Abstracts: Research Society on Alcoholism, 26th Annual Meeting, Ft. Lauderdale, Florida

(MUSC) Medical University at South Carolina

Gender Differences in Ethanol Intake Prior to and Following Repeated Chronic Ethanol Exposure and Withdrawal in C57BL/6J Mice

Lopez, MF and Becker, HC; Charleston Alcohol Research Center, Medical University of South Carolina and Department of Veterans Affairs, Charleston, SC 29425

The present study was design to assess possible sex-related differences in ethanol (EtOH) intake following repeated chronic EtOH exposure and withdrawal in mice. Adult male and female C57BL/6J mice were trained to drink EtOH using a sucrose-fading procedure in a limited access situation. Mice were allowed daily 2 hr access to EtOH vs. Water in their home cage starting at the beginning of the dark cycle. Mice were not food or water deprived at any time. Once stable daily baseline intake of the final EtOH solution (15% v/v without sucrose) was achieved, mice were separated into chronic EtOH exposure/withdrawal and control groups. The mice were exposed to four cycles of 16 hr EtOH vapor (or air for controls) followed by 8 hr periods of withdrawal. Starting at 32 hr after final withdrawal, all mice were tested for EtOH intake during five daily limited access sessions. Then, mice were exposed to a second set of four cycles of chronic EtOH (or air) exposure and withdrawal followed by five more days of limited access testing for EtOH intake. Analysis of the EtOH intake during baseline indicated that female mice consumed more EtOH than males (3.5 g/kg vs. 2.5 g/kg). Analysis of volume of EtOH intake, as well as the total fluid intake (EtOH+Water) during baseline indicated a higher consumption level in females. EtOH intake following chronic EtOH exposure/withdrawal was significantly increased above baseline levels in male and female mice. Both male and female mice showed heightened EtOH intake compared to the air-exposed controls. The magnitude of this increase in EtOH consumption was proportionally similar in both sexes. These results indicate that despite initial sex differences in EtOH intake, both male and female mice exhibit a similar increase in voluntary EtOH intake after repeated chronic EtOH exposure/withdrawal experience. Supported by NIAAA grants AA10761, AA13509, and VA Medical Research.

Stress Related to Repeated Chronic Ethanol Exposure and Withdrawal: Effects on Ethanol Intake in C57BL/6J Mice

Lopez, MF; Juarez, LE, Becker, HC; Charleston Alcohol Research Center, Medical University of South Carolina and the Department of Veterans Affairs, Charleston, SC 2942

The present study examined whether stress associated with repeated chronic EtOH exposure/withdrawal experience plays a role in influencing subsequent voluntary EtOH drinking behavior. Adult male C57BL/6J mice were trained to drink EtOH using a sucrose-fading procedure. Mice were allowed daily limited access (2 hr) to EtOH vs. Water in their home cage starting at the beginning of the dark cycle. Mice were not food or water deprived at any time. Once stable baseline intake of the final EtOH solution (15% v/v without sucrose) was achieved (~2.7 g/kg/day), mice were divided into chronic EtOH exposure/withdrawal and Control groups. Mice were exposed to four cycles of 16 hr EtOH vapor (or air for controls) separated by 8 hr periods of withdrawal. At various times after final withdrawal, blood was collected from separate groups of mice to determine plasma corticosterone (CORT) levels. Some mice were allowed 2 hr access to EtOH starting at 30 hr post-withdrawal. Results indicated that EtOH-treated mice had higher CORT levels than controls at 0, 8, 24, and 30 hr after the last (fourth) EtOH exposure. In mice given the opportunity to drink, EtOH intake was significantly greater than baseline levels for EtOH-treated mice (3.5 g/kg) but not controls (2.8 g/kg). Immediately after this limited access session (32 hr), plasma CORT levels decreased compared to levels just prior to the drinking session in EtOH-treated mice, while an opposite effect was observed for controls. These results suggest that: (a) repeated chronic EtOH exposure/withdrawal experience increases plasma CORT levels; (b) changes in stress levels may be related to heightened EtOH self-administration; and (c) the effects of voluntary EtOH intake on CORT levels are different in EtOH-dependent and non-dependent mice. Supported by NIAAA grants AA10761, AA13509, and VA Medical Research.

