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. 1986 Mar 1;102(3):863–877. doi: 10.1083/jcb.102.3.863

Nerve-induced remodeling of muscle basal lamina during synaptogenesis

PMCID: PMC2114107  PMID: 3005339

Abstract

To identify mechanisms that regulate the deposition of the junctional basal lamina during synaptogenesis, immunocytochemical experiments were carried out on cultured nerve and muscle cells derived from Xenopus laevis embryos. In some experiments successive observations were made on individual muscle cells after pulse-labeling with a fluorescent monoclonal antibody specific for a basal lamina proteoglycan. In others, old and new proteoglycan molecules were differentially labeled with antibody conjugated to contrasting fluorochromes. These observations revealed that surface deposits of antibody-labeled proteoglycan remain morphologically stable for several days on developing muscle cells. Over the same period, however, new sites of proteoglycan accumulation formed that contained primarily those antigenic sites recently exposed at the cell surface. When muscle cells became innervated by cholinergic neurites, new proteoglycan accumulations were induced at the developing neuromuscular junctions, and these too were composed almost exclusively of recently deposited antigen. In older muscle cultures, where many cells possessed relatively high background concentrations of antigen over their surfaces, developing neuromuscular junctions initially showed a markedly reduced proteoglycan site-density compared with the adjacent, extrajunctional muscle surface. Much of this perineural region eventually became filled with dense, nerve induced proteoglycan plaques at later stages of synapse development. Motoneurons thus appear to have two, superficially paradoxical effects on muscle basal lamina organization. They first cause the removal of any existing, extrajunctional proteoglycan from the path of cell contact, and then induce the deposition of dense plaques of recently synthesized proteoglycan within the developing junctional basal lamina. This observation suggests that the proteolytic enzyme systems that have already been implicated in tissue remodeling may also contribute to the inductive interaction between nerve and muscle cells during synaptogenesis.

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Selected References

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  1. Aggeler J., Frisch S. M., Werb Z. Collagenase is a major gene product of induced rabbit synovial fibroblasts. J Cell Biol. 1984 May;98(5):1656–1661. doi: 10.1083/jcb.98.5.1656. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Anderson M. J., Cohen M. W. Fluorescent staining of acetylcholine receptors in vertebrate skeletal muscle. J Physiol. 1974 Mar;237(2):385–400. doi: 10.1113/jphysiol.1974.sp010487. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Anderson M. J., Cohen M. W. Nerve-induced and spontaneous redistribution of acetylcholine receptors on cultured muscle cells. J Physiol. 1977 Jul;268(3):757–773. doi: 10.1113/jphysiol.1977.sp011880. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Anderson M. J., Fambrough D. M. 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. 1983 Nov;97(5 Pt 1):1396–1411. doi: 10.1083/jcb.97.5.1396. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Anderson M. J., Kidokoro Y., Gruener R. Correlation between acetylcholine receptor localization and spontaneous synaptic potentials in cultures of nerve and muscle. Brain Res. 1979 Apr 20;166(1):185–190. doi: 10.1016/0006-8993(79)90662-0. [DOI] [PubMed] [Google Scholar]
