Archives of Toxicology. Jan2014, Vol. 88 Issue 1, p109-113. 5p.
NEURONS, METHYLMERCURY, TOXICITY testing, MESSENGER RNA, LABORATORY rats, and CEREBELLUM
Methylmercury (MeHg), an environmental neurotoxicant, induces site-specific toxicity in the brain. Although oxidative stress has been demonstrated with MeHg toxicity, the site-specific toxicity is not completely understood. Among the cerebellar neurons, cerebellar granule cells (CGCs) appear vulnerable to MeHg, whereas Purkinje cells and molecular layer neurons are resistant. Here, we use a MeHg-intoxicated rat model to investigate these cerebellar neurons for the different causes of susceptibility to MeHg. Rats were exposed to 20 ppm MeHg for 4 weeks and subsequently exhibited neuropathological changes in the cerebellum that were similar to those observed in humans. We first isolated the three cerebellar neuron types using a microdissection system and then performed real-time PCR analyses for antioxidative enzymes. We observed that expression of manganese-superoxide dismutase (Mn-SOD), glutathione peroxidase 1 (GPx1), and thioredoxin reductase 1 (TRxR1) was significantly higher in Purkinje cells and molecular layer neurons than in CGCs. Finally, we performed immunohistochemical analyses on the cerebellum. Immunohistochemistry showed increased expression of Mn-SOD, GPx1, and TRxR1 in Purkinje cells and molecular layer neurons, which was coincident with the mRNA expression patterns. Considering Mn-SOD, GPx1, and TRxR1 are critical for protecting cells against MeHg intoxication, the results indicate that low expression of these antioxidative enzymes increases CGCs vulnerability to MeHg toxicity. [ABSTRACT FROM AUTHOR]
Abstract: The ability of docosahexaenoic acid (DHA) to modulate methylmercury (MeHg)-induced neurotoxicity was investigated in primary astrocytes and neurons from the cerebellum. Gas chromatography measurements indicated increased DHA content in both cell types after 24h supplementation. After individual or combined treatment with MeHg (10μM) and DHA (30 and 90μM), the cell-associated MeHg measurements were done using 14C-labelled MeHg. In addition, mitochondrial activity was evaluated by MTT reduction, glutathione (GSH) content was measured with the fluorescent indicator monochlorobimane (MCB) and reactive oxygen species (ROS) were detected with the fluorescent indicator—chloro methyl derivative of di-chloro di-hydro fluorescein diacetate (CMH2DCFDA). For all the tested treatments, i.e. DHA, MeHg or DHA+MeHg treatment, the neurons differed significantly (p <0.001) from astrocytes exhibiting increased ROS production and decreased MTT activity. After MeHg and 30μM DHA treatment there were no changes in MTT or GSH content but significant decrease (p <0.001) in ROS was observed in both the cell types when compared to MeHg alone. The cell-associated MeHg measurements indicated reduced MeHg-accumulation in both cell types (p <0.05) upon 30μM DHA exposure. Taken together, this study, for the first time establishes that DHA pretreatment effectively reduces cell-associated MeHg and prooxidant response from MeHg in both cerebellar astrocytes and neurons and thus supports the hypothesis that fish-derived nutrients offer possible neuroprotection from MeHg. [Copyright &y& Elsevier]