Acrolein, a product of lipid peroxidation, inhibits glucose and glutamate uptake in primary neuronal cultures

Abstract

Oxidative stress has been implicated in the pathogenesis of several neurodegenerative disorders including Alzheimer’s disease (AD). Increased lipid peroxidation, decreased levels of polyunsaturated fatty acids, and increased levels of 4-hydroxynonenal (HNE), F₂-isoprostanes, and F₄-neuroprostanes are present in the brain in patients with AD. Acrolein, an α,β-unsaturated aldehydic product of lipid peroxidation has been demonstrated to be approximately 100 times more reactive than HNE and is present in neurofibrillary tangles in the brain in AD. We recently demonstrated statistically significant elevated concentrations of extractable acrolein in the hippocampus/parahippocampal gyrus and amygdala in AD compared with age-matched control subjects. Concentrations of acrolein were two to five times those of HNE in the same samples. Treatment of hippocampal cultures with acrolein led to a time- and concentration-dependent decrease in cell survival as well as a concentration-dependent increase in intracellular calcium. In cortical neuron cultures, we now report that acrolein causes a concentration-dependent impairment of glutamate uptake and glucose transport in cortical neuron cultures. Treatment of cortical astrocyte cultures with acrolein led to the same pattern of impairment of glutamate uptake as observed in cortical neuron cultures. Collectively, these data demonstrate neurotoxicity mechanisms of arolein that might be important in the pathogenesis of neuron degeneration in AD.

Introduction

Increasing evidence supports the potential role of oxidative stress in neuronal degeneration in Alzheimer’s disease (AD). Recent studies show an increase in the redox active metal iron [1], [2], [3] as well as increased levels of lipid peroxidation, [4] a decline in membrane polyunsaturated fatty acids, [5], [6] and increased protein oxidation [7], [8] and DNA oxidation [9], [10], [11] in the brain in patients with AD. Other studies from our laboratory demonstrate increased levels of oxidatively modified DNA in AD ventricular cerebrospinal fluid (CSF) concomitant with decreased levels of the free repair product, [12] and increased levels of 4-hydroxynonenal (HNE), a neurotoxic marker of lipid peroxidation, in AD brain [13] and ventricular CSF [14]. Elevated levels of F₂-isoprostanes and F₄-neuroprostanes, markers of lipid peroxidation, are found in AD CSF, [15], [16] and increased levels of F₂-isoprostanes are present in the AD brain [17]. Markers of oxidative stress in neurofibrillary tangles (NFT) and senile plaques (SP) are present in the brain in AD [18], [19], [20], [21], [22], [23]. Peroxidation of lipids leads to the formation of a number of aldehydic by-products, including malondialdehyde, C₃-C₁₀ straight chain aldehydes, and α,β-unsaturated aldehydes such as HNE and acrolein [18], [24], [25].

Acrolein (CH₂CHCHO) occurs in the environment as a ubiquitous pollutant that is generated as a by-product of overheated organic materials. In vivo, acrolein is formed in the metal-catalyzed oxidation of polyunsaturated fatty acids, including arachidonic acid [26]. Acrolein is the strongest electrophile among the unsaturated aldehydes and shows the highest reactivity with nucleophiles, including sulfhydryl groups of cysteine, histidine, and lysine.[24] Acrolein formed in vivo, through iron-catalyzed oxidation of arachidonic and docosahexenoic acids, exhibits facile reactivity with various biomolecules, including proteins and phospholipids, has the potential to inhibit many enzymes, and quickly depletes cellular glutathione levels [27]. It is postulated that acrolein may inactivate the reductase responsible for the reduction of vitamin E radicals [27] and, coupled with the depletion of glutathione, leads to further lipid peroxidation. Acrolein is capable of modifying DNA bases with the formation of exocyclic adducts [28], [29]. Studies of Uchida et al. [26], [30] and Esterbauer et al. [24] demonstrated that acrolein is rapidly incorporated into proteins and generates carbonyl derivatives. More recently, Uchida et al. [26] demonstrated that acrolein preferentially reacts with lysine residues that are prominent components of tau, and Calingasan et al. [31] described the presence of acrolein adducts in NFT and dystrophic neurites surrounding SP. We recently demonstrated statistically significant elevations of extractable acrolein in AD amygdala (2.5 ± 0.9 nmol/mg of protein) compared with control amygdala (0.3 ± 0.5 nmol/mg of protein), and AD hippocampus and parahippocampal gyrus (5.0 ± 1.6 nmol/mg of protein) compared with age-matched control AD hippocampus and parahippocampal gyrus (0.7 ± 0.1 nmol/mg of protein) [32]. Our study demonstrated a significant time- and concentration-dependent decrease in cell survival in hippocampal neuron cultures treated with acrolein. Acrolein also led to a concentration-dependent increase in intracellular calcium concentration in hippocampal cultures.

The present report demonstrates that acrolein toxicity may be mediated through the inhibition of glucose transport and glutamate uptake.