Skip to main content
NCBI home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1983 Jan 1;96(1):240–247. doi: 10.1083/jcb.96.1.240

Changes in the amounts of cytoskeletal proteins within the perikarya and axons of regenerating frog motoneurons

PMCID: PMC2112238  PMID: 6402517

Abstract

Changes in the amounts of tubulin, actin, and neurofilament polypeptides were found in regenerating motoneurons of grass frogs during the period of axonal elongation. Ventral roots 9 and 10 were transected unilaterally about 7 mm from the spinal cord. 35 d later, [3H]colchicine binding had decreased in the proximal stumps to approximately one-half of contralateral control values, well before the regenerating motor axons had reinnervated skeletal muscles of the hind limb. [3H]colchicine binding did not change significantly in the operated halves of the 9th and 10th spinal cord segments over a 75-d period. The relative amounts of actin, tubulin, and neurofilament polypeptides in the operated ventral roots were measured by quantitative densitometry of stained two-dimensional electrophoretic gels. Alpha-tubulin, beta-tubulin, and the 68,000 molecular weight subunit of neurofilaments (NF68) decreased within the transected ventral roots to 78%, 57%, and less than 15% of control values, respectively. The amount of actin increased to 132% of control values within the operated ventral roots, although this change was not statistically significant. Opposite changes were found within motoneuronal cell bodies isolated from the spinal cord. The relative amounts of alpha-tubulin, beta-tubulin and NF68 within axotomized perikarya increased, respectively, to 191%, 146%, and 144% of that in control perikarya isolated from the contralateral side of the spinal cord. Thus, the changes in NF68 and tubulin did not occur uniformly throughout the injured cells. The possible structural and functional consequences of these changes are discussed.

Full Text

The Full Text of this article is available as a PDF (1.9 MB).

