Journal of Molecular Histology

, Volume 39, Issue 1, pp 5–13 | Cite as

Semaphorin 6C expression in innervated and denervated skeletal muscle

  • Anna Svensson
  • Rolf Libelius
  • Sven TågerudEmail author
Original Paper


Semaphorins are secreted or transmembrane proteins important for axonal guidance and for the structuring of neuronal systems. Semaphorin 6C, a transmembrane Semaphorin, has growth cone collapsing activity and is expressed in adult skeletal muscle. In the present study the expression of Semaphorin 6C mRNA and immunoreactivity has been compared in innervated and denervated mouse hind-limb and hemidiaphragm muscles. Microscopic localization of immunoreactivity was studied in innervated and denervated rat skeletal muscle. The results show that Semaphorin 6C mRNA expression and immunoreactivity on Western blots are down-regulated following denervation. The mRNA of Semaphorin 6C as well as immunoreactivity determined by Western blots are expressed in extrasynaptic as well as perisynaptic regions of muscle. Immunohistochemical studies, however, show Semaphorin 6C-like immunoreactivity to be concentrated at neuromuscular junctions. The results suggest a role for Semaphorin 6C in neuromuscular communication.


Semaphorin 6C  Skeletal muscle  Denervation  Neuromuscular junction  Endplate 



We are grateful to Dr. Lennart Mellblom and Anita Jäderberg for providing access to cryostats at the County Hospital in Kalmar and to Dr. Caroline Magnusson for cloning the β-actin cDNA fragment. This work was supported by grants from the Faculty of Natural Sciences and Technology, University of Kalmar, the Crafoord Foundation, the Knowledge Foundation (KK-stiftelsen) and from Umeå University Hospital, Clinical Neuroscience Research Fund, Sweden.


