Skilled forelimb reaching in Wistar rats: Evaluation by means of Montoya staircase test

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Abstract

Experimental animals have been used as models for several neurological disorders; their performance in behavioral tests is useful in determining the success of lesion repair procedures and assessing functional recovery. The staircase test is a behavioral test that consists in reaching for food inside a special box and allows for a sensitive measure of skilled reaching by each limb in an independent manner. In most laboratories in the south of Brazil, Wistar rats are used for the study of experimental stroke, hypoxia and peripheral neuropathy, but most studies with the staircase test have used other strains such as Sprague–Dawley and Long–Evans. Because skilled reaching, grasping and performance can differ among strains, the purpose of the present study was to characterize the performance of Wistar rats in the staircase test and determine the effect of median and ulnar nerve crush. Our results with Wistar rats on the staircase test showed that: similar to other strains, Wistar animals can display high performance after 2 weeks of training; the number of animals that attained the inclusion criterion increased by 10% with longer times of training; the stricter criterion of 15 pellets taken can be adopted as study inclusion criterion; the test has an unquestionable value in assessing lateralized deficits, as evidenced by the lack of performance deficit of the non-manipulated forelimb at any time point. These results extend the understanding about the performance of Wistar rats in the staircase test, which will be used for the best training and research using this strain.

Introduction

Rodents, including rats and mice, have been used as models for several neurological diseases. Their performance in behavioral tests are widely used to evaluate the success of surgical procedures that cause impairments to motor control, as well as to assess the pattern of functional recovery after different types of treatment (Biernaskie and Corbett, 2001, Windle et al., 2006).

The need to assess goal-directed movement abilities in animal models has led to the development of a variety of tests. The Montoya staircase test was developed to evaluate lateralized forepaw ability (the animal must reach with the left or right forepaw independently), and it allows a sensitive and quantitative measure of skilled reaching and grasping in rats and mice (Montoya et al., 1991, Whishaw et al., 1997, Baird et al., 2001, Clarke et al., 2007). This skilled forelimb task is largely used to evaluate lesion-induced impairments and recovery after nervous system lesion in different strains. The staircase test has been mostly applied in Sprague–Dawley rats for assessment of recovery from brain ischemia after training, or in Long–Evans rats to evaluate either toxicological effects of drugs (Samsam et al., 2004) or motor cortical electrical stimulation (Adkins-Muir and Jones, 2003). Besides its use for the study of central nervous injury, the staircase test has been recently employed to evaluate recovery after peripheral nerve injury in Lister–Hooded rats (Galtrey and Fawcett, 2007).

Although Wistar rats have been rarely used in studies of skilled forelimb reaching, compared to Sprague–Dawley and Long–Evans rats, they have been widely used in the south of Brazil in most research laboratories studying experimental stroke, hypoxia and ischemia (Valentim et al., 1999, Pagnussat et al., 2007, Pereira et al., 2007) and peripheral neuropathy (Rodrigues-Filho et al., 2003). Skilled reaching, grasping acquisition and performance can, however, vary with rat strain (Nikkhah et al., 1998). Galtrey and Fawcett (2007) reported a poor performance of Lewis rats in the staircase test. Differences in motor behavior have been described in a task of reaching for food from a shelf (single pellet reaching) either between Long–Evans and Fischer-344 rats (VandenBerg et al., 2002) or between Long–Evans and Sprague–Dawley rats (Whishaw et al., 2003). Little has been reported about Wistar rats’ reaching performance (Metz et al., 2005), particularly using the staircase test.

Furthermore, we can find in the existing literature several differences in training protocols (number of trials, number of steps, number of pellets in well, time of test and inclusion criteria). Most studies use three pellets in each step, 2 weeks of training and 15 min for the test (Biernaskie and Corbett, 2001, Clarke et al., 2007). For a total of 21 pellets per side, Hewlett and Corbett (2006) adopted a criterion of a minimum of 12 pellets reached on the last eight trials, with a standard deviation less than ±2 pellets. However, because rats could access the food pellets on each bottom stair with their tongue (Hoane et al., 2004) a stricter criterion can be adopted.

To be confident in assessing the efficacy of a treatment approach in Wistar rats, we must know if skilled forelimb use, in terms of scores of pellets taken and performance at the end of training, is similar to those described for other rats. The brachial plexus lesion provides a good experimental model to assess the control of muscle function, examine the mechanisms underlying functional recovery, and test the effects of treatments to enhance recovery (Bertelli et al., 2005).

Therefore, the aims of this study were: (1) to characterize acquisition of the staircase test for inbred albino Wistar rats in terms of pellets taken; (2) to compare final performance of inbred albino Wistar rats after two different periods of training (completing 20 and 50 trials); (3) to evaluate performance before and after brachial plexus avulsion.

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Animals

Adult Wistar rats (obtained from a local breeder-Central Animal House of the Institute of Basic Health Sciences, Universidade Federal do Rio Grande do Sul), weighing approximately 300 g at the beginning of the experiment, were used. Animals were housed in groups of four or five in Plexiglas cages under standard conditions with a 12 h light/dark cycle (lights off at 7:30 p.m.) and controlled temperature (22 ± 2 °C).

Water and standard laboratory chow were provided ad libitum except during behavioral

Results

All animals learned to collect pellets from the staircases in the first two days of training. The success score in reaching progressively improved over the days, as evidenced by the number of pellets eaten. The learning curve of pellets taken and eaten with the right and left forepaw is illustrated in Fig. 2. At the end of the training period (2 weeks), Wistar rats could take on average 15.3 ± 0.5 pellets with the right and 14.2 ± 0.6 with the left paw (mean ± S.E.M.).

Discussion

Since its development, the staircase test has been widely used to evaluate the performance of forelimb reach in a variety of studies (Montoya et al., 1990, Whishaw et al., 1997, Kloth et al., 2006, Milman et al., 2006, Ago et al., 2007, Clarke et al., 2007). This is a standardized method to assess the functional integrity of forelimbs by means of reaching and grasping movements in rodents (Montoya et al., 1991, Baird et al., 2001). To obtain the food, the animal must reach for, grasp, and

Acknowledgments

The authors thank Ana Luisa Jeanty de Seixas, Erica do Espírito Santo Hermel and Jocemar Ilha and for their technical assistance. This study was supported by the Brazilian Funding Agency CNPq. Carlos Alexandre Netto is a CNPq investigator and Aline de Souza Pagnussat has a CNPq Ph.D. fellowship. Dr. A. Leyva provided English editing of the manuscript.

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