Selected References

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

  1. Autilio-Gambetti L., Velasco M. E., Sipple J., Gambetti P. Immunochemical characterization of antisera to rat neurofilament subunits. J Neurochem. 1981 Nov;37(5):1260–1265. doi: 10.1111/j.1471-4159.1981.tb04676.x. [DOI] [PubMed] [Google Scholar]
  2. Aycock B. F., Weil D. E., Sinicropi D. V., McIlwain D. L. Television-based densitometric analysis of proteins separated by two-dimensional gel electrophoresis. Comput Biomed Res. 1981 Aug;14(4):314–326. doi: 10.1016/0010-4809(81)90003-3. [DOI] [PubMed] [Google Scholar]
  3. Barron K. D., Dentinger M. P., Nelson L. R., Mincy J. E. Ultrastructure of axonal reaction in red nucleus of cat. J Neuropathol Exp Neurol. 1975 May;34(3):222–248. doi: 10.1097/00005072-197505000-00002. [DOI] [PubMed] [Google Scholar]
  4. Bensadoun A., Weinstein D. Assay of proteins in the presence of interfering materials. Anal Biochem. 1976 Jan;70(1):241–250. doi: 10.1016/s0003-2697(76)80064-4. [DOI] [PubMed] [Google Scholar]
  5. Burrell H. R., Heacock A. M., Water R. D., Agranoff B. W. Increased tubulin messenger RNA in the goldfish retina during optic nerve regeneration. Brain Res. 1979 Jun 8;168(3):628–632. doi: 10.1016/0006-8993(79)90318-4. [DOI] [PubMed] [Google Scholar]
  6. CRAGG B. G., THOMAS P. K. Changes in conduction velocity and fibre size proximal to peripheral nerve lesions. J Physiol. 1961 Jul;157:315–327. doi: 10.1113/jphysiol.1961.sp006724. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Carpenter S. Proximal axonal enlargement in motor neuron disease. Neurology. 1968 Sep;18(9):841–851. doi: 10.1212/wnl.18.9.841. [DOI] [PubMed] [Google Scholar]
  8. Castle A. G., Crawford N. The [3H]colchicine-binding properties of platelet tubulin. Int J Biochem. 1978;9(6):439–447. doi: 10.1016/0020-711x(78)90058-7. [DOI] [PubMed] [Google Scholar]
  9. Chiu F. C., Korey B., Norton W. T. Intermediate filaments from bovine, rat, and human CNS: mapping analysis of the major proteins. J Neurochem. 1980 May;34(5):1149–1159. doi: 10.1111/j.1471-4159.1980.tb09954.x. [DOI] [PubMed] [Google Scholar]
  10. Cork L. C., Griffin J. W., Munnell J. F., Lorenz M. D., Adams R. J. Hereditary canine spinal muscular atrophy. J Neuropathol Exp Neurol. 1979 May;38(3):209–221. doi: 10.1097/00005072-197905000-00002. [DOI] [PubMed] [Google Scholar]
  11. Farel P. B. Reflex activity of regenerating frog spinal motoneurons. Brain Res. 1978 Dec 15;158(2):331–341. doi: 10.1016/0006-8993(78)90679-0. [DOI] [PubMed] [Google Scholar]
  12. Friede R. L., Samorajski T. Axon caliber related to neurofilaments and microtubules in sciatic nerve fibers of rats and mice. Anat Rec. 1970 Aug;167(4):379–387. doi: 10.1002/ar.1091670402. [DOI] [PubMed] [Google Scholar]
  13. GRAY E. G. The spindle and extrafusal innervation of a frog muscle. Proc R Soc Lond B Biol Sci. 1957 May 7;146(924):416–430. doi: 10.1098/rspb.1957.0021. [DOI] [PubMed] [Google Scholar]
  14. GUTH L. Regeneration in the mammalian peripheral nervous system. Physiol Rev. 1956 Oct;36(4):441–478. doi: 10.1152/physrev.1956.36.4.441. [DOI] [PubMed] [Google Scholar]
  15. Giller E., Jr, Schwartz J. H. Choline acetyltransferase in identified neurons of abdominal ganglion of Aplysia californica. J Neurophysiol. 1971 Jan;34(1):93–107. doi: 10.1152/jn.1971.34.1.93. [DOI] [PubMed] [Google Scholar]
  16. Giulian D., Des Ruisseux H., Cowburn D. Biosynthesis and intra-axonal transport of proteins during neuronal regeneration. J Biol Chem. 1980 Jul 10;255(13):6494–6501. [PubMed] [Google Scholar]
  17. Gozes I., Richter-Landsberg C. Identification of tubulin associated with rat brain myelin. FEBS Lett. 1978 Nov 1;95(1):169–172. doi: 10.1016/0014-5793(78)80076-3. [DOI] [PubMed] [Google Scholar]
  18. Grafstein B., Forman D. S. Intracellular transport in neurons. Physiol Rev. 1980 Oct;60(4):1167–1283. doi: 10.1152/physrev.1980.60.4.1167. [DOI] [PubMed] [Google Scholar]
