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. 1973 Nov 1;59(2):456–470. doi: 10.1083/jcb.59.2.456

EFFECTS OF CALCIUM ION CONCENTRATION ON THE DEGENERATION OF AMPUTATED AXONS IN TISSUE CULTURE

W W Schlaepfer 1, R P Bunge 1
PMCID: PMC2109098  PMID: 4805010

Abstract

Light and electron microscope studies were conducted on the nature of the degenerative changes in amputated nerve fibers of cultured rat sensory ganglia and on the effects of media with differing calcium concentrations upon these changes. With glucose-enriched Eagle's media (MEM) containing 1.6 mM calcium, the amputated myelinated and unmyelinated axons undergo a progressive granular disintegration of their axoplasm with collapse and fragmentation of myelin sheaths between 6 and 24 h after transection. With MEM containing only 25–50 µM calcium, the granular axoplasmic degeneration does not occur in transected fibers and they retain their longitudinal continuity and segmental myelin ensheathment for at least 48 h. Addition of 6 mM EGTA to MEM (reducing the estimated Ca++ below 0.3 µM) results in the structural preservation of both microtubules and neurofilaments within transected axons. A transient focal swelling of amputated axons occurs, however, in cultures with normal and reduced calcium. These observations suggest that an alteration in the permeability of the axolemma is a crucial initiating event leading to axonal degenerative changes distal to nerve transection. The loss of microtubules and neurofilaments and the associated granular alterations of the axoplasm in transected fibers appears to result from the influx of calcium into the axoplasm.

