Obesity, Diabetes, Coffee, Tea, and Cannabis Use Alter Risk for Alcohol-Related Cirrhosis in 2 Large Cohorts of High-Risk Drinkers.

INTRODUCTION: Sustained high alcohol intake is necessary but not sufficient to produce alcohol-related cirrhosis. Identification of risk factors, apart from lifetime alcohol exposure, would assist in discovery of mechanisms and prediction of risk. METHODS: We conducted a multicenter case-control study (GenomALC) comparing 1,293 cases (with alcohol-related cirrhosis, 75.6% male) and 754 controls (with equivalent alcohol exposure but no evidence of liver disease, 73.6% male). Information confirming or excluding cirrhosis, and on alcohol intake and other potential risk factors, was obtained from clinical records and by interview. Case-control differences in risk factors discovered in the GenomALC participants were validated using similar data from 407 cases and 6,573 controls from UK Biobank. RESULTS: The GenomALC case and control groups reported similar lifetime alcohol intake (1,374 vs 1,412 kg). Cases had a higher prevalence of diabetes (20.5% (262/1,288) vs 6.5% (48/734), P = 2.27 × 10-18) and higher premorbid body mass index (26.37 ± 0.16 kg/m2) than controls (24.44 ± 0.18 kg/m2, P = 5.77 × 10-15). Controls were significantly more likely to have been wine drinkers, coffee drinkers, smokers, and cannabis users than cases. Cases reported a higher proportion of parents who died of liver disease than controls (odds ratio 2.25 95% confidence interval 1.55-3.26). Data from UK Biobank confirmed these findings for diabetes, body mass index, proportion of alcohol as wine, and coffee consumption. DISCUSSION: If these relationships are causal, measures such as weight loss, intensive treatment of diabetes or prediabetic states, and coffee consumption should reduce the risk of alcohol-related cirrhosis.

Genome-wide Association Study and Meta-analysis on Alcohol-Associated Liver Cirrhosis Identifies Genetic Risk Factors.

BACKGROUND AND AIMS: Only a minority of heavy drinkers progress to alcohol-associated cirrhosis (ALC). The aim of this study was to identify common genetic variants that underlie risk for ALC. APPROACH AND RESULTS: We analyzed data from 1,128 subjects of European ancestry with ALC and 614 heavy-drinking subjects without known liver disease from Australia, the United States, the United Kingdom, and three countries in Europe. A genome-wide association study (GWAS) was performed, adjusting for principal components and clinical covariates (alcohol use, age, sex, body mass index, and diabetes). We validated our GWAS findings using UK Biobank. We then performed a meta-analysis combining data from our study, the UK Biobank, and a previously published GWAS. Our GWAS found genome-wide significant risk association of rs738409 in patatin-like phospholipase domain containing 3 (PNPLA3) (odds ratio [OR] = 2.19 [G allele], P = 4.93 × 10-17 ) and rs4607179 near HSD17B13 (OR = 0.57 [C allele], P = 1.09 × 10-10 ) with ALC. Conditional analysis accounting for the PNPLA3 and HSD17B13 loci identified a protective association at rs374702773 in Fas-associated factor family member 2 (FAF2) (OR = 0.61 [del(T) allele], P = 2.56 × 10-8 ) for ALC. This association was replicated in the UK Biobank using conditional analysis (OR = 0.79, P = 0.001). Meta-analysis (without conditioning) confirmed genome-wide significance for the identified FAF2 locus as well as PNPLA3 and HSD17B13. Two other previously known loci (SERPINA1 and SUGP1/TM6SF2) were also genome-wide significant in the meta-analysis. GeneOntology pathway analysis identified lipid droplets as the target for several identified genes. In conclusion, our GWAS identified a locus at FAF2 associated with reduced risk of ALC among heavy drinkers. Like the PNPLA3 and HSD17B13 gene products, the FAF2 product has been localized to fat droplets in hepatocytes. CONCLUSIONS: Our genetic findings implicate lipid droplets in the biological pathway(s) underlying ALC.

