Spatio-Temporal Changes in Neurofilament Proteins Immunoreactivity Following Kainate-Induced Cerebellar Lesion in Rats

Abstract

1. Spatio-temporal changes in phosphorylated (pNFP) and nonphosphorylated (npNFP) neurofilament proteins were assessed immunocytochemicaly in adult rat cerebe- llum, 2–30 days following unilateral injection of kainic acid (KA) or physiological saline (SC).

2. Analysis of the staining intensity and pattern demonstrated that injection of both KA and physiological saline elicited significant and long-lasting increase of pNFP and npNFP immunoreactivity, at the ipsilateral, and to lesser extent at the contralateral side of lesion.

3. Kainate intoxication induced abundant expression of pNFP and npNFP in cerebellar white matter, as well as in all layers of perilesioned cortex. Higher pNFP expression was evidenced in the Purkinje cell layer, particularly at cell bodies, initial segments, and proximal dendrites, which normally do not contain pNFP. In addition, synaptophysin immunocytochemistry was used as a marker of synaptogenesis and plasticity.

4. Spatio-temporal pattern of NFP and synaptophysin expression suggests that perilesioned cortex undergoes dynamic changes following brain demage and possess a reparative capacity to abridge the consequences of brain trauma.

This is a preview of subscription content, access via your institution.

REFERENCES

  1. Angaut, P., Alvarado-Mallart, R. M., and Sotelo, C. (1982). Ultrastructural evidence for compensatory sprouting of climbing and mossy afferents to the cerebellar hemisphere after ipsilateral pedunculotomy in the newborn rat. J. Comp. Neurol. 205:101-111.

    Google Scholar 

  2. Azizi, S. A., and Woodward, D. J. (1987). Inferior olivary nuclear complex of the rat: Morphology and comments on the principle of organization within the olivocerebellar system. J. Comp. Neurol. 263:467-484.

    Google Scholar 

  3. Bidmon, H. J., Jancsik, V., Schleicher, A., Hagemann, G., Witte, O. W., Woodhams, P., and Zilles, K. (1997). Structural alterations and changes in cytoskeletal proteins and proteoglycans after focal cortical ischemia. Neuroscience 82:397-420.

    Google Scholar 

  4. Black, M. M., and Lee, V. M. Y. (1988). Phosphorylation of neurofilament proteins in intact neurons: Demonstration of phosphorylation in cell bodies and axons. J. Neurosci. 8:3296-3305.

    Google Scholar 

  5. Chen, S., and Hillman, E. D. (1982). Plasticity of the parallel fiber-Purkinje cell synapse takeover and new synapse formation in the adult rat. Brain Res. 240:205-220.

    Google Scholar 

  6. Chen, S., Pickard, J. D., and Harris, N. G. (2003). Time course of cellular pathology after control cortical impact injury. Exp. Neurol. 182:87-102.

    Google Scholar 

  7. Deller, T., Haas, C. A., and Frotscher, M. (2000). Reorganization of the rat fascia dentate after unilateral entorhinal cortex lesion. Role of the extracellular matrix. Ann. N. Y. Acad. Sci. 911:207-220.

    Google Scholar 

  8. De Waegh, S. M., Lee, V. M. Y., and Brady, S. T. (1992). Local modulation of neurofilament phosphorylation, axonal calibre and slow axonal transport by myelinating Schwann cells. Cell 68:451-463.

    Google Scholar 

  9. Elhanany, E., Jaffe, H., Link, W. T., Shelly, D. M., Gainer, H., and Pant, H. C. (1994). Identification of endogenously phosphorylated KSP sites in the high molecular weight rat neurofilament protein. J. Neurochem. 63:2324-2335.

    Google Scholar 

  10. Foster, A. C., and Roberts, J. P. (1980). Morphological and biochemical changes in the cerebellum induced kainic acid in vivo. J. Neurochem. 34:1191-1200.

    Google Scholar 

  11. Grant, P., Sharma, P., and Pant, H. C. (2001). Cyclin-dependent protein kinase 5 (Cdk5) and the regulation of neurofilament metabolism. Eur. J. Biochem. 268:1534-1546.

    Google Scholar 

  12. Hayes, R. L., Yang, K., Whitson, J. S., and Postmantur, R. (1995). Cytoskeletal derangements following central nervous system injury: Modulation by neurotrophic gene transfection. J. Neurotrauma 12:933-941.

    Google Scholar 

  13. Hicks, R. R., Smith, H. D., and McIntosh, K. T. (1995). Temporal response and effects of excitatory amino acid antagonism on MAP2 protein immunoreactivity following experimental brain injury in rats. Brain Res. 678:151-160.

