Cellular and Molecular Neurobiology

, Volume 25, Issue 2, pp 441–450 | Cite as

Repair and Regeneration of Functional Synaptic Connections: Cellular and Molecular Interactions in the Leech

  • Yuanli Duan
  • Joseph Panoff
  • Brian D. Burrell
  • Christie L. Sahley
  • Kenneth J. Muller


A major problem for neuroscience has been to find a means to achieve reliable regeneration of synaptic connections following injury to the adult CNS. This problem has been solved by the leech, where identified neurons reconnect precisely with their usual targets following axotomy, re-establishing in the adult the connections formed during embryonic development.

It cannot be assumed that once axons regenerate specific synapses, function will be restored. Recent work on the leech has shown following regeneration of the synapse between S-interneurons, which are required for sensitization of reflexive shortening, a form of non-associative learning, the capacity for sensitization is delayed.

The steps in repair of synaptic connections in the leech are reviewed, with the aim of understanding general mechanisms that promote successful repair. New results are presented regarding the signals that regulate microglial migration to lesions, a first step in the repair process. In particular, microglia up to 900 μm from the lesion respond within minutes by moving rapidly toward the injury, controlled in part by nitric oxide (NO), which is generated immediately at the lesion and acts via a soluble guanylate cyclase (sGC). The cGMP produced remains elevated for hours after injury. The relationship of microglial migration to axon outgrowth is discussed.


nerve regeneration microglia nitric oxide cell migration time-lapse video neuronal plasticity non-associative learning sensitization 