  6. Anderson M. J., Klier F. G., Tanguay K. E. Acetylcholine receptor aggregation parallels the deposition of a basal lamina proteoglycan during development of the neuromuscular junction. J Cell Biol. 1984 Nov;99(5):1769–1784. doi: 10.1083/jcb.99.5.1769. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Bauer H. C., Daniels M. P., Pudimat P. A., Jacques L., Sugiyama H., Christian C. N. Characterization and partial purification of a neuronal factor which increases acetylcholine receptor aggregation on cultured muscle cells. Brain Res. 1981 Mar 30;209(2):395–404. doi: 10.1016/0006-8993(81)90161-x. [DOI] [PubMed] [Google Scholar]
  8. Baxter D. A., Johnston D., Strittmatter W. J. Protease inhibitors implicate metalloendoprotease in synaptic transmission at the mammalian neuromuscular junction. Proc Natl Acad Sci U S A. 1983 Jul;80(13):4174–4178. doi: 10.1073/pnas.80.13.4174. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Bayne E. K., Anderson M. J., Fambrough D. M. Extracellular matrix organization in developing muscle: correlation with acetylcholine receptor aggregates. J Cell Biol. 1984 Oct;99(4 Pt 1):1486–1501. doi: 10.1083/jcb.99.4.1486. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Bernfield M., Banerjee S. D., Koda J. E., Rapraeger A. C. Remodelling of the basement membrane: morphogenesis and maturation. Ciba Found Symp. 1984;108:179–196. doi: 10.1002/9780470720899.ch12. [DOI] [PubMed] [Google Scholar]
  11. Bloch R. J., Geiger B. The localization of acetylcholine receptor clusters in areas of cell-substrate contact in cultures of rat myotubes. Cell. 1980 Aug;21(1):25–35. doi: 10.1016/0092-8674(80)90111-7. [DOI] [PubMed] [Google Scholar]
  12. Bloch R. J., Hall Z. W. Cytoskeletal components of the vertebrate neuromuscular junction: vinculin, alpha-actinin, and filamin. J Cell Biol. 1983 Jul;97(1):217–223. doi: 10.1083/jcb.97.1.217. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Buckley K. M., Schweitzer E. S., Miljanich G. P., Clift-O'Grady L., Kushner P. D., Reichardt L. F., Kelly R. B. A synaptic vesicle antigen is restricted to the junctional region of the presynaptic plasma membrane. Proc Natl Acad Sci U S A. 1983 Dec;80(23):7342–7346. doi: 10.1073/pnas.80.23.7342. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Burden S. J., Sargent P. B., McMahan U. J. Acetylcholine receptors in regenerating muscle accumulate at original synaptic sites in the absence of the nerve. J Cell Biol. 1979 Aug;82(2):412–425. doi: 10.1083/jcb.82.2.412. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Burden S. Identification of an intracellular postsynaptic antigen at the frog neuromuscular junction. J Cell Biol. 1982 Sep;94(3):521–530. doi: 10.1083/jcb.94.3.521. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Burrage T. G., Lentz T. L. Ultrastructural characterization of surface specializations containing high-density acetylcholine receptors on embryonic chick myotubes in vivo and in vitro. Dev Biol. 1981 Jul 30;85(2):267–286. doi: 10.1016/0012-1606(81)90259-1. [DOI] [PubMed] [Google Scholar]
  17. Carlson S. S., Kelly R. B. A highly antigenic proteoglycan-like component of cholinergic synaptic vesicles. J Biol Chem. 1983 Sep 25;258(18):11082–11091. [PubMed] [Google Scholar]
  18. Christian C. N., Daniels M. P., Sugiyama H., Vogel Z., Jacques L., Nelson P. G. A factor from neurons increases the number of acetylcholine receptor aggregates on cultured muscle cells. Proc Natl Acad Sci U S A. 1978 Aug;75(8):4011–4015. doi: 10.1073/pnas.75.8.4011. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Cohen M. W., Weldon P. R. Localization of acetylcholine receptors and synaptic ultrastructure at nerve-muscle contacts in culture: dependence on nerve type. J Cell Biol. 1980 Aug;86(2):388–401. doi: 10.1083/jcb.86.2.388. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Fallon J. R., Nitkin R. M., Reist N. E., Wallace B. G., McMahan U. J. Acetylcholine receptor-aggregating factor is similar to molecules concentrated at neuromuscular junctions. Nature. 1985 Jun 13;315(6020):571–574. doi: 10.1038/315571a0. [DOI] [PubMed] [Google Scholar]
