Inhibition of neuroinflammatory nitric oxide signalling supresses protein glycation and recovers neuronal dysfunction in prion disease

Abstract Background: Several neurodegenerative diseases associated with protein misfolding (Alzheimer’s, Parkinson’s disease) exhibit oxidative and nitrergic stress following initiation of neuroinflammatory pathways. Associated nitric oxide (NO)-mediated post-translational modifications impact upon protein functions that can exacerbate pathology. Non-enzymatic and irreversible glycation signalling has been implicated as an underlying pathway that promotes protein misfolding, but the direct interactions between both pathways are poorly understood. Methods: Here we investigated the potential therapeutic effects of supressing neurotoxic NO signalling during early progression of prion disease. Tg37 mice aged 3-4 weeks were inoculated by intracerebral injection with either 1% brain homogenate of Rocky Mountain Laboratory (RML) scrapie prion protein or control normal brain homogenate (NBH). Hippocampal gene and protein expression levels of oxidative and nitrergic stress markers were analysed and electrophysiological characterisations of pyramidal neurons were performed in 6-10 weeks old RML and NBH mice. Mice were injected with a NO synthase (NOS) inhibitor and the time course of disease markers was compared to controls. Electrophysiology, immunoblotting and immunocytochemistry studies were performed to identify the effects of NOS inhibition on neurophysiology, glycation, prion protein misfolding and cell death. Statistical analyses employed two-tailed unpaired Student’s t-test, one-way or two-way ANOVA as required and data were considered significant with P<0.05.Results: Increased neuroinflammatory signalling was observed in mice between 6 and 10 weeks post inoculation (w.p.i.) with scrapie prion protein which was characterised by enhanced nitrergic stress and associated with a decline in hippocampal neuronal function by 9 w.p.i.. Daily in vivo administration of the NOS inhibitor L-NAME between 6 and 9 w.p.i. at 20 mg/kg abolished the functional degeneration of hippocampal neurons in prion mice. We further found that this intervention in diseased mice ameliorated 3-nitrotyrosination of triose-phosphate isomerase, an enzyme involved in the formation of disease-associated glycation signalling. Furthermore, L-NAME application reduced the expression of the receptor for advanced glycation end products and the accumulation of hippocampal prion misfolding. Conclusions: Our data suggest that alleviating nitrergic stress during early phases of neurodegeneration reduces neurotoxic post-translational NO signalling and glycation-assisted prion misfolding in the hippocampus, a mechanism which might be applicable to other protein misfolding neurodegenerative conditions.