Skip to main content
Log in

Electrical Stimulation Accelerates and Enhances Expression of Regeneration-Associated Genes in Regenerating Rat Femoral Motoneurons

  • Published:
Cellular and Molecular Neurobiology Aims and scope Submit manuscript

Abstract

1. In this study we investigated whether electrical stimulation accelerates the upregulation of Tα1-tubulin and GAP-43 (regeneration-associated genes; RAGs) and the downregulation of the medium-molecular-weight neurofilament (NFM), in concert with stimulation-induced acceleration of BDNF and trkB gene expression and axonal regener- ation.

2. Two weeks prior to unilateral femoral nerve transection and suture, fluorogold (Fluorochrome Inc., Denver) or fluororuby (Dextran tetramethylrhodamine, Mol. Probes, D-1817, Eugene, OR) was injected into quadriceps muscles of the left and right hindlimbs to label the femoral motoneuron pools as previously described. Over a period of 7 days, fresh spinal cords were processed for semiquantitation of mRNA by using in situ hybridi- zation.

3. There was an increase in Tα1-tubulin and GAP-43 mRNA and a decline in the NFM mRNA at 7 days after nerve suture and sham stimulation but not in intact nerves. In contrast, 1-h stimulation of sutured but not intact nerves dramatically accelerated the changes in gene expression: mRNA levels of Tα1-tubulin and GAP-43 were significantly elevated above control levels by 2 days while NFM mRNA was significantly reduced by 2 days in the sutured nerves. Thereby, the neurofilament/tubulin expression ratio was reduced at 2 days after suture and stimulation, possibly allowing more tubulin to be transported faster into the growing axons to accelerate the elongation rate following stimulation. Importantly, the changes in RAGs and NFM gene expression were delayed relative to the accelerated upregulation of BDNF and trkB mRNA by electrical stimulation.

4. The temporal sequence of upregulation of BDNF and trkB, altered gene expression of RAGs and NFM, and accelerated axonal outgrowth from the proximal nerve stump are consistent with a key role of BDNF and trkB in mediating the altered expression of RAGs and, in turn, the promotion of axonal outgrowth after electrical stimulation.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

REFERENCES

  • Aigner, L., Arber, S., Kapfhammer, J. P., Laux, T., Schneider, C., Botteri, F., Brenner, H. R., and Catoni, P. (1995). Overexpression of the neural growth-associated protein GAP-43 induces nerve sprouting in the adult nervous system of transgenic mice. Cell 83:269-278.

    Google Scholar 

  • Aigner, L., and Caroni, P. (1993). Depletion of 43-kD growth-associated protein in primary sensory neurons leads to diminished formation and spreading of growth cones. J. Cell Biol. 123:417-429.

    Google Scholar 

  • Aigner, L., and Caroni, P. (1995). Absence of persistent spreading, branching, and adhesion in GAP-43-depleted growth cones. J. Cell Biol. 128:647-660.

    Google Scholar 

  • Al-Majed, A. A., Brushart, T. M., and Gordon, T. (2000a). Electrical stimulation accelerates and increases expression of BDNF and trkB mRNA in regenerating rat femoral motoneurons. Eur. J. Neurosci. 12:1-11.

    Google Scholar 

  • Al-Majed, A. A., Neumann, C. M., Brushart, T. M., and Gordon, T. (2000b). Brief electrical stimulation promotes the speed and accuracy of motor axonal regeneration. J. Neurosci. 20:2602-2608.

    Google Scholar 

  • Ausubel, F. M., Brent, R., Kingston, R. E., Moore, D. D., Seidman, J. G., Smith, K. A., and Struhl, K. (1987). Current Protocols in Molecular Biology, Wiley-Interscience, New York.

    Google Scholar 

  • Basi, G. S., Jacobson, R. D., Virag, I., Schilling, J., and Skene, J. H. P. (1987). Primary structure and transcriptional regulation of GAP-43. A protein associated with nerve growth. Cell 236:597-600.

    Google Scholar 

  • Bisby, M. A., and Tetzlaff, W. (1992). Changes in cytoskeletal protein synthesis following axon injury and during axon regeneration. Mol. Neurobiol. 6:107-123.

    Google Scholar 

  • Boyd, J. G., and Gordon, T. (2004). Signaling and function of neurotrophic factors in the normal and injured peripheral nervous system. Invited Rev.: Mol. Neurobiol. (in press).