CIS-Acting Allele-Specific Expression Differences Induced by Alcohol and Impacted by Sex as Well as Parental Genotype of Origin.

BACKGROUND: Alcohol use disorders (AUDs) are influenced by complex interactions between the genetics of the individual and their environment. We have previously identified hundreds of polygenic genetic variants between the selectively bred high- and low-alcohol drinking (HAD and LAD) rat lines. Here, we report allele-specific expression (ASE) differences, between the HAD2 and LAD2 rat lines. METHODS: The HAD2 and LAD2 rats, which have been sequenced, were reciprocally crossed to generate 10 litters of F1 progeny. For 5 of these litters, the sire was HAD2, and for the other 5 litters, the sire was a LAD2. From these 10 litters, 2 males and 2 females were picked from each F1 litter (N = 40 total). The F1 pups were divided, balancing for sex and direction of cross, into an alcohol (15%) versus a water control group. Alcohol drinking started in the middle of adolescence (~postnatal day 35) and lasted 9 weeks. At the end of these treatments, rats were euthanized, the nucleus accumbens was dissected, and RNA was processed for RNA-sequencing and ASE analyses. RESULTS: Analyses revealed that adolescent ethanol (EtOH) drinking, individual EtOH drinking levels, parentage, and sex-of-animal affected ASEs of about 300 genes. The identified genes included those associated with EtOH metabolism (e.g., Aldh2); neuromodulatory function (e.g., Cckbr, Slc6a7, and Slc1a1); ion channel activity (e.g., Kcnc3); and other synaptic and epigenetic functions. CONCLUSIONS: These data indicate that EtOH drinking differentially amplified paternal versus maternal allelic contribution to the transcriptome. We hypothesize that this was due, at least in part, to EtOH-induced changes in cis-regulation of polymorphisms previously identified between the HAD2 and LAD2 rat lines. This report highlights the complexity of gene-by-environment interactions mediating a genetic predisposition for, and/or the active development of, AUDs.

Evaluation of laboratory tests for cirrhosis and for alcohol use, in the context of alcoholic cirrhosis.

Laboratory tests can play an important role in assessment of alcoholic patients, including for evaluation of liver damage and as markers of alcohol intake. Evidence on test performance should lead to better selection of appropriate tests and improved interpretation of results. We compared laboratory test results from 1578 patients between cases (with alcoholic cirrhosis; 753 men, 243 women) and controls (with equivalent lifetime alcohol intake but no liver disease; 439 men, 143 women). Comparisons were also made between 631 cases who had reportedly been abstinent from alcohol for over 60 days and 364 who had not. ROC curve analysis was used to estimate and compare tests' ability to distinguish patients with and without cirrhosis, and abstinent and drinking cases. The best tests for presence of cirrhosis were INR and bilirubin, with areas under the ROC curve (AUCs) of 0.91 ± 0.01 and 0.88 ± 0.01, respectively. Confining analysis to patients with no current or previous ascites gave AUCs of 0.88 ± 0.01 for INR and 0.85 ± 0.01 for bilirubin. GGT and AST showed discrimination between abstinence and recent drinking in patients with cirrhosis, including those without ascites, when appropriate (and for GGT, sex-specific) limits were used. For AST, a cut-off limit of 85 units/L gave 90% specificity and 37% sensitivity. For GGT, cut-off limits of 288 units/L in men and 138 units/L in women gave 90% specificity for both and 40% sensitivity in men, 63% sensitivity in women. INR and bilirubin show the best separation between patients with alcoholic cirrhosis (with or without ascites) and control patients with similar lifetime alcohol exposure. Although AST and GGT are substantially increased by liver disease, they can give useful information on recent alcohol intake in patients with alcoholic cirrhosis when appropriate cut-off limits are used.

High Resolution Genomic Scans Reveal Genetic Architecture Controlling Alcohol Preference in Bidirectionally Selected Rat Model.

