P60: H2S production in peripheral neurons is mediated by cysteine aminotransferase and mercaptopyruvate sulfurtransferase

Abstract

Hydrogen sulfide (H2S) is reported to be generated by cystathionine gamma-lyase (CSE), cystathionine beta-synthase (CBS) and mercaptopyruvate sulfurtransferase (MPST) coupled with cysteine aminotransferase (CAT) in mammalian cells. In this study, we investigated the relative contributions of these three pathways to H2S production in peripheral neurons, PC12 cells (rat pheochromocytoma-derived cell-lines) and the rat dorsal root ganglion (DRG) neurons. Expressions of H2S-producing enzymes and the amount of H2S production were examined by western blot analysis and methylene blue assay, respectively. In PC12 cells and the rat DRG, CBS, CAT and MPST, but not CSE, were expressed, and appreciable amounts of H2S were produced from L-cysteine in the presence of a co-substrate for CAT, alpha-ketoglutarate, together with a reducing agent, dithiothreitol. The production of H2S was inhibited by a CAT inhibitor (aminooxyacetate), competitive CAT substrates (L-aspartate and oxaloacetate), and RNA interference against MPST. Immunocytochemistry revealed a mitochondrial expression of MPST. H2S production by CAT/MPST was increased by alkalization, and was much higher at pH 8.0, a mitochondrial matrix pH, than at pH 7.5, a cytosolic pH. These results indicate that H2S production in peripheral neurons is primarily mediated by CAT and MPST, and that the production of H2S may be influenced by regulation of mitochondrial metabolism.