(UM) University of Memphis, Tennessee

Effects of Mild Footshock Stress on Anxiety and Ethanol Drinking in Three Mouse Strains

D.B. Matthews¹, A.L. Morrow², G. Mittleman¹, M Ferkin³ R.B. Berry¹, J.M. Silvers¹, C. Brittingham¹ & A. Esquivel^{1 1}Dept. of Psych., Univ. of Memphis, Memphis TN, 38152; ²Bowles Ctr. Alcohol Studies, UNC-Chapel Hill, Chapel Hill NC 27599; ³Dept. of Bio., Univ. of Memphis, Memphis TN, 38152

Stress has long been cited as an underlying factor leading to anxiety and ethanol consumption. In laboratory studies, acute stress has been shown to produce anxiety-like behaviors and increases in plasma corticosterone and brain allopregnanolone levels. Furthermore, acute stress can re-instate ethanol self-administration. However, the relationship between acute stress, anxiety and ethanol self-administration has yet to be completely elucidated. To further examine these relationships, the vocalization to a mild footshock stressor was determined in male and female C57BL/6J, A/J and DBA/2J mice. Following vocalization determination, mice received either 15 min of inescapable footshock (shock level yoked to vocalization level) or no shock prior to measures of anxiety (elevated plus maze, open field), biochemical measures (plasma corticosterone, brain allopregnanolone) or ethanol self-administration (two bottle choice). Preliminary analysis reveals that mild footshock stress produces stain and gender differences in measures of anxiety and allopregnanolone. Additional studies are determining the relationship between anxiogenic effect of footshock and ethanol self-administration. Supported by AA13509 (DBM), INIA Core (ALM).

(UNC-CH) University of North Carolina at Chapel Hill

Effects of Reduced Steroid Derivatives of 11-Deoxycortisol and Cortisol on GABA_A Receptor-Mediated Chloride Flux in Rat Cortical Synaptoneurosomes

AL Morrow

The tetrahydro-reduced derivatives of progesterone and deoxycorticosterone, allopregnanolone (ALLO) and tetrahydrodeoxycorticosterone (THDOC), are potent positive modulators of GABA_A receptors. ALLO and THDOC stimulate increased chloride flux through GABA_A receptors causing hyperpolarization of neurons. Because of this function, these steroids help to regulate the hypothalamic-pituitary-adrenal (HPA) axis and maintain normal brain homeostasis in rodents. Alterations in ALLO and THDOC levels have been demonstrated in several major psychiatric disorders including alcoholism and major depression. However, in humans, the major steroids that regulate the HPA axis are 11-deoxycortisol (11-DOC) and cortisol (CORT). We hypothesized that reductions of 11-DOC and CORT at C3 and C5 would generate steroids active at GABA_A receptors. We performed chloride flux assays in rat cortical synaptoneurosomes to determine if any C3 or C5-reduced derivatives of 11-DOC or CORT modulate GABAergic activity. 11-DOC and CORT did not alter basal chloride flux or potentiate muscimol-stimulated chloride flux, while THDOC displayed potent positive modulatory activity. Additionally, these steroids did not antagonize THDOC-stimulated chloride flux. Furthermore, 5α-dihydrocortisol, 3α,5α-reduced CORT, 3α,5α-reduced 11-DOC, and 3α,5β-reduced 11-DOC had no modulatory activity at GABA_A receptors. 3α,5β-reduced CORT and androsterone had weak dose-dependent antagonist properties on both THDOC and muscimol-stimulated chloride flux. These data suggest that cortisol derivatives do not possess GABAergic activity, and that changes in HPA axis activity are not regulated via this mechanism. (Supported by AA10564)

(UTHSC) University of Tennessee Health Science Center

Web-Based QTL Mapping and Genetic Correlation Analysis of Gene Expression and Alcohol-Related Traits in BXD Strains

¹Chesler, E.J.; ²Wang, J.; ¹Lu, L.; ¹Peirce, J.L.; ¹Qu, Y.; ²Manly, K.F.; ¹Williams, R.W. ¹Dept. of Anatomy & Neurobiology and Center for Genomics & Bioinformatics, Univ. of Tennessee Health Science Center, Memphis TN. ²Roswell Park Cancer Institute, Buffalo, NY.