Investigations on the influence of nature vs. nurture on Alcoholism (Alcohol Use Disorder) in human have yet to provide a clear view on potential genomic etiologies. To address this issue, we sequenced a replicated animal model system bidirectionally-selected for alcohol preference (AP). This model is uniquely suited to map genetic effects with high reproducibility, and resolution. The origin of the rat lines (an 8-way cross) resulted in small haplotype blocks (HB) with a corresponding high level of resolution. We sequenced DNAs from 40 samples (10 per line of each replicate) to determine allele frequencies and HB. We achieved ~46X coverage per line and replicate. Excessive differentiation in the genomic architecture between lines, across replicates, termed signatures of selection (SS), were classified according to gene and region. We identified SS in 930 genes associated with AP. The majority (50%) of the SS were confined to single gene regions, the greatest numbers of which were in promoters (284) and intronic regions (169) with the least in exon's (4), suggesting that differences in AP were primarily due to alterations in regulatory regions. We confirmed previously identified genes and found many new genes associated with AP. Of those newly identified genes, several demonstrated neuronal function involved in synaptic memory and reward behavior, e.g. ion channels (Kcnf1, Kcnn3, Scn5a), excitatory receptors (Grin2a, Gria3, Grip1), neurotransmitters (Pomc), and synapses (Snap29). This study not only reveals the polygenic architecture of AP, but also emphasizes the importance of regulatory elements, consistent with other complex traits.

Npy deletion in an alcohol non-preferring rat model elicits differential effects on alcohol consumption and body weight.

Neuropeptide Y (NPY) is widely expressed in the central nervous system and influences many physiological processes. It is located within the rat quantitative trait locus (QTL) for alcohol preference on chromosome 4. Alcohol-nonpreferring (NP) rats consume very little alcohol, but have significantly higher NPY expression in the brain than alcohol-preferring (P) rats. We capitalized on this phenotypic difference by creating an Npy knockout (KO) rat using the inbred NP background to evaluate NPY effects on alcohol consumption. Zinc finger nuclease (ZNF) technology was applied, resulting in a 26-bp deletion in the Npy gene. RT-PCR, Western blotting and immunohistochemistry confirmed the absence of Npy mRNA and protein in KO rats. Alcohol consumption was increased in Npy(+/-) but not Npy(-/-) rats, while Npy(-/-) rats displayed significantly lower body weight when compared to Npy(+/+) rats. In whole brain tissue, expression levels of Npy-related and other alcohol-associated genes, Npy1r, Npy2r, Npy5r, Agrp, Mc3r, Mc4r, Crh and Crh1r, were significantly greater in Npy(-/-) rats, whereas Pomc and Crhr2 expressions were highest in Npy(+/-) rats. These findings suggest that the NPY-system works in close coordination with the melanocortin (MC) and corticotropin-releasing hormone (CRH) systems to modulate alcohol intake and body weight.

Brief report: genetics of alcoholic cirrhosis-GenomALC multinational study.

BACKGROUND: The risk of alcohol-related liver cirrhosis increases with increasing alcohol consumption, but many people with very high intake escape from liver disease. We postulate that susceptibility to alcoholic cirrhosis has a complex genetic component and propose that this can be dissected through a large and sufficiently powered genomewide association study (GWAS). METHODS: The GenomALC Consortium comprises researchers from Australia, France, Germany, Switzerland, United Kingdom, and United States, with a joint aim of exploring the genetic and genomic basis of alcoholic cirrhosis. For this National Institutes of Health/National Institute on Alcohol Abuse and Alcoholism funded study, we are recruiting high-risk drinkers who are either cases (with alcoholic cirrhosis) or controls (drinking comparable amounts over similar time, but free of significant liver disease). Extensive phenotypic data are obtained using semistructured interviews and patient records, and blood samples are collected. RESULTS: We have successfully recruited 859 participants including 538 matched case-control samples as of September 2014, using study-specific inclusion-exclusion criteria and data collection protocols. Of these, 580 are cases (442 men and 138 women) and 279 are controls (205 men and 74 women). Duration of excessive drinking was slightly greater in cases than controls and was significantly less in women than men. Cases had significantly lower lifetime alcohol intake than controls. Both cases and controls had a high prevalence of reported parental alcohol problems, but cases were significantly more likely to report that a father with alcohol problems had died from liver disease (odds ratio 2.53, 95% confidence interval 1.31 to 4.87, p = 0.0055). CONCLUSIONS: Recruitment of participants for a GWAS of alcoholic cirrhosis has proved feasible across countries with multiple sites. Affected patients often consume less alcohol than unaffected ones, emphasizing the existence of individual vulnerability factors. Cases are more likely to report liver disease in a father with alcohol problems than controls, consistent with a potential genetic component to the risk of alcoholic cirrhosis.

A snapshot of the hepatic transcriptome: ad libitum alcohol intake suppresses expression of cholesterol synthesis genes in alcohol-preferring (P) rats.

Research is uncovering the genetic and biochemical effects of consuming large quantities of alcohol. One prime example is the J- or U-shaped relationship between the levels of alcohol consumption and the risk of atherosclerotic cardiovascular disease. Moderate alcohol consumption in humans (about 30 g ethanol/d) is associated with reduced risk of coronary heart disease, while abstinence and heavier alcohol intake is linked to increased risk. However, the hepatic consequences of moderate alcohol drinking are largely unknown. Previous data from alcohol-preferring (P) rats showed that chronic consumption does not produce significant hepatic steatosis in this well-established model. Therefore, free-choice alcohol drinking in P rats may mimic low risk or nonhazardous drinking in humans, and chronic exposure in P animals can illuminate the molecular underpinnings of free-choice drinking in the liver. To address this gap, we captured the global, steady-state liver transcriptome following a 23 week free-choice, moderate alcohol consumption regimen (∼ 7.43 g ethanol/kg/day) in inbred alcohol-preferring (iP10a) rats. Chronic consumption led to down-regulation of nine genes in the cholesterol biosynthesis pathway, including HMG-CoA reductase, the rate-limiting step for cholesterol synthesis. These findings corroborate our phenotypic analyses, which indicate that this paradigm produced animals whose hepatic triglyceride levels, cholesterol levels and liver histology were indistinguishable from controls. These findings explain, at least in part, the J- or U-shaped relationship between cardiovascular risk and alcohol intake, and provide outstanding candidates for future studies aimed at understanding the mechanisms that underlie the salutary cardiovascular benefits of chronic low risk and nonhazardous alcohol intake.

The alcohol-preferring (P) and high-alcohol-drinking (HAD) rats--animal models of alcoholism.

The objective of this article is to review the literature on the utility of using the selectively bred alcohol-preferring (P) and high-alcohol-drinking (HAD) lines of rats in studies examining high alcohol drinking in adults and adolescents, craving-like behavior, and the co-abuse of alcohol with other drugs. The P line of rats meets all of the originally proposed criteria for a suitable animal model of alcoholism. In addition, the P rat exhibits high alcohol-seeking behavior, demonstrates an alcohol deprivation effect (ADE) under relapse drinking conditions, consumes amounts of ethanol during adolescence equivalent to those consumed in adulthood, and co-abuses ethanol and nicotine. The P line also exhibits excessive binge-like alcohol drinking, attaining blood alcohol concentrations (BACs) of 200 mg% on a daily basis. The HAD replicate lines of rats have not been as extensively studied as the P rats. The HAD1,2 rats satisfy several of the criteria for an animal model of alcoholism, e.g., these rats will voluntarily consume ethanol in a free-choice situation to produce BACs between 50 and 200 mg%. The HAD1,2 rats also exhibit an ADE under repeated relapse conditions, and will demonstrate similar levels of ethanol intake during adolescence as seen in adults. Overall, the P and HAD1,2 rats have characteristics attributed to an early onset alcoholic, and can be used to study various aspects of alcohol use disorders.

Gene expression within the extended amygdala of 5 pairs of rat lines selectively bred for high or low ethanol consumption.

