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Alcohol abuse can result in significant alterations to the structure of the brain and ultimately to behavioral dysfunctions. Epidemiological studies have shown that alcoholism is closely associated with impaired memory and judgment. However, the degree of deficit (brain injury) depends on factors such as the age of onset, duration of heavy drinking, continuous versus periodic (binge) drinking and the typical amount consumed per session. In recent years, neuroinflammation has been proposed as one of the alcoholism-induced neuropathological mechanisms, since increased levels of microglial markers are observed in the brains of both post-mortem human alcoholics and various alcohol-treated animals, from newborn or adolescent rodents to adult rodents. Many studies have investigated how microglia modulate alcohol-induced behavioral changes such as cognitive deficits, abnormal locomotor activity, motor impairment and mood disturbance. Importantly, we try to characterize and compare the distinct features in different ethanol (EtOH)-induced neurodegenerative disease (NDD) models. Moreover, mounting evidence indicates that in response to certain environmental toxins, microglia can become over-activated under oxidative stress, releasing pro-inflammatory mediators that cause central nervous system (CNS) disease. The molecular mechanisms involve free radical formation and the release of pro-inflammatory cytokines that are detrimental to neighboring neurons and interfere with the molecules regulating cell-cell interactions. The identification and understanding of the cellular and molecular mechanisms of microglial activation are described, as well as multiple downstream targets, in different alcohol-treated animal models. This review might contribute to the development of treatments and/or therapeutic agents that can reduce or eliminate the deleterious effects of alcohol-induced NDD., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Microglia function as the primary immune effector cells in the brain and play a pivotal role in the neuroinflammatory processes which are critical component of neurodegenerative diseases. Alcohol abuse has been considered as one of the common reasons for neurodegeneration although the causative factors are poorly understood. Here, we investigated whether activated microglia were implicated in neurodegeneration and cognitive dysfunctions in adult rats after intermittent alcohol abuse. Rats were given orally a priming dose of 5 g/kg ethanol and then 3g/kg every 8h for 4 days, followed by a 3-day ethanol-withdrawal period. These 4 days of ethanol treatments were repeated four times intermittently to simulate the binge drinking of human alcoholics. Neurodegeneration and microglial activation were detected by Fluoro-Jade B staining, Golgi staining, immunohistochemistry and ELISA, respectively, while cognitive function was assessed by Morris water maze and novel object recognition. The results showed that microglial activation and inflammatory cytokine expression were obvious in the parietal association cortex, entorhinal cortex and hippocampus accompanied by neurodegeneration following ethanol treatment. Moreover, learning and memory abilities also declined following ethanol treatments. However, the hypertrophied microglia disappeared accompanied by the decrease of inflammatory cytokines levels on day 4, and ramified microglial proliferated significantly on day 14 after ethanol withdrawal, along with a recovery from neuronal damage and cognitive impairment. Thus, the present study indicated that activated microglia might be involved in neurodegeneration and cognitive dysfunctions induced by intermittent ethanol exposure, and neurotrophic microglia appear to have a contribution to the recovery during abstinence., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Previous studies have shown that ethanol induces oxidative DNA damage in human peripheral lymphocytes. In the present study, protective