Forced limb-use effects on the behavioral and neurochemical effects of 6-hydroxydopamine

Abstract

Rats with unilateral depletion of striatal dopamine (DA) show marked preferential use of the ipsilateral forelimb. Previous studies have shown that implementation of motor therapy after stroke improves functional outcome (Taub et al., 1999). Thus, we have examined the impact of forced use of the impaired forelimb during or soon after unilateral exposure to the DA neurotoxin 6-hydroxydopamine (6-OHDA). In one group of animals, the nonimpaired forelimb was immobilized using a cast, which forced exclusive use of the impaired limb for the first 7 d after infusion. The animals that received a cast displayed no detectable impairment or asymmetry of limb use, could use the contralateral (impaired) forelimb independently for vertical and lateral weight shifting, and showed no contralateral turning to apomorphine. The behavioral effects were maintained throughout the 60 d of observation. In addition to the behavioral sparing, these animals showed remarkable sparing of striatal DA, its metabolites, and the expression of the vesicular monoamine transporter, suggesting a decrease in the extent of DA neuron degeneration. Behavioral and neurochemical sparing appeared to be complete when the 7 d period of immobilization was initiated immediately after 6-OHDA infusion, only partial sparing was evident when immobilization was initiated 3 d postoperatively, and no sparing was detected when immobilization was initiated 7 d after 6-OHDA treatment. These results suggest that physical therapy may be beneficial in Parkinson's disease.

Fig. 1.

Example of the appropriate positioning of a rat during the test for forelimb akinesia. The hindquarters of an animal are suspended while the animal supports its weight on only one forelimb. The animal is allowed to initiate stepping movements in a 10 sec period for one forelimb and then the other in a balanced order.

Fig. 2.

Limb-use asymmetry was prevented with early forced use, but not in animals in a delayed forced-use condition, which showed behavior similar to that of noncast animals. Animals that received the cast on days 3–9 also showed no significant group difference from shams. Bonferroni post hoc analysis indicated significant differences (*) between sham controls (sham) and lesion + no cast (no cast) and lesion + late cast (c 7–13) groups.

Fig. 3.

Forced use of the impaired forelimb on days 1–7 and 3–9 after 6-OHDA exposure ameliorated akinesia. Those animals that did not receive the cast until days 7–13 displayed strong akinetic tendencies as did the animals in the no cast group. Bonferronipost hoc analysis indicated significant differences (*) between sham controls and lesion + no cast and lesion + late cast groups.

Fig. 4.

Limb-placing deficits were prevented by early (days 1–7) and intermediate (days 3–9) forced use of the impaired forelimb. The no treatment and late (days 7–13) treatment groups were significantly different (*) from the sham groups with an increased limb-placing deficit.

Fig. 5.

The number of contralateral rotations for sham groups was significantly different (*) from that of both the lesioned + no cast group and the lesioned + late cast group. Animals that received casts on days 1–7 or 3–9 did not differ from sham animals. Contralateral rotations were determined by counting the number of contralateral rotations in a 90 min period after apomorphine administration.

Fig. 6.

Animals receiving early casts (days 1–7) and sham animals do not show significant differences in DA, DOPAC, or HVA levels. Animals receiving casts on days 3–9 show intermediate DA levels, although still not significantly different from sham. Animals with late casts (days 7–13) and animals not receiving casts show significantly lower (*) DA levels when compared with sham-treated animals.

Fig. 7.

DOPAC/DA ratio is seen as a measure of DA turnover. The ratio is increased in lesioned plus no cast animals and in lesioned plus cast on days 3–9 (d3) and lesioned plus cast on days 7–13 (d7) groups, possibly reflecting a compensatory change in these animals. Animals that received a cast on days 1–7 (d1) do not show an increase in the DOPAC/DA ratio when compared with shams. Bonferronipost hoc analysis indicated significant trends (+) between sham controls and no cast animals, as well as between sham and lesion plus late cast animals. In addition, there was a trend toward significance (+) when comparing lesion plus intermediate (days 3–9) and sham animals.

Fig. 8.

Vesicular monoamine transporter expression in the striatum was measured using Western blot techniques. An overall analysis of group revealed a significant difference in the amount of VMAT2 remaining in the striatum after lesioning between those animals that did not receive a cast (Lesion No Cast) and those animals that were cast for 7 d immediately after surgery (Lesion Cast D1–7). Each striatum was divided into lateral (Lat) and medial (Med) sections. Shown are the ipsilateral or lesioned side (i) versus the contralateral or control side (c). No significant differences in VMAT expression or depletion were seen between these two sections, and the data were pooled for densitometric analysis. This is a representative blot of four separate experiments.