Elsevier

Experimental Neurology

Volume 195, Issue 2, October 2005, Pages 524-539
Experimental Neurology

Regular Article
Assessing behavioural function following a pyramidotomy lesion of the corticospinal tract in adult mice

https://doi.org/10.1016/j.expneurol.2005.06.017Get rights and content

Abstract

We have developed a pyramidotomy model in mice to lesion the corticospinal tract at the level of the brainstem pyramidal tract, and evaluated the resultant impairments in motor function in a series of behavioural tests. Adult C57BL/6 mice received a unilateral pyramidotomy and a control group of mice underwent sham surgery. We studied the effects of this lesion on forepaw function using five behavioural paradigms, some of which have been widely used in rat studies but have not been fully explored in mice. The tests used were: a rearing test, which assesses forepaw use for weight support during spontaneous vertical exploration of a cylinder; a grid walking test, which assesses the ability to accurately place the forepaws during exploration of an elevated grid; a tape-removal test, which measures both sensory and motor function of the forepaw; a CatWalk automated gait analysis, which provides a number of quantitative measures including stride length and stride width during locomotion; and a staircase reaching task, which assesses skilled independent forepaw use. All tests revealed lesion effects on forepaw function with the tape removal, grid walking, rearing and CatWalk tests demonstrating robust effects throughout the testing period. The development of a pyramidotomy lesion model in mice, together with behavioural tests which can reliably measure functional impairments, will provide a valuable tool for assessing therapeutic strategies to promote regeneration and plasticity.

Introduction

Following injury to the adult mammalian central nervous system (CNS), axonal regeneration and plasticity is extremely limited. Most of the functional deficits resulting from injury to the spinal cord are caused by the interruption of descending and ascending axonal tracts, and the persistence of these deficits is due to their failure to regenerate (Schwab and Bartholdi, 1996, Houle and Tessler, 2003, Selzer, 2003). This failure is thought to be due to the combination of a limited inherent regenerative capacity of the axons (Schwab and Bartholdi, 1996, Neumann and Woolf, 1999); an insufficiency of trophic factors (Widenfalk et al., 2001, Jones et al., 2001) and the presence of growth inhibitory molecules, such as chondroitin sulphate proteoglycans (Fawcett and Asher, 1999, Silver and Miller, 2004) and myelin-associated neurite growth inhibitors (Schwab, 2002, McGee and Strittmatter, 2003, Filbin, 2003). Although a number of recent studies have demonstrated a capacity for some spinal systems to spontaneously sprout in the adult (Raineteau and Schwab, 2001, Weidner et al., 2001, Bareyre et al., 2004), the fact is that there is limited functional recovery following CNS injury and there is a great potential for enhancing CNS plasticity with the aim of promoting functional improvements.

One recent focus has been to study intact systems following CNS injury, with the aim of enhancing plasticity of spared fibres. A number of studies have investigated whether intact corticospinal tract (CST) projections can be induced to sprout into the denervated side of the spinal cord following a unilateral pyramidotomy lesion. This lesion transects one half of the CST at the level of the pyramids in the brainstem, rostral to the spinal cord. The reactions of the uninjured tract projecting in the spinal cord can then be studied following various intervention strategies aimed at promoting plasticity. Sprouting of the intact CST has been observed in this lesion model following treatments which neutralise inhibitory factors associated with CNS myelin (Vanek et al., 1998, Thallmair et al., 1998, Z'Graggen et al., 1998, Raineteau et al., 1999, Raineteau et al., 2002, Blochlinger et al., 2001, Bareyre et al., 2002) and treatments which promote growth, such as delivery of neurotrophic factors (Zhou and Shine, 2003, Zhou et al., 2003). Pyramidotomy results in deficits in voluntary motor function of the forepaws (Steward et al., 2004, Lacroix et al., 2004) and impairments in forelimb function such as limb use asymmetry (Thallmair et al., 1998, Z'Graggen et al., 1998), precise stepping (Z'Graggen et al., 1998, Metz and Whishaw, 2002), somatosensory function (Thallmair et al., 1998), locomotion and gait (Metz et al., 1998, Fanardjian et al., 2001) and skilled paw reaching (Whishaw et al., 1993, Thallmair et al., 1998, Z'Graggen et al., 1998, Weidner et al., 2001). However, all these studies have used a rat model and there has been a distinct lack of studies describing this lesion in adult mice, other than to look at the glial reaction caused by this injury (Leong et al., 1995) and no one, to our knowledge, has investigated behavioural deficits after pyramidotomy in mice. Given the increasing use of genetically modified mice, it would be beneficial to develop a murine pyramidotomy model and to characterise a number of reliable behavioural outcome measures for assessing functional deficits. This model would be valuable for future studies examining the effects of gene deletion or other strategies aimed at promoting plasticity following CNS injury.

