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. 1981 Feb;78(2):1245–1249. doi: 10.1073/pnas.78.2.1245

Na+-channel-associated scorpion toxin receptor sites as probes for neuronal evolution in vivo and in vitro.

Y Berwald-Netter, N Martin-Moutot, A Koulakoff, F Couraud
PMCID: PMC319985  PMID: 6262759

Abstract

Purified neurotoxin II of the scorpion Androctonus australis Hector (ScTx) has previously been shown to bind specifically to the Na+-ionophore-associated, voltage-sensitive receptor sites of excitable cells. We have conducted binding studies, using high-specific-activity 125I-labeled ScTx, to detect and quantify the Na+-channel receptors on cells of the developing fetal mouse brain. In vivo, the onset of detectable specific binding is at 12 fetal days. The rate of receptor appearance is initially slow but increases sharply as of the 16th day of mouse ontogenesis. The mean number of receptors at 12 and 19 days is 120 and 20,000 per cell, respectively (i.e., 0.5 and 80 per square micrometer). When corrected for the fraction of cell population corresponding to putative neuroblasts and neurons, identified by immunofluorescence as tetanus toxin binding cells, these values are, respectively, 1040 and 33,900 ScTx receptors per tetanus toxin binding cell or 4.2 and 136 per square micrometer. At all stages, the toxin binds to a single class of noninteracting sites; Kd = 0.1-0.5 nM. Similar findings in terms of ScTx-receptor properties and quantitative evolution were obtained in vitro. Specific 125I-labeled ScTx binding the presence of tetanus toxin binding cells. In cultures of central nervous system glia without neurons, only nonspecific low-level ScTx binding was detected. These results suggest that the high-affinity scorpion toxin receptors may be used as quantitative markers of neuronal differentiation.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Beneski D. A., Catterall W. A. Covalent labeling of protein components of the sodium channel with a photoactivable derivative of scorpion toxin. Proc Natl Acad Sci U S A. 1980 Jan;77(1):639–643. doi: 10.1073/pnas.77.1.639. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bernard P., Couraud F. Electrophysiological studies on embryonic heart cells in culture. Scorpion toxin as a tool to reveal latent fast sodium channel. Biochim Biophys Acta. 1979 May 3;553(1):154–168. doi: 10.1016/0005-2736(79)90037-3. [DOI] [PubMed] [Google Scholar]
  3. Bernard P., Couraud F., Lissitzky S. Effects of a scorpion toxin from Androctonus australis venom on action potential of neuroblastoma cells in culture. Biochem Biophys Res Commun. 1977 Jul 25;77(2):782–788. doi: 10.1016/s0006-291x(77)80046-6. [DOI] [PubMed] [Google Scholar]
  4. Catterall W. A. Activation of the action potential Na+ ionophore by neurotoxins. An allosteric model. J Biol Chem. 1977 Dec 10;252(23):8669–8676. [PubMed] [Google Scholar]
  5. Catterall W. A. Membrane potential-dependent binding of scorpion toxin to the action potential Na+ ionophore. Studies with a toxin derivative prepared by lactoperoxidase-catalyzed iodination. J Biol Chem. 1977 Dec 10;252(23):8660–8668. [PubMed] [Google Scholar]
  6. Catterall W. A. Neurotoxins that act on voltage-sensitive sodium channels in excitable membranes. Annu Rev Pharmacol Toxicol. 1980;20:15–43. doi: 10.1146/annurev.pa.20.040180.000311. [DOI] [PubMed] [Google Scholar]
