Endurance exercise promotes cardiorespiratory rehabilitation without neurorestoration in the chronic mouse model of parkinsonism with severe neurodegeneration

Abstract

Physical rehabilitation with endurance exercise for patients with Parkinson's disease has not been well established, although some clinical and laboratory reports suggest that exercise may produce a neuroprotective effect and restore dopaminergic and motor functions. In this study, we used a chronic mouse model of Parkinsonism, which was induced by injecting male C57BL/6 mice with 10 doses of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (25 mg/kg) and probenecid (250 mg/kg) over 5 weeks. This chronic parkinsonian model displays a severe and persistent loss of nigrostriatal neurons, resulting in robust dopamine depletion and locomotor impairment in mice. Following the induction of Parkinsonism, these mice were able to sustain an exercise training program on a motorized rodent treadmill at a speed of 18 m/min, 0 degrees of inclination, 40 min/day, 5 days/week for 4 weeks. At the end of exercise training, we examined and compared their cardiorespiratory capacity, behavior, and neurochemical changes with that of the probenecid-treated control and sedentary parkinsonian mice. The resting heart rate after 4 weeks of exercise in the chronic parkinsonian mice was significantly lower than the rate before exercise, whereas the resting heart rate at the beginning and 4 weeks afterward in the control or sedentary parkinsonian mice was unchanged. Exercised parkinsonian mice also recovered from elevated electrocardiogram R-wave amplitude that was detected in the parkinsonian mice without exercise for 4 weeks. The values of oxygen consumption, carbon dioxide production, and body heat generation in the exercised parkinsonian mice before and during the Bruce maximal exercise challenge test were all significantly lower than that of their sedentary counterparts. Furthermore, the exercised parkinsonian mice demonstrated a greater mass in the left ventricle of the heart and an increased level of citrate synthase activity in the skeletal muscles. The amphetamine-induced, dopamine release-dependent locomotor activity was markedly inhibited in the sedentary parkinsonian mice and was also inhibited in the exercised parkinsonian mice. Finally, neuronal recovery from the loss of nigrostriatal tyrosine hydroxylase expression and dopamine levels in the severe parkinsonian mice after exercise was not evident. Taken all together, these data suggest that 4 weeks of treadmill exercise promoted physical endurance, resulting in cardiorespiratory and metabolic adaptations in the chronic parkinsonian mice with severe neurodegeneration without demonstrating a restorative potential for the nigrostriatal dopaminergic function.

Figure 1

Movement deficit in chronic PK mice. (A) Comparison of approximate entropy (ApEn) values between the chronic probenecid-treated and chronic MPTP/probenecid-treated mice 7 days after treatment (N=5 per group). *Significantly higher than the probenecid-treated control animals (mean ± S.E.M., p<0.05). A larger ApEn value denotes a less consistent movement pattern in the chronic PK mice. Digitized positions of a representative chronic probenecid-treated control mouse (B) and a chronic MPTP/probenecid-treated PK mouse (C) 7 days after treatment. Qualitatively it is apparent that the control mouse explored the perimeter of the cage with a more consistent movement pattern than the chronic PK mouse.

Figure 2

ECG analysis of cardiac responses to endurance exercise in chronic PK mice. (A) The resting heart rate in beats per min (bpm) was recorded in each experimental group (N=5−6/group) at the beginning (Week 0) and end of 4-week period with or without exercise (Week 4) and their values (mean ± S.E.M.) were compared. There were no significant intra-group differences in the resting heart rate among the three experimental groups at Week 0. After 4 weeks without exercise, the resting heart rate of the probenecid-treated control and sedentary PK group was not changed from their respective value at Week 0. However, the resting heart rate in the 4-week exercised PK mice was significantly lower than their rate at the beginning of exercise (*p<0.007). (B) The amplitude of the ECG R wave was also analyzed in the three groups of animals at the beginning of the study (Week 0) and again 4 weeks later (Week 4). There were no significant intra-group differences in the R wave amplitude in all 3 groups of mice at Week 0. There was a significant increase in the R wave amplitude in the sedentary PK mice at Week 4 comparing to Week 0 (*p<0.01). The R wave amplitude in the exercised PK mice at Week 4 was not different from the probenecid-treated control animals; however, it was significantly lower than the 4 week sedentary PK mice (*p<0.001).