  21. Fertuck H. C., Salpeter M. M. Quantitation of junctional and extrajunctional acetylcholine receptors by electron microscope autoradiography after 125I-alpha-bungarotoxin binding at mouse neuromuscular junctions. J Cell Biol. 1976 Apr;69(1):144–158. doi: 10.1083/jcb.69.1.144. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Fischbach G. D., Cohen S. A. The distribution of acetylcholine sensitivity over uninnervated and innervated muscle fibers grown in cell culture. Dev Biol. 1973 Mar;31(1):147–162. doi: 10.1016/0012-1606(73)90326-6. [DOI] [PubMed] [Google Scholar]
  23. Frank E., Fischbach G. D. Early events in neuromuscular junction formation in vitro: induction of acetylcholine receptor clusters in the postsynaptic membrane and morphology of newly formed synapses. J Cell Biol. 1979 Oct;83(1):143–158. doi: 10.1083/jcb.83.1.143. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Froehner S. C., Gulbrandsen V., Hyman C., Jeng A. Y., Neubig R. R., Cohen J. B. Immunofluorescence localization at the mammalian neuromuscular junction of the Mr 43,000 protein of Torpedo postsynaptic membranes. Proc Natl Acad Sci U S A. 1981 Aug;78(8):5230–5234. doi: 10.1073/pnas.78.8.5230. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Galloway W. A., Murphy G., Sandy J. D., Gavrilovic J., Cawston T. E., Reynolds J. J. Purification and characterization of a rabbit bone metalloproteinase that degrades proteoglycan and other connective-tissue components. Biochem J. 1983 Mar 1;209(3):741–752. doi: 10.1042/bj2090741. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Godfrey E. W., Nitkin R. M., Wallace B. G., Rubin L. L., McMahan U. J. Components of Torpedo electric organ and muscle that cause aggregation of acetylcholine receptors on cultured muscle cells. J Cell Biol. 1984 Aug;99(2):615–627. doi: 10.1083/jcb.99.2.615. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Hall Z. W., Lubit B. W., Schwartz J. H. Cytoplasmic actin in postsynaptic structures at the neuromuscular junction. J Cell Biol. 1981 Sep;90(3):789–792. doi: 10.1083/jcb.90.3.789. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Hooper J. E., Carlson S. S., Kelly R. B. Antibodies to synaptic vesicles purified from Narcine electric organ bind a subclass of mammalian nerve terminals. J Cell Biol. 1980 Oct;87(1):104–113. doi: 10.1083/jcb.87.1.104. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Jessell T. M., Siegel R. E., Fischbach G. D. Induction of acetylcholine receptors on cultured skeletal muscle by a factor extracted from brain and spinal cord. Proc Natl Acad Sci U S A. 1979 Oct;76(10):5397–5401. doi: 10.1073/pnas.76.10.5397. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Kalcheim C., Vogel Z., Duksin D. Embryonic brain extract induces collagen biosynthesis in cultured muscle cells: involvement in acetylcholine receptor aggregation. Proc Natl Acad Sci U S A. 1982 May;79(10):3077–3081. doi: 10.1073/pnas.79.10.3077. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Kidokoro Y., Anderson M. J., Gruener R. Changes in synaptic potential properties during acetylcholine receptor accumulation and neurospecific interactions in Xenopus nerve-muscle cell culture. Dev Biol. 1980 Aug;78(2):464–483. doi: 10.1016/0012-1606(80)90347-4. [DOI] [PubMed] [Google Scholar]