    Google Scholar 

  14. Hugon, J., and Vallat, J. M. (1990). Abnormal distribution of phosphorylated neurofilaments in neuronal degeneration induced by kainic acid. Neurosci. Lett. 119:45-48.

    Google Scholar 

  15. Krapfenbauer, K., Berger, M., Lubec, G., and Fountoulakis, M. (2001). Changes in the brain protein levels following administration of kainic acid. Electrophoresis 22:2086-2091.

    Google Scholar 

  16. Laskawi, R., and Wolff, J. R. (1996). Changes in the phosphorylation of neurofilament proteins in facial motoneurons following various types of nerve lesion. ORL J. Otorhinolaryngol. Relat. Spec. 58:13-22.

    Google Scholar 

  17. Lees, J. F., Shneidman, P. S., Skuntz, S. F., Carden, M. J., and Lazzarini, R. A. (1988). The gene structure of the largest human neurofilament subunit (NF-H) predicts a protein with multiple repeated sites for in vivo phosphorylation. EMBO J. 7:1947-1955.

    Google Scholar 

  18. Leterrier, J. F., Hartwig, J., Kas, J., Vegners, R., and Janmey, P. A. (1996). Mechanical effects of neurofilament cross bridges: Modulation by phosphorylation, lipids and interactions with F-actin. J. Biol. Chem. 271:15687-15694.

    Google Scholar 

  19. Li, B.-S., Daniels, M. P., and Pant, H. C. (2001). Integrins stimulate phosphorylation of neurofilament NF-M subunit KSP repeats through activation of extracellular regulated-kinases (Erk1/Erk2) in cultured motoneurons and transfected NIH 3T3 cells. J. Neurochem. 76:703-710.

    Google Scholar 

  20. Li, B.-S., Veeranna, G. J., Grant, P., and Pant, H. C. (1999). Activation of mitogen-activated protein kinases (Erk1 and erk2) cascade results in phosphorylation of NF-M tail domains in transfected NIH 3T3 cells. Eur. J. Biochem. 262:211-217.

    Google Scholar 

  21. Liu, M. H., Lei, D. L., and Yang, L. D. (1996). Kainate-induced brain lesion: Similar local and remote histopathologic and molecular changes as in ischemic brain infarct. J. Neuropathol. Exp. Neurol. 55:787-797.

    Google Scholar 

  22. Nakagawa, T., Chen, J., Zhang, Z., Kanai, Y., and Hirokawa, N. (1995). Two distinct functions of the carboxyl-terminal tail domain of NF-M upon neurofilament assembly: Cross-bridge formation and longitudinal elongation of filaments. J. Cell Biol. 129:387-395.

    Google Scholar 

  23. O'Ferrall, E. K., Robertson, J., and Mushynski, W. E. (2000). Inhibition of aberrant and constitutive phosphorylation of the high-molecular-mass neurofilament subunit by CEP-1347 (KT7515), an inhibitor of the stress-activated protein kinase signaling pathway. J. Neurochem. 75:2358-2367.

    Google Scholar 

  24. Paxinos, G., and Watson, C. (1983). The Rat Brain in Stereotaxic Coordinates, Academic Press, New York.

    Google Scholar 

  25. Petzold, A., Baker, D., Pryce, G., Kier, G., Thompson, E. J., and Giovannoni, G. (2003). Quantification of neurodegeneration by measurement of brain-specific proteins. J. Neuroimmun. 138:45-48.

    Google Scholar 

  26. Pollanen, M. S., Bergeron, C., and Weyer, L. (1994). Characterisation of a shared epitope in cortical Lewy body fibrils and Alzheimer paired helical filaments. Acta Neuropathol. 88:1-6.

    Google Scholar 

  27. Popa-Wagner, A., Schroeder, E., Schmoll, H., Walker, L. C., and Kessler, C. (1999). Upregulation of MAP1B and MAP2 in the rat brain after middle cerebral artery occlusion: Effect of age. J. Cereb. Blood Flow Metab. 19:425-434.

    Google Scholar 

  28. Roberts-Lewis, J. M., Savage, M. J., Marcy, V. R., Pinsker, L. R., and Siman, R. (1994). Immunolocalisation of calpain I-mediated spectrin degradation to vulnerable neurons in the ischemic gerbil brain. J. Neurosci. 14:3934-3944.

    Google Scholar 

  29. Rossi, F., Borsello, T., and Strata, P. (1993b). Exposure to kainic acid mimics the effects of axotomy in cerebellar Purkinje cells of the adult rat. Eur. J. Neurosci. 6:392-402.

    Google Scholar 

  30. Rossi, F., Borsello, T., Vaudano, E., and Strata, P. (1993a). Regressive modifications of climbing fibers following Purkinje cell degeneration in the cerebellar cortex of the adult rat. Neuroscience 53:759-778.

