Table 1.
Number, design and location of studies examining effects of paraquat and MPTP on C57BL/6J and C57BL/6NHsd male mice (aged 9 or 16 weeks)1.
Fig 1.
Stereological assessment of the mean number of TH+ neurons in the SNpc following PQ or MPTP treatment in C57BL/6J and C57BL/6NHsd male mice.
Five different groups (G1-G5) of animals, varying in age and site of experiment were injected with saline, paraquat or MPTP and the extent of TH+ neuron loss was assessed by design-based or model-based stereology. ** significantly different from control mice (p ≤ 0.01). Syngenta-sourced PQ was used to treat mice G1 to G3 mice, whereas Sigma Chemical PQ was used to treat mice in groups G4 and G5.
Fig 2.
Stereological assessment of the mean number of resting and activated microglial cells in the SNpc of C57BL/6J or C57BL/6NHsd male mice treated with saline, PQ or MPTP.
No change in resting or activated microglia number was seen in PQ-treated mice of either C57BL/6 substrain, while both C57BL/6 substrains demonstrated increased numbers of activated and total microglia 7 days following MPTP treatment. * p ≤ 0.05, **p ≤ 0.01.
Fig 3.
Appearance of microglia in PQ- and MPTP-treated mice.
Representative photomicrographs of microglia (Iba-1+) cells in PQ-treated (Panels A-D) or MPTP-treated (Panel E-H) in the SNpc of C57BL/6J or C57BL/6NHsd male mice aged 9- or 16-weeks at the time of the 1st dose. The boxes in each panel indicate the region shown in the adjacent box. Red arrow in D shows an example of resting microglia; characterized by a small cell body and thin processes. Red arrows in H show the typical appearance of activated microglia seen in MPTP-treated mice where the cell body is increased in size compared to resting microglia and the processes are shortened and thickened. Scale bars A,E = 100 μm, B,F = 40 μm, C,G = 20 μm, D-H = 10 μm.
Fig 4.
Correlation between the number of activated microglia in the SNpc and the number of TH+ neurons.
The correlation between the number of activated microglia in the SNpc and the number of TH+ neurons was assessed in the same animal in two studies on C57BL/6J mice (A, B) and one study using C57BL/6NHsd (C) performed at two different sites (WIL, A) and SJCRH (B, C). A significant negative inverse correlation between TH+, DA neuron number and activated microglia was observed in MPTP-treated mice, while no correlation was seen in PQ- or saline treated mice.
Fig 5.
Pathological assessment of SNpc and striatum in PQ- or MPTP-treated mice.
Microscopic appearance of the SNpc (G-L) and striatum (A-F) in control (column 1), PQ- (column 2) or MPTP-treated (column 3) mice, 48 hours after dosing. Tyrosine hydroxylase (TH) immunostaining was decreased in the striatum (C) and SNpc (I) of the MPTP-treated animal but was unchanged in the PQ-treated mouse (B, H) compared to the control (A, G). Amino cupric silver (AmCuAg) staining was used to reveal degenerating neurons in the SNpc (J-L) and degenerating fibers in the striatum (D-F). There were no differences in AmCuAg staining in either the SNpc or striatum after PQ treatment compared to the control (D,J). AmCuAg staining was increased in the SNpc and striatum of the MPTP-treated mouse (F,L). The scale bar shown in Panel L represents 400 μm in panels A-C, G-I and 40 μm in panels D-F and J-L.
Fig 6.
Appearance of microglia and astrocytes in SNpc and striatum of PQ- and MPTP-treated mice.
Microscopic appearance of Iba-1-stained microglia (A-F) and GFAP-stained astroctyes (G-L) in the SNpc of mice 48 hours after PQ (column 2) or MPTP treatment (column 3) compared to a control (column 1). Iba-1 immunostaining of microglia was increased in the MPTP-treated mouse (C, F) but was not different in the PQ-treated mouse (B, E) compared to the control (A, D). Similarly, increased GFAP immunostaining of astrocytes was noted in the MPTP-treated mouse (I, L) but not in the PQ-treated mouse (H, K) compared to the control (G, J). Scale bar A-L 40 μm.
Fig 7.
Mean histopathological severity scores in control, paraquat and MPTP-treated groups of C57BL/6J male mice.
Mice were 16 weeks of age at the time of treatment initiation. Mice were administered 10 mg/kg/dose PQ·Cl2 by ip injection, twice a week for 3 weeks and were sacrificed 8, 16, 24, 48, 96 or 168 hours after the last dose. Control mice were given the vehicle while MPTP-treated mice received four injections of MPTP (16 mg/kg/dose; expressed as free base) at 2-hour intervals, and then euthanized 48 hours after the final dose. Serial sections through the SNpc were evaluated qualitatively and the group mean severity grades are plotted. Grades 0 to 5 reflect increasing intensity of staining for Iba-1, AmCuGg, GFAP and decreased staining intensity of TH.