  19. Grafstein B. The nerve cell body response to axotomy. Exp Neurol. 1975 Sep;48(3 Pt 2):32–51. doi: 10.1016/0014-4886(75)90170-3. [DOI] [PubMed] [Google Scholar]
  20. Hall M. E., Wilson D. L., Stone G. C. Changes in synthesis of specific proteins following axotomy: detection with two-dimensional gel electrophoresis. J Neurobiol. 1978 Sep;9(5):353–366. doi: 10.1002/neu.480090503. [DOI] [PubMed] [Google Scholar]
  21. Heacock A. M., Agranoff B. W. Enhanced labeling of a retinal protein during regeneration of optic nerve in goldfish. Proc Natl Acad Sci U S A. 1976 Mar;73(3):828–832. doi: 10.1073/pnas.73.3.828. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Hester S., Capps-Covey P., McIlwain D. L. The plasma membrane of bulk-isolated mature spinal neurons. Brain Res. 1978 Dec 22;159(1):41–54. doi: 10.1016/0006-8993(78)90108-7. [DOI] [PubMed] [Google Scholar]
  23. Hoffman P. N., Lasek R. J. Axonal transport of the cytoskeleton in regenerating motor neurons: constancy and change. Brain Res. 1980 Dec 8;202(2):317–333. doi: 10.1016/0006-8993(80)90144-4. [DOI] [PubMed] [Google Scholar]
  24. Härkönen M. H., Kauffman F. C. Metabolic alterations in the axotomized superior cervical ganglion of the rat. II. The pentose phosphate pathway. Brain Res. 1974 Jan 4;65(1):141–157. doi: 10.1016/0006-8993(74)90341-2. [DOI] [PubMed] [Google Scholar]
  25. Kreutzberg G. W., Schubert P. Volume changes in the axon during regeneration. Acta Neuropathol. 1971;17(3):220–226. doi: 10.1007/BF00685055. [DOI] [PubMed] [Google Scholar]
  26. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  27. Lazarides E. Intermediate filaments as mechanical integrators of cellular space. Nature. 1980 Jan 17;283(5744):249–256. doi: 10.1038/283249a0. [DOI] [PubMed] [Google Scholar]
  28. Lieberman A. R. The axon reaction: a review of the principal features of perikaryal responses to axon injury. Int Rev Neurobiol. 1971;14:49–124. doi: 10.1016/s0074-7742(08)60183-x. [DOI] [PubMed] [Google Scholar]
  29. Luine V. N., Kauffman F. C. Triphosphopyridine nucleotide-dependent enzymes in the developing spinal cord of the rabbit. J Neurochem. 1971 Jun;18(6):1113–1124. doi: 10.1111/j.1471-4159.1971.tb12040.x. [DOI] [PubMed] [Google Scholar]
  30. MACKEY E. A., SPIRO D., WIENER J. A STUDY OF CHROMATOLYSIS IN DORSAL ROOT GANGLIA AT THE CELLULAR LEVEL. J Neuropathol Exp Neurol. 1964 Jul;23:508–526. doi: 10.1097/00005072-196407000-00008. [DOI] [PubMed] [Google Scholar]
  31. Murray M., Forman D. S. Fine structural changes in goldfish retinal ganglion cells during axonal regeneration. Brain Res. 1971 Sep 24;32(2):287–298. doi: 10.1016/0006-8993(71)90325-8. [DOI] [PubMed] [Google Scholar]
  32. Murray M. Regeneration of retinal axons into the goldfish optic tectum. J Comp Neurol. 1976 Jul 15;168(2):175–195. doi: 10.1002/cne.901680202. [DOI] [PubMed] [Google Scholar]
  33. Nathaniel E. J., Nathaniel D. R. Electron microscopic studies of spinal ganglion cells following crushing of dorsal roots in adult rat. J Ultrastruct Res. 1973 Nov;45(3):168–182. doi: 10.1016/s0022-5320(73)80045-0. [DOI] [PubMed] [Google Scholar]
  34. Norton W. T., Autilio L. A. The lipid composition of purified bovine brain myelin. J Neurochem. 1966 Apr;13(4):213–222. doi: 10.1111/j.1471-4159.1966.tb06794.x. [DOI] [PubMed] [Google Scholar]
  35. Oakley B. R., Kirsch D. R., Morris N. R. A simplified ultrasensitive silver stain for detecting proteins in polyacrylamide gels. Anal Biochem. 1980 Jul 1;105(2):361–363. doi: 10.1016/0003-2697(80)90470-4. [DOI] [PubMed] [Google Scholar]
  36. Perry G. W., Wilson D. L. Protein synthesis and axonal transport during nerve regeneration. J Neurochem. 1981 Nov;37(5):1203–1217. doi: 10.1111/j.1471-4159.1981.tb04671.x. [DOI] [PubMed] [Google Scholar]