  1. Anderson MJ, Fambrough DM (1983) Aggregates of acetylcholine receptors are associated with plaques of a basal lamina heparan sulfate proteoglycan on the surface of skeletal muscle fibers. J Cell Biol 97:1396–1411PubMedCrossRefGoogle Scholar
  2. Bernhardt RR, Schachner M (2000) Chondroitin sulfates affect the formation of the segmental motor nerves in Zebrafish embryos. Dev Biol 221:206–219PubMedCrossRefGoogle Scholar
  3. Billard C, Delaire S, Raffoux E, Bensussan A, Boumsell L (2000) Switch in the protein tyrosine phosphatase associated with human CD100 semaphorin at terminal B-cell differentiation stage. Blood 95:965–972PubMedGoogle Scholar
  4. Burgaya F, Fontana X, Martínez A, Montolio M, Mingorance A, Simó S, del Rio JA, Soriano E (2006) Semaphorin 6C leads to GSK-3-dependent growth cone collapse and redistributes after entorhino-hippocampal axotomy. Mol Cell Neurosci 33:321–334PubMedCrossRefGoogle Scholar
  5. Burkhardt C, Müller M, Badde A, Garner CC, Gundelfinger ED, Püschel AW (2005) Semaphorin 4B interacts with the post-synaptic density protein PSD-95/SAP90 and is recruited to synapses through a C-terminal PDZ-binding motif. FEBS Lett 579:3821–3828PubMedCrossRefGoogle Scholar
  6. Cohen S, Funkelstein L, Livet J, Rougon G, Henderson CE, Castellani V, Mann F (2005) A semaphorin code defines subpopulations of spinal motor neurons during mouse development. Eur J Neurosci 21:1767–1776PubMedCrossRefGoogle Scholar
  7. De Winter F, Vo T, Stam FJ, Wisman LAB, Bär PR, Niclou SP, van Muiswinkel FL, Verhaagen J (2006) The expression of the chemorepellent Semaphorin 3A is selectively induced in terminal Schwann cells of a subset of neuromuscular synapses that display limited anatomical plasticity and enhanced vulnerability in motor neuron disease. Mol Cell Neurosci 32:102–117PubMedCrossRefGoogle Scholar
  8. De Wit J, Verhaagen J (2003) Role of semaphorins in the adult nervous system. Prog Neurobiol 71:249–267PubMedCrossRefGoogle Scholar
  9. De Wit J, De Winter F, Klooster J, Verhaagen J (2005) Semaphorin 3A displays a punctate distribution on the surface of neuronal cells and interacts with proteoglycans in the extracellular matrix. Mol Cell Neurosci 29:40–55PubMedCrossRefGoogle Scholar
  10. Eckhardt F, Behar O, Calautti E, Yonezawa K, Nishimoto I, Fishman MC (1997) A novel transmembrane semaphorin can bind c-src. Mol Cell Neurosci 9:409–419PubMedCrossRefGoogle Scholar
  11. Feng T-P, Lu D-X (1965) New lights on the phenomenon of transient hypertrophy in the denervated hemidiaphragm of the rat. Sci Sin 14:1772–1784PubMedGoogle Scholar
  12. Fertuck HC, Salpeter MM (1974) Localization of acetylcholine receptor by 125I-labelled α-bungarotoxin binding at mouse motor endplates. Proc Natl Acad Sci USA 71:1376–1378PubMedCrossRefGoogle Scholar
  13. Festoff BW, Rao JS, Hantaï D (1991) Plasminogen activators and inhibitors in the neuromuscular system: III. The serpin protease nexin 1 is synthesized by muscle and localized at neuromuscular synapses. J Cell Physiol 147:76–86PubMedCrossRefGoogle Scholar
  14. Fiore R, Püschel AW (2003) The function of semaphorins during nervous system development. Front Biosci 8:484–499CrossRefGoogle Scholar
  15. Godenschwege TA, Hu H, Shan-Crofts X, Goodman CS, Murphey RK (2002) Bi-directional signaling by semaphorin 1a during central synapse formation in Drosophila. Nat Neurosci 5:1294–1301PubMedCrossRefGoogle Scholar
  16. Gutmann E, Haníková M, Hájek I, Klicpera M, Syrovy I (1966) The postdenervation hypertrophy of the diaphragm. Physiol Bohemoslov 15:508–524PubMedGoogle Scholar
  17. Hantaï D, Rao JS, Festoff BW (1988) Serine proteases and serpins: their possible roles in the motor system. Rev Neurol 144:680–687PubMedGoogle Scholar
  18. Huber AB, Kania A, Tran TS, Gu C, De Marco Garcia N, Lieberam I, Johnson D, Jessell TM, Ginty DD, Kolodkin AL (2005) Distinct roles for secreted semaphorin signaling in spinal motor axon guidance. Neuron 48:949–964PubMedCrossRefGoogle Scholar
  19. Inagaki S, Ohoka Y, Sugimoto H, Fujioka S, Amazaki M, Kurinami H, Miyazaki N, Tohyama M, Furuyama T (2001) Sema4C, a transmembrane semaphorin, interacts with a post-synaptic density protein, PSD-95. J Biol Chem 276:9174–9181PubMedCrossRefGoogle Scholar
  20. Kantor DB, Chivatakarn O, Peer KL, Oster SF, Inatani M, Hansen MJ, Flanagan JG, Yamaguchi Y, Sretavan DW, Giger RJ, Kolodkin AL (2004) Semaphorin 5A is a bifunctional axon guidance cue regulated by heparan and chondroitin sulfate proteoglycans. Neuron 44:961–975PubMedCrossRefGoogle Scholar
