Linking Binge Alcohol-Induced Neurodamage to Brain Edema and Potential Aquaporin-4 Upregulation: Evidence in Rat Organotypic Brain Slice Cultures and In Vivo

CoMentis Inc., Oklahoma City, Oklahoma, USA.
Journal of neurotrauma (Impact Factor: 3.71). 03/2009; 26(2):261-73. DOI: 10.1089/neu.2008.0682
Source: PubMed


Brain edema and derived oxidative stress potentially are critical events in the hippocampal-entorhinal cortical (HEC) neurodegeneration caused by binge alcohol (ethanol) intoxication and withdrawal in adult rats. Edema's role is based on findings that furosemide diuretic antagonizes binge alcohol-dependent brain overhydration and neurodamage in vivo and in rat organotypic HEC slice cultures. However, evidence that furosemide has significant antioxidant potential and knowledge that alcohol can cause oxidative stress through non-edemic pathways has placed edema's role in question. We therefore studied three other diuretics and a related non-diuretic that, according to our oxygen radical antioxidant capacity (ORAC) assays or the literature, possess minimal antioxidant potential. Acetazolamide (ATZ), a carbonic anhydrase inhibitor/diuretic with negligible ORAC effectiveness and, interestingly, an aquaporin-4 (AQP4) water channel inhibitor, prevented alcohol-dependent tissue edema and neurodegeneration in HEC slice cultures. Likewise, in binge alcohol-intoxicated rats, ATZ suppressed brain edema while inhibiting neurodegeneration. Torasemide, a loop diuretic lacking furosemide's ORAC capability, also prevented alcohol-induced neurodamage in HEC slice cultures. However, bumetanide (BUM), a diuretic blocker of Na(+)-K(+)-2Cl(-) channels, and L-644, 711, a nondiuretic anion channel inhibitor--both lacking antioxidant capabilities as well as reportedly ineffective against alcohol-dependent brain damage in vivo--reduced neither alcohol-induced neurotoxicity nor (with BUM) edema in HEC slices. Because an AQP4 blocker (ATZ) was neuroprotective, AQP4 expression in the HEC slices was examined and found to be elevated by binge alcohol. The results further indicate that binge ethanol-induced brain edema/swelling, potentially associated with AQP4 upregulation, may be important in consequent neurodegeneration that could derive from neuroinflammatory processes, for example, membrane arachidonic acid mobilization and associated oxidative stress.