  37. Peters A., Vaughn J. E. Microtubules and filaments in the axons and astrocytes of early postnatal rat optic nerves. J Cell Biol. 1967 Jan;32(1):113–119. doi: 10.1083/jcb.32.1.113. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Price D. L., Porter K. R. The response of ventral horn neurons to axonal transection. J Cell Biol. 1972 Apr;53(1):24–37. doi: 10.1083/jcb.53.1.24. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Sims K. L., Kauffman F. C., Johnson E. C., Pickel V. M. Cytochemical localization of brain nicotinamide adenine dinucleotide phosphate (oxidized)-dependent dehydrogenases. Qualitative and quantitative distributions. J Histochem Cytochem. 1974 Jan;22(1):7–19. doi: 10.1177/22.1.7. [DOI] [PubMed] [Google Scholar]
  40. Sinicropi D. V., Kauffman F. C., Burt D. R. Axotomy in rat sympathetic ganglia: reciprocal effects on muscarinic receptor binding and 6-phosphogluconate dehydrogenase activity. Brain Res. 1979 Feb 9;161(3):560–565. doi: 10.1016/0006-8993(79)90688-7. [DOI] [PubMed] [Google Scholar]
  41. Sinicropi D. V., Kauffman F. C. Retrograde alteration of 6-phosphogluconate dehydrogenase in axotomized superior cervical ganglia of the rat. J Biol Chem. 1979 Apr 25;254(8):3011–3017. [PubMed] [Google Scholar]
  42. Sinicropi D. V., Michels K., McIlwain D. L. Acetylcholinesterase distribution in axotomized frog motoneurons. J Neurochem. 1982 Apr;38(4):1099–1105. doi: 10.1111/j.1471-4159.1982.tb05353.x. [DOI] [PubMed] [Google Scholar]
  43. Skene J. H., Willard M. Changes in axonally transported proteins during axon regeneration in toad retinal ganglion cells. J Cell Biol. 1981 Apr;89(1):86–95. doi: 10.1083/jcb.89.1.86. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Skene J. H., Willard M. Electrophoretic analysis of axonally transported proteins in toad retinal ganglion cells. J Neurochem. 1981 Jul;37(1):79–87. doi: 10.1111/j.1471-4159.1981.tb05293.x. [DOI] [PubMed] [Google Scholar]
  45. Turner J. E., Delaney R. K., Powell R. E. Retinal ganglion cell response to axotomy in the regenerating visual system of the newt (Triturus viridescens): an ultrastructural morphometric analysis. Exp Neurol. 1978 Nov;62(2):444–462. doi: 10.1016/0014-4886(78)90067-5. [DOI] [PubMed] [Google Scholar]
  46. Vance W. H., Clifton G. L., Applebaum M. L., Willis W. D., Jr, Coggeshall R. E. Unmyelinated preganglionic fibers in frog ventral roots. J Comp Neurol. 1975 Nov 1;164(1):117–125. doi: 10.1002/cne.901640110. [DOI] [PubMed] [Google Scholar]
  47. Waehneldt T. V., Malotka J. Comparative electrophoretic study of the Wolfgram proteins in myelin from several mammalia. Brain Res. 1980 May 12;189(2):582–587. doi: 10.1016/0006-8993(80)90373-x. [DOI] [PubMed] [Google Scholar]
  48. Watson W. E. Alteration of the adherence of glia to neurons following nerve injury. J Neurochem. 1966 Jun;13(6):536–537. doi: 10.1111/j.1471-4159.1966.tb09869.x. [DOI] [PubMed] [Google Scholar]
  49. Weil D. E., Busby W. H., Jr, McIlwain D. L. Choline acetyltransferase activity in large ventral spinal neurons. J Neurochem. 1977 Nov;29(5):847–852. doi: 10.1111/j.1471-4159.1977.tb10727.x. [DOI] [PubMed] [Google Scholar]
  50. Weil D. E., McIlwain D. L. Distribution of soluble proteins within spinal motoneurons: a quantitative two-dimensional electrophoretic analysis. J Neurochem. 1981 Jan;36(1):242–250. doi: 10.1111/j.1471-4159.1981.tb02400.x. [DOI] [PubMed] [Google Scholar]
  51. Yamada K. M., Spooner B. S., Wessells N. K. Ultrastructure and function of growth cones and axons of cultured nerve cells. J Cell Biol. 1971 Jun;49(3):614–635. doi: 10.1083/jcb.49.3.614. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Yen S. H., Fields K. L. Antibodies to neurofilament, glial filament, and fibroblast intermediate filament proteins bind to different cell types of the nervous system. J Cell Biol. 1981 Jan;88(1):115–126. doi: 10.1083/jcb.88.1.115. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES