Ample research indicates that age-related neuronal-behavioral decrements are the result of oxidative stress that may be ameliorated by antioxidants. Our previous study had shown that rats given dietary supplements of fruit and vegetable extracts with high antioxidant activity for 8 months beginning at 6 months of age retarded age-related declines in neuronal and cognitive function. The present study showed that such supplements (strawberry, spinach, or blueberry at 14.8, 9.1, or 18.6 gm of dried aqueous extract per kilogram of diet, respectively) fed for 8 weeks to 19-month-old Fischer 344 rats were also effective in reversing age-related deficits in several neuronal and behavioral parameters including: oxotremorine enhancement of K(+)-evoked release of dopamine from striatal slices, carbachol-stimulated GTPase activity, striatal Ca(45) buffering in striatal synaptosomes, motor behavioral performance on the rod walking and accelerod tasks, and Morris water maze performance. These findings suggest that, in addition to their known beneficial effects on cancer and heart disease, phytochemicals present in antioxidant-rich foods may be beneficial in reversing the course of neuronal and behavioral aging.

https://www.jneurosci.org/content/jneuro/19/18/8114.full.pdf

Reversals of Age-Related Declines in Neuronal Signal Transduction, Cognitive, and Motor Behavioral Deficits with Blueberry, Spinach, or Strawberry Dietary Supplementation

Oxidative stress results from a cell or tissue failing to detoxify the free radicals that are produced during metabolic activity. Diabetes is characterized by chronic hyperglycemia that produces dysregulation of cellular metabolism. This review explores the concept that diabetes overloads glucose metabolic pathways, resulting in excess free radical production and oxidative stress. Evidence is presented to support the idea that both chronic and acute hyperglycemia cause oxidative stress in the peripheral nervous system that can promote the development of diabetic neuropathy. Proteins that are damaged by oxidative stress have decreased biological activity leading to loss of energy metabolism, cell signaling, transport, and, ultimately, to cell death. Examination of the data from animal and cell culture models of diabetes, as well as clinical trials of antioxidants, strongly implicates hyperglycemia-induced oxidative stress in diabetic neuropathy. We conclude that striving for superior antioxidative therapies remains essential for the prevention of neuropathy in diabetic patients.

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Oxidative stress in the pathogenesis of diabetic neuropathy.

Using the fluorescent dye 2′,7′-dichlorodihydrofluorescein (DCF-H2) we investigated the role of glutamate in the production of reactive oxygen species (ROS) in cultured neurons from fetal rat forebrain. The addition of an excitotoxic concentration of glutamate (100 microM) produced a generalized decrease in cellular DCF fluorescence accompanied by local areas of increased fluorescence around the margins of the cell body that could be observed within 2–4 min of glutamate exposure. Increases in fluorescence were dependent on NMDA receptor activation and Ca2+ entry and were blocked by the mitochondrial proton ionophore carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP). Additional studies suggested that the generalized decrease in fluorescence was due to intracellular acidification. These studies suggest a critical role for mitochondria in the production of ROS in association with glutamate excitotoxicity, and additionally demonstrate the feasibility of measuring the production of ROS at the level of the single cell.

https://www.jneurosci.org/content/jneuro/15/5/3318.full.pdf

Glutamate induces the production of reactive oxygen species in cultured forebrain neurons following NMDA receptor activation

Dichlorodihydrofluorescein and dihydrorhodamine 123 are sensitive indicators of peroxynitrite in vitro: implications for intracellular measurement of reactive nitrogen and oxygen species.

Exposure of cultured rat hippocampal neurons to glutamate resulted in accumulation of cellular peroxides (measured using the dye 2,7-dichlorofluorescein). Peroxide accumulation was prevented by an N-methyl-D-aspartate (NMDA) receptor antagonist and by removal of extracellular Ca2+, indicating the involvement of NMDA receptor-induced Ca2+ influx in peroxide accumulation. Glutamate-induced reactive oxygen species contributed to loss of Ca2+ homeostasis and excitotoxic injury because antioxidants (vitamin E, propyl gallate, and N-tert-butyl-alpha-phenylnitrone) suppressed glutamate-induced elevation of intracellular Ca2+ concentration ([Ca2+]i) and cell death. Basic fibroblast growth factor (bFGF), nerve growth factor (NGF), and brain-derived neurotrophic factor (BDNF), but not ciliary neurotrophic factor, each suppressed accumulation of peroxides induced by glutamate and protected neurons against excitotoxicity. bFGF, NGF, and BDNF each increased (to varying degrees) activity levels of superoxide dismutases and glutathione reductase. NGF increased catalase activity, and BDNF increased glutathione peroxidase activity. The ability of the neurotrophic factors to suppress glutamate toxicity and glutamate-induced peroxide accumulation was attenuated by the tyrosine kinase inhibitor genistein, indicating the requirement for tyrosine phosphorylation in the neuro-protective signal transduction mechanism. The data suggest that glutamate toxicity involves peroxide production, which contributes to loss of Ca2+ homeostasis, and that induction of antioxidant defense systems is a mechanism underlying the [Ca2+]i-stabilizing and excitoprotective actions of neurotrophic factors.

Neurotrophic factors attenuate glutamate-induced accumulation of peroxides, elevation of intracellular Ca2+ concentration, and neurotoxicity and increase antioxidant enzyme activities in hippocampal neurons.

Characterization of 2',7'-dichlorofluorescin fluorescence in dissociated mammalian brain neurons: estimation on intracellular content of hydrogen peroxide.

Toshiko Ueha

Papers

Characterization of 2',7'-dichlorofluorescin fluorescence in dissociated mammalian brain neurons: estimation on intracellular content of hydrogen peroxide.

Ginkgo biloba extract protects brain neurons against oxidative stress induced by hydrogen peroxide

Ca2+-Induced Increase in Oxidative Metabolism of Dissociated Mammalian Brain Neurons. Effect of Extract of Ginkgo Biloba Leaves.

Flow cytometric estimation of the effect of Ginkgo biloba extract on the content of hydrogen peroxide in dissociated mammalian brain neurons.

Change in Membrane Permeability Induced by Amyloid β-Protein Fragment 25 - 35 in Brain Neurons Dissociated from Rats