  • Brushart, T. M., Hoffman, P. M., Royall, R. M., Murinson, B. B., Witzel, C., and Gordon, T. (2002). Electrical stimulation promotes motoneuron regeneration without increasing its speed or conditioning the neuron. J. Neurosci. 22:6631-6638.

    Google Scholar 

  • Brushart, T. M., and Seiler, W. A. (1987). Selective reinnervation of distal motor stumps by peripheral motor axons. Exp. Neurol. 97:290-300.

    Google Scholar 

  • Caroni, P. (1997). Intrinsic neuronal determinants that promote axonal sprouting and elongation. Bioessays 19:767-775.

    Google Scholar 

  • Caroni, P., Aigner, L., Arber, S., Botteri, F., Kapfhammer, J., and Brenner, H. R. (1995). GAP-43 and CAP-32 induce nerve sprouting in the adult nervous system of transgenic mice. Soc. Neurosci. Abstr. 21:14.

    Google Scholar 

  • Caroni, P., and Becker, M. (1992). The downregulation of growth-associated proteins in motoneurons at the onset of synapse elimination is controlled by muscle activity and IGF1. J. Neurosci. 12:3849-3861.

    Google Scholar 

  • Fawcett, J. W., Mathews, G., Housden, E., Goedert, M., and Matus, A. (1994). Regenerating sciatic nerve axons contain the adult rather than embryonic pattern of microtubule associated protein. Neurosci. 61:789-804.

    Google Scholar 

  • Fernandes, K. J. L., Fan, D. P., Tsui, B. J., Cassar, S. L., and Tetzlaff, W. (1999). Influence of the axotomy to cell body distance in rat rubrospinal and spinal motoneurons: Differential regulation of GAP-43, tubulin, and neurofilament-M. J. Comp. Neurol. 414:495-510.

    Google Scholar 

  • Fernandes, K. J. L., Jasmin, B. J., and Tetzlaff, W. (1995). Effect of neurotrophins on mRNA levels in axotomized adult facial motoneurons. Soc. Neurosci. Abstr. 21:1534.

    Google Scholar 

  • Fournier, A. E., Beer, J., Arregui, C. O., Essagian, C., Aguayo, A. J., and McKerracher, L. (1997). Brain-derived neurotrophic factor modulates GAP-43 but not Tα1-tubulin expression in injured retinal ganglion cells of adult rats. J. Neurosci. Res. 47:561-572.

    Google Scholar 

  • Fu, S. Y., and Gordon, T. (1997). The cellular and molecular basis of peripheral nerve regeneration. Mol. Neurobiol. 14:67-116.

    Google Scholar 

  • Herdegen, T., Skene, P., and Bahr, M. (1997). The c-jun transcription factor-biopotential mediator of neuronal death, survival and regeneration. Trends Neurosci. 20:227-231.

    Google Scholar 

  • Hoffman, P. N., Cleveland, D. W., Griffin, J. W., Landes, N. J., and Price, D. L. (1987). Neurofilament gene expression: A major determinant of axonal caliber. Proc. Natl. Acad. U.S.A. 84:3472-3476.

    Google Scholar 

  • Hoffman, P. N., and Lasek, R. J. (1975). The slow component of axonal transport: Identification of major structural polypeptide of the axon and their generality among mammalian neurons. J. Cell Biol. 66:351-366.

    Google Scholar 

  • Hoffman, P. N., and Lasek, R. J. (1980). Axonal transport of the cytoskeleton in regenerating motor neurons: Constancy and change. Brain Res. 202:317-333.

    Google Scholar 

  • Hoffman, P. N., Thompson, G. W., Griffin, J. W., and Price, D. L. (1985). Changes in neurofilament transport coincide temporally with alterations in the caliber of axons in regenerating motor fibers. J. Cell Biol. 101:1332-1340.

    Google Scholar 

  • Jacob, J. M., and McQuarrie, I. G. (1993). Acceleration of axonal outgrowth in rat sciatic nerve at one week after axotomy. J. Neurobiol. 24:356-367

    Google Scholar 

  • Julien, J. P., Meyer, D., Flavell, D., Hurst, J., and Grosveld, F. (1986). Cloning and developmental expression of the murine neurofilament gene family. Brain Res. 387:243-250.

    Google Scholar 

  • Kobayashi, N. R., Fan, D. P., Giehl, K. M., Bedard, A. M., Wiegand, S. J., and Tetzlaff, W. (1997). BDNF and NT-4/5 prevent atrophy of rat rubrospinal neurons after cervical axotomy, stimulate GAP-43 and Tα1-Tubulin mRNA expression, and promote axonal regeneration. J. Neurosci. 17:9583-9595.

