Elsevier

Neuroscience

Volume 177, 17 March 2011, Pages 114-126
Neuroscience

Cognitive, Behavioral, and Systems Neuroscience
Research Paper
The time course of serotonin 2A receptor expression after spinal transection of rats: an immunohistochemical study

https://doi.org/10.1016/j.neuroscience.2010.12.062Get rights and content

Abstract

Hyperexcitability of motoneurons is one of the key mechanism that has been demonstrated to underlie the pathogenesis of spasticity after spinal injury. Serotonin (5-HT) denervation supersensitivity is one of the mechanisms underlying this increased motoneuron excitability. In this study, to examine whether the supersensitivity is caused by 5-HT receptor upregulation we investigated changes in levels of 5-HT2A receptor immunoreactivity (5-HT2AR-IR) following a spinal transection in the sacral spinal cord of rats at seven different time points post injury: 2, 8, 16 h, and 1, 2, 7 and 28 days, respectively. 5-HT2AR-IR density was analyzed in motoneurons (regions containing their somata and dendrites) in the spinal segments below the lesion. The results showed no significant changes in 5-HT2AR-IR in the motoneurons up to 16 h following the transection. After 1-day, however the levels of 5-HT2AR-IR increased in the motoneurons and their dendrites, with the density level being 3.4-fold higher in spinalized rats than in sham-operated rats. The upregulation increased progressively until a maximal level was reached at 28 days post-injury. We also investigated 5-HT and 5-HT transporter expressions at five different post injury time points: 1, 2, 7, 21 and 60 days and they showed concurrent down-regulation changes after 2 days. These results suggest that the upregulation of 5-HT2ARs may at least partly underlie the development of 5-HT denervation supersensitivity in spinal motoneurons following spinal injury and thereby implicates their involvement in the pathogenesis of the subsequent development of spasticity.

Research highlights

▶5-HT2A receptor expression was investigated in spinalized rats at 7 time points. ▶5-HT2A receptor started to upregulate in spinal motoneurons 1 day after lesion. ▶5-HT2A receptor upregulation reached a plateau level at around 28 days. ▶5-HT2A receptor upregulation is implicated in 5-HT denervation supersensitivity.

Section snippets

Animal operation and tissue preparations

All experiments were conducted in accordance with the guidelines of the EU Directive 86/609/EEC and were approved by the Danish Animal Experiments Inspectorate. All efforts were made to minimize the number of animals used and their suffering. Initially a total of 132 adult male Wistar rats were used with a body weight of 200–250 g. Due to the loss of animals during the post-operative period and the exclusion of those animals with incorrect lesion sites 117 remaining rats were used in this study

Spinal lesion sites in the spinalized rats

In most cases, especially in those with a long post-operation survival time, it could be confirmed by surgical microscope that the spinal cord was completely transected at S2–S3 spinal segments, where a 2–3 mm space could be clearly observed between the rostral and the caudal stumps (Fig. 1A, B). In the cases whose spinal transection sites were processed with Fast Blue and Thionin the results showed that all of the spinal cords were completely transected with no neurons or nerve fibers passing

Discussion

Using immunohistochemistry our present study has revealed the time course of 5-HT2AR expression changes following a complete spinal transection. We have demonstrated that 5-HT2AR upregulation starts at a time point before 1 day after spinalization and reaches a plateau at approximately the fourth week. Our previous findings have demonstrated that this upregulation persists for at least 2 months—the longest time we have investigated so far (Kong et al., 2010). We also investigated the changes of

Conclusion

In this study we have shown that 1 day after spinalization the levels of 5-HT2AR-IR increased in the motoneuron somata and their dendrites and this increase persisted for an indefinite long period. Furthermore, the time course of 5-HT2AR upregulation was significantly correlated with the development of 5-HT denervation supersensitivity. These results indicate that the upregulation of 5-HT2ARs at least partly underlies the development of 5-HT denervation supersensitivity in spinal motoneurons

Acknowledgments

We are grateful to Lillian Grøndahl for her unfailing assistance in all phases of this investigation. We thank Dr. Claire Meehan for critical reading of the manuscript. This project was supported by the Lundbeck Foundation, the Danish Multiple Sclerosis Foundation, the Ludvig and Sara Elsass' Foundation, Sofus Friis Foundation and the Danish Medical Research Council.

References (53)

  • B.C. Hains et al.

    Changes in serotonin, serotonin transporter expression and serotonin denervation supersensitivity: involvement in chronic central pain after spinal hemisection in the rat

    Exp Neurol

    (2002)
  • Y. Hayashi et al.

    5-HT precursor loading, but not 5-HT receptor agonists, increases motor function after spinal cord contusion in adult rats

    Exp Neurol

    (2010)
  • X.Y. Kong et al.

    Robust upregulation of serotonin 2A receptors after chronic spinal transection of rats: an immunohistochemical study

    Brain Res

    (2010)
  • J.K. Lee et al.

    Up-regulation of 5-HT2 receptors is involved in the increased H-reflex amplitude after contusive spinal cord injury

    Exp Neurol

    (2007)
  • B.W. Newton et al.

    The morphology and distribution of rat serotoninergic intraspinal neurons: an immunohistochemical study

    Brain Res Bull

    (1988)
  • B.J. Schmidt et al.

    The role of serotonin in reflex modulation and locomotor rhythm production in the mammalian spinal cord

    Brain Res Bull

    (2000)
  • H.W. Steinbusch

    Distribution of serotonin-immunoreactivity in the central nervous system of the rat-cell bodies and terminals

    Neuroscience

    (1981)
  • N.E. Anden et al.

    The time course of the disappearance of noradrenaline and 5-hydroxytryptamine in the spinal cord after transection

    Acta Physiol Scand

    (1964)
  • M. Antri et al.

    Locomotor recovery in the chronic spinal rat: effects of long-term treatment with a 5-HT2 agonist

    Eur J Neurosci

    (2002)
  • H. Barbeau et al.

    Action of cyproheptadine in spastic paraparetic patients

    J Neurol Neurosurg Psychiatry

    (1982)
  • D.J. Bennett et al.

    Spasticity in rats with sacral spinal cord injury

    J Neurotrauma

    (1999)
  • D.J. Bennett et al.

    Evidence for plateau potentials in tail motoneurons of awake chronic spinal rats with spasticity

    J Neurophysiol

    (2001)
  • D.J. Bennett et al.

    Plateau potentials in sacrocaudal motoneurons of chronic spinal rats, recorded in vitro

    J Neurophysiol

    (2001)
  • D.J. Bennett et al.

    Spastic long-lasting reflexes in the awake rat after sacral spinal cord injury

    J Neurophysiol

    (2004)
  • R.D. Blakely et al.

    Molecular physiology of norepinephrine and serotonin transporters

    J Exp Biol

    (1994)
  • P. Boulenguez et al.

    Down-regulation of the potassium-chloride cotransporter KCC2 contributes to spasticity after spinal cord injury

    Nat Med

    (2010)
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