1-3-dihydroxy-4-4-5-5-tetramethyl-2-(4-carboxyphenyl)tetrahydroimidazole and Nerve-Degeneration

Nitric oxide has been investigated widely both during neurodevelopment and in neurological diseases. However, whilst it has been established that nitric oxide-producing enzymes of nitric oxide synthase family are expressed in cerebellar Purkinje neurons, the effects of nitric oxide on the viability and morphology of these neurons remain unknown. Here, we have demonstrated that the activity of neuronal nitric oxide synthase, but not the inducible or endothelial forms of this enzyme, is required to support the survival of a proportion of cerebellar Purkinje neurons in vitro. We discovered that donation of high concentrations of exogenous nitric oxide reduces Purkinje neuron survival in culture and that peroxynitrite is also toxic to these cells. Finally, we demonstrated that exogenous nitric oxide and peroxynitrite reduce both the magnitude and the complexity of the neurite arbour extended by cerebellar Purkinje neurons. Taken together, these findings reveal that whilst a low level of endogenous nitric oxide, released by the activity of neuronal nitric oxide synthase, is beneficial to cerebellar Purkinje neurons in vitro, high levels of exogenous nitric oxide and peroxynitrite are detrimental to both the survival of these neurons and to their ability to extend processes and form functional neural networks.

Topics: Animals; Benzoates; Cell Count; Cells, Cultured; Cerebellar Cortex; Citrulline; Female; Imidazoles; Isothiuronium; Mice; Mice, Inbred C57BL; Nerve Degeneration; Nerve Tissue Proteins; Neurites; Nitric Oxide; Nitric Oxide Donors; Nitric Oxide Synthase; Nitroso Compounds; Peroxynitrous Acid; Pregnancy; Purkinje Cells; Reactive Nitrogen Species; Thiourea

Nitric oxide (NO) has been implicated as a potential contributor to neural cell death in a variety of neurological conditions. Cultured glial cells were exposed to extracellular superoxide generated by the action of xanthine oxidase on xanthine. In this experimental paradigm, both C6 glioma cells and primary astrocytes from rat cerebral cortex produced a rapid release of nitric oxide, measured using an NO specific electrode, in response to the applied superoxide stimulus. Application of a superoxide scavenger, or over-expression of Cu/Zn superoxide dismutase decreased the observed NO release. Authenticity of the NO signal was confirmed by the addition of the NO scavenger 2-(carboxyphenyl)-4,4,5,5-tetramethyllimidazoline-1-oxyl 3-oxide (carboxy-PTIO), which abolished the observed NO release without affecting simultaneously measured superoxide. Therefore, we suggest that glial cells may produce NO under free radical stimulation, which may be relevant to several neurological disorders where superoxide radicals are generated in the vicinity of glia. This would be predicted to result in the release of NO, which may exert toxic effects on neighbouring cells.

Topics: Animals; Astrocytes; Benzoates; Extracellular Space; Free Radical Scavengers; Gene Expression Regulation; Genetic Vectors; Imidazoles; Metalloporphyrins; Nerve Degeneration; Neurodegenerative Diseases; Neurons; Nitric Oxide; Nitric Oxide Synthase; Oxidative Stress; Rats; RNA, Messenger; Superoxide Dismutase; Superoxide Dismutase-1; Superoxides; Transfection; Tumor Cells, Cultured; Xanthine; Xanthine Oxidase