Here, we develop a mouse pyramidotomy lesion model and assess functional impairments in the following behavioural paradigms: rearing, grid walking, tape removal, CatWalk gait analysis and staircase pellet reaching. The rearing test is specifically designed to encourage use of the forelimbs for vertical exploration of a cylinder in rats (Napieralski et al., 1998, Liu et al., 1999, Schallert et al., 2000, Soblosky et al., 2001, MacLellan et al., 2002, Woodlee et al., 2005), and mice (Baskin et al., 2003, Wells et al., 2005). This test assesses the animals' voluntary use of the forelimbs for upright postural weight bearing support (Baskin et al., 2003) and thus highlights limb use asymmetries. The grid walking test assesses deficits in voluntary motor control and limb movements involved in precise stepping, co-ordination and accurate paw placement (Z'Graggen et al., 1998, Napieralski et al., 1998, Metz et al., 2000, Soblosky et al., 2001, Ma et al., 2001, Schucht et al., 2002, MacLellan et al., 2002, Selak and Fritzler, 2004). Grid walking also requires sensory feedback for limb placement (Baskin et al., 2003) and thus assesses deficits in sensorimotor function (Z'Graggen et al., 1998, Merkler et al., 2001, Zhang et al., 2002, Menet et al., 2003). The tape removal test is particularly sensitive to somatosensory deficits (Thallmair et al., 1998) and has been used in both rats (Thallmair et al., 1998, Schallert et al., 2000, Bradbury et al., 2002, Onifer et al., 2005) and mice (Wells et al., 2005). The CatWalk is a novel test of gait which has the benefit of measuring a number of locomotor-related assessments simultaneously (Hamers et al., 2001) and has been used in various experimental paradigms with rats (Hamers et al., 2001, Lankhorst et al., 2001, van Meeteren et al., 2003, Vrinten and Hamers, 2003, Vogelaar et al., 2004) and mice (Vogelaar et al., 2004). The staircase pellet reaching test assesses accurate forepaw reaching and forelimb motor control and has a history of use in rats (Montoya et al., 1991, Whishaw et al., 1997, Nikkhah et al., 1998, Colbourne et al., 2000, MacLellan et al., 2002, Samsam et al., 2004) but less in mice (Baird et al., 2001). This task was chosen because unilateral lesions of the CST are known to abolish directed forepaw retrieval by the ipsilateral paw (Keyvan-Fouladi et al., 2003).

Here, we report the development of a unilateral pyramidotomy model in mice and the methodology for a number of reliable behavioural tests to assess functional deficits following this lesion.

Section snippets

Animals and surgery

Adult male C57BL/6 mice (Harlan UK Ltd., 20–25 g, 6–8 weeks old) were used in these studies and all surgical procedures were performed in accordance with U.K. Home Office regulations (Animals Scientific Procedures Act, 1986). Mice were anaesthetised with a mixture of medetomidine (0.5 mg/kg) and ketamine (75 mg/kg) and sterile precautions were used throughout. Surgical procedures were adapted from previous studies using rats to perform a pyramidotomy lesion in mice, whereby a unilateral lesion

Histological assessment of lesion

Gross anatomy revealed a transection in the right pyramidal tract of the brainstem in lesioned mice (Fig. 1A). Apart from a small ventral uncrossed component, the CST decussates below the level of the lesion; therefore, a lesion of the right pyramidal tract will denervate the CST on the left side of the spinal cord (illustrated in schematic, Fig. 1B). The extent of the pyramidotomy lesion was determined using immunostaining for protein kinase Cγ (PKCγ) in the brainstem and spinal cord (Fig. 1).

Discussion

In the present study, we have established a consistent and reliable pyramidotomy lesion model in adult mice. We selected a number of behavioural tests to evaluate effects on both motor and sensory behaviours and assessed them on their ability to consistently measure deficits in forelimb behaviours following this lesion. We conclude that the rearing, tape removal, grid walking and CatWalk tests translate well to mice.

Conclusion

We have developed a robust, reproducible and consistent model for assessing a unilateral pyramidotomy lesion of the CST in mice. We have also developed sensitive and reliable behavioural outcome measures for assessing deficits following this lesion. The majority of tests revealed lesion effects on forepaw function, to varying degrees, with the rearing, grid walking, tape removal and CatWalk tests in particular revealing these effects throughout the testing period. The development of a

Acknowledgments

This work was supported by the Medical Research Council, The International Spinal Research Trust, and the Wellcome Trust. The authors would like to thank Prof. Patrick Doherty for invaluable input and John Grist and Jonathan Ramsey for technical support.

