BHT Blocks NF-jB activation and Ethanol-Induced Brain Damage Fulton Crews, Kimberly Nixon, Daniel Kim, James Joseph, Barbara Shukitt-Hale, Liya Qin, and Jian Zou Background: Binge ethanol administration causes corticolimbic brain damage that models alco- holic neurodegeneration. The mechanism of binge ethanol-induced degeneration is unknown, but is not simple glutamate–N-methyl-D-aspartate (NMDA) excitotoxicity. To test the hypothesis that oxidative stress and inflammation are mechanisms of binge ethanol-induced brain damage, we administered 4 antioxidants, e.g., butylated hydroxytoluene (BHT), ebselen (Eb), vitamin E (VE), and blueberry (BB) extract, during binge ethanol treatment and assessed various measures of neuro- degeneration. Methods: Adult Sprague–Dawley rats were treated with intragastric ethanol 3 times per day (8–12 g/ kg/d) alone or in combination with antioxidants or isocaloric diet for 4 days. Animals were killed, and brains were perfused and extracted for histochemical silver stain determination of brain damage, mark- ers of neurogenesis, or other immunohistochemistry. Some animals were used for determination of nuclear factor k B (NF-kB)–DNA binding by electrophoretic mobility shift assay (EMSA) or for reverse transcription–polymerase chain reaction (RT-PCR) of cyclooxygenase 2 (COX2). Results: Binge ethanol induced corticolimbic brain damage and reduced neurogenesis. Treatment with BHT reversed binge induced brain damage and blocked ethanol inhibition of neurogenesis in all regions studied. Interestingly, the other antioxidants studied, e.g., Eb, VE, and BB, did not protect against binge-induced brain damage. Binge ethanol treatment also caused microglia activation, increased NF-kB–DNA binding and COX2 expression. Butylated hydroxytoluene reduced binge- induced NF-kB–DNA binding and COX2 expression. Conclusions: Binge-induced brain damage and activation of NF-kB–DNA binding are blocked by BHT. These studies support a neuroinflammatory mechanism of binge ethanol-induced brain damage. Key Words: Antioxidants, Neurodegeneration, Neurogenesis, NF-kB, Neuroinflammation. O XIDATIVE STRESS AND neuroinflammation have been linked to a number of neurodegenerative diseases including alcoholism (Crews et al., 2005a). Alco- hol dependence affects over 14% of the United States population and is the second leading cause of dementia (Kessler et al., 1994), with 50% to 75% of sober, detoxified former alcoholics having cognitive impairments (Eckardt and Martin, 1986). Both magnetic resonance imaging (MRI) and postmortem studies of alcoholic brain have found both gray and white matter loss in corticolimbic regions including the hippocampus (Crews et al., 2005b). Although the occurrence of alcoholic dementia and neu- rodegeneration are well supported by multiple studies, the mechanisms of neurotoxicity are poorly understood and likely contribute to the severity of alcohol dependence (Bowden et al., 2001). Alcoholic neurodegeneration can be modeled in animals with a 4-day binge ethanol administration treatment that causes neuronal death in corticolimbic regions as well as long-lasting relearning impairments (Obernier et al., 2002a, 2002b). Some studies have indicated that chronic ethanol can increase sensitivity to N-methyl-D-aspartate (NMDA) neurotoxicity in neur- onal cultures (Chandler et al., 1993) and up-regulate NMDA receptor proteins and calcium responses (Hu and Ticku, 1995), suggesting that glutamate neurotoxicity during ethanol withdrawal induces degeneration through supersensitive NMDA receptors (Chandler et al., 1995; Lovinger, 1993). However, studies have not found that binge ethanol treatment increases NMDA receptor density (Rudolph and Crews, 1996), and multiple studies investi- gating the effects of glutamate antagonists on binge ethanol-induced neuronal cell death have failed to sup- port a role of NMDA receptor excitotoxicity in binge From the Bowles Center for Alcohol Studies, UNC Chapel Hill, Chapel Hill, North Carolina (FC, DK, LQ, JZ); the Department of Pharmaceutical Sciences, The University of Kentucky, Lexington, Kentucky (KN); and the Neuroscience Laboratory, Tufts University, Boston, Massachusetts (JJ, BS). Received for publication April 28, 2006; accepted July 31, 2006. This research is supported by Grants from NIAAA. Reprint requests: Dr. Fulton T., Bowles Center for Alcohol Studies, The University of North Carolina at Chapel Hill, 1021 Thurston Bowles Building, CB 7178 Chapel Hill, NC 27599-7178; Fax: 919-966-5679; E-mail: ftcrews@med.unc.edu Copyright r 2006 by the Research Society on Alcoholism. DOI: 10.1111/j.1530-0277.2006.00239.x Alcohol Clin Exp Res, Vol 30, No 11, 2006: pp 1938–1949 1938 ALCOHOLISM:CLINICAL AND EXPERIMENTAL RESEARCH Vol. 30, No. 11 November 2006