    Google Scholar 

  31. Rossi, F., JankovskI, A., and Sotelo, C. (1995). Differential regenerative capacity in two systems of injured adult cerebellar axons confronted to embryonic grafts. J. Comp. Neurol. 359:663-677.

    Google Scholar 

  32. Rossi, F., and Strata, P. (1995). Reciprocal trophic interactions in the adult climbing fiber-Purkinje cell system. Prog. Neurobiol. 47:341-369.

    Google Scholar 

  33. Schgroeder, E., Vogelgesang, S., Popa-Wagner, A., and Kessler, C. (2003). Neurofilament expression in the rat brain after cerebral infarctation: Effect of age. Neurobiol. Aging 24:135-145.

    Google Scholar 

  34. Shetty, K. A., and Turner, A. D. (1995). Intracerebroventricular kainic acid administration in adult rat alters hippocampal calbindin and non-phosphorylated neurofilament expression. J. Comp. Neurol. 363:581-599.

    Google Scholar 

  35. Shinozaki, H. (1994). Neuron damage induced by some potent kainoids and neuroprotective action of new agonist for metabotropic glutamate receptors. Eur. Neurol. 34:2-9.

    Google Scholar 

  36. Sternberger, L. A., and Sterneberger, N. H. (1983). Monoclonal antibodies distinguish phosphorylated and non-phosphorylated forms of neurofilaments in situ. Proc. Natl. Acad. Sci. U.S.A. 80:6126-6130.

    Google Scholar 

  37. Stone, J. D., Peterson, A. P., Eyer, J., Oblak, G., and Sickles, D. W. (2001). Neurofilaments are nonessential to the pathogenesis of toxicant-induced axonal degeneration. J. Neurosci. 21:2278-2287.

    Google Scholar 

  38. Terro, F., Lesort, M., Dussartre, C., Barthe, D., and Hugon, J. (1996). Phosphorylated neurofilament expression and resistance to kainate toxicity. Brain Res. Bull. 41:231-235.

    Google Scholar 

  39. Trojanowski, J. Q., Obrocka, M. A., and Lee, V. M.-Y. (1985). Distribution of neurofilament subunits in neurons and neuronal processes. Immunocytochemical studies of bovine cerebellum with subunit specific monoclonal antibodies. J. Histochem. Cytochem. 33:557-563.

    Google Scholar 

  40. Vartiainen, N., Tikka, T., Keinanen, R., Chan, P. H., and Koistinaho, J. (1999). Glutamatergic receptors regulate expression, phosphorylation and accumulation of neurofilaments in spinal cord neurons. Neuroscience 93:1123-1133.

    Google Scholar 

  41. Vickers, J. C., and Costa, M. (1992). The neurofilament triplets are present in distinct subpopulations of neurons in the central nervous system of the guinea-pig. Neuroscience 49:73-100.

    Google Scholar 

  42. Wang, S., Hamberger, A., Ding, M., and Haglid, K. G. (1992). In vivo activation of kainate receptors induces dephosphorylation of the heavy neurofilament subunit. J. Neurochem. 59:1975-1978.

    Google Scholar 

  43. Wang, S., Hamberger, A., Yang, Q., and Haglid, K. G. (1994). Changes in neurofilament protein NF-L and NF-H immunoreactivity following kainic acid-induced seizures. J. Neurochem. 62:739-748.

    Google Scholar 

  44. Xu, S., Cork, L. C., Griffin, J. W., and Cleveland, D. W. (1993). Increased expression of neurofilaments subunit NF-L produces morphological alterations that resembles the pathology of human motor neuron disease. Cell 73:23-33.

    Google Scholar 

  45. Yang, Q., Wang, S., Hamberger, A., and Haglid, K. S. (1996). Plasticity of granule cell-mossy fiber system following kainic acid induced seizures: An immunocytochemical study on neurofilament proteins. Neurosci. Res. 26:57-64.

    Google Scholar 

  46. Zhang, H., Sternberger, N. H., Rabinstein, L. J., Herman, M. H., Binder, L. I., and Sternberger, L. A. (1989). Abnormal processing of multiple proteins in Alzheimer disease. Proc. Natl. Acad. Sci. U.S.A. 86:8045-8049.

    Google Scholar 

Download references

Author information

Affiliations

Authors

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Milenkovic, I., Filipovic, R., Nedeljkovic, N. et al. Spatio-Temporal Changes in Neurofilament Proteins Immunoreactivity Following Kainate-Induced Cerebellar Lesion in Rats. Cell Mol Neurobiol 24, 367–378 (2004). https://doi.org/10.1023/B:CEMN.0000022769.44211.2b

Download citation

  • cerebellum
  • immunocytochemistry
  • kainic acid
  • neurofilament proteins
  • plasticity
  • synaptophysin