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Available from: Michael A Collins,
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    • "Apoptosis as a chief neurodegenerative pathway was ruled out by the absence of TUNEL staining [10]. Evidence that diuretics such as furosemide and acetazolamide prevent ethanol-induced elevations in rat brain water content while reducing or blocking neurodegeneration indicated that brain edema was important [11], [12]. Cellular edema, a major and often intractable clinical problem that exacerbates neuropathology in brain ischemia and trauma, thus might elicit neuroinflammatory processes in binge ethanol abuse. "
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    ABSTRACT: Evidence that brain edema and aquaporin-4 (AQP4) water channels have roles in experimental binge ethanol-induced neurodegeneration has stimulated interest in swelling/edema-linked neuroinflammatory pathways leading to oxidative stress. We report here that neurotoxic binge ethanol exposure produces comparable significant effects in vivo and in vitro on adult rat brain levels of AQP4 as well as neuroinflammation-linked enzymes: key phospholipase A2 (PLA2) family members and poly (ADP-ribose) polymerase-1 (PARP-1). In adult male rats, repetitive ethanol intoxication (3 gavages/d for 4 d, ∼9 g/kg/d, achieving blood ethanol levels ∼375 mg/dl; "Majchrowicz" model) significantly increased AQP4, Ca+2-dependent PLA2 GIVA (cPLA2), phospho-cPLA2 GIVA (p-cPLA2), secretory PLA2 GIIA (sPLA2) and PARP-1 in regions incurring extensive neurodegeneration in this model-hippocampus, entorhinal cortex, and olfactory bulb-but not in two regions typically lacking neurodamage, frontal cortex and cerebellum. Also, ethanol reduced hippocampal Ca+2-independent PLA2 GVIA (iPLA2) levels and increased brain "oxidative stress footprints" (4-hydroxynonenal-adducted proteins). For in vitro studies, organotypic cultures of rat hippocampal-entorhinocortical slices of adult age (∼60 d) were ethanol-binged (100 mM or ∼450 mg/dl) for 4 d, which augments AQP4 and causes neurodegeneration (Collins et al. 2013). Reproducing the in vivo results, cPLA2, p-cPLA2, sPLA2 and PARP-1 were significantly elevated while iPLA2 was decreased. Furthermore, supplementation with docosahexaenoic acid (DHA; 22:6n-3), known to quell AQP4 and neurodegeneration in ethanol-treated slices, blocked PARP-1 and PLA2 changes while counteracting endogenous DHA reduction and increases in oxidative stress footprints (3-nitrotyrosinated proteins). Notably, the PARP-1 inhibitor PJ-34 suppressed binge ethanol-dependent neurodegeneration, indicating PARP upstream involvement. The results with corresponding models support involvement of AQP4- and PLA2-associated neuroinflammatory pro-oxidative pathways in the neurodamage, with potential regulation by PARP-1 as well. Furthermore, DHA emerges as an effective inhibitor of these binge ethanol-dependent neuroinflammatory pathways as well as associated neurodegeneration in adult-age brain.
    PLoS ONE 07/2014; 9(7):e101223. DOI:10.1371/journal.pone.0101223 · 3.23 Impact Factor
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    • "Early in cytotoxic edema, AQP4 facilitates edema fluid formation, but in vasogenic brain edema, AQP4 increases the rate of edema fluid elimination [6]. Sripathirathan et al. suggest binge ethanol-induced brain edema is potentially associated with AQP4 upregulation [7]. The expression of AQP4 after TBI is time-dependent, region-specific, and possibly implicated in the formation and resolution of TBI-induced cerebral edema [8]. "
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    ABSTRACT: The aim of this study is to assess the effects of alcohol on traumatic brain injury by using diffusion tensor imaging (DTI) and evaluate aquaporin-4(AQP4) expression changes in rat brainstems following acute alcohol intoxication with diffuse axonal injury (DAI). We further investigated the correlation between the AQP4 expression and DTI in the brain edema. Eighty-five rats were imaged before and after injury at various stages. DTI was used to measure brainstem apparent diffusion coefficient (ADC) and fractional anisotropy (FA), with immunostaining being used to determine AQP4 expression. After acute alcoholism with DAI, ADC values of the brainstem first decreased within 6 h and then elevated. FA values began to decline by 1 h, reaching a minimum at 24 h after trauma. There was a negative correlation between ADC values and brainstem AQP4 expression at 6 h and positive correlation at 6 h to 24 h. Changes of ADC and FA values in DAI with acute alcoholism indicate the effects of ethanol on brain edema and the severity of axonal injury. The correlations between ADC values and the brainstem AQP4 expression at different time points suggest that AQP4 expression follows an adaptative profile to the severity of brain edema.
    10/2013; 2013(1):798261. DOI:10.1155/2013/798261
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    ABSTRACT: Three aquaporins are expressed in the brain. AQP4, the predominant brain water channel, is expressed in astrocyte endfeet facing brain capillaries, perisynaptic spaces, and nodes of Ranvier. It is implicated in brain edema formation and resolution. It is also believed to assist clearance of K(+) released during neuronal activity. AQP1 is expressed in epithelial cells of choroid plexus and is implicated in cerebrospinal fluid formation. AQP9, which has been reported to be present in astrocytes and in subpopulations of neurons, is implicated in the brain energy metabolism. All three brain AQPs are strongly upregulated in brain tumors and in injured brain tissue. Water and solute transport via AQPs depends on concentration gradients across the membrane, but the magnitude of the transport is to a large extent determined by the single channel permeability of AQPs and by their abundance in the cell membrane. The future therapies will have to address not only the forces driving the water and solute transport (e.g. as mannitol infusion does in the treatment of brain edema), but also the regulation of AQPs, which provide the means for water entry to the brain, for water exit from the brain, and for redistribution of water and solutes within the brain compartments. This review summarizes the data concerning structure, permeability, role in the brain, short-term and long-term regulation of the three AQPs.
    Neurochemistry International 04/2010; 57(4):468-88. DOI:10.1016/j.neuint.2010.03.022 · 3.09 Impact Factor
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