The Alzheimer's disease (AD) brain is characterized by plaques containing ␤-amyloid (A␤) protein surrounded by astrocytes and reactive microglia. Activation of microglia by A␤ initiates production of reactive oxygen species (ROS) by the plasmalemmal NADPH oxidase; the resultant oxidative stress is thought to contribute to neurodegeneration in AD. We have previously shown that A␤ upregulates a chloride current mediated by the chloride intracellular channel 1 (CLIC1) protein in microglia. We now demonstrate that A␤ promotes the acute translocation of CLIC1 from the cytosol to the plasma membrane of microglia, where it mediates a chloride conductance. Both the A␤ induced Cl Ϫ conductance and ROS generation were prevented by pharmacological inhibition of CLIC1, by replacement of chloride with impermeant anions, by an anti-CLIC1 antibody and by suppression of CLIC1 expression using siRNA. Thus, the CLIC1-mediated Cl Ϫ conductance is required for A␤-induced generation of neurotoxic ROS by microglia. Remarkably, CLIC1 activation is itself dependent on oxidation by ROS derived from the activated NADPH oxidase. We therefore propose that CLIC1 translocation from the cytosol to the plasma membrane, in response to redox modulation by NADPH oxidase-derived ROS, provides a feedforward mechanism that facilitates sustained microglial ROS generation by the NAPDH oxidase.

Edited by Adam SzewczykKeywords: Intracellular chloride channel 1 Microglia Reactive oxygen species Nicotinamide adenine dinucleotide phosphate oxidase Alzheimer disease Charge compensation a b s t r a c t Oxidative stress, characterized by overproduction of reactive oxygen species (ROS), is a major feature of several pathological states. Indeed, many cancers and neurodegenerative diseases are accompanied by altered redox balance, which results from dysregulation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. In this review, we consider the role of the intracellular chloride channel 1 (CLIC1) in microglial cells during oxidative stress. Following microglial activation, CLIC1 translocates from the cytosol to the plasma membrane where it promotes a chloride conductance. The resultant anionic current balances the excess charge extruded by the active NADPH oxidase, supporting the generation of superoxide by the enzyme. In this scenario, CLIC1 could be considered to act as both a second messenger and an executor. Ó 2010 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved. CLIC1 and oxidative stressReactive oxygen species (ROS), may act as compounds that impair cell and protein function, but they may also act as second messengers in cellular processes that involve changes in the cellular redox state, including migration, differentiation, and cell replication. Indeed, many proteins have redox-sensitive motifs, such as cystein residues and metal co-factors, that are altered by redox state. Kinases such as mitogen-activated protein kinase (MAPK), protein kinase C (PKC) and protein kinase B (PKB), are also activated by cell oxidation [1]. Cell homeostatic mechanisms establish a balance between ROS production and their removal by antioxidant systems. The overwhelming of antioxidant defences by ROS generation results in a condition of oxidative stress. Several pathological conditions are characterized by changes in cellular redox state, in particular chronic inflammatory states, oncologic conditions [2,3] and degenerative process [4][5][6].Alzheimer's disease (AD) is a progressive neurodegenerative disease that affects millions of people every year and is the main cause of dementia in the elderly for which an effective therapy is yet to be found. The AD brain is characterized by the presence of intraneuronal neurofibrillary tangles constituted by the hyperphosphorilated form of the cytoskeleton protein Tau and deposits of the amyloid beta (Ab) protein, also known as senile plaques [7]. Ab is the aberrant form of the transmembrane protein APP (amyloid precursor protein), resulting from the cleavage by gamma and beta secretases [8]. Although for decades the presence of amyloid plaques in the central nervous system (CNS) have been thought to be the main causative factor for neurodegeneration, recent studies propose that soluble oligomers can be more dangerous for neurons than the actual plaques [9]. Indeed, the neurological deficits in AD patients do not always correlat...

Glutathione (GSH) is one of the major antioxidants in the brain. GSH is secreted by astrocytes and this extracellular GSH is used by neurones to maintain and increase their intracellular GSH levels. For efficient GSH trafficking between astrocytes and neurones, GSH needs to be maintained in the reduced form. In model systems, GSH trafficking has been shown to be essential for neuroprotection against a variety of stress conditions. Previously we and others have shown that GSH and thiols are unstable in cell culture media and are easily oxidised. In the present study it is shown that nanomolar concentrations of copper (II) ions can cause decay of GSH in cell culture media. Increased free or redox active copper has been implicated in a variety of diseases and degradation of extracellular GSH is a possible mechanism by which it exerts its harmful effects. Rat astrocytes, a human astrocytoma cell line and astrocyte-conditioned media, in the absence of cells, are able to retard this copper-catalysed decay of GSH and maintain GSH in its reduced form. The protective effect of astrocytes appears to be a combination of copper removing and antioxidant mechanisms. The importance of these protective mechanisms is discussed with regards to neurodegenerative diseases.

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