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The present study tested the effects of EUK-134, a synthetic superoxide dismutase/catalase mimetic, on several indices of oxidative stress and neuropathology produced in the rat limbic system as a result of seizure activity elicited by systemic kainic acid (KA) administration. Pretreatment of rats with EUK-134 did not modify the latency for or duration of KA-induced seizure activity. It did produce a highly significant reduction in increased protein nitration, activator protein-1- and NF-[Kappa]B-binding activity, and spectrin proteolysis as well as in neuronal damage resulting from seizure activity in limbic structures. These results support the hypothesis that kainate-induced excitotoxicity is caused, at least in part, by the action of reactive oxygen species. Furthermore, they suggest that synthetic superoxide dismutase/catalase mimetics such as EUK-134 might be used to prevent excitotoxic neuronal injury.

Abstract: The identification of mRNAs that have restricted expression patterns in the brain represents powerful tools with which to characterize and manipulate the nervous system. Here, we describe a strategy using microarray technology (Affymetrix Mouse Genome 430 2.0 Arrays) to identify mRNA transcripts that are candidate markers of cerebellar Purkinje neurons. Initially, gene expression profiles were compared between cerebella of 4-month-old Purkinje cell degeneration (pcd3J) mice, in which most Purkinje cells had already degenerated and wild-type littermates with a normal complement of Purkinje neurons. Of 14,563 probe sets expressed in wild-type cerebellum, 797 showed a significant (p<0.0001) reduction in pcd3J mice. These probes could represent transcripts with varying levels of specificity for Purkinje cells as well as transcripts in other cell types that decline as a secondary consequence of Purkinje cell loss. Ranking of the probe signals revealed that well-known Purkinje cell-specific transcripts such as calbindin and L7/pcp2 clustered in a group that was <33% of wild-type levels. Therefore, to identify potentially new Purkinje cell-specific transcripts that cluster with the known markers, more stringent selection criteria were applied (<33% of wild-type signal and p<0.0001). With these criteria, 55 independent transcripts were identified of which 33 were annotated genes and 22 were ESTs and RIKEN cDNAs. A literature search revealed that 25 of the 33 annotated genes were expressed in Purkinje cells, with no data being available on the other 8. Thus, the additional 8 annotated and 22 un-annotated genes are clustered with many genes expressed in Purkinje cells making them candidate markers. To confirm the microarray data, eight representative annotated genes were selected including five reported to be in Purkinje neurons and three for which no data was available. Semi-quantitative RT-PCR demonstrated reduced expression of all eight transcripts in cerebella from pcd3J mice. The promoters of genes expressed selectively in subsets of neurons can be used to direct heterologous gene expression in transgenic mice and the more restricted the expression pattern the greater their utility. Therefore, microarray analysis was used to assess expression levels of all 55 transcripts in cerebral cortex, striatum, substantia nigra and ventral tegmental area. This permitted the identification of a set of genes whose promoters might have utility for selectively targeting gene expression to cerebellar Purkinje cells. [Copyright &y& Elsevier]