The objectives of this study were to determine innate differences in gene expression in 2 regions of the extended amygdala between 5 different pairs of lines of male rats selectively bred for high or low ethanol consumption: a) alcohol-preferring (P) vs. alcohol-non-preferring (NP) rats, b) high-alcohol-drinking (HAD) vs. low-alcohol-drinking (LAD) rats (replicate line-pairs 1 and 2), c) ALKO alcohol (AA) vs. nonalcohol (ANA) rats, and d) Sardinian alcohol-preferring (sP) vs. Sardinian alcohol-nonpreferring (sNP) rats, and then to determine if these differences are common across the line-pairs. Microarray analysis revealed up to 1772 unique named genes in the nucleus accumbens shell (AcbSh) and 494 unique named genes in the central nucleus of the amygdala (CeA) that significantly differed [False Discovery Rate (FDR) = 0.10; fold-change at least 1.2] in expression between the individual line-pairs. Analysis using Gene Ontology (GO) and Ingenuity Pathways information indicated significant categories and networks in common for up to 3 or 4 line-pairs, but not for all 5 line-pairs. However, there were almost no individual genes in common within these categories and networks. ANOVAs of the combined data for the 5 line-pairs indicated 1014 and 731 significant (p < 0.01) differences in expression of named genes in the AcbSh and CeA, respectively. There were 4-6 individual named genes that significantly differed across up to 3 line-pairs in both regions; only 1 gene (Gsta4 in the CeA) differed in as many as 4 line-pairs. Overall, the findings suggest that a) some biological categories or networks (e.g., cell-to-cell signaling, cellular stress response, cellular organization, etc.) may be in common for subsets of line-pairs within either the AcbSh or CeA, and b) regulation of different genes and/or combinations of multiple biological systems may be contributing to the disparate alcohol drinking behaviors of these line-pairs.

Animal models for medications development targeting alcohol abuse using selectively bred rat lines: neurobiological and pharmacological validity.

The purpose of this review paper is to present evidence that rat animal models of alcoholism provide an ideal platform for developing and screening medications that target alcohol abuse and dependence. The focus is on the 5 oldest international rat lines that have been selectively bred for a high alcohol-consumption phenotype. The behavioral and neurochemical phenotypes of these rat lines are reviewed and placed in the context of the clinical literature. The paper presents behavioral models for assessing the efficacy of pharmaceuticals for the treatment of alcohol abuse and dependence in rodents, with particular emphasis on rats. Drugs that have been tested for their effectiveness in reducing alcohol/ethanol consumption and/or self-administration by these rat lines and their putative site of action are summarized. The paper also presents some current and future directions for developing pharmacological treatments targeting alcohol abuse and dependence.

Gene expression in the ventral tegmental area of 5 pairs of rat lines selectively bred for high or low ethanol consumption.

The objective of this study was to determine if there are common innate differences in gene expression or gene pathways in the ventral tegmental area (VTA) among 5 different pairs of rat lines selectively bred for high (HEC) or low (LEC) ethanol consumption: (a) alcohol-preferring (P) vs. alcohol-non-preferring (NP) rats; (b) high-alcohol-drinking (HAD) vs. low-alcohol-drinking (LAD) rats (replicate line pairs 1 and 2); (c) ALKO alcohol (AA) vs. nonalcohol (ANA) rats; and (d) Sardinian alcohol-preferring (sP) vs. alcohol-nonpreferring (sNP) rats. Microarray analysis revealed between 370 and 1340 unique named genes that significantly differed in expression between the individual line-pairs. Analysis using Gene Ontology (GO) and Ingenuity Pathways information indicated significant categories and networks in common for up to 3 line-pairs, but not for all 5 line-pairs; moreover, there were few genes in common in these categories and networks. ANOVA of the combined data for the 5 line-pairs indicated 1295 significant (p<0.01) differences in expression of named genes. Although no individual named gene was significant across all 5 line-pairs, there were 22 genes that overlapped in the same direction in 3 or 4 of the line-pairs. Overall, the findings suggest that (a) some biological categories or networks may be in common for subsets of line-pairs; and (b) regulation of different genes and/or combinations of multiple biological systems (e.g., transcription, synaptic function, intracellular signaling and protection against oxidative stress) within the VTA (possibly involving dopamine and glutamate) may be contributing to the disparate alcohol drinking behaviors of these line-pairs.