    Google Scholar 

  • Lund, L. M., Machado, V. M., and McQuarrie, I. G. (2002). Increased Actin and tubulin polymerization in regrowing axons: Relationship to the conditioning lesion effect. Exp. Neurol. 178:306-312.

    Google Scholar 

  • McQuarrie, I. G. (1983). Role of the cytoskeleton in the regenerating nervous system. In Seil, F. J. (ed.), Nerve, Organ and Tissue Regeneration, Academic Press, New York and London, pp. 51-88.

    Google Scholar 

  • McQuarrie, I. G., and Grafstein, B. (1982). Protein synthesis and axonal transport in goldfish retinal ganglion cells during regeneration accelerated by a conditioning lesion. Brain Res. 251:25-37.

    Google Scholar 

  • McQuarrie, I. G., and Jacob, J. M. (1991). Conditioning nerve crush accelerates cytoskeletal protein transport in sprouts that form after a subsequent crush. J. Comp. Neurol. 305:139-147.

    Google Scholar 

  • Nothias, F., Boyne, L., Murrary, M., Tessler, A., and Fischer, I. (1995). The expression and distribution of tau proteins and messenger RNA in rat dorsal root ganglion neurons during development and regeneration. Neuroscience 66:707-719.

    Google Scholar 

  • Oblinger, M. M., Brady, S. T., McQuarrie, I. G., and Lasek, R. J. (1987). Cytotypic differences in the protein composition of the axonally transported cytoskeleton in mammalian neurons. J. Neurosci. 7:453-462.

    Google Scholar 

  • Petrov, T., You, S., Cassar, S. L., Tetzlaff, W., and Gordon, T. (1996). Cytoskeletal protein expression in long-term axotomized facial and sciatic. Soc. Neurosci. Abstr. 22:674.

    Google Scholar 

  • Richmond, F. J. R., Gladdy, R., Creasy, J. L., Kitamura, S., Smits, E., and Thomas, D. B. (1994). Efficacy of seven retrograde tracers, compared in multiple-labelling studies of feline motoneurons. J. Neurosci. Methods 53:35-46.

    Google Scholar 

  • Tetzlaff, W., Alxander, S. W., Miller, F. D., and Bisby, M. A. (1991). Response of facial and rubrospinal neurons to axotomy: Changes in mRNA expression for cytoskeletal proteins and GAP-43. J. Neurosci. 11:2528-2544.

    Google Scholar 

  • Tetzlaff, W., Bisby, M. A., and Kreutzberg, G. M. (1988). Changes in cytoskeletal proteins in the rat facial nucleus following axotomy. J. Neurosci. 8:3181-3189.

    Google Scholar 

  • Tetzlaff, W., Leonard, C., Krekoski, C. A., Parhad, I. M., and Bisby, M. (1996). Reduction in otoneuronal neurofilament synthesis by successive axotomies: A possible explanation for the conditioning lesion effect on axon regeneration. Exp. Neurol. 139:95-106.

    Google Scholar 

  • Tetzlaff, W., Zwiers, H., Lederis, K., Cassar, L., and Bisby, M. A. (1989). Axonal transport and localization of B50/GAP-43-like immunoreactivity in regenerating sciatic and facial nerves of the rat. J. Neurosci. 9:1303-1313.

    Google Scholar 

  • Verge, V. M., Merlio, J. P., Grondin, J., Ernfors, P., Persson, H., Riopelle, R. J., Hokfelt, T., and Richardson, P. M. (1992). Colocalization of NGF receptor mRNA in primary sensory neurons: Responses to injury and infusion of NGF. J. Neurosci. 12:4011-4022.

    Google Scholar 

  • Widmer, F., and Caroni, P. (1990). Identification, localization and primary structure of CAP-23, a particle-bound cytosolic protein of early development. J. Cell Biol. 111:3035-3047.

    Google Scholar 

  • Wujek, J. R., and Lasek, R. J. (1983). Correlation of axonal regeneration and slow component B in two branches of a single axon. J. Neurosci. 3:243-251.

    Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Al-Majed, A.A., Tam, S.L. & Gordon, T. Electrical Stimulation Accelerates and Enhances Expression of Regeneration-Associated Genes in Regenerating Rat Femoral Motoneurons. Cell Mol Neurobiol 24, 379–402 (2004). https://doi.org/10.1023/B:CEMN.0000022770.66463.f7

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1023/B:CEMN.0000022770.66463.f7

Navigation