Selective neurotoxicity of ruthenium red in primary cultures

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Abstract
The inorganic dye ruthenium red (RuR) has been shown to be neurotoxic in vivo when injected intracerebrally. In this work toxicity of RuR was compared in primary cultures of rat cotical neurons, cerebellar granule neurons and cerebellar astroglia. Microscopic examination of the cultures revealed that RuR penetrates the somata of both types of neurosn used and produces vacuolization and loss and fragmentation of neurites. In contrast, no RuR was seen inside cultured astrocytes and no morphological signs of damage were observed in these cells. RuR toxicity wa also assessed by immunocytochemistry of α-tubulin and by biochemical measurement of the reduction of (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) by the cultured cells. The morphological alteratios in the neuros were closely correlated with loss of tubulin immunoreactivity and partivular with a notable decrement in the ability to reduce MTT. Using the latter parameter, it was found that neuronal damage was independent of the age of the cultures, augmented progressively with time of incubation with RuR, from 8 to 24 h, and showed a clear dose-response curve from 20 to 100 μM RuR. Astrocytes showed only a slight decrease in MTT reduction after 24 h of incubation with 100 μM RuR. It is concluded that RuR seems to be toxic for neurons but not for astroglia, and that this selectivity is probably related to the ability of the neurons to internalize the dye. The possible mechanisms ofRuR penetration and neuronal damage are discussed.