Gliotoxins are a group of amino acids that are toxic to astrocytes, and are substrates of high-affinity sodium-dependent glutamate transporters. In the present study, C6 glioma cells were preincubated for 20 h in the presence of 400 microM L-alpha-aminoadipate, L-serine-O-sulphate, D-aspartate or L-cysteate, as well as in the presence of the poorly transported L-glutamate uptake inhibitor, L-anti-endo-methanopyrrolidine dicarboxylate. In experiments following [3-13C]alanine metabolism, all toxins caused a decreased incorporation of label into glutamate. Production of labelled lactate changed only when cells were incubated in the presence of L-alpha-aminoadipate or L-serine-O-sulphate. Incubation with L-anti-endo-methanopyrrolidine dicarboxylate caused no change in the amount of label incorporated into either glutamate or lactate. When glutathione production was followed using 1 mM [2-13C]glycine, differential effects of the gliotoxins were revealed. Most notably, both L-serine-O-sulphate and L-alpha-aminoadipate caused significant increases in labelling of glutathione. Once again, L-anti-endo-methanopyrrolidine dicarboxylate was without effect. Overall, we have shown that the gliotoxins cause disruption to alanine metabolism and glutathione production in C6 glioma cells, but that there are notable differences in their mechanisms of action. In the absence of any disruption to metabolism by L-anti-endo-methanopyrrolidine dicarboxylate, it is concluded that their mode of action involves more than inhibition of glutamate transport.
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