effect of resveratrol and ascorbic acid on ethanol-induced oxidative DNA damage in human peripheral lymphocytes in vitro were comparatively investigated. Pretreatments with resveratrol at 5, 25, and 50μM, which were in the concentration range of in vitro research, significantly inhibited ethanol-induced oxidative DNA damage in 24h, whereas ascorbic acid showed such DNA protective activity only in 1h. Further study showed that both compounds could directly scavenge hydroxyl radical produced during ethanol metabolism. Resveratrol significantly inhibited ethanol metabolism by regulating alcohol dehydrogenase 1B (ADH1B) and acetaldehyde dehydrogenase 2 (ALDH2) mRNA expressions. Moreover, resveratrol also activated the base excision repair (BER) system in mRNA and protein levels in DNA auto-repair process. However, ascorbic acid showed no effect on ethanol metabolic pathway and BER system. Thus, the present study provided the first evidence that even though both resveratrol and ascorbic acid are anti-oxidants, they possessed differential mechanisms of action in protection against ethanol-induced oxidative DNA damage in human peripheral lymphocytes., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Background: Ethanol abuse, especially binge drinking, can be toxic to human organs. However, there have been few studies on the genotoxicity induced by ethanol in human peripheral lymphocytes under binge drinking conditions. The purpose of this study was to investigate the oxidative DNA damage induced by ethanol in human peripheral lymphocytes in vitro and the possible mechanism associated with ethanol metabolism. The concentrations of ethanol investigated in this study were 50 and 100 mM, which are equal to the ethanol concentrations in blood after binge drinking. The maximum concentration we used was 150 mM although it is not the typical blood ethanol concentration seen during binge drinking, and most people may die at such a high concentration. The purpose of using this maximum concentration was to obtain more detailed evidence about the genotoxicity induced by ethanol. The DNA repair process was also studied., Methods: Peripheral lymphocytes were isolated from donors who were nonsmokers and not ethanol drinkers. Oxidative DNA damage, possible metabolic pathways of ethanol in human peripheral lymphocytes, and the repair system involved in the DNA auto-repair process were examined by comet assay, flow cytometry, time-of-flight mass spectrometry (TOF-MS), reverse transcription-polymerase chain reaction (RT-PCR), and western blotting., Results: The results showed that ethanol at the concentrations of 50, 100, and 150 mM significantly induced the oxidative DNA damage in human peripheral lymphocytes in vitro, which was accompanied by a parallel increase in the generation of 8-OHdG, intracellular hydroxyl radical, and reactive oxygen species (iROS). The DNA damage induced by ethanol could be attenuated by alcohol dehydrogenase 1B (ADH1B) or acetaldehyde dehydrogenase 2 (ALDH2) inhibitor, and the mRNA expression levels of ADH1B and ALDH2 were increased markedly by ethanol. The inhibitor of cytochrome P450 2E1 (CYP2E1) had no effect on ethanol-induced DNA damage, and CYP2E1 mRNA expression was not affected by ethanol. Furthermore, ethanol-induced DNA damage could be auto-repaired by lymphocytes. The expression of 8-oxoguanine DNA glycosylase 1 (OGG1) and the X-ray repair cross-complementation group 1 (XRCC1), 2 core enzymes in the base excision repair (BER) system, were increased in both of transcriptional and protein levels after ethanol treatment., Conclusions: This study provides direct evidence that ethanol can induce oxidative DNA damage in human peripheral lymphocytes in vitro, and its mechanism may be associated with the metabolism of ethanol by the ADH1B/ALDH2 pathway. Moreover, ethanol-induced DNA damage can be auto-repaired by human peripheral lymphocytes possibly mediated by the BER system., (Copyright © 2010 by the Research Society on Alcoholism.)