BXD recombinant inbred (RI) mice are isogenic strains made by inbreeding progeny of an intercross between C57BL/6J and DBA/2J. Numerous alcohol-related phenotypes have been studies using 26 BXD RIs strains. We have quantified mRNA expression levels in these strains and integrated molecular, neuropharmacological, and behavioral traits into a single web-accessible interface called WebQTL (http://webqtl.roswellpark.org, Wang et al, 2003). WebQTL allows clients to publicly share new trait data and re-analyze existing data with improved high-density marker maps and a wealth of gene expression data. WebQTL supports marker regression, full genome interval mapping and composite analysis with facilities for bootstrapping and permutation analysis. Genetic correlation analysis, also incorporated in WebQTL, associates phenotypes in similar or identical genetic background to evaluate shared genetic mediation of traits. Gene expression was assayed in mouse forebrain samples using the Affymetrix U74Av2 mouse microarray with a minimum of three replicate arrays per strain each hybridized with pooled samples from three mice. Previously reported alcohol related BXD strain phenotypes were obtained from the literature and from personal communication with authors. The correlation of phenotypic values of gene expression and other traits in combination with transcriptome QTL analysis can be used to narrow down the many hundreds of genes in a quantitative trait locus (QTL) to a handful of promising candidates for further evaluation. The assembly of data for the study of shared trait mediation will cumulatively enhance the utility of these strains as a behavior genetic resource. Our computational tools and database structures will form the framework for expansion of the existing resources to include the LXS strain set developed specifically for alcoholism research.

Wang, J., Williams, R.W., Manly, K.F. (in press) WebQTL: Web based complex trait analysis. Our thanks to John Crabbe and John Belknap and colleagues for an extensive file of error-corrected alcohol traits.

Use of Consomic Strains as a Systematic, Whole Genome, High Throughput First Pass Screen for the Localization of Genes Relative to Alcohol-Related Phenotypes

¹D. Goldowitz, ²D. Matthews and ¹K.M. Hamre. ¹Dept. of Anat. & Neurobiol., Univ. Tenn. Health Sci. Center, Memphis, TN; ²Dept. of Psychol., Univ. of Memphis, Memphis, TN.

Initially a QTL is typically defined or confirmed in recombinant inbred (RI) strains such as the AXB series (A/J X C57BL/6J). A novel means to associate alcohol-related traits with genetic territory is the use of recently created lines of consomic mice, where each individual chromosome from a donor inbred strain (e.g., A/J) has been introgressed into a parental background (C57BL/6J) (Nadeau JH et al., Nat Genet 24:221-5, 2000). This present study evaluates several lines of consomic mice to assess how the A/J genome affects the phenotype of traits that have previously been mapped as QTLs. Consomic lines for Chr. 1, 2, 4, 9, 11, 14, and 16 mice are being tested for the following behavioral measures: two bottle choice test, handling induced convulsions (HIC), stress vocalization measures and plus maze activity. Our most recent analysis of vocalization threshold produced by a mild footshock stress revealed that C57BL/6J mice have significantly lower vocalization thresholds compared to A/J mice. Furthermore, consomic mice carrying one of the following A/J chromosomes 2, 4 or 16 have similar vocalization phenotype as the C57BL/6J parental strain (Two-way ANOVA, strain X gender; main effect of strain, F = 10.12, df[4,37], p < 0.0001; Dunnett post-hoc test, A/J only significantly different from C57BL/6J, q = 5.45, p < 0.01).