Comparison of the effect of the GABAΒ receptor agonist, baclofen, and the positive allosteric modulator of the GABAB receptor, GS39783, on alcohol self-administration in 3 different lines of alcohol-preferring rats.

BACKGROUND: Administration of the GABA(B) receptor agonist, baclofen, and positive allosteric modulator, GS39783, has been repeatedly reported to suppress multiple alcohol-related behaviors, including operant oral alcohol self-administration, in rats. This study was designed to compare the effect of baclofen and GS39783 on alcohol self-administration in 3 lines of selectively bred, alcohol-preferring rats: Indiana alcohol-preferring (P), Sardinian alcohol-preferring (sP), and Alko Alcohol (AA). METHODS: Rats of each line were initially trained to respond on a lever, on a fixed ratio (FR) 4 (FR4) schedule of reinforcement, to orally self-administer alcohol (15%, v/v) in daily 30-minute sessions. Once responding reached stable levels, rats were exposed to a sequence of experiments testing baclofen (0, 1, 1.7, and 3 mg/kg; i.p.) and GS39783 (0, 25, 50, and 100 mg/kg; i.g.) on FR4 and progressive ratio (PR) schedules of reinforcement. Finally, to assess the specificity of baclofen and GS39783 action, rats were slightly food-deprived and trained to lever-respond for food pellets. RESULTS: The rank of order of the reinforcing and motivational properties of alcohol was P>sP>AA rats. Under both FR and PR schedules of reinforcement, the rank of order of potency and efficacy of baclofen and GS39783 in suppressing alcohol self-administration was P>sP>AA rats. Only the highest dose of baclofen reduced lever-responding for food pellets; this effect was common to all 3 rat lines. Conversely, no dose of GS39783 altered lever-responding for food in any rat line. CONCLUSIONS: These results suggest that: (i) the strength of the reinforcing and motivational properties of alcohol differ among P, sP, and AA rats; (ii) the reinforcing and motivational properties of alcohol in P, sP, and AA rats are differentially sensitive to treatment with baclofen and GS39783; (iii) the heterogeneity in sensitivity to baclofen and GS39783 of alcohol self-administration in P, sP, and AA rats may resemble the differential effectiveness of pharmacotherapies among the different typologies of human alcoholics; and (iv) the GABA(B) receptor is part of the neural substrate mediating the reinforcing and motivational properties of alcohol.

Behavioral profiling of multiple pairs of rats selectively bred for high and low alcohol intake using the MCSF test.

Genetic aspects of alcoholism have been modeled using rats selectively bred for extremes of alcohol preference and voluntary alcohol intake. These lines show similar alcohol drinking phenotypes but have different genetic and environmental backgrounds and may therefore display diverse behavioral traits as seen in human alcoholics. The multivariate concentric square field™ (MCSF) test is designed to provoke exploration and behaviors associated with risk assessment, risk taking and shelter seeking in a novel environment. The aim was to use the MCSF to characterize behavioral profiles in rat lines from selective breeding programs in the United States (P/NP, HAD1/LAD1, HAD2/LAD2), Italy (sP/sNP) and Finland (AA/ANA). The open field and elevated plus maze tests were used as reference tests. There were substantial differences within some of the pairs of selectively bred rat lines as well as between all alcohol-preferring rats. The most pronounced differences within the pairs of lines were between AA and ANA rats and between sP and sNP rats followed by intermediate differences between P and NP rats and minor differences comparing HAD and LAD rats. Among all preferring lines, P, HAD1 and HAD2 rats shared similar behavioral profiles, while AA and sP rats were quite different from each other and the others. No single trait appeared to form a common 'pathway' associated with a high alcohol drinking phenotype among all of the alcohol-preferring lines of rats. The marked behavioral differences found in the different alcohol-preferring lines may mimic the heterogeneity observed among human alcoholic subtypes.

