The effect of low dose amphetamine in rotenone-induced toxicity in a mice model of Parkinson's disease

Abstract

Objectives: The effects of low dose amphetamine on oxidative stress and rotenone-induced neurotoxicity and liver injury were examined in vivo in a mice model of Parkinson's disease.

Materials and methods: Male mice were treated with rotenone (1.5 mg/kg, every other day for two weeks, subcutaneously). Mice received either the vehicle or amphetamine intraperitoneally at doses of 0.5, 1.0, or 2.0 mg/kg. Oxidative stress was assessed by measurement of the lipid peroxidation product malondialdehyde (MDA), nitric oxide (NO), total anti-oxidant capacity (TAC), and paraoxonase-1 (PON-1) activity in the brain and liver. In addition, brain concentrations of nuclear factor kappa B (NF-κB) and tyrosine hydroxylase were determined and histopathology and Bax/Bcl-2 immunohistochemistry were performed.

Results: The levels of lipid peroxidation and NO were increased and TAC and PON-1 were decreased significantly compared with vehicle-injected control mice. There were also significantly increased NF-κB and decreased tyrosine hydroxylase in the brain following rotenone administration. These changes were significantly attenuated by amphetamine. Rotenone caused neurodegenerative changes in the substantia nigra, cerebral cortex, and hippocampus. The liver showed degenerative changes in hepatocytes and infiltration of Kupffer cells. Bax/Bcl2 ratio was significantly increased in brain and liver tissues. Amphetamine prevented these histopathological changes and the increase in apoptosis evoked by rotenone.

Conclusion: These results suggest that low dose amphetamine exerts anti-oxidant and anti-apoptotic effects, protects against rotenone-induced neurodegeneration, and could prevent neuronal cell degeneration in Parkinson's disease.

Keywords: Amphetamine; Anti-oxidant capacity; Neuroprotection; Parkinson’s disease; Reactive oxygen species; Rotenone.

Figure 1

Effect of amphetamine treatment on brain malondialdehyde (MDA), nitric oxide, total anti-oxidant capacity (TAC), and paraoxonase-1 (PON-1) activity in rotenone-treated mice. *: P<0.05 vs vehicle. +: P<0.05 vs rotenone control. #: P<0.05 vs rotenone +amphetamine 0.5 mg/kg

Figure 2

Effect of amphetamine treatment on the rotenone-induced changes in nuclear factor kappa-B (NF-κB) and tyrosine hydroxylase in mice brain. *: P<0.05 vs vehicle. +: P<0.05 vs rotenone control

Figure 3

Effect of amphetamine treatment on liver malondialdehyde (MDA), nitric oxide, total antioxidant capacity (TAC), and paraoxonase-1 (PON-1) activity in rotenone-treated mice. *: P<0.05 vs vehicle. +: P<0.05 vs rotenone control. #: P<0.05 vs rotenone +amphetamine 0.5 mg/kg

Figure 4

Representative photomicrographs of sections of the substantia nigra from (A) Vehicle. (B) Rotenone only. (C) Rotenone +amphetamine 0.5 mg/kg. (D) Rotenone+amphetamine 1 mg/kg. (E) Rotenone+amphetamine 2 mg/kg. Black arrows; degenerated neuronal cells. Green arrows; healthy dendritic pigmented neurons (H&E x 200 & 400)

Figure 5

Representative photomicrographs of sections of the cerebral cortex from (A) Vehicle. (B) Rotenone only. (C) Rotenone+amphetamine 0.5 mg/kg. (D) Rotenone+amphetamine 1 mg/kg. (E) Rotenone+amphetamine 2 mg/kg. Abbreviations: ML; molecular layer, PL; pyramidal layer. Black arrows; normal neuronal cells. Green arrows; Purkinje cells. Yellow arrows; pyknotic deeply stained nuclei, Blue arrows; cells with nuclear karyolysis. Red arrows; congested cerebral vessels (H&E 200x & 400x)

Figure 6

Representative photomicrographs of sections of hippocampus region from (A) Vehicle. (B) Rotenone only. (C) Rotenone+amphetamine 0.5 mg/kg. (D) Rotenone+amphetamine 1 mg/kg. (E) Rotenone+amphetamine 2 mg/kg. Black arrow heads; pyknotic nuclei. Black arrows; normally granular cells with rounded vesicular nuclei (H&E x400)

Figure 7

Representative photomicrographs of Bax immunoreactivities in the cortex from (A) Vehicle. (B) Rotenone only. (C) Rotenone+amphetamine 0.5 mg/kg. (D) Rotenone+amphetamine 1 mg/kg. (E) Rotenone+amphetamine 2 mg/kg (BAX 400x)

Figure 8

Representative photomicrographs of sections of hippocampus region stained by BAX from (A) Vehicle. (B) Rotenone only. (C) Rotenone+amphetamine 0.5 mg/kg. (D) Rotenone+amphetamine 1 mg/kg. (E) Rotenone+amphetamine 2 mg/kg (BAX 200x &400x)

Figure 9

Representative photomicrographs of Bcl2 immunoreactivities in the cortex from (A) Vehicle. (B) Rotenone only. (C) Rotenone+amphetamine 0.5 mg/kg. (D) Rotenone+amphetamine 1 mg/kg. (E) Rotenone+amphetamine 2 mg/kg (Bcl 400x)

Figure 10

Representative photomicrographs of sections of liver tissue from (A) Vehicle. (B) Rotenone only. (C) Rotenone+amphetamine 0.5 mg/kg. (D) Rotenone+amphetamine 1 mg/kg. (E) Rotenone+amphetamine 2 mg/kg. Abbreviations: CV; central vein, CHV; congested hepatic vein, S; dilated congested sinusoids, IN; inflammatory cells. Yellow arrows; Kupffer cells. Blue arrows; binucleated hepatocytes. Black arrows; ductular hyperplasia (H&E x200)

Figure 11.

Representative photomicrographs of Bax immunoreactivities in the liver from (A) Vehicle. (B) Rotenone only. (C) Rotenone+amphetamine 0.5 mg/kg. (D) Rotenone+amphetamine 1 mg/kg. (E) Rotenone+amphetamine 2 mg/kg (Bax 200x)

Figure 12

Representative photomicrographs of Bcl2 immunoreactivities in the liver from (A) Vehicle. (B) Rotenone only. (C) Rotenone+amphetamine 0.5 mg/kg. (D) Rotenone+amphetamine 1 mg/kg. (E) Rotenone+amphetamine 2 mg/kg (Bcl2 200x)