It is suggested that reactive oxygen species produced during ethanol intake may induce oxidative DNA damage. The present study, by the use of single cell gel electrophoresis (comet assay), investigated the potential genotoxicity of acute and long-term ethanol administration in mouse peripheral leucocytes. Urinary 8-hydroxy-2'-deoxyguanosine and total antioxidant capacity and reactive oxygen species in whole blood were also detected. Acute or long-term administration of ethanol, at the dose of 2.5 or 5.0 g/kg intraperitoneally, for 30 days, could induce significant DNA damage in peripheral leucocytes, which could be repaired at least 3 days after ethanol withdrawal in long-term ethanol treatment. A significant increase of urinary 8-hydroxy-2'-deoxyguanosine and generations of reactive oxygen species in whole blood after ethanol administration were observed, which demonstrated the ethanol-induced oxidative DNA damage. Interestingly, it was found that the total antioxidant capacity was significantly increased in whole blood after long-term ethanol administration compared to the control group, which suggested enhancement in the activities of the antioxidative defence system against oxidative tissue damage caused by excessive ethanol consumption. Thus, the present studies provide a clear evidence that ethanol induced DNA damage in peripheral leucocytes, which might result from ethanol-induced oxidative stress in the body.

The promyelocytic leukemia (PML) gene is a tumor suppressor gene associated with cell apoptosis, cell proliferation, and senescence. However, the role of PML in the ethanol-induced apoptosis is not fully-known. In this study, using wild-type mouse embryo fibroblasts (MEF) and PML null MEF cells, we found that (1) ethanol (100 mM and 200 mM) could obviously induce apoptosis of wild-type MEF cells, whereas, in PML null MEF cells, the pro-apoptotic function of ethanol was partially blocked; (2) the expression levels of phosphorylated p53 and two of its target genes, p21 and Bax, could be significantly up-regulated by ethanol (200 mM) in wild-type MEF cells in a time-dependent manner, but not in PML null MEF cells. These results indicate that PML plays an important role in ethanol-induced apoptosis, and p53-dependent apoptotic pathway may be involved in this process.

Ethanol is a principle ingredient of alcoholic beverages with potential neurotoxicity and carcinogenicity, and the ethanol-associated oxidative DNA damage in the central nervous system is well documented. The present work studied the possible protective effects of grape seed oligomer and polymer procyanidin fractions against ethanol-induced toxicity and compared these with resveratrol and other well-known antioxidants (ascorbic acid and vitamin E). By using the single cell gel electrophoresis (comet assay), a simple and sensitive technique for genotoxicity studies, the potential genotoxicity of acute and chronic ethanol administration in the different brain regions was investigated. Acute ethanol administration, at the dose of 2.5 or 5.0 g kg(-1) i.p., could induce significant DNA damage in cerebellum and hippocampus. Chronic administration of ethanol at the dose of 2.5 or 5.0 g kg-1 p.o. for 30 days could induce significant DNA damage in cerebellum, hippocampus, hypothalamus, and cortex, which could be auto-repaired at least 3 days after ethanol withdrawal. Oral administration of grape seed oligomer and polymer procyanidins and resveratrol (25, 50, and 100 mg kg(-1)) for 3 days before acute ethanol (5.0 g kg(-1), i.p.) or repeated administration of these substances together with ethanol (5.0 g kg(-1), p.o.) for 30 consecutive days could significantly inhibit DNA damage in brain cells induced by ethanol. As compared, ascorbic acid (50, 100, and 200 mg kg(-1)) and vitamin E (100, 200, and 400 mg kg(-1)) could also present protective effects on ethanol-induced DNA damage. Furthermore, the concentrations of ethanol and acetaldehyde in brain regions of the mice were detected by gas chromatography after administration of ethanol plus antioxidants. All of the results indicated that ethanol could induce region-specific oxidative DNA damage in which the cerebellum and hippocampus were more vulnerable, but intake of grape seed procyanidins or other natural antioxidants could protect the brain against ethanol-induced genotoxicity.

It is controversial regarding to the roles of acetaldehyde and ethanol in the central nervous system. In the present study, the effects of acetaldehyde and ethanol on extracellular levels of glutamate, taurine and GABA in the anterior cingulate cortex (ACC) of freely moving rats were investigated by using the microdialysis technique coupled to high performance liquid chromatography (HPLC) with fluorescent detection. The result showed that glutamate levels were significantly decreased after acute administration of acetaldehyde (AcH, 20 and 100 mg/kg, i.p.), while taurine levels were significantly increased after the higher dose of acetaldehyde (100 mg/kg, i.p.). GABA levels had no changes at any doses of acetaldehyde tested. Interestingly, similar changes of these amino acids were induced by ethanol (EtOH, 3 g/kg, i.p.) when sodium azide (NaN3, 10 mg/kg, i.p.), a catalase inhibitor that can reduce brain ethanol metabolism, was used simultaneously. These findings suggest that acetaldehyde and ethanol have the similar effects on the extracellular output of glutamate, taurine and GABA in the ACC.

Previous studies have shown that acute systemic administration of ethanol induced ascorbic acid (AA) release in mouse striatum and prefrontal cortex. Clozapine and olanzapine showed similar effects on ethanol-induced AA release in mouse striatum. However, their effects on ethanol-induced AA release in mouse prefrontal cortex have not been reported. Thus, their effects on this neurochemical event were further investigated in the present study. The results showed that ethanol (4.0 gkg i.p.) significantly stimulated AA release in the prefrontal cortex by about 200 of baseline in mice. Clozapine and olanzapine, at the dose of 1.0 mgkg s.c., had no effect on basal AA or ethanol-induced AA release. However, both drugs, at the dose of 10 mgkg s.c., significantly inhibited ethanol-induced AA release. The present study demonstrated for the first time that similar actions were exhibited by clozapine and olanzapine for the regulation of ethanol-induced AA release in the mouse prefrontal cortex.

