  • Colquhoun D, Sigworth FJ (1983) Fitting and statistical analysis of single-channel records. In: Sakmann B, Neher E (eds) Single-channel recording. Plenum Press, New York, pp 191–263

    Chapter  Google Scholar 

  • Frankenhaeuser B (1960) Quantitative description of sodium currents in myelinated nerve fibres of Xenopus laevis. J Physiol 151:491–501

    Article  CAS  Google Scholar 

  • French CR, Gage PW (1985) A thereshold sodium current in pyramidal cells in rat hippocampus. Neurosci Lett 56:289–293

    Article  CAS  Google Scholar 

  • Green MR, Maniatis T, Melton DA (1983) Human β-globin pre-mRNA synthesized in vitro is accurately spliced in Xenopus oocyte nuclei. Cell 32:681–694

    Article  CAS  Google Scholar 

  • Gundersen CB, Miledi R, Parker I (1983) Voltage-operated channels induced by foreign messenger RNA in Xenopus oocytes. Proc R Soc London B 220:131–140

    Article  CAS  Google Scholar 

  • Gundersen CB, Miledi R, Parker I (1984) Messenger RNA from human brain induces drug— and voltage-operated channels in Xenopus oocytes. Nature 308:421–424

    Article  CAS  Google Scholar 

  • Gurdon JB (1974) The control of gene expression in animal development. Clarendon Press, Oxford

    Google Scholar 

  • Hamill OP, Marty A, Neher E, Sakmann B, Sigworth FJ (1981) Improved patch clamp techniques for high-resolution current recording from cells and cell-free membrane patches. Pflügers Arch 391:85–100

    Article  CAS  Google Scholar 

  • Hirono C, Yamagishi S, Ohara R, Hisanaga Y, Nakayama T, Sugiyama H (1985) Characterization of mRNA responsible for induction of functional sodium channels in Xenopus oocytes. Brain Res 359:57–64

    Article  CAS  Google Scholar 

  • Hodgkin AL, Huxley AF (1952a) A quantitative description of membrane current and its application to conduction and excitation in nerve. J Physiol 117:500–544

    Article  CAS  Google Scholar 

  • Hodgkin AL, Huxley AF (1952b) The dual effect of membrane potential on sodium conductance in the giant axon of loligo. J Physiol 116:497–506

    Article  CAS  Google Scholar 

  • Leonard J, Snutch T, Lubbert H, Davidson N, Lester HA (1986) Macroscopic Na currents with gigaohm seals on mRNA-injected Xenopus oocytes. Biophys J 49:386a

    Google Scholar 

  • Melton DA, Krieg PA, Rebagliati MR, Maniatis T, Sinn K, Green MR (1984) Efficient in vitro synthesis of biologically active RNA and RNA hybridization probes from plasmids containing a bacteriophage SP6 promoter. Nucleic Acids Res 12:7035–7056

    Article  CAS  Google Scholar 

  • Methfessel C, Witzemann V, Takahashi T, Mishina M, Numa S, Sakmann B (1986) Patch clamp measurements on Xenopus laevis oocytes: Currents through endogenous channels and implanted acetylcholine receptor and sodium channels. Pflügers Arch (in press)

  • Mishina M, Takai T, Imoto K, Noda M, Takahashi T, Numa S, Methfessel C, Sakmann B (1986) Molecular distinction between fetal and adult forms of muscle acetylcholine receptor. Nature 321:406–411

    Article  CAS  Google Scholar 

  • Neumcke B, Stämpfli R (1982) Sodium currents and sodiumcurrent fluctuations in rat myelinated nerve fibres. J Physiol 329:163–384

    Article  CAS  Google Scholar 

  • Neumcke B, Schwarz W, Stämpfli R (1979) Slow actions of hyperpolarization on sodium channels in the membrane of myelinated nerve. Biochim Biophys Acta 558:113–118

    Article  CAS  Google Scholar 

  • Noda M, Shimizu S, Tanabe T, Takai T, Kayano T, Ikeda T, Takahashi H, Nakayama H, Kanaoka Y, Minamino N, Kangawa K, Matsuo H, Raftery MA, Hirose T, Inayama S, Hayashida H, Miyata T, Numa S (1984) Primary structure of Electrophorus electricus sodium channel deduced from cDNA sequence. Nature 312:121–127

    Article  CAS  Google Scholar 

  • Noda M, Ikeda T, Kayano T, Suzuki H, Takeshima H, Kurasaki M, Takahashi H, Numa S (1986a) Existence of distinct sodium channel messenger RNAs in rat brain. Nature 320:188–192

    Article  CAS  Google Scholar 

  • Noda M, Ikeda T, Suzuki H, Takeshima H, Takahashi T, Kuno M, Numa S (1986b) Expression of functional sodium channels from cloned cDNA. Nature 322:826–828

    Article  CAS  Google Scholar 

  • Pappone PA (1980) Voltage-clamp experiments in normal and denervated mammalian skeletal muscle fibres. J Physiol 306:377–410

    Article  CAS  Google Scholar 

  • Ruff RL, Simoncini L, Stühmer W (1987) Comparison between slow sodium channel inactivation in rat slow and fast twitch muscle. J Physiol 383:339–348

    Article  CAS  Google Scholar 

  • Sakmann B, Neher E (1983) Geometric parameters of pipettes and membrane patches. In: Sakmann B, Neher E (eds) Single-channel recording. Plenum Press, New York, pp 37–51

    Chapter  Google Scholar 

  • Sakmann B, Methfessel C, Mishina M, Takahashi T, Takai T, Kurasaki M, Fukuda K, Numa S (1985) Role of acetylcholine receptor subunits in gating of the channel. Nature 318:538–543

    Article  CAS  Google Scholar 

  • Sigworth FJ (1980) The variance of sodium current fluctuations at the node of Ranvier. J Physiol 307:97–129

    Article  CAS  Google Scholar 

  • Sigworth FJ, Neher E (1980) Single Na+ channel currents observed in cultured rat muscle cells. Nature 287:447–449

    Article  CAS  Google Scholar 

  • Simoncini L, Stühmer W (1987) Slow sodium inactivation in rat fast twitch muscle. J Physiol 383:327–337

    Article  CAS  Google Scholar 

  • Stühmer W, Methfessel C, Witzemann V, Sakmann B (1986) Rat brain sodium channels recorded from Xenopus oocytes. In: Lüttgau HC (ed) Membrane control of cellular activity, vol 33. G Fischer, Stuttgart, pp 155–159

    Google Scholar 

  • Sumikawa K, Parker I, Miledi R (1984) Partial purification and functional expression of brain mRNAs coding for neurotransmitter receptors and voltage operated channels. Proc Natl Acad Sci USA 81:7994–7998

    Article  CAS  Google Scholar 

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Stühmer, W., Methfessel, C., Sakmann, B. et al. Patch clamp characterization of sodium channels expressed from rat brain cDNA. Eur Biophys J 14, 131–138 (1987). https://doi.org/10.1007/BF00253837

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  • DOI: https://doi.org/10.1007/BF00253837

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