  21. Kikuchi K, Chédotal A, Hanafusa H, Ujimasa Y, de Castro F, Goodman CS, Kimura T (1999) Cloning and characterization of a novel class VI semaphorin, semaphorin Y. Mol Cell Neurosci 13:9–23PubMedCrossRefGoogle Scholar
  22. Klostermann A, Lutz B, Gertler F, Behl C (2000) The orthologous human and murine semaphorin 6A-1 proteins (SEMA6A-1/Sema6A-1) bind to the enabled/vasodilator-stimulated phosphoprotein-like protein (EVL) via a novel carboxyl-terminal zyxin-like domain. J Biol Chem 275:39647–39653PubMedCrossRefGoogle Scholar
  23. Magnusson C, Högklint L, Libelius R, Tågerud S (2001) Expression of mRNA for plasminogen activators and protease nexin-1 in innervated and denervated mouse skeletal muscle. J Neurosci Res 66:457–463PubMedCrossRefGoogle Scholar
  24. Magnusson C, Libelius R, Tågerud S (2003) Nogo (reticulon 4) expression in innervated and denervated mouse skeletal muscle. Mol Cell Neurosci 22:298–307PubMedCrossRefGoogle Scholar
  25. Matthes DJ, Sink H, Kolodkin AL, Goodman CS (1995) Semaphorin II can function as a selective inhibitor of specific synaptic arborizations. Cell 81:631–639PubMedCrossRefGoogle Scholar
  26. Nagase T, Nakayama M, Nakajima D, Kikuno R, Ohara O (2001) Prediction of the coding sequence of unidentified human genes. XX. The complete sequence of 100 new cDNA clones from brain which code for large proteins in vitro. DNA Res 8:85–95PubMedCrossRefGoogle Scholar
  27. Pasterkamp RJ, Giger RJ, Verhaagen J (1998) Regulation of semaphorin III/collapsin-1 gene expression during peripheral nerve regeneration. Exp Neurol 153:313–327PubMedCrossRefGoogle Scholar
  28. Qu X, Wei H, Zhai Y, Que H, Chen Q, Tang F, Wu Y, Xing G, Zhu Y, Liu S, Fan M, He F (2002) Identification, characterization, and functional study of the two novel human members of the semaphorin gene family. J Biol Chem 277:35574–35585PubMedCrossRefGoogle Scholar
  29. Roncarati R, Di Chio M, Sava A, Terstappen GC, Fumagalli G (2001) Presynaptic localization of the small conductance calcium-activated potassium channel SK3 at the neuromuscular junction. Neuroscience 104:253–262PubMedCrossRefGoogle Scholar
  30. Roos M, Schachner M, Bernhardt RR (1999) Zebrafish semaphorin Z1b inhibits growing motor axons in vivo. Mech Dev 87:103–117PubMedCrossRefGoogle Scholar
  31. Schultze W, Eulenburg V, Lessmann V, Herrmann L, Dittmar T, Gundelfinger ED, Heumann R, Erdmann KS (2001) Semaphorin4F interacts with the synapse-associated protein SAP90/PSD-95. J Neurochem 78:482–489PubMedCrossRefGoogle Scholar
  32. Semaphorin Nomenclature Committee (1999) Unified nomenclature for the semaphorins/collapsins. Semaphorin Nomenclature Committee. Cell 97:551–552CrossRefGoogle Scholar
  33. Sola OM, Martin AW (1953) Denervation hypertrophy and atrophy of the hemidiaphragm of the rat. Am J Physiol 172:324–332PubMedGoogle Scholar
  34. Varela-Echavarría A, Tucker A, Püschel AW, Guthrie S (1997) Motor axon subpopulations respond differentially to the chemorepellents netrin-1 and semaphorin D. Neuron 18:193–207PubMedCrossRefGoogle Scholar
  35. Winberg ML, Mitchell KJ, Goodman CS (1998) Genetic analysis of the mechanisms controlling target selection: complementary and combinatorial functions of netrins, semaphorins, and IgCAMs. Cell 93:581–591PubMedCrossRefGoogle Scholar
  36. Wood SJ, Slater CR (1998) β-spectrin is colocalized with both voltage-gated sodium channels and ankyrinG at the adult rat neuromuscular junction. J Cell Biol 140:675–684PubMedCrossRefGoogle Scholar
  37. Yu H-H, Araj HH, Ralls SA, Kolodkin AL (1998) The transmembrane semaphorin sema I is required in Drosophila for embryonic motor and CNS axon guidance. Neuron 20:207–220PubMedCrossRefGoogle Scholar
  38. Zhan W-Z, Sieck GC (1992) Adaptations of diaphragm and medial gastrocnemius muscles to inactivity. J Appl Physiol 72:1445–1453PubMedGoogle Scholar
  39. Zhan W-Z, Farkas GA, Schroeder MA, Gosselin LE, Sieck GC (1995) Regional adaptations of rabbit diaphragm muscle fibers to unilateral denervation. J Appl Physiol 79:941–950PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2007

Authors and Affiliations

  1. 1.School of Pure and Applied Natural SciencesUniversity of KalmarKalmarSweden
  2. 2.Department of Pharmacology and Clinical Neuroscience, Division of Clinical NeurophysiologyUmeå University HospitalUmeåSweden

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