Research reportIncreased gait variability in mice with small cerebellar cortex lesions and normal rotarod performance
Highlights
► Bilateral lesions of the cerebellar cortex were electrolytically produced in mice. ► Cerebellar lesioned mice showed no overt signs of ataxia or abnormal ambulation. ► Accelerating rotarod did not distinguish cerebellar lesioned mice from sham controls. ► Treadmill gait analysis distinguished cerebellar lesioned mice from sham controls. ► Cerebellar lesions evoked an increase of gait variability in mice.
Introduction
Different behavioural assays have been described to evaluate neuromotor performance in rodents, most of which involve the ability to balance on a rotating cylinder [1]. The rotarod test has been used for more than 50 years as an easy way to test effects of drugs, brain damage or disease states [2]. Most protocols not only assess motor coordination proper, but various other aspects of neuromotor functioning as well, such as motor learning manifested by improved performance over sequential trials (e.g., [3], [4], [5]). However, several authors have questioned the sensitivity of rotarod procedures to detect subtle alterations in neuromotor function, especially in the early stages of progressive disease (e.g., [6], [7]).
Footprint gait analysis procedures have been used to quantify motor function in rodents for many years as well. Initial manual methods to obtain footprint patterns included dipping or painting the animal's paws, and having them walk down a paper-covered aisle (for an extensive overview, see [8]). Originally, Rushton et al. [9] smeared the paws with vaseline, which was later replaced by ink ([10], [11]), a method still frequently used today (e.g. [12], [13], [14]). The need for automated and objective behavioural measurements [15] inspired the development of computerized footprint recording techniques during spontaneous [7], [16] or forced locomotion [17], [18]. Forced locomotion on a treadmill may mask deficits in movement initiation [19], but the possibility to manipulate ambulatory speed allows experimenters to measure the influence of velocity on gait parameters sensitively and precisely [20], [21]. Although still less thoroughly validated, gait analysis may therefore be the method of choice for detailed assessment of neuromotor performance [22]. Much more sensitive than any rotarod procedure, automated analysis of walking patterns has documented neuromotor impairment in rodent models of stroke, Huntington's disease, amyotrophic lateral sclerosis and metachromatic leukodystrophy [7], [23], [24].
Since rotarod procedures are still widely accepted to assess cerebellum-based performance [25], we presently compare treadmill walking and an accelerating rotarod task in their ability to detect consequences of cerebellar cortex lesions in mice. The electrolytic methods used here resulted in small lesions that did not induce overt signs of ataxia or abnormal ambulatory activity. The cerebellum processes ascending sensory information and descending motor impulses, and projects to structures involved in the execution of movement to control motor coordination and equilibrium [25]. Accordingly, impaired cerebellar function results in gait disturbances and lack of coordination of voluntary movements, and mice with cerebellar defects display alterations in footprint and rotarod measures [21], [26], [27], [28], [29], [30]. In view of the putative differences in sensitivity between these two methods, we presently hypothesize treadmill gait analysis is actually more likely to detect subtle motor alterations in cerebellar-lesioned mice than rotarod procedures. In addition, we define a subset of sensitive gait parameters that can be used to assess the effects of cerebellar defects in laboratory mice, and relate to clinical neurological signs in humans with cerebellar lesions.
Section snippets
Animals and surgical procedure
A total of 23 adult female C57Bl/6 mice, approximately 2 months of age, were purchased from Elevage Janvier (Le-Genest-Saint-Isle, France) and were group housed (4–7 mice per cage) at standard laboratory conditions (12 h light/dark cycle, constant room temperature and humidity). Behavioural testing took place during the light phase of the cycle. Food and water were available at libitum. All procedures were approved by the ethical research committee of the university in accordance with the
Location and size of lesion
Fig. 2 illustrates the size and location of the lesion in cerebellar cortex area viewed in a coronal section. Following histological analysis, the mean extent of the lesion along the axis of the brain was determined to be between 5.40 mm and 7.30 mm posterior to bregma for the largest lesion and between 5.50 mm and 6.30 mm posterior to bregma for the smallest lesion. Cerebellar lesions were located between 1.3 mm and 2.5 mm lateral to bregma and 1.5–3 mm ventral to bregma. Lesions in all animals were
Discussion
Although the rotarod assay remains a practical and well-validated tool for assessing neuromotor performance in rodents, it may lack the sensitivity to detect subtle motor deficits. Conversely, automated footprint-based gait procedures assess neuromotor performance objectively, precisely and sensitively. In the present study, we compared accelerating rotarod and treadmill gait procedures in mice with bilateral lesions of the cerebellar cortex. A carefully tuned electrolytic technique produced
Acknowledgements
The authors would like to thank Leen Van Aerschot for excellent technical assistance. Financial support was received from EU FP7 grant ALPHA-MAN and MM Delacroix Foundation.
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