  7. Catterall W. A., Nirenberg M. Sodium uptake associated with activation of action potential ionophores of cultured neuroblastoma and muscle cells. Proc Natl Acad Sci U S A. 1973 Dec;70(12):3759–3763. doi: 10.1073/pnas.70.12.3759. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Catterall W. A., Ray R., Morrow C. S. Membrane potential dependent binding of scorpion toxin to action potential Na+ ionophore. Proc Natl Acad Sci U S A. 1976 Aug;73(8):2682–2686. doi: 10.1073/pnas.73.8.2682. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Chalazonitis A., Greene L. A., Nirenberg M. Electrophysiological chracteristics of chick embryo sympathetic neurons in dissociated cell culture. Brain Res. 1974 Mar 22;68(2):235–252. doi: 10.1016/0006-8993(74)90393-x. [DOI] [PubMed] [Google Scholar]
  10. Couraud F., Rochat H., Lissitzky S. Binding of scorpion and sea anemone neurotoxins to a common site related to the action potential Na+ ionophore in neuroblastoma cells. Biochem Biophys Res Commun. 1978 Aug 29;83(4):1525–1530. doi: 10.1016/0006-291x(78)91394-3. [DOI] [PubMed] [Google Scholar]
  11. Couraud F., Rochat H., Lissitzky S. Binding of scorpion neurotoxins to chick embryonic heart cells in culture and relationship to calcium uptake and membrane potential. Biochemistry. 1980 Feb 5;19(3):457–462. doi: 10.1021/bi00544a009. [DOI] [PubMed] [Google Scholar]
  12. Dichter M. A., Fischbach G. D. The action potential of chick dorsal root ganglion neurones maintained in cell culture. J Physiol. 1977 May;267(2):281–298. doi: 10.1113/jphysiol.1977.sp011813. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Dimpfel W., Huang R. T., Habermann E. Gangliosides in nervous tissue cultures and binding of 125I-labelled tetanus toxin, a neuronal marker. J Neurochem. 1977 Aug;29(2):329–334. doi: 10.1111/j.1471-4159.1977.tb09626.x. [DOI] [PubMed] [Google Scholar]
  14. Fields K. L., Brockes J. P., Mirsky R., Wendon L. M. Cell surface markers for distinguishing different types of rat dorsal root ganglion cells in culture. Cell. 1978 May;14(1):43–51. doi: 10.1016/0092-8674(78)90299-4. [DOI] [PubMed] [Google Scholar]
  15. Fukuda J., Kameyama M. Enhancement of Ca spikes in nerve cells of adult mammals during neurite growth in tissue culture. Nature. 1979 Jun 7;279(5713):546–548. doi: 10.1038/279546a0. [DOI] [PubMed] [Google Scholar]
  16. Godfrey E. W., Nelson P. G., Schrier B. K., Breuer A. C., Ransom B. R. Neurons from fetal rat brain in a new cell culture system: a multidisciplinary analysis. Brain Res. 1975 Jun 6;90(1):1–21. doi: 10.1016/0006-8993(75)90679-4. [DOI] [PubMed] [Google Scholar]
  17. Jover E., Martin-Moutot N., Couraud F., Rochat H. Binding of scorpion toxins to rat brain synaptosomal fraction. Effects of membrane potential, ions, and other neurotoxins. Biochemistry. 1980 Feb 5;19(3):463–467. doi: 10.1021/bi00544a010. [DOI] [PubMed] [Google Scholar]
  18. Jover E., Martin-Moutot N., Couraud F., Rochat H. Scorpion toxin: specific binding to rat synaptosomes. Biochem Biophys Res Commun. 1978 Nov 14;85(1):377–382. doi: 10.1016/s0006-291x(78)80053-9. [DOI] [PubMed] [Google Scholar]
  19. Matsuda Y., Yoshida S., Yonezawa T. Tetrodotoxin sensitivity and Ca component of action potentials of mouse dorsal root ganglion cells cultured in vitro. Brain Res. 1978 Oct 6;154(1):69–82. doi: 10.1016/0006-8993(78)91052-1. [DOI] [PubMed] [Google Scholar]
  20. Miranda F., Kupeyan C., Rochat H., Rochat C., Lissitzky S. Purification of animal neurotoxins. Isolation and characterization of eleven neurotoxins from the venoms of the scorpions Androctonus australis hector, Buthus occitanus tunetanus and Leiurus quinquestriatus quinquestriatus. Eur J Biochem. 1970 Nov;16(3):514–523. doi: 10.1111/j.1432-1033.1970.tb01111.x. [DOI] [PubMed] [Google Scholar]
