Viral-toxin interactions and Parkinson's disease: poly I:C priming enhanced the neurodegenerative effects of paraquat

Abstract

Background: Parkinson's disease (PD) has been linked with exposure to a variety of environmental and immunological insults (for example, infectious pathogens) in which inflammatory and oxidative processes seem to be involved. In particular, epidemiological studies have found that pesticide exposure and infections may be linked with the incidence of PD. The present study sought to determine whether exposure to a viral mimic prior to exposure to pesticides would exacerbate PD-like pathology.

Methods: Mice received a supra-nigral infusion of 5 μg of the double-stranded RNA viral analog, polyinosinic: polycytidylic acid (poly(I:C)), followed 2, 7 or 14 days later by administration of the pesticide, paraquat (nine 10 mg/kg injections over three weeks).

Results: As hypothesized, poly(I:C) pre-treatment enhanced dopamine (DA) neuron loss in the substantia nigra pars compacta elicited by subsequent paraquat treatment. The augmented neuronal loss was accompanied by robust signs of microglial activation, and by increased expression of the catalytic subunit (gp91) of the NADPH oxidase oxidative stress enzyme. However, the paraquat and poly(I:C) treatments did not appreciably affect home-cage activity, striatal DA terminals, or subventricular neurogenesis.

Conclusions: These findings suggest that viral agents can sensitize microglial-dependent inflammatory responses, thereby rendering nigral DA neurons vulnerable to further environmental toxin exposure.

Figure 1

Total home-cage locomotor activity was monitored for 24 hours at three separate time points: 15 and 22 days after poly(I:C) infusion, and 24 hours before animals were killed. No significant differences were seen between treatment groups at any of the time intervals. However, there was a trend towards reduced activity at the middle time point (22 days). Data are expressed as a mean ± SEM; n = 8 to 15 Analysis of pole-test performance was used to investigate any PD-like hindlimb and forelimb coordination deficits that might have been induced by paraquat with or without poly(I:C) priming. The pole test was conducted once, during the second week after saline/poly(I:C) infusion. Latency to turn and latency to pole descent were both recorded for three trials. No behavioral difference were found for either of the measures across treatment groups (Fs_(5,42) < 1, P > 0.05; data not shown).

Figure 2

Substania nigra pars compacta (SNc) tyrosine hydroxylase (TH) + neurons were quantified from multiple bregma levels. Paraquat and poly(I:C) alone caused a modest, but non-significant TH + reduction relative to saline-treated controls. However, pre-treatment with poly(I:C) followed by paraquat induced a significant reduction in TH + neuronal counts at all bregma levels and at all re-exposure intervals. (A) Representative photomicrographs showing the degree of nigral TH + neuron loss in paraquat, poly(I:C), and the combination treatment groups compared with saline-treated controls. (B) Quantification of SNc TH + neuronal counts depicted in bar graphs. *P < 0.05, relative to saline-treated controls. All data expressed as mean ± SEM, n = 4 to 8.

Figure 3

Cresyl violet-stained tyrosine hydroxylase (TH)- neurons of the SNc from bregma level −3.08 to −3.40 were quantified as a measure of non-dopaminergic neurons. Treatment with paraquat or poly(I:C) alone did not affect TH- neuron counts, but poly(I:C) pre-treatment followed 2 days later by paraquat administration induced a significant decrease in TH- neuron counts at bregma −3.28. *P < 0.05, relative to saline-treated controls. Data expressed as mean ± SEM, n = 4 to 7.

Figure 4

Substania nigra pars compacta (SNc) sections were stained with anti-Cd11b and rated on morphological appearance as marker for microglial state. Paraquat or poly(I:C) alone promoted a modest but non-significant elevation of microglial activation state relative to saline-treated controls. However, paralleling the TH + changes, poly(I:C) priming followed by later paraquat exposure significantly augmented signs of microglial activation. (A) Representative photomicrographs showing the degree of nigral microglial immunoreactivity induced by paraquat and poly(I:C) (note that the poly(I:C)-PQ image was taken from the group that received paraquat 2 days after poly(I:C) pre-treatment). (B) Quantification of ratings (arbitrary units) for microglial activation state. *P < 0.05, relative to saline-treated controls. Data expressed as mean ± SEM, n = 4 to 5.

Figure 5

Substania nigra pars compacta (SNc) sections were stained with anti-gp91^PHOXas a marker for signs of oxidative stress. In agreement with the CD11b ratings, poly(I:C) priming followed by later paraquat treatment resulted in significantly increased expression of gp91^PHOX immunostaining relative to saline-treated controls. (A) Representative photomicrographs showing the degree of gp91^PHOX immunoreactivity in paraquat, poly(I:C), and the combination treatment groups, (note that the poly(I:C)-PQ image was taken from the group that received paraquat 2 days after poly(I:C) pre-treatment), (B) Quantification of ratings (arbitrary scale units) of gp91^PHOX immunoreactivity in the SNc. * P < 0.05, relative to saline-treated controls. Data expressed as mean ± SEM, n = 5–8.