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  1. Arechiga, H. (1993). Circadian rhythms. Curr. Opin. Neurobiol. 3:1005–1010.PubMedGoogle Scholar
  2. Arechiga, H., and Rodriguez-Sosa, L. (1998). Circadian clock function in isolated eyestalk tissue of crayfish. Proc. Roy. Soc. Lond. B Biol. Sci. 265:1819–1823.Google Scholar
  3. Baccus, S. A., Burrell, B. D., Sahley, C. L., and Muller, K. J. (2000). Action potential reflection and failure at axon branch points cause stepwise changes in EPSPs in an interneuron essential for learning. J. Neurophysiol. 83:1693–1700.PubMedGoogle Scholar
  4. Baccus, S. A., Sahley, C. L., and Muller, K. J. (2001). Multiple sites of action potential initiation increase neuronal firing rate. J. Neurophysiol. 86:1226–1236.PubMedGoogle Scholar
  5. Boulis, N. M., and Sahley, C. L. (1988). A behavioral analysis of habituation and sensitization of shortening in the semi-intact leech. J. Neurosci. 8:4621–4627.PubMedGoogle Scholar
  6. Burrell, B. D., Sahley, C. L., and Muller, K. J. (2001). Non-associative learning and serotonin induce similar bi-directional changes in excitability of a neuron critical for learning in the medicinal leech. J. Neurosci. 21:1401–1412.PubMedGoogle Scholar
  7. Burrell, B. D., Sahley, C. L., and Muller, K. J. (2002). Differential effects of serotonin enhance activity of an electrically coupled neural network. J. Neurophysiol. 87:2889–2895.PubMedGoogle Scholar
  8. Burrell, B. D., Sahley, C. L., and Muller, K. J. (2003). Progressive recovery of learning during regeneration of a single synapse in the medicinal leech. J. Comp. Neurol. 457:67–74.PubMedGoogle Scholar
  9. Camhi, J. M. (1993). Neural mechanisms of behavior. Curr. Opin. Neurobiol. 3:1011–1019.PubMedGoogle Scholar
  10. Carew, T. J., and Sahley, C. L. (1986). Learning in invertebrates: From behavior to molecules. Ann. Rev. Neurosci. 9:435–487.PubMedGoogle Scholar
  11. Chen, A., Kumar, S. M., Sahley, C. L., and Muller, K. J. (2000). Nitric oxide influences injury-induced microglial migration and accumulation in the leech CNS. J. Neurosci. 20:1036–1043.PubMedGoogle Scholar
  12. Duan, Y., Haugabook, S. J., Sahley, C. L., and Muller, K. J. (2003). Methylene blue blocks cGMP production and disrupts directed migration of microglia to nerve lesions in the leech CNS. J. Neurobiol.Google Scholar
  13. Ehrlich, J. S., Boulis, N. M., Karrer, T., and Sahley, C. L. (1992). Differential effects of serotonin depletion on sensitization and dishabituation in the leech, Hirudo medicinalis. J. Neurobiol. 23:270–279.Google Scholar
  14. Fernandez-de-Miguel, F., and Drapeau, P. (1995). Synapse formation and function: Insights from identified leech neurons in culture. J. Neurobiol. 27:367–379.PubMedGoogle Scholar
  15. Fernandez-de-Miguel, F., and Vargas, J. (1997). Different determinants on growth and synapse formation in cultured neurons. NeuroReport 8:761–765.PubMedGoogle Scholar
  16. Filbin, M. T. (2003). Myelin-associated inhibitors of axonal regeneration in the adult mammalian CNS. Nat. Rev. Neurosci. 4:703–713.PubMedGoogle Scholar
  17. Gu, X., Macagno, E. R., and Muller, K. J. (1989). Laser microbeam axotomy and conduction block show that electrical transmission at a central synapse is distributed at multiple contacts. J. Neurobiol. 20:422–434.PubMedGoogle Scholar
  18. Kimmel, A. R., and Parent, C. A. (2003). The signal to move: D. discoideum go orienteering. Science 300:1525–1527.PubMedGoogle Scholar
  19. Koeberle, P. D., and Bähr, M. (2004). Growth and guidance cues for regenerating axons: Where have they gone? J. Neurobiol. 59:162–180.PubMedGoogle Scholar
  20. Kristan, W. B., Jr., Lockery, S. R., and Lewis, J. E. (1995). Using reflexive behaviors of the medicinal leech to study information processing. J. Neurobiol. 27:380–389.PubMedGoogle Scholar
  21. Kumar, S. M., Porterfield, D. M., Muller, K. J., Smith, P. J., and Sahley, C. L. (2001). Nerve injury induces a rapid efflux of nitric oxide (NO) detected with a novel NO microsensor. J. Neurosci. 21:215–220.PubMedGoogle Scholar
  22. Kuno, M., and Llinás, R. (1970). Alterations of synaptic action on chromatolysed motoneurones of the cat. J. Physiol. (Lond.) 210:823–828.Google Scholar
  23. Lewis, J. E., and Kristan, W. B., Jr. (1998). A neuronal network for computing population vectors in the leech. Nature 391:76–79.PubMedGoogle Scholar
  24. Luebke, A. E., Dickerson, I. M., and Muller, K. J. (1995). In situ hybridization reveals transient laminin B-chain expression by individual glial and muscle cells in embryonic leech CNS. J. Neurobiol. 27:1–14.PubMedGoogle Scholar
  25. Mar, A., and Drapeau, P. (1996). Modulation of conduction block in leech mechanosensory neurons. J. Neurosci. 16:4335–4343.PubMedGoogle Scholar
  26. Mason, A., and Muller, K. J. (1996). Accurate synapse regeneration despite ablation of the distal axon segment. Eur. J. Neurosci. 8:11–20.PubMedGoogle Scholar
  27. Masuda-Nakagawa, L. M., Muller, K. J., and Nicholls, J. G. (1990). Accumulation of laminin and microglial cells at sites of injury and regeneration in the central nervous system of the leech. Proc. Roy. Soc. Lond. B 241:201–206.Google Scholar
  28. Masuda-Nakagawa, L. M., Muller, K. J., and Nicholls, J. G. (1993). Axonal sprouting and laminin appearance after destruction of glial sheaths. Proc. Natl. Acad. Sci. USA 90:4966–4970.PubMedGoogle Scholar
