Summary
We have studied the changes in the distribution of three intrinsic axonal components during the growth and maturation of sproutsin vivo. Neurofilaments, tubulin and synaptophysin, a synaptic vesicle protein, were visualized in motor axons and their sprouts using immunocytochemical staining of frozen longitudinal sections of muscle. We examined changes in these elements in sprouts regenerating after axonal crush injury and in those evoked from intact axons by denervation changes in muscle. Our results show that intrinsic axonal components move into newly formed motor axon sprouts in different temporal patterns. Based on the patterns of reorganization of staining of intrinsic axonal components, two types of outgrowth can be distinguished. One type, synaptic elaboration, is manifest by short, broad axonal processes that produce enlargement of the synaptic zone (synaptophysin staining) with little change in the distribution of intrinsic cytoskeletal elements. A second type of outgrowth, axonal elongation, occurs during axonal regeneration and ultraterminal sprouting and is longitudinal in form. In these sprouts there is a sequential appearance of neurofilament and then, several days later, tubulin immunostaining. Synaptophysin only accumulates in these sprouts after two weeks at points of synaptic contact with a muscle fibre.
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References
AGUAYO, A. J. (1985) Axonal regeneration from injured neurons in the adult mammalian central nervous system. InSynaptic Plasticity (edited by COTMAN, C. W.), pp. 457–84. New York: Guildford Press.
ALDERSON, K., PESTRONK, A., YEE, W. C. & DRACHMAN, D. B. (1989) Silver cholinesterase immunocytochemistry: a new neuromuscular junction stain.Muscle & Nerve 12, 9–14.
BAMBURG, J. R., BRAY, D. & CHAPMAN, K. (1986) Assembly of microtubules at the tip of growing axons.Nature 321, 788–90.
BENOWITZ, L. I., SHASHOUA, V. A. & YOON, M. G. (1981) Specific changes in transported proteins during regeneration of the goldfish optic nerve.Journal of Neuroscience 1, 300–7.
BJORNSKOV, E. K., NORRIS, F. H. & MOWER-KUBY, J. (1984) Quantitative axon terminal and end-plate morphology in amyotrophic lateral sclerosis.Archives of Neurology 41, 527–30.
BROWN, M. C. & HOLLAND, R. L. (1981) Motor nerve sprouting.Annual Review of Neuroscience 4, 17–42.
CARDASIS, C. A. & LAFONTAINE, D. M. (1987) Aging rat neuromuscular junctions: a morphometric study of cholinesterase-stained whole mounts and ultrastructure.Muscle & Nerve 10, 200–13.
CARLSEN, R. C., KIFF, J. & RYUGO, K. (1982) Suppression of the cell body response in axotomized frog spinal neurons does not prevent initiation of nerve regeneration.Brain Research 234, 11–25.
DANIELS, M. P. (1973) Fine structural changes in neurons and nerve fibres associated with colchicine inhibition of nerve fibre formationin vitro.Journal of Cell Biology 58, 463–70.
DUCHEN, L. W. (1970) Changes in motor innervation and cholinesterase localization induced by botulinum toxin in skeletal muscle of the mouse: differences between fast and slow muscles.Journal of Neurology, Neurosurgery and Psychiatry 33, 40–54.
DUCHEN, L. W. & STRITCH, S. J. (1968) The effects of botulinum toxin on the pattern of innervation of skeletal muscle in the mouse.Quarterly Journal of Experimental Physiology 53, 84–9.
FRIEDE, R. L. & BISCHHAUSEN, R. (1980) The fine structure of stumps of transected nerve fibres in subserial sections.Journal of the Neurological Sciences 44, 181–203.
GALLANT, P. R., PANT, H. C., PRUSS, R. M. & GAINER, H. (1986) Calcium-activated proteolysis of neurofilament proteins in the squid giant neuron.Journal of Neurochemistry 46, 1573–81.
HOFFMAN, P. N. (1988) Distinct roles of neurofilament and tubulin gene expression in axonal growth. InPlasticity of the Neuromuscular System, Ciba Foundation Symposium (edited by EVERED, D. & WHELAN, J.), pp. 192–204. Chichester, UK: John Wiley & Sons.
HOPKINS, W. G. & BROWN, M. C. (1985) Growth and remodelling of the neuromuscular synapse. InSynaptic Plasticity (edited by COTMAN, C. W.), pp. 169–99. New York: Guildford Press.
HOPKINS, W. G., BROWN, M. C. & KEYNES, R. J. (1985) Postnatal growth of motor nerve terminals in muscles of the mouse.Journal of Neurocytology 14, 525–40.
LASEK, R. J. & HOFFMAN, P. N. (1976) The neuronal cytoskeleton, axonal transport and axonal growth.Cold Spring Harbour Conference on Cell Proliferation 3, 1021–49.
LASEK, R. J. & KATZ, M. J. (1987) Mechanisms at the axon tip regulate metabolic processes critical to axonal elongation.Progress in Brain Research 71, 49–60.
LETINSKY, M. S., FISCHBECK, K. H. & MCMAHAN, U. J. (1976) Precision of reinnervation of original postsynaptic sites in frog muscle after a nerve crush.Journal of Neurocytology 5, 691–718.
LETOURNEAU, P. C. & RESSLER, A. H. (1984) Inhibition of neurite initiation and growth by taxol.Journal of Cell Biology 98, 1355–62.
MATUS, A., RIEDERER, B. & HUBER, G. (1987) Influence of monoclonal antibodies on microbutule assembly.Journal of Neurochemistry 49, 714–20.
MC KEITHAN, T. W. & ROSENBAUM, J. L. (1984) The biochemistry of microtubules: a review.Cell-Muscle Motility 5, 255–88.