Our overall analysis is focused at three levels: Level 1 - traits mapped in AXB and BXA recombinant inbred mice to confirm previously mapped loci as a positive control (i.e., two-bottle choice; Gill et al., Mamm Gen 9:929-35,1998), Level 2 - QTLs mapped using other RI lines - to test the QTL in these consomic lines (i.e, HIC), and Level 3 - The analysis of traits that have not previously been mapped (i.e., vocalization).

This approach shows the promise of inquiring, one chromosome at a time, the genomic involvement in alcohol-related behaviors. Furthermore, these lines of mice provide the reagents for ENU-based, phenotype-driven screens to identify the quantitative trait genes that underlie QTLs. We thank Joe Nadeau for the easy availability of the consomic lines. Support: 1UO1AA13503 (DG, KMH ), AA13509 (DM).

Examination of the Role of the NR1 Subunit of the NMDA Receptor in Ethanol Related Phenotypes Using Chimeric Mice

¹K.M. Hamre, ¹A.J. Elberger, ²D. Matthews, and ¹X.P. Du. ¹Dept. of Anat. & Neurobiol., Univ. Tenn. Health Sci. Center, Memphis, TN; ²Dept. of Psychol., Univ. of Memphis, Memphis, TN.

The NR1 subunit of the NMDA receptor has been hypothesized to play a role in numerous ethanol related phenotypes and responses. However, the ability to assess this hypothesis is compromised by the fact that NR1 knock-out mice are neonatal lethal and die within 24 hours after birth. To circumvent this problem, experimental mouse chimeras were generated. The advantage of the use of chimeric mice is that each cell maintains its original genotype so that genotypically NR1-/- cells can be examined. Chimeric mice are generated by combining two 4-8 cell embryos. One embryo is the normal transgenic ROSA/ROSA embryo that expresses β-galactosidase (β-gal) in most cells in the body. The second embryo is an NR1 embryo generated by mating heterozygous parents. At the conclusion of behavioral testing, single cell PCR of β-gal negative liver cells is done to distinguish the homozygous knock-out chimeras from heterozygous or wild-type. All chimeric mice are tested behaviorally for handling induced convulsions (HIC) using standard protocols following a dose of 4.0 g/kg of ethanol. Mice are tested between 60 and 90 days of age. Initial behavioral results demonstrate that the baseline scores of the HIC do not differ among the groups but that a range of phenotypes is seen following the ethanol injection. Subsequent behavioral testing includes examination of ethanol preference using a standard two-bottle choice test. Histological examination of the first chimeric mice demonstrates that NR1-/-<-->ROSA/ROSA chimeric mice are able to be generated and to survive until adulthood. More importantly, examination of the brains of the chimeric mice demonstrates that genotypically NR1-/- neurons do colonize the brain and will thus contribute to any behavioral phenotype. Supported by: AA13511 (KMH), AA12163 (AJE) and AA13509 (DM).

Identification of Pedigrees with Abnormal Ethanol-Induced Locomotor Activation Using ENU Muatagenesis

¹K.M. Hamre, ¹S. Wilkinson, ¹D. Goldowitz, and ²D. Matthews. ¹Dept. of Anat. & Neurobiol., Univ. Tenn. Health Sci. Center, Memphis, TN; ²Dept. of Psychol., Univ. of Memphis, Memphis, TN.