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Selected References

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  1. Baker P. F., Foster R. F., Gilbert D. S., Shaw T. I. Sodium transport by perfused giant axons of Loligo. J Physiol. 1971 Dec;219(2):487–506. doi: 10.1113/jphysiol.1971.sp009674. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Baker P. F., Hodgkin A. L., Ridgway E. B. Depolarization and calcium entry in squid giant axons. J Physiol. 1971 Nov;218(3):709–755. doi: 10.1113/jphysiol.1971.sp009641. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Birnberger A. C., Birnberger K. L., Eliasson S. G., Simpson P. C. Effect of cyanide and electrical stimulation on phosphoinositide metabolism in lobster nerves. J Neurochem. 1971 Jul;18(7):1291–1298. doi: 10.1111/j.1471-4159.1971.tb00228.x. [DOI] [PubMed] [Google Scholar]
  4. Blaustein M. P., Hodgkin A. L. The effect of cyanide on the efflux of calcium from squid axons. J Physiol. 1969 Feb;200(2):497–527. doi: 10.1113/jphysiol.1969.sp008704. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Borisy G. G., Olmsted J. B. Nucleated assembly of microtubules in porcine brain extracts. Science. 1972 Sep 29;177(4055):1196–1197. doi: 10.1126/science.177.4055.1196. [DOI] [PubMed] [Google Scholar]
  6. Bunge R. P., Wood P. Studies on the transplantation of spinal cord tissue in the rat. I. The development of a culture system for hemisections of embryonic spinal cord. Brain Res. 1973 Jul 27;57(2):261–276. doi: 10.1016/0006-8993(73)90135-2. [DOI] [PubMed] [Google Scholar]
  7. CHENG S. C. Metabolism of frog nerve during activity and recovery. J Neurochem. 1961 Aug;7:278–288. doi: 10.1111/j.1471-4159.1961.tb13514.x. [DOI] [PubMed] [Google Scholar]
  8. DAHL N. A., SAMSON F. E., Jr, BALFOUR W. M. ADENOSINE TRIPHOSPHATE AND ELECTRICAL ACTIVITY IN CHICKEN VAGUS. Am J Physiol. 1964 Apr;206:818–822. doi: 10.1152/ajplegacy.1964.206.4.818. [DOI] [PubMed] [Google Scholar]
  9. Dawson R. M. Metabolism and function of polyphosphoinositides in nervous tissue. Ann N Y Acad Sci. 1969 Oct 17;165(2):774–783. [PubMed] [Google Scholar]
  10. Durell J., Garland J. T. Acetylcholine-stimulated phosphodiesteratic cleavage of phosphoinositides: hypothetical role in membrane depolarization. Ann N Y Acad Sci. 1969 Oct 17;165(2):743–754. [PubMed] [Google Scholar]
  11. EAGLE H. Amino acid metabolism in mammalian cell cultures. Science. 1959 Aug 21;130(3373):432–437. doi: 10.1126/science.130.3373.432. [DOI] [PubMed] [Google Scholar]
  12. GREENGARD P., BRINK F., Jr, COLOWICK S. P. Some relationships between action potential, oxygen consumption and coenzyme content in degenerating peripheral axons. J Cell Physiol. 1954 Dec;44(3):395–420. doi: 10.1002/jcp.1030440304. [DOI] [PubMed] [Google Scholar]
  13. GREENGARD P., STRAUB R. W. Effect of frequency of electrical stimulation on the concentration of intermediary metabolites in mammalian non-myelinated fibres. J Physiol. 1959 Oct;148:353–361. doi: 10.1113/jphysiol.1959.sp006292. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. HODGKIN A. L., KEYNES R. D. Movements of labelled calcium in squid giant axons. J Physiol. 1957 Sep 30;138(2):253–281. doi: 10.1113/jphysiol.1957.sp005850. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Heuser J., Katz B., Miledi R. Structural and functional changes of frog neuromuscular junctions in high calcium solutions. Proc R Soc Lond B Biol Sci. 1971 Sep 28;178(1053):407–415. doi: 10.1098/rspb.1971.0072. [DOI] [PubMed] [Google Scholar]
  16. Holtzman E., Novikoff A. B. Lysomes in the rat sciatic nerve following crush. J Cell Biol. 1965 Dec;27(3):651–669. doi: 10.1083/jcb.27.3.651. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Huneeus F. C., Davison P. F. Fibrillar proteins from squid axons. I. Neurofilament protein. J Mol Biol. 1970 Sep 28;52(3):415–428. doi: 10.1016/0022-2836(70)90410-9. [DOI] [PubMed] [Google Scholar]
  18. JOHNSON A. C., McNABB A. R., ROSSITER R. J. Chemical studies of peripheral nerve during Wallerian degeneration; lipids. Biochem J. 1949;45(4):500–508. doi: 10.1042/bj0450500. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. KEYNES R. D., LEWIS P. R. The intracellular calcium contents of some invertebrate nerves. J Physiol. 1956 Nov 28;134(2):399–407. doi: 10.1113/jphysiol.1956.sp005652. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Kai M., Hawthorne J. N. Physiological significance of polyphosphoinositides in brain. Ann N Y Acad Sci. 1969 Oct 17;165(2):761–773. [PubMed] [Google Scholar]
  21. Katz B., Miledi R. A study of synaptic transmission in the absence of nerve impulses. J Physiol. 1967 Sep;192(2):407–436. doi: 10.1113/jphysiol.1967.sp008307. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Katz B., Miledi R. Tetrodotoxin-resistant electric activity in presynaptic terminals. J Physiol. 1969 Aug;203(2):459–487. doi: 10.1113/jphysiol.1969.sp008875. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Katz B., Miledi R. The effect of prolonged depolarization on synaptic transfer in the stellate ganglion of the squid. J Physiol. 1971 Jul;216(2):503–512. doi: 10.1113/jphysiol.1971.sp009537. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. LEE J. C. Electron microscopy of Wallerian degeneration. J Comp Neurol. 1963 Feb;120:65–79. doi: 10.1002/cne.901200107. [DOI] [PubMed] [Google Scholar]
  25. MANNELL W. A. Wallerian degeneration in the rat; a chemical study. Can J Med Sci. 1952 Jun;30(3):173–179. doi: 10.1139/cjms52-025. [DOI] [PubMed] [Google Scholar]
  26. Mire J. J., Hendelman W. J., Bunge R. P. Observations on a transient phase of focal swelling in degenerating unmyelinated nerve fibers. J Cell Biol. 1970 Apr;45(1):9–22. doi: 10.1083/jcb.45.1.9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. NATHANIEL E. J. PEASE DC: DEGENERATIVE CHANGES IN RAT DORSAL ROOTS DURING WALLERIAN DEGENERATION. J Ultrastruct Res. 1963 Dec;52:511–532. doi: 10.1016/s0022-5320(63)80082-9. [DOI] [PubMed] [Google Scholar]
  28. OHMI S. Electron microscopic study of wallerian degeneration of the peripheral nerve. Z Zellforsch Mikrosk Anat. 1961;54:39–67. doi: 10.1007/BF00384198. [DOI] [PubMed] [Google Scholar]
  29. Okada Y., McDougal D. B., Jr Physiological and biochemical changes in frog sciatic nerve during anoxia and recovery. J Neurochem. 1971 Dec;18(12):2335–2353. doi: 10.1111/j.1471-4159.1971.tb00189.x. [DOI] [PubMed] [Google Scholar]
  30. PETERSON E. R., MURRAY M. R. PATTERNS OF PERIPHERAL DEMYELIMINATION IN VITRO. Ann N Y Acad Sci. 1965 Mar 31;122:39–50. [PubMed] [Google Scholar]
  31. PORTZEHL H., CALDWELL P. C., RUEEGG J. C. THE DEPENDENCE OF CONTRACTION AND RELAXATION OF MUSCLE FIBRES FROM THE CRAB MAIA SQUINADO ON THE INTERNAL CONCENTRATION OF FREE CALCIUM IONS. Biochim Biophys Acta. 1964 May 25;79:581–591. doi: 10.1016/0926-6577(64)90224-4. [DOI] [PubMed] [Google Scholar]
  32. Redman C. M. Phospholipid metabolism in intact and modified erythrocyte membranes. J Cell Biol. 1971 Apr;49(1):35–49. doi: 10.1083/jcb.49.1.35. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Redman C. M. Proteolipid involvement in human erythrocyte membrane function. Biochim Biophys Acta. 1972 Sep 1;282(1):123–134. doi: 10.1016/0005-2736(72)90316-1. [DOI] [PubMed] [Google Scholar]
  34. Schlaepfer W. W. Experimental alterations of neurofilaments and neurotubules by calcium and other ions. Exp Cell Res. 1971 Jul;67(1):73–80. doi: 10.1016/0014-4827(71)90622-7. [DOI] [PubMed] [Google Scholar]
  35. Schlaepfer W. W. Vincristine-induced axonal alterations in rat peripheral nerve. J Neuropathol Exp Neurol. 1971 Jul;30(3):488–505. doi: 10.1097/00005072-197107000-00012. [DOI] [PubMed] [Google Scholar]
  36. Stewart M. A., Passonneau J. V., Lowry O. H. Substrate changes in peripheral nerve during ischaemia and Wallerian degeneration. J Neurochem. 1965 Aug;12(8):719–727. doi: 10.1111/j.1471-4159.1965.tb06786.x. [DOI] [PubMed] [Google Scholar]
  37. VIAL J. D. The early changes in the axoplasm during wallerian degeneration. J Biophys Biochem Cytol. 1958 Sep 25;4(5):551–555. doi: 10.1083/jcb.4.5.551. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Weddell G., Glees P. The early stages in the degeneration of cutaneous nerve fibres. J Anat. 1941 Oct;76(Pt 1):65–93. [PMC free article] [PubMed] [Google Scholar]
  39. Weisenberg R. C. Microtubule formation in vitro in solutions containing low calcium concentrations. Science. 1972 Sep 22;177(4054):1104–1105. doi: 10.1126/science.177.4054.1104. [DOI] [PubMed] [Google Scholar]
  40. Weisenberg R. C., Timasheff S. N. Aggregation of microtubule subunit protein. Effects of divalent cations, colchicine and vinblastine. Biochemistry. 1970 Oct 13;9(21):4110–4116. doi: 10.1021/bi00823a012. [DOI] [PubMed] [Google Scholar]
  41. Wilson L., Bryan J., Ruby A., Mazia D. Precipitation of proteins by vinblastine and calcium ions. Proc Natl Acad Sci U S A. 1970 Jul;66(3):807–814. doi: 10.1073/pnas.66.3.807. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Zelená J., Lubińska L., Gutmann E. Accumulation of organelles at the ends of interrupted axons. Z Zellforsch Mikrosk Anat. 1968;91(2):200–219. doi: 10.1007/BF00364311. [DOI] [PubMed] [Google Scholar]

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