  32. Kleinman H. K., McGarvey M. L., Liotta L. A., Robey P. G., Tryggvason K., Martin G. R. Isolation and characterization of type IV procollagen, laminin, and heparan sulfate proteoglycan from the EHS sarcoma. Biochemistry. 1982 Nov 23;21(24):6188–6193. doi: 10.1021/bi00267a025. [DOI] [PubMed] [Google Scholar]
  33. Ko P. K., Anderson M. J., Cohen M. W. Denervated skeletal muscle fibers develop discrete patches of high acetylcholine receptor density. Science. 1977 Apr 29;196(4289):540–542. doi: 10.1126/science.850796. [DOI] [PubMed] [Google Scholar]
  34. Krystosek A., Seeds N. W. Peripheral neurons and Schwann cells secrete plasminogen activator. J Cell Biol. 1984 Feb;98(2):773–776. doi: 10.1083/jcb.98.2.773. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Liotta L. A., Rao N. C., Barsky S. H., Bryant G. The laminin receptor and basement membrane dissolution: role in tumour metastasis. Ciba Found Symp. 1984;108:146–162. doi: 10.1002/9780470720899.ch10. [DOI] [PubMed] [Google Scholar]
  36. Markelonis G. J., Oh T. H., Eldefrawi M. E., Guth L. Sciatin: a myotrophic protein increases the number of acetylcholine receptors and receptor clusters in cultured skeletal muscle. Dev Biol. 1982 Feb;89(2):353–361. doi: 10.1016/0012-1606(82)90324-4. [DOI] [PubMed] [Google Scholar]
  37. Moody-Corbett F., Cohen M. W. Localization of cholinesterase at sites of high acetylcholine receptor density on embryonic amphibian muscle cells cultured without nerve. J Neurosci. 1981 Jun;1(6):596–605. doi: 10.1523/JNEUROSCI.01-06-00596.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Mullins D. E., Rohrlich S. T. The role of proteinases in cellular invasiveness. Biochim Biophys Acta. 1983 Dec 29;695(3-4):177–214. doi: 10.1016/0304-419x(83)90011-2. [DOI] [PubMed] [Google Scholar]
  39. Orida N., Poo M. M. Electrophoretic movement and localisation of acetylcholine receptors in the embryonic muscle cell membrane. Nature. 1978 Sep 7;275(5675):31–35. doi: 10.1038/275031a0. [DOI] [PubMed] [Google Scholar]
  40. Peng H. B., Cheng P. C., Luther P. W. Formation of ACh receptor clusters induced by positively charged latex beads. Nature. 1981 Aug 27;292(5826):831–834. doi: 10.1038/292831a0. [DOI] [PubMed] [Google Scholar]
  41. Peng H. B., Froehner S. C. Association of the postsynaptic 43K protein with newly formed acetylcholine receptor clusters in cultured muscle cells. J Cell Biol. 1985 May;100(5):1698–1705. doi: 10.1083/jcb.100.5.1698. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Pittman R. N. Release of plasminogen activator and a calcium-dependent metalloprotease from cultured sympathetic and sensory neurons. Dev Biol. 1985 Jul;110(1):91–101. doi: 10.1016/0012-1606(85)90067-3. [DOI] [PubMed] [Google Scholar]
  43. Podleski T. R., Axelrod D., Ravdin P., Greenberg I., Johnson M. M., Salpeter M. M. Nerve extract induces increase and redistribution of acetylcholine receptors on cloned muscle cells. Proc Natl Acad Sci U S A. 1978 Apr;75(4):2035–2039. doi: 10.1073/pnas.75.4.2035. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Ravdin P., Axelrod D. Fluorescent tetramethyl rhodamine derivatives of alpha-bungarotoxin: preparation, separation, and characterization. Anal Biochem. 1977 Jun;80(2):585–592. doi: 10.1016/0003-2697(77)90682-0. [DOI] [PubMed] [Google Scholar]
  45. Rubin L. L., Schuetze S. M., Fischbach G. D. Accumulation of acetylcholinesterase at newly formed nerve--muscle synapases. Dev Biol. 1979 Mar;69(1):46–58. doi: 10.1016/0012-1606(79)90273-2. [DOI] [PubMed] [Google Scholar]