The genetics of the individual has a significant impact on the degree of the ethanol-related response exhibited by that individual. However, the genes that are responsible for many of these responses are just beginning to be elucidated. One strategy for identifying mice that exhibit abnormal ethanol responses is through a regional mutagenesis approach. In this approach, single base pair mutations are made with the mutagen ethyl-n-nitrosourea (ENU). Importantly, our regional mutagenesis approach identifies the 30-40cM region of the genome that contains the mutation. Mutagenized adult mice were tested in a screen for a number of alcohol induced responses including ataxia as measured on a rotorod, alterations in core body temperature, the anxiolytic response as measured in the elevated plus maze, locomotor activation as measured in an activity chamber and ethanol metabolism gauged with/by blood alcohol concentrations. Additionally, mice are also tested for their ethanol preference using the two-bottle choice test. Two pedigrees have been identified that exhibit accentuated locomotor activation in the activity chamber following ethanol exposure. This abnormality is specific to the locomotor response as these pedigrees are normal in all other measures obtained in the ethanol screen. In both pedigrees, the mutation is located on the distal one-half of chromosome 15 beginning at approximately 43.4 cM demonstrating that there is at least one gene that is critical for ethanol-induced locomotor activation in this region of the genome. In this continuing effort, other pedigrees are being screened with mutations localized to one of the following: chromosome 7, 10, 14, or 19 or on the X chromosome. The progress of this mutagenesis effort can be followed by visiting the website "http://tnmouse.org". Supported by: 5 UO1 MH61971 to DG, KMH and DM.

(VU) Vanderbilt University School of Medicine

Electrophysiological Characterization of Bed Nucleus of the Stria Terminalis Neurons: Membrane Properties, Synaptic Transmission and Neuromodulation by Beta Adrenergic Receptors

R.E. Egli; D.G. Winder Vanderbilt University School of Medicine, Nashville, TN 37232

The bed nucleus of the stria terminalis (BNST) has recently been identified as a critical component of affective responses to substances of abuse. The BNST is also the site of one of the densest noradrenergic inputs in the brain, and it has been shown that this noradrenergic input plays a key role in mediating the BNST's role in substance abuse. To begin to address the mechanisms underlying the role of the BNST in substance abuse, we are utilizing intracellular and field recordings in an in vitro slice preparation of C57Bl/6J adult male mouse BNST. We sampled both dorsal and ventral subdivisions of the BNST because of the differential NE input to these two regions. We find that resting membrane potentials and input resistances are comparable across vBNST and dBNST, and a proportion of cells in both regions possess hyperpolarization activated (I_h) current as well as a low-threshold spike (likely mediated by T-type Ca²⁺ channel activity). Interestingly, a greater number of cells display I_h current in dBNST than in vBNST, and a greater number of cells in vBNST display the low-threshold spike than in dBNST. We have also begun to examine synaptic transmission in these cells and find that local afferent stimulation results in an EPSP as well as an IPSP. Based on the importance of noradrenergic transmission in the BNST in substance abuse, we have begun to explore the potential effects of noradrenergic transmission on BNST neurons. There does not appear to be a profound effect of β-adrenergic receptor activation on the basic membrane properties of cells in either dBNST or vBNST. We have however found that application of 1 μM isoproterenol (a β-adrenergic receptor agonist) in dBNST leads to a substantial enhancement of synaptic transmission. We intend to further characterize noradrenergic modulation of BNST neurons by studying the effects of application of norepinephrine as well as other noradrenergic agents.

NMDA-Receptor Dependent Long-Term Potentiation of Glutamatergic Synaptic Transmission in the Mouse Bed Nucleus of the Stria Terminalis

Carl Weitlauf, Regula E. Egli and Danny G. Winder

The Bed Nucleus of the Stria Terminalis (BNST) serves to relay information from several components of the limbic system and stress axis (such as the hippocampal formation, amygdala, and caudal medulla) to areas involved in both the stress response (i.e. the periventricular nucleus of the hypothalamus) as well as those involved in the reward response [i.e., the ventral tegmental area (VTA) and nucleus accumbens (NAc)]. This anatomy suggests that the BNST may be important in the relationship between stress and drug abuse. Indeed, data from behavioral studies support the role of the BNST in stress-induced reinstatement of drug seeking.

The pathological state of alcohol/drug addiction is believed to involve long-term changes in the physiology of key brain structures relevant to the rewarding effects of these drugs of addiction. One such example of a neuroadaptation, NMDA receptor dependent long-term potentiation (LTP), has been reported in the VTA and NAc, although this plasticity is very modest in these regions. We report here that robust LTP can be induced in the BNST with a simple high frequency stimulation (HFS) protocol. Furthermore, using the NMDA receptor antagonist AP5, we find an NMDA receptor-dependent component of this LTP. We are currently characterizing the molecular substrates underlying LTP in the BNST. The discovery of LTP in this brain region provides a plausible adaptable substrate for some of the long-term adaptations that underlie stress/alcohol interactions.