References (70)

  • A.W. McGee et al.

    The Nogo-66 receptor: focusing myelin inhibition of axon regeneration

    Trends Neurosci.

    (2003)
  • G.A. Metz et al.

    Cortical and subcortical lesions impair skilled walking in the ladder rung walking test: a new task to evaluate fore- and hindlimb stepping, placing, and co-ordination

    J. Neurosci. Methods

    (2002)
  • G.A. Metz et al.

    The effects of unilateral pyramidal tract section on hindlimb motor performance in the rat

    Behav. Brain Res.

    (1998)
  • G.A. Metz et al.

    Efficient testing of motor function in spinal cord injured rats

    Brain Res.

    (2000)
  • C.P. Montoya et al.

    The “staircase test”: a measure of independent forelimb reaching and grasping abilities in rats

    J. Neurosci. Methods

    (1991)
  • G.D. Muir et al.

    Complete locomotor recovery following corticospinal tract lesions: measurement of ground reaction forces during overground locomotion in rats

    Behav. Brain Res.

    (1999)
  • J.A. Napieralski et al.

    Motor and somatosensory deficits following uni- and bilateral lesions of the cortex induced by aspiration or thermocoagulation in the adult rat

    Exp. Neurol.

    (1998)
  • S. Neumann et al.

    Regeneration of dorsal column fibers into and beyond the lesion site following adult spinal cord injury

    Neuron

    (1999)
  • G. Nikkhah et al.

    Differences in acquisition and full performance in skilled forelimb use as measured by the ‘staircase test’ in five rat strains

    Behav. Brain Res.

    (1998)
  • S.M. Onifer et al.

    Adult rat forelimb dysfunction after dorsal cervical spinal cord injury

    Exp. Neurol.

    (2005)
  • T.E. Samsam et al.

    Toxicological evaluation of the staircase test for assessing fine motor movements

    Neurotoxicol. Teratol.

    (2004)
  • T. Schallert et al.

    CNS plasticity and assessment of forelimb sensorimotor outcome in unilateral rat models of stroke, cortical ablation, parkinsonism and spinal cord injury

    Neuropharmacology

    (2000)
  • P. Schucht et al.

    Anatomical correlates of locomotor recovery following dorsal and ventral lesions of the rat spinal cord

    Exp. Neurol.

    (2002)
  • M.E. Schwab

    Increasing plasticity and functional recovery of the lesioned spinal cord

    Prog. Brain Res.

    (2002)
  • M.E. Selzer

    Promotion of axonal regeneration in the injured CNS

    Lancet Neurol.

    (2003)
  • J.S. Soblosky et al.

    Graded unilateral cervical spinal cord injury in the rat: evaluation of forelimb recovery and histological effects

    Behav. Brain Res.

    (2001)
  • C.F. Vogelaar et al.

    Sciatic nerve regeneration in mice and rats: recovery of sensory innervation is followed by a slowly retreating neuropathic pain-like syndrome

    Brain Res.

    (2004)
  • D.H. Vrinten et al.

    ‘CatWalk’ automated quantitative gait analysis as a novel method to assess mechanical allodynia in the rat; a comparison with von Frey testing

    Pain

    (2003)
  • I.Q. Whishaw

    An endpoint, descriptive, and kinematic comparison of skilled reaching in mice (Mus musculus) with rats (Rattus norvegicus)

    Behav. Brain Res.

    (1996)
  • I.Q. Whishaw

    Loss of the innate cortical engram for action patterns used in skilled reaching and the development of behavioral compensation following motor cortex lesions in the rat

    Neuropharmacology

    (2000)
  • I.Q. Whishaw et al.

    Proximal and distal impairments in rat forelimb use in reaching follow unilateral pyramidal tract lesions

    Behav. Brain Res.

    (1993)
  • I.Q. Whishaw et al.

    Analysis of limb use by control rats and unilateral DA-depleted rats in the Montoya staircase test: movements, impairments and compensatory strategies

    Behav. Brain Res.

    (1997)
  • M.T. Woodlee et al.

    Testing forelimb placing “across the midline” reveals distinct, lesion-dependent patterns of recovery in rats

    Exp. Neurol.

    (2005)
  • L. Zhang et al.

    A test for detecting long-term sensorimotor dysfunction in the mouse after focal cerebral ischemia

    J. Neurosci. Methods

    (2002)
  • F.M. Bareyre et al.

    Long-lasting sprouting and gene expression changes induced by the monoclonal antibody IN-1 in the adult spinal cord

    J. Neurosci.

    (2002)
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