  46. Sanes J. R., Feldman D. H., Cheney J. M., Lawrence J. C., Jr Brain extract induces synaptic characteristics in the basal lamina of cultured myotubes. J Neurosci. 1984 Feb;4(2):464–473. doi: 10.1523/JNEUROSCI.04-02-00464.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Sanes J. R., Hall Z. W. Antibodies that bind specifically to synaptic sites on muscle fiber basal lamina. J Cell Biol. 1979 Nov;83(2 Pt 1):357–370. doi: 10.1083/jcb.83.2.357. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Sanes J. R. Laminin, fibronectin, and collagen in synaptic and extrasynaptic portions of muscle fiber basement membrane. J Cell Biol. 1982 May;93(2):442–451. doi: 10.1083/jcb.93.2.442. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Sanes J. R., Marshall L. M., McMahan U. J. Reinnervation of muscle fiber basal lamina after removal of myofibers. Differentiation of regenerating axons at original synaptic sites. J Cell Biol. 1978 Jul;78(1):176–198. doi: 10.1083/jcb.78.1.176. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Sellers A., Murphy G. Collagenolytic enzymes and their naturally occurring inhibitors. Int Rev Connect Tissue Res. 1981;9:151–190. doi: 10.1016/b978-0-12-363709-3.50010-3. [DOI] [PubMed] [Google Scholar]
  51. Sellers A., Reynolds J. J., Meikle M. C. Neutral metallo-proteinases of rabbit bone. Separation in latent forms of distinct enzymes that when activated degrade collagen, gelatin and proteoglycans. Biochem J. 1978 May 1;171(2):493–496. doi: 10.1042/bj1710493. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Stadler H., Dowe G. H. Identification of a heparan sulphate-containing proteoglycan as a specific core component of cholinergic synaptic vesicles from Torpedo marmorata. EMBO J. 1982;1(11):1381–1384. doi: 10.1002/j.1460-2075.1982.tb01326.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Sytkowski A. J., Vogel Z., Nirenberg M. W. Development of acetylcholine receptor clusters on cultured muscle cells. Proc Natl Acad Sci U S A. 1973 Jan;70(1):270–274. doi: 10.1073/pnas.70.1.270. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Tryggvason K., Pihlajaniemi T., Salo T. Studies on the molecular composition and degradation of type IV procollagen. Ciba Found Symp. 1984;108:117–129. doi: 10.1002/9780470720899.ch8. [DOI] [PubMed] [Google Scholar]
  55. Vaes G., Eeckhout Y., Lenaers-Claeys G., François-Gillet C., Druetz J. E. The simultaneous release by bone explants in culture and the parallel activation of procollagenase and of a latent neutral proteinase that degrades cartilage proteoglycans and denatured collagen. Biochem J. 1978 May 15;172(2):261–274. doi: 10.1042/bj1720261. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Vater C. A., Nagase H., Harris E. D., Jr Purification of an endogenous activator of procollagenase from rabbit synovial fibroblast culture medium. J Biol Chem. 1983 Aug 10;258(15):9374–9382. [PubMed] [Google Scholar]
  57. Vogel Z., Christian C. N., Vigny M., Bauer H. C., Sonderegger P., Daniels M. P. Laminin induces acetylcholine receptor aggregation on cultured myotubes and enhances the receptor aggregation activity of a neuronal factor. J Neurosci. 1983 May;3(5):1058–1068. doi: 10.1523/JNEUROSCI.03-05-01058.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Weldon P. R., Cohen M. W. Development of synaptic ultrastructure at neuromuscular contacts in an amphibian cell culture system. J Neurocytol. 1979 Apr;8(2):239–259. doi: 10.1007/BF01175564. [DOI] [PubMed] [Google Scholar]
  59. Werb Z., Dingle J. T., Reynolds J. J., Barrett A. J. Proteoglycan-degrading enzymes of rabbit fibroblasts and granulocytes. Biochem J. 1978 Sep 1;173(3):949–958. doi: 10.1042/bj1730949. [DOI] [PMC free article] [PubMed] [Google Scholar]

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