(WFUHS) Wake Forest University Health Sciences

Assessing Binge Drinking in Monkeys

Kathleen A. Grant. Wake Forest University School of Medicine, Medical Center Blvd

We have established a model of oral ethanol self-administration in monkeys that results in a high proportion of monkeys (approximately 35%) drinking 3.0-6.0 g/kg day (12-24 drink equivalent) and attaining blood ethanol concentrations between 100-400 mg/dl when measured on a weekly basis. An interesting pattern of behavior emerges in these heavy-drinking monkeys following 6 months of daily drinking where there is an increase in the variability of total ethanol consumed on a day-to-day basis (i.e., heavy drinking days followed by low drinking days). These "binges" of ethanol self-administration are defined as 1-3 days of increased ethanol intake (>20% above average) followed by 1-3 days of decreased ethanol intake (<20% below average). To address the issue of craving prior to and during a binge, we have analyzed patterns of intake that are predictive of a binge. Initial statistical analysis indicate that the monkeys are likely to have increased drinking episodes during the night, larger intakes during the morning, and increased volumes of intake during their first drinking bout of the next day. These patterns of intake suggest that ethanol self-administration becomes dysregulated in the heavy-drinking macaque monkey and may suggest a critical time for treatment intervention to prevent binge drinking and an escalation of ethanol-related problems.

Extensive Characterization of HPA Response to Social Housing in Monkeys

N. Maners, P. Pierre, S. Thornton, H. Green, L. Rogers, C. Craig , K.A. Grant. Wake Forest University School of Medicine, Winston-Salem, N.C. 27157-1083

Stress has been suggested as a risk factor for excessive alcohol consumption in humans and laboratory animals. The current study is designed to extensively examine the HPA response to social stress in monkeys prior to ethanol self-administration. Adult male cynomolgus monkeys (n=12) were characterized for HPA response following several acute pharmacological challenges, including dexamethasone, ACTH, CRF, naloxone (125 & 375 μg/kg) and ethanol (1.0 &1.5 g/kg), and blood samples were taken, depending on the challenge, to capture either a peak response or a time course (up to 6 samples over 2 hours). Monkeys were assessed twice, first after living individually for 6 months and second after living in a social group of 4 for 3 months. Average baseline cortisol values ranged from 4-8 μg/dl. Dexamethasone suppressed average cortisol and all other challenges increased average cortisol levels. Overall, social housing increased average and within-subject cortisol levels. Ranking of subjects for HPA activity was consistent across housing conditions, with correlations of 0.85-0.99 for each assay. Overall, the data show that HPA response, as characterized by a number of pharmacological challenges, is a stable between-subject measure in the macaque monkey. Thus, HPA response appears to have trait-like characteristics that can be used to assess socially-derived stress as a risk factor for excessive alcohol consumption.

Effects of Chronic Ethanol Ingestion on Rat NMDA Receptor Subunit Expression within Brain Regions Regulating Axniety

