In vitro mitochondrial failure and oxidative stress mimic biochemical features of Alzheimer disease

• Published in: Neurochemistry international Vol. 63; no. 2; pp. 112 - 120
• Main Authors: Selvatici, Rita, Marani, Luca, Marino, Silvia, Siniscalchi, Anna
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 01.08.2013
• Subjects: Alzheimer Disease - metabolism; Alzheimer Disease - physiopathology; Animals; Antioxidants - pharmacology; BACE1; Glycogen Synthase Kinase 3 - metabolism; Glycogen Synthase Kinase 3 beta; GSK3; Hydrogen peroxide; In Vitro Techniques; Mitochondria - drug effects; Mitochondria - physiology; Neuroprotective Agents - pharmacology; Nitrogen - pharmacology; Oxidative Stress; p35/25; Phosphorylation; Rats; Rats, Sprague-Dawley; Sodium azide; Tau; tau Proteins - metabolism; BACE1; Tau; Hydrogen peroxide; Sodium azide; GSK3; p35/25
• ISSN: 0197-0186; 1872-9754
• DOI: 10.1016/j.neuint.2013.05.005

•Neuronal suffering similar to initial stage of AD was obtained by NaN3 and H2O2.•As in AD, tau phosphorylation and the protein level of p35/25 and BACE1 were raised.•The neurotoxic treatment depressed the acetylcholine release in rat hippocampal slices.•Antioxidant drugs and memantine prevented neurodegeneration induced by NaN3 and H2O2.•This protocol could be suitable to test the efficacy of neuroprotective treatments. Primary cortical neurons exposed to the mitochondrial toxin NaN3 (0.1–3mM) were submitted to oxidative stress with H2O2 (30–150μM), to mimic conditions observed in neurodegenerative disorders. The effects of such treatment on a series of parameters useful in characterizing neuronal damage were investigated: (i) the basal release of glutamate, evaluated as 3H-d-Aspartate efflux, was sharply, concentration-dependently, increased; (ii) the phosphorylation status of intracellular markers known to be involved in the neurodegenerative processes, in particular in Alzheimer disease: tau and GSK3β were increased, as well as the protein level of β-secretase (BACE1) and p35/25 evaluated by Western blotting, while (iii) the cell metabolic activity, measured with the MTT method, was reduced, in a concentration- and time-dependent manner. The latter effect, as well as tau hyperphosphorylation, was prevented both by a mixture of antioxidant drugs (100μM ascorbic acid, 10μM trolox, 100μM glutathione) and by the anti-Alzheimer drug, memantine, 20μM. Since it is well known that hippocampal cholinergic neurons are particularly affected in Alzheimer disease, the effects of NaN3 and H2O2 were also studied in electrically stimulated rat hippocampal slices, evaluating the 3H-Choline efflux, as an index of acetylcholine release. The neurotoxic treatment depressed the neurosecretory function and the mixture of antioxidant drugs, as well as memantine, were able to restore it. The neuronal damage induced by the in vitro protocol adopted in the present work displays peculiarities of neurodegenerative disorders, e.g. Alzheimer disease, underlining the role of mitochondrial failure and oxidative stress, which appear to occur upstream the neurodegenerative process; such protocol could be utilized to test the efficacy of neuroprotective treatments.