  29. McGlade-McCulloh, E., Morrissey, A. M., Norona, F., and Muller, K. J. (1989). Individual microglia move rapidly and directly to nerve lesions in the leech central nervous system. Proc. Natl. Acad. Sci. USA 86:1093–1097.PubMedGoogle Scholar
  30. McMahan, U. J., Edgington, D. R., and Kuffler, D. P. (1980). Factors that influence regeneration of the neuromuscular junction. J. Exp. Biol. 89:31–42.PubMedGoogle Scholar
  31. Modney, B. K., Sahley, C. L., and Muller, K. J. (1997). Regeneration of a central synapse restores non-associative learning. J. Neurosci. 17:6478–6482.PubMedGoogle Scholar
  32. Morgese, V. J., Elliott, E. J., and Muller, K. J. (1983). Microglial movement to sites of nerve lesion in the leech CNS. Brain Res. 272:166–170.PubMedGoogle Scholar
  33. Muller, K. J., and Carbonetto, S. (1979). The morphological and physiological properties of a regenerating synapse in the C.N.S. of the leech. J. Comp. Neurol. 185:485–516.PubMedGoogle Scholar
  34. Nakajima, K., and Kohsaka, S. (2004). Microglia: Neuroprotective and neurotrophic cells in the central nervous system. Curr. Drug Targets Cardiovasc. Haematol. Disord. 4:65–84.PubMedGoogle Scholar
  35. Nicholls, J. G. (1987). The Search for Connections: Study of Regeneration in the Nervous System of the Leech. Magnes Lecture Series: Vol. II, Sinauer Associates Inc., Sunderland, Massachusetts, 84 pp.Google Scholar
  36. Nicholls, J. G., Martin, A. R., Wallace, B. G., and Fuchs, P. A. (2001). From Neuron to Brain, Sinauer Associates, Sunderland, MA, 580 pp.Google Scholar
  37. Perry, V. H., and Gordon, S. (1997). Microglia and macrophages. In Keane, R. W., and Hickey, W. F. (eds.), Immunology of the Nervous System, Oxford University Press, New York, pp. 155–172.Google Scholar
  38. Rotshenker, S. (2003). Microglia and macrophage activation and the regulation of complement-receptor-3 (CR3/MAC-1)-mediated myelin phagocytosis in injury and disease. J. Mol. Neurosci. 21:65–72.PubMedGoogle Scholar
  39. Sahley, C. L., Modney, B. K., Boulis, N. M., and Muller, K. J. (1994). The S cell: An interneuron essential for sensitization and full dishabituation of leech shortening. J. Neurosci. 14:6715–6721.PubMedGoogle Scholar
  40. Sahley, C. L., and Ready, D. F. (1988). Associative learning modifies two behaviors in the leech, Hirudo medicinalis. J. Neurosci. 8:4612–4620.Google Scholar
  41. Sanes, D. H., Reh, T. A., and Harris, W. A. (2000). Development of the Nervous System, Academic Press, San Diego, 500 pp.Google Scholar
  42. Scott, S. A., and Muller, K. J. (1980). Synapse regeneration and signals for directed axonal growth in the C.N.S. of the leech. Dev. Biol. 80:345–363.PubMedGoogle Scholar
  43. Shafer, O. T., Chen, A., Kumar, S. M., Muller, K. J., and Sahley, C. L. (1998). Injury-induced expression of endothelial nitric oxide synthase by glial and microglial cells in the leech central nervous system within minutes after injury. Proc. Roy. Soc. Lond. B 265:2171–2175.Google Scholar
  44. Shaw, B. K., and Kristan, W. B., Jr. (1995). The whole-body shortening reflex of the medicinal leech: Motor pattern, sensory basis, and interneuronal pathways. J. Comp. Physiol. (A) 177:667–681.Google Scholar
  45. Shaw, B. K., and Kristan, W. B., Jr. (1999). Relative roles of the S cell network and parallel interneuronal pathways in the whole-body shortening reflex of the medicinal leech. J. Neurophysiol. 82:1114–1123.PubMedGoogle Scholar
  46. Smith, P. J. S., Howes, E. A., and Treherne, J. E. (1987). Mechanisms of glial regeneration in an insect central nervous system. J. Exp. Biol. 132:59–78.PubMedGoogle Scholar
  47. Varga, Z. M., Schwab, M. E., and Nicholls, J. G. (1995). Myelin-associated neurite growth-inhibitory proteins and suppression of regeneration of immature mammalian spinal cord in culture. Proc. Natl. Acad. Sci. USA 92:10959–10963.PubMedGoogle Scholar
  48. von Bernhardi, R., and Muller, K. J. (1995). Repair of the central nervous system: Lessons from lesions in leeches. J. Neurobiol. 27:353–366.PubMedGoogle Scholar
  49. Wu, J. Y., Cohen, L. B., and Falk, C. X. (1994). Neuronal activity during different behaviors in Aplysia: A distributed organization? Science 263:820–823.PubMedGoogle Scholar
  50. Zecevic, D., Wu, J.-Y., Cohen, L. B., London, J. A., Höpp, H.-P., and Falk, C. X. (1989). Hundreds of neurons in the Aplysia abdominal ganglion are active during the gill-withdrawal reflex. J. Neurosci. 9:3681–3689.PubMedGoogle Scholar

Copyright information

© Springer Science + Business Media, Inc. 2005

Authors and Affiliations

  • Yuanli Duan
    • 1
  • Joseph Panoff
    • 1
  • Brian D. Burrell
    • 2
  • Christie L. Sahley
    • 3
  • Kenneth J. Muller
    • 1
    • 4
    • 5
  1. 1.Department of Physiology & BiophysicsUniversity of Miami School of MedicineMiami
  2. 2.Neuroscience Group, Division of Basic Biomedical SciencesUniversity of South Dakota School of MedicineVermillion
  3. 3.Department of Biological SciencesPurdue UniversityWest Lafayette
  4. 4.Neuroscience ProgramUniversity of Miami School of MedicineMiami
  5. 5.RMSB 5089, Department of Physiology and BiophysicsUniversity of Miami School of MedicineMiami

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