  21. Mirsky R., Wendon L. M., Black P., Stolkin C., Bray D. Tetanus toxin: a cell surface marker for neurones in culture. Brain Res. 1978 Jun 9;148(1):251–259. doi: 10.1016/0006-8993(78)90399-2. [DOI] [PubMed] [Google Scholar]
  22. Munson R., Jr, Westermark B., Glaser L. Tetrodotoxin-sensitive sodium channels in normal human fibroblasts and normal human glia-like cells. Proc Natl Acad Sci U S A. 1979 Dec;76(12):6425–6429. doi: 10.1073/pnas.76.12.6425. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Nelson P. G., Peacock J. H. Electrical activity in dissociated cell cultures from fetal mouse cerebellum. Brain Res. 1973 Oct 26;61:163–174. doi: 10.1016/0006-8993(73)90525-8. [DOI] [PubMed] [Google Scholar]
  24. Pado C. H., Munson R., Glaser L., Gottlieb D. I. Evidence for ionic channels in cultured chick embryonic CNS cells. Brain Res. 1980 Mar 3;185(1):187–191. doi: 10.1016/0006-8993(80)90682-4. [DOI] [PubMed] [Google Scholar]
  25. Palfrey C., Littauer U. Z. Sodium-dependent efflux of K+ and Rb+ through the activated sodium channel of neuroblastoma cells. Biochem Biophys Res Commun. 1976 Sep 7;72(1):209–215. doi: 10.1016/0006-291x(76)90981-5. [DOI] [PubMed] [Google Scholar]
  26. Peacock J. H., Nelson P. G., Goldstone M. W. Electrophysiologic study of cultured neurons dissociated from spinal cords and dorsal root ganglia of fetal mice. Dev Biol. 1973 Jan;30(1):137–152. doi: 10.1016/0012-1606(73)90053-5. [DOI] [PubMed] [Google Scholar]
  27. Pouysségur J., Jacques Y., Lazdunski M. Identification of a tetrodotoxin-sensitive Na+ channel in a variety in fibroblast lines. Nature. 1980 Jul 10;286(5769):162–164. doi: 10.1038/286162a0. [DOI] [PubMed] [Google Scholar]
  28. Raff M. C., Fields K. L., Hakomori S. I., Mirsky R., Pruss R. M., Winter J. Cell-type-specific markers for distinguishing and studying neurons and the major classes of glial cells in culture. Brain Res. 1979 Oct 5;174(2):283–308. doi: 10.1016/0006-8993(79)90851-5. [DOI] [PubMed] [Google Scholar]
  29. Ray R., Morrow C. S., Catterall W. A. Binding of scorpion toxin to receptor sites associated with voltage-sensitive sodium channels in synaptic nerve ending particles. J Biol Chem. 1978 Oct 25;253(20):7307–7313. [PubMed] [Google Scholar]
  30. Ritchie J. M., Rogart R. B. Density of sodium channels in mammalian myelinated nerve fibers and nature of the axonal membrane under the myelin sheath. Proc Natl Acad Sci U S A. 1977 Jan;74(1):211–215. doi: 10.1073/pnas.74.1.211. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Rochat H., Bernard P., Couraud F. Scorpion toxins: chemistry and mode of action. Adv Cytopharmacol. 1979;3:325–334. [PubMed] [Google Scholar]
  32. Rochat H., Tessier M., Miranda F., Lissitzky S. Radioiodination of scorpion and snake toxins. Anal Biochem. 1977 Oct;82(2):532–548. doi: 10.1016/0003-2697(77)90192-0. [DOI] [PubMed] [Google Scholar]
  33. Romey G., Chicheportiche R., Lazdunski M., Rochat H., Miranda F., Lissitzky S. Scorpion neurotoxin - a presynaptic toxin which affects both Na+ and K+ channels in axons. Biochem Biophys Res Commun. 1975 May 5;64(1):115–121. doi: 10.1016/0006-291x(75)90226-0. [DOI] [PubMed] [Google Scholar]
  34. Spitzer N. C. Ion channels in development. Annu Rev Neurosci. 1979;2:363–397. doi: 10.1146/annurev.ne.02.030179.002051. [DOI] [PubMed] [Google Scholar]
  35. Tang C. M., Strichartz G. R., Orkand R. K. Sodium channels in axons and glial cells of the optic nerve of Necturus maculosa. J Gen Physiol. 1979 Nov;74(5):629–642. doi: 10.1085/jgp.74.5.629. [DOI] [PMC free article] [PubMed] [Google Scholar]

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