Effects of commissural de- and remyelination on motor skill behaviour in the cuprizone mouse model of multiple sclerosis
Introduction
Multiple sclerosis (MS), an inflammatory disorder of the central nervous system (CNS), is often associated with a relapsing-remitting disease course in which cycles of newly clinical deficits emerge that resolve within a few days or weeks. The clinical outcome in MS ultimately depends on the balance of damage inflicted on the CNS and the extent of repair that can counter it (Frohman et al., 2006). Among several compensatory mechanisms, repair of myelin (“remyelination”) has been postulated for the observed functional recovery in MS and in animal models of this disease, and remyelination has been identified frequently in postmortem brain analyzes of MS patients (Prineas and Connell, 1979, Bruck et al., 2003, Barkhof et al., 2003). As an animal model of MS, feeding of neurotoxicant cuprizone (bis-cyclohexanone-oxaldihydrazone) at a low dose of 0.2% (wt/wt) to young adult C57/BL6 mice induces consistent demyelination of the corpus callosum (Hiremath et al., 1998). Furthermore, after 5–6 weeks of exposure, its removal from the diet results in extensive remyelination (Arnett et al., 2001, Merkler et al., 2005). Together with the availability of knockout and transgenic mice, the cuprizone model has developed into an attractive tool to study the events related to myelin damage and repair (Matsushima and Morell, 2001). Nevertheless, testing for overt behavioural impairments related to cuprizone-induced demyelination in the CNS have so far not been reported. Analogously, the genetic absence of corpus callosum (inbred line derived from Balb/cWah1 and 129ReJ) or its surgical transection is associated only with subtle behaviour deficits (Schalomon and Wahlsten, 2002). This has limited the practicability of the cuprizone model in the analysis of functional impairments and studies have so far focused exclusively on molecular and morphological analysis.
The primary aim of this study was therefore to develop a motor test that allows the sensitive detection of latent central motor impairments related to cuprizone-induced demyelination and functional recovery following remyelination. We thus developed the motor skill sequence paradigm (MOSS), which is based on a sequence of voluntary running performances. In a first step, animals were habituated to a running wheel with regularly spaced crossbars (training wheel) for two weeks, until maximal wheel-running performance was achieved. After this training step, which ensures cardiopulmonary and musculoskeletal exercise, we introduced the animals for an additional week to a wheel with irregularly spaced crossbars (complex wheel), which demands highly complex bihemispherical motor coordination. This two-step approach of MOSS was completely automated and reliably revealed central motor deficits during the phase of cuprizone-induced demyelination and partial recovery after remyelination. Thus, the MOSS test system represents an extremely sensitive tool to include functional aspects related to myelin damage and repair in the cuprizone model and putatively in other animal models of human CNS diseases with latent central motor deficits.
Section snippets
Animals and cuprizone feeding
The experiments were conducted in accordance with German animal protection laws and were approved by the Government of Lower Saxony. The experiments were performed on 48 female C57BL/6 mice (8 weeks, 16.5 ± 1.2 g). The animals were housed individually under standard laboratory conditions with food and water ad libitum. They were divided randomly into two treatment (n = 2 × 12) and two control groups (n = 2 × 12). The mice of the treatment groups were fed for 6 weeks with 0.2% (wt/wt) cuprizone (Sigma, St
Callosal de- and remyelination in the cuprizone mouse model
The admixture of 0.2% cuprizone to the diet induces almost complete neuro-inflammation and demyelination within 4–6 weeks in the corpus callosum of C57BL/6 mice (Hiremath et al., 1998, Mason et al., 2001). After withdrawal of cuprizone from the diet, an almost complete remyelination occurs within a time frame of weeks (Matsushima and Morell, 2001). To verify that our treatment regimen with cuprizone was effective in our experimental setup, histological examination of PFA-fixed brain tissue was
Discussion
In the present study, we applied the cuprizone C57/Bl6 mouse model of MS to investigate the feasibility of a novel murine motor task, that is, the motor skill sequence (MOSS). We show that MOSS represents a sensitive tool to characterize and quantify behavioural correlates of callosal demyelination and remyelination. In the cuprizone mice model, motor impairments have so far been difficult to detect, despite extensive demyelination in the corpus callosum. The wheel-running protocol of MOSS
Acknowledgments
We thank C. Crozier for improving the English. We also thank M. Schedensack for expert technical assistance. This study was supported by the IMSF (Gemeinnützige Hertie Stiftung, Germany, Grant-No: 1.01.1/02/007) and Walter and Ilse Rose Stiftung (Grant No.:T298/14375/2004).
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