Genetic propensities to increase ethanol intake in response to stress: studies with selectively bred swim test susceptible (SUS), alcohol-preferring (P), and non-preferring (NP) lines of rats.

RATIONALE: Swim test susceptible (SUS) rats selectively bred for reduced struggling in the forced swim test (FST) following stress show high voluntary ethanol intake like alcohol-preferring (P) rats selectively bred for ethanol preference. It is unknown whether stress enhances drinking in SUS rats or FST behavior in P and non-preferring (NP) rats. OBJECTIVES: The aim of this study was to assess the response to stress in male SUS, Sprague-Dawley (SD), P, and NP rats on 10% ethanol drinking and FST behavior. METHODS: In experiment 1, SUS and SD rats had limited access to ethanol and water following white noise, rehousing, and forced swim stress. In experiment 2, P and NP rats received footshock, white noise, restraint, or no stress prior to the FST. Rats then had continuous access to ethanol and water, and the effects of weekly exposures to stress were measured. RESULTS: SUS rats drank more ethanol (M = 2.98 g/kg) than SD rats (M = 1.26 g/kg) at baseline. Stress produced sustained increases (~33% of baseline) in ethanol intake in SUS rats. NP rats spent twice as much time immobile as P rats in the FST. Stress did not alter FST behavior in P or NP rats. Only footshock produced an increase (~29%) in ethanol intake in P rats. CONCLUSIONS: Selection for stress-induced depressive-like behavior in SUS rats is associated with enhanced stress-induced ethanol drinking. However, the selection for alcohol preference is not associated with stress-induced depressive-like behavior but is associated with footshock stress-induced ethanol drinking. In these experiments, relationships among stress, depressive-like behavior, and alcohol preference were not symmetrical.

Hepatic and extrahepatic cancer in cirrhosis: a longitudinal cohort study.

OBJECTIVES: We conducted a retrospective cohort study in cirrhotic patients to understand (i) the risk of developing hepatocellular carcinoma (HCC) after an initial negative screening computed tomography (CT) scan and its relationship with underlying etiology and (ii) the risk of extrahepatic cancers (EHCs). METHODS: Our cohort consisted of 952 cirrhotics who had at least one contrast-enhanced CT scan over a 5-year period from 1997 to 2002. We assessed their risk of HCC and EHC until the study closure (31 December 2007). Using data from the Indiana State Cancer Registry (ISCR), standardized incidence ratios (SIRs) were calculated for HCC and EHC. RESULTS: The cohort's follow-up was 4.7±3.0 years. The frequency of HCC at baseline and during follow-up was 6.9 and 7.2%, respectively. The 1-, 3-, and 5-year HCC incidence after an initial negative CT scan was 1.2, 4.4, and 7.8%, respectively. The 1-, 3-, and 5-year EHC incidence was 2.2, 4.5, and 6.8%, respectively. The most common EHCs were breast, lung, and lymphoma. Incidence of both HCC (P=0.016) and EHC (P=0.004) varied significantly by the etiology of underlying cirrhosis. The SIRs for HCC and EHC were 186 (95% confidence interval (CI) 140-238) and 1.83 (95% CI 1.36-2.36), respectively. Compared with adjusted ISCR data, cirrhosis due to alcohol (SIR 2.73, 95% CI 1.14-4.33) but not other etiologies had significantly higher incidence of EHC. CONCLUSIONS: This study furthers our understanding of HCC and EHC risk in cirrhosis. If confirmed by other studies, these data will assist in developing optimal strategies for monitoring of cancer in individuals with cirrhosis.

Effects of sazetidine-A, a selective alpha4beta2 nicotinic acetylcholine receptor desensitizing agent on alcohol and nicotine self-administration in selectively bred alcohol-preferring (P) rats.

RATIONALE: Manipulations of nicotinic cholinergic receptors have been shown to influence both alcohol and nicotine