Abstract
Nitrative stress is a key component of the pathogenic process in Parkinson’s disease (PD), but the relative roles of constitutive neuronal nitric oxide synthase (n-NOS) and inducible nitric oxide synthase (i-NOS) in glial cells remain unresolved. We have investigated the effects of a range of concentrations of the selective n-NOS inhibitor ARR17477, and the selective i-NOS inhibitor 1400W, on MPP+-induced cell death in foetal ventral mesencephalic (VM) dopaminergic cultures. MPP+ induced a loss of TH-positive neurones accompanied by an increase in immunoreactivity for GFAP and OX-6 as markers of astrocytes and activated microglia, respectively, and induced i-NOS immunoreactivity. Unexpectedly, MPP+ treatment did not induce 3-NT immunoreactivity in the cultures. ARR17477 and 1400W alone had no effect on the number of TH-positive cells or on the number of GFAP or OX-6 positive cells. ARR17477 did not prevent the MPP+-induced decrease in TH-positive neurones and had no effect on the increased number of GFAP- and OX-6-positive cells. By contrast, 1400W caused a concentration-dependent preservation of TH-positive neurones in the presence of MPP+. It also significantly reduced the number of OX-6-immunoreactive cells and there was a small reduction in GFAP immunoreactivity. The results suggest a major role for i-NOS-mediated nitrative stress in microglia in MPP+-induced dopaminergic cell death and this may have important implications for developing neuroprotective strategies for PD.
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Abbreviations
- 3-NT:
-
3-Nitrotyrosine
- BSA:
-
Bovine serum albumin
- DAB:
-
3,3′-Diaminobenzidine tetrahydrochloride
- DI:
-
Deionised
- DIV:
-
Days in vitro
- D-MEM:
-
Dulbecco’s modified Eagle medium
- DPX:
-
Di-n-butylphthalate in xylene
- DS:
-
Donkey serum
- GFAP:
-
Glial fibrillary acidic protein
- i-NOS:
-
Inducible nitric oxide synthase
- MPP+ :
-
4-Methylpyridinium
- n-NOS:
-
Neuronal nitric oxide synthase
- OX-6:
-
Major histocompatibility complex class II
- PBS:
-
Phosphate buffered saline
- PD:
-
Parkinson’s disease
- PSN:
-
Penicillin, streptomycin and neomycin solution
- TH:
-
Tyrosine hydroxylase
- VM:
-
Ventral mesencephalic
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This research was funded by Proximagen Neurosciences Ltd., London, UK. There is no actual or potential conflict of interest in relation to this article.
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Brzozowski, M.J., Jenner, P. & Rose, S. Inhibition of i-NOS but not n-NOS protects rat primary cell cultures against MPP+-induced neuronal toxicity. J Neural Transm 122, 779–788 (2015). https://doi.org/10.1007/s00702-014-1334-8
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DOI: https://doi.org/10.1007/s00702-014-1334-8