B.A. McCool & D.W. Floyd

Alcohol abuse and anxiety are intimately associated behaviors. For example, anxiety during withdrawal from chronic alcohol exposure is likely to contribute significantly to drug-seeking and relapse in alcoholics. We therefore hypothesize that brain regions regulating fear/anxiety behaviors are likely to be altered by chronic ethanol exposure. We have recently shown that chronic ethanol ingestion up-regulates NMDA-gated currents and enhances their ifenprodil sensitivity in isolated lateral/basolateral amygdala neurons. Since amygdala NMDA receptors play a significant role in the expression fear/anxiety, we hope to determine the mechanism for this functional adaptation and to investigate whether similar changes are apparent within additional brain regions that, along with the amygdala, form the 'anxiety circuit' regulating such behaviors. To this end, we have utilized real-time RT-PCR to measure NMDA receptor subunit mRNA expression. In our first series of experiments, total RNA was isolated from lateral/basolateral amygdala dissected from individual rats exposed to either a control liquid diet (n=5) or an alcohol-containing liquid diet (n = 6) for 10-12 days; and cDNAs synthesized using standard procedures. cDNAs were then subjected to the polymerase chain reaction using NR2 gene-specific primers and a fluorogenic detection system that utilizes the inherent 5'-3' exonuclease activity in Taq DNA polymerase. The production of PCR products was measured in real-time following each cycle; and, the cycle after which product was initially detected was used to provide a relative measure of gene expression as represented in the original cDNA population. We found that mRNA expression for NR2B, 2C, and 2D was not appreciably affected by chronic ethanol exposure. There was however a trend for decreased NR2A expression in the chronic ethanol animals, although this did not reach statistical significance. We can conclude that enhanced NMDA receptor function in the lateral/basolateral amygdala may not be related to significant alterations in NR2 subunit mRNA expression in this brain region. We are currently examining the effects of chronic alcohol on the relative expression of the remaining NMDA receptor subunit mRNAs. Regardless, real-time RT-PCR provides an excellent measure of gene expression on an individual-by-individual basis in samples derived from relatively small brain regions. Supported by AA13120.

Cerebellar Gene Expression in a Non-Human Primate Model of Chronic Alcohol Consumption

S.J. Walker; D.P. Friedman; J.T. Graves; J. B. Daunais; K.E. Vrana; K.A. Grant. Center for the Neurobehavioral Study of Alcohol and Integrative Neuroscience Initiative on Alcoholism Department of Physiology and Pharmacology, Wake Forest University, School of Medicine, Winston-Salem NC 27156

The most commonly acquired degenerative condition affecting the cerebellum is chronic alcoholism. While ataxia of stance and gait are the most frequently observed clinical signs of cerebellar dysfunction in alcoholics, recent reports of cerebellar neuropathology associated with certain psychiatric disorders also point to a role of the cerebellum in cognition and emotion. These findings suggest that alcohol-induced cerebellar pathology may affect a broader range of behaviors than previously thought. To explore the basis of alcohol-induced neuropathology, we profiled gene expression in cerebellar tissue in a non-human primate model of heavy ethanol consumption. Total RNA was prepared from cerebellar tissue obtained from two adult male cynomolgus monkeys with histories of heavy ethanol self-administration. These monkeys had been drinking ethanol ad libitum in daily 22 hour sessions for at least 12 months. Each had a lifetime total ethanol consumption in excess of 12,000 grams. For comparison, total RNA was also prepared from cerebellar tissue obtained from two alcohol naïve male cynomolgus monkeys. The RNA samples (4 total) were each labeled and used to query an individual Affymetrix U133A Human GeneChip® that contains features representing 22,000 well-annotated human genes. Data analysis revealed that all four samples gave hybridization signal (i.e. a "present" call) to between 27-30% of the genes on the chip. Fewer than 10 genes out of the more than 6000 overall scored as "present" were differentially expressed between the two control animals. When alcohol-consuming animals were compared to controls, however, there were more than 300 gene expression differences, representing both up- and down-regulation. A number of receptor genes were upregulated (e.g. GABAB, GABRA6 and GABRD). In addition, the gene for cerebellar degeneration-related (CDR) protein (autoantibodies against CDR have been identified in some subjects with paraneoplastic cerebellar degeneration) and the upregulation of several proteins known to be associated with Alzheimer disease (microtubule-associated protein tau (MAPT), and alpha & beta synuclein) were also upregulated. Increased levels of tau have been shown to be detrimental to neurons, and the synucleins are found in Alzheimer disease neurofibrillary lesions. Activation of these genes suggests mechanisms by which ethanol may damage the cerebellum. (Supported by grants: AA13510 & AA11997 [KAG, DPF, and SJW]).