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Polyamines are ubiquitous cations that are essential for cell growth, regeneration and differentiation. Increases in polyamine metabolism have been implicated in several neuropathological conditions, including excitotoxicity. However, the precise role of polyamines in neuronal degeneration is still unclear. To investigate mechanisms by which polyamines could contribute to excitotoxic neuronal death, the present study examined the role of the polyamine interconversion pathway in kainic acid (KA) neurotoxicity using organotypic hippocampal slice cultures. Treatment of cultures with N1,N(2)-bis(2,3-butadienyl)-1,4-butanediamine (MDL 72527), an irreversible inhibitor of polyamine oxidase, resulted in a partial but significant neuronal protection, especially in CA1 region. In addition, this pre-treatment also attenuated KA-induced increase in levels of lipid peroxidation, cytosolic cytochrome C release and glial cell activation. Furthermore, pre-treatment with a combination of cyclosporin A (an inhibitor of the mitochondrial permeability transition pore) and MDL 72527 resulted in an additive and almost total neuronal protection against KA toxicity, while the combination of MDL 72527 and EUK-134 (a synthetic catalase/superoxide dismutase mimetic) did not provide additive protection. These data strongly suggest that the polyamine interconversion pathway partially contributes to KA-induced neurodegeneration via the production of reactive oxygen species.

Free iron has been assumed to potentiate oxygen toxicity by generating reactive oxygen species (ROS) via the iron-catalyzed Haber-Weiss reaction, leading to oxidative stress. ROS-mediated iron cytotoxicity may trigger apoptotic cell death. In the present study, we used iron treatment of organotypic cultures of hippocampal slices to study potential mechanisms involved in iron-induced neuronal damage. Exposure of mature hippocampal slices to ferrous sulfate resulted in concentration- and time-dependent cell death. After iron treatment, markers of ROS formation and lipid peroxidation, i.e. intensity of dichlorofluorescein (DCF) fluorescence and levels of thiobarbiturate reactive substances (TBARS), were significantly increased. Levels of cytochrome c were increased while levels of pro-caspase-9 and pro-caspase-3 were decreased in cytosolic fractions of iron-treated hippocampal slice cultures. Treatment of cultured slices with a synthetic catalytic ROS scavenger, EUK-134, provided between 50 and 70% protection against various parameters of cell damage and markers of oxidative stress. In addition, inhibition of caspase-3 activity by Ac-DEVDcho partially protected cells from iron toxicity. The combination of EUK-134 and Ac-DEVDcho resulted in an almost complete blockade of iron-induced damage. These results indicate that iron elicits cellular damage predominantly by oxidative stress, and that ROS-mediated iron toxicity may involve cytochrome c- and caspase-3-dependent apoptotic pathways.

Free iron has been assumed to potentiate oxygen toxicity by generating reactive oxygen species (ROS) via the iron-catalyzed Haber-Weiss reaction, leading to oxidative stress. ROS-mediated iron cytotoxicity may trigger apoptotic cell death. In the present study, we used iron treatment of organotypic cultures of hippocampal slices to study potential mechanisms involved in iron-induced neuronal damage. Exposure of mature hippocampal slices to ferrous sulfate resulted in concentration- and time-dependent cell death. After iron treatment, markers of ROS formation and lipid peroxidation, i.e. intensity of dichlorofluorescein (DCF) fluorescence and levels of thiobarbiturate reactive substances (TBARS), were significantly increased. Levels of cytochrome c were increased while levels of pro-caspase-9 and pro-caspase-3 were decreased in cytosolic fractions of iron-treated hippocampal slice cultures. Treatment of cultured slices with a synthetic catalytic ROS scavenger, EUK-134, provided between 50 and 70% protection against various parameters of cell damage and markers of oxidative stress. In addition, inhibition of caspase-3 activity by Ac-DEVDcho partially protected cells from iron toxicity. The combination of EUK-134 and Ac-DEVDcho resulted in an almost complete blockade of iron-induced damage. These results indicate that iron elicits cellular damage predominantly by oxidative stress, and that ROS-mediated iron toxicity may involve cytochrome c- and caspase-3-dependent apoptotic pathways.