Previous studies have shown that acute systemic administration of ethanol-induced striatal ascorbic acid (AA) release in mice and rats. In the present study, in vivo brain microdialysis coupled with high performance liquid chromatography (HPLC) with electrochemical detection (ECD) was used to comparatively evaluate the effects of clozapine on ethanol-induced AA release in mouse and rat striatum. The results showed that clozapine, at the dose of 15 mg/kg i.p., had no effect on basal AA or ethanol-induced AA release in rat striatum. The potentiating effect of clozapine on ethanol-induced striatal AA release was still observed in rats, at the higher dose of 30 mg/kg. In contrast, clozapine significantly inhibited ethanol-induced AA release in mouse striatum, at the dose of 15 and 30 mg/kg, without affecting basal AA release. The present study suggested that clozapine differentially regulated ethanol-induced AA release in the mouse and rat striatum.

The effects of clozapine, olanzapine and haloperidol on ethanol-induced striatal ascorbic acid (AA) release in mice were compared by using microdialysis coupled to high performance liquid chromatography with electrochemical detection. Ethanol (4.0 g/kg i.p.) significantly stimulated striatal AA release by about 200% of baseline in mice. Clozapine and olanzapine, two atypical neuroleptic drugs, at the dose of 1.0 mg/kg s.c., had no effect on basal AA or ethanol-induced AA release. However, both drugs, at the dose of 10 mg/kg s.c., significantly inhibited ethanol-induced AA release. In contrast, haloperidol, a typical neuroleptic drug, at the doses of 0.1-2.0 mg/kg, had no effect on both basal and ethanol-induced AA release. The present study demonstrated that similar actions were exhibited by clozapine and olanzapine, but not by haloperidol, for the regulation of ethanol-induced AA release in the mouse striatum.

The effect of exogenous ascorbic acid (AA) on acute ethanol-induced decrease in extracellular nitrites contents in striatum of freely moving mice was investigated by using microdialysis coupled to high performance liquid chromatography with electrochemical detection. The results showed that exogenous AA had no effect on basal extracellular levels of nitrites, but significantly increased striatal extracellular contents of AA in a dose-depended manner. Co-administration of AA with ethanol significantly antagonized the ethanol-induced decrease in extracellular nitrites contents, but did not affect ethanol-induced release of AA. These data provided the first evidence of an antagonistic action of AA on ethanol-induced decrease in extracellular nitrites contents in striatum of freely moving mice.

Abstract: The mechanism of ethanol, morphine, methamphetamine (MAP), and nicotine-induced ascorbic acid (AA) release in striatum, and nucleus accumbens (NAc) is not well understood. Our previous study showed that the glutamatergic system was involved in the addictive drug-induced AA release in NAc and striatum. Furthermore, frontal decortication eliminates drug-induced ascorbic acid release in the striatum but not in the NAc. In the present study, the roles of the hippocampus in drug-induced AA release in the striatum and NAc were studied by using microdialysis coupled to high performance liquid chromatography with electrochemical detection (HPLC-ECD). Ethanol (3.0 g/kg, i.p.), methamphetamine (3.0 mg/kg, i.p.), and nicotine (1.5 mg/kg, i.p.) significantly stimulated AA release in the striatum and NAc, respectively. Morphine (20 mg/kg, i.p.) significantly stimulated AA release in the striatum, but not in the NAc. After hippocampal lesion by kainic acid, AA release induced by ethanol, methamphetamine, and nicotine could be eliminated in NAc, but not in the striatum. These results suggest that the hippocampus might be a common and necessary area in addictive drug-induced AA release in the NAc, which also imply that different pathways might be involved in drug-induced AA release in the striatum and the NAc of the rats. [Copyright &y& Elsevier]