MORRIS, J. R. & LASEK, R. J. (1982) Stable polymers of the axonal cytoskeleton: the axoplasmic ghost.Journal of Cellular Biology 92, 192–8.
NAVONE, F., JAHN, R., DIGIOIA, G., STUKENBROK, H., GREENGARD, P. & DE CAMILLI, P. (1987) Protein p38: an integral membrane protein specific for small vesicles of neurons and neuroendocrine cells.Journal of Cell Biology 106, 2511–27.
PAGGI, P. & LASEK, R. J. (1987) Axonal transport of cytoskeletal proteins in oculomotor axons and their residence times in the axon terminals.Journal of Neuroscience 7, 2397–411.
PERRY, G. W., BURMEISTER, D. W. & GRAFSTEIN, B. (1987) Fast axonally transported proteins in regenerating goldfish optic axons.Journal of Neuroscience 7, 792–806.
PESTRONK, A. & DRACHMAN, D. B. (1978a) Motor nerve sprouting and acetylcholine receptors.Science 199, 1223–5.
PESTRONK, A. & DRACHMAN, D. B. (1978b) A new stain for quantitative measurement of sprouting at neuromuscular junctions.Muscle & Nerve 1, 70–4.
PESTRONK, A. & DRACHMAN, D. B. (1984) Regulation of motor nerve outgrowth. InNeuromuscular Diseases (edited by SERRATRICE, G., CROS, D., DESNUELLE, C., GASTAUT, J.-L., PELLISIER, J.-F., POUGET, J. & SCHIANO, A.), pp. 407–13. New York: Raven Press.
PESTRONK, A. & DRACHMAN, D. B. (1985) Motor nerve terminal outgrowth and acetylcholine receptors: Inhibition of terminal outgrowth byα-bungarotoxin and anti-acetylcholine receptor antibody.Journal of Neuroscience 5, 751–8.
PESTRONK, A. & DRACHMAN, D. B. (1988) Motor nerve outgrowth: reduced capacity for sprouting in the terminals of longer axons.Brain Research 463, 218–22.
PESTRONK, A., DRACHMAN, D. B. & GRIFFIN, J. W. (1980) Effects of aging on nerve sprouting and regeneration.Experimental Neurology 70, 65–82.
PIPERNO, G., LEDIZET, M. & CHANG, X. (1987) Microtubules containing acetylated β-tubulin in mammalian cells in culture.Journal of Cell Biology 104, 289–302.
PURVES, D., VOYVODIC, J. T., MAGRASSI, L. & YAWO, H. (1987) Nerve terminal remodelling visualized in living mice by repeated examination of the same neuron.Science 238, 1122–6.
RAMONY CAJAL, S. (1928)Degeneration and Regeneration of the Nervous System. Translated by MAY, R. M.. Cambridge UK: Oxford University Press.
REH, T. A., REDSHAW, J. D. & BISBY, M. A. (1987) Axons of the pyramidal tract do not increase their transport of growth-associated proteins after axotomy.Molecular Brain Research 2, 1–6.
ROBBINS, N. & FAHIM, M. A. (1985) Progression of age changes in mature mouse motor nerve terminals and its relation to locomotor activity.Journal of Neurocytology 14, 1019–36.
ROOTS, B. I. (1983) Neurofilament accumulation induced in synapses by leupeptin.Science 221, 971–2.
ROSENHEIMER, J. L. & SMITH, D. O. (1985) Differential changes in the end-plate architecture of functionally diverse muscles during aging.Journal of Neurophysiology 53, 1567–81.
SANAKA, M., SHIBASAKU, T. & LEDERIS, K. (1987) Improved fixation and Cobalt-Glucose Oxidase-Diaminobenzidine intensification for immunohistochemical demonstration of corticotropin-releasing factor in rat brain.Journal of Histochemistry and Cytochemistry 35, 207–12.
SANES, J. R., MARSHALL, L. M. & MCMAHAN, U. J. (1978) Reinnervation of muscle fibre basal lamina after removal of myofibres. Differentiation of regenerating axons at original synaptic sites.Journal of Cell Biology 78, 176–98.
SKENE, J. H. P. & WILLARD, M. (1981) Axonally transported proteins associated with growth in rabbit central and peripheral nervous system.Journal of Cell Biology 89, 96–103.
SLACK, J. R. & POCKET, S. (1981) Terminal sprouting is a local response to a local stimulus.Brain Research 217, 368–74.
TUFFERY, A. R. (1971) Growth and degeneration of motor end-plates in normal cat hind limb muscles.Journal of Anatomy 110, 221–47.
VAN HARREVELD, A. (1945) Reinnervation of denervated muscle fibres by adjacent-functioning motor units.American Journal of Physiology 144, 477–93.
WIEDENMANN, B., FRANKE, W. W., KUHN, C., MOLL, R. & GOULD, V. E. (1986) Synaptophysin: a marker protein for neuroendocrine cells and neoplasms.Proceedings of the National Academy of Sciences, USA 83, 3500–4.
YEE, W. C., PESTRONK, A., ALDERSON, K. & YUAN, C. M. (1988) Regional heterogeneity in the distal motor axon: three zones with distinctive intrinsic components.Journal of Neurocytology 17, 649–56.
YEE, W. C. & PESTRONK, A. (1987) Mechanisms of postsynaptic plasticity: Remodelling of the junctional acetylcholine receptor cluster induced by motor nerve terminal outgrowth.Journal of Neuroscience 7, 2019–24.
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Alderson, K., Yee, W.C. & Pestronk, A. Reorganization of intrinsic components in the distal motor axon during outgrowth. J Neurocytol 18, 541–552 (1989). https://doi.org/10.1007/BF01474549
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DOI: https://doi.org/10.1007/BF01474549