Figure 3

Effect of exercise on myocardial left ventricle mass in the chronic PK mice. The left ventricle of the heart from each experimental animal was isolated and weighed. The left ventricle mass was expressed in mg per g of body weight raised to the power of 0.78, an index factor for estimating the lean body mass (Kemi et al., 2002). The left ventricle mass (mean ± S.E.M.) of the exercised PK mice (N=4) was significantly greater than the mass of either the probenecid-treated controls (N=5) or the sedentary PK mice (N=5) (*p<0.001) suggesting that 4 weeks of exercise induced cardiac hypertrophy in the chronic PK mice.

Figure 4

Effect of exercise on respiratory and thermal responses. At the end of 4-week session with or without exercise, O₂ consumption (VO₂), CO₂ production (VCO₂), and body heat generation were measured in all 3 experimental groups of animals (N=5−6/group) in an enclosed treadmill/calorimetry apparatus before and during the Bruce maximal exercise test. (A) The sedentary PK mice had a significantly higher before-test O₂ consumption than either probenecid-treated or exercised PK mice (*p<0.05). During the maximal exercise test, VO₂ consumption was expectedly elevated in all groups of animals while running at 6 m/min (0° inclination) (^ə p<0.05). No further increase of VO₂ was detected while running maximally at 15 m/min (15° inclination), which might be an indication of physical fatigue. Comparing VO₂ at 6 m/min (0° inclination) and 15 m/min (15° inclination), the exercised PK mice consumed less O₂ than the sedentary counterparts (ϕp<0.05). (B) The before-test CO₂ production in all 3 groups of animals was statistically indifferent. During the maximal exercise test, VCO₂ levels were elevated in all groups of animals when running at 6 m/min (0° inclination) and at 15 m/min (15° inclination) (^*p<0.05). Running at 6 m/min (0° inclination), VCO₂ produced by the exercised PK mice was significantly lower than that produced by the sedentary PK mice (^əp<0.03). (C) During the maximal exercise test, all groups of animals generated slightly higher but not statistically significant body heat when running at 6 m/min (0° inclination) and at 15 m/min (15° inclination). The values were indifferent between probenecid-treated controls and sedentary PK mice. However, body heat production by the exercised PK mice before and during the maximal exercise test were significantly lower than the sedentary PK mice (*p<0.01).

Figure 5

Effect of exercise on citrate synthase activity in the gastrocnemius muscle. The citrate synthase activity (mean ± S.E.M.) in the gastrocnemius muscle of the 4-week exercised PK mice (N=6) was significantly higher than the activity obtained from the probenecid-treated (N=5) or sedentary PK mice (N=6) (*p<0.001).

Figure 6

Effect of exercise on amphetamine-induced locomotor activity. The horizontal movement expressed as cumulative distance traveled by each animal was recorded over 30 min. The amphetamine (3 mg/kg, i.p.)-induced locomotor activity in the sedentary PK mice was significantly inhibited after 4 weeks without exercise when compared with the probenecid-treated control mice (*p<0.01). The amphetamine-induced locomotor activity remained to be inhibited after four weeks of exercise in the chronic PK mice when compared with the control mice (**p<0.02) and not significantly different from the sedentary PK mice.

Figure 7

Western blot analysis of TH protein expression in (A) the striatum and (B) substantia nigra of chronic probenecid-treated (N=5), sedentary PK (N=6), and exercised PK mice (N=6). The TH content was expressed as an OD ratio with reference to that of β-tubulin. Western blots revealed that there was a marked reduction of TH content in both striatum and SN of the sedentary PK mice 4 weeks after treatment when compared with the probenecid-treated mice (*P<0.001). The TH protein expressions in the striatum and SN of 4-week exercised PK mice were significantly lower than the probenecid-treated controls (**p<0.03) but not significantly higher than the sedentary counterparts.

Figure 8

Striatal DA (A) and DOPAC (B) contents in chronic probenecid-treated (N=5), sedentary PK (N=6), and exercised PK mice (N=6). Striatal DA and DOPAC levels in the sedentary PK mice 4 weeks after treatment were significantly lower than the probenecid-treated mice (*P<0.0001). The striatal DA and DOPAC levels in 4-week exercised PK mice were significantly lower than the probenecid-treated controls (**p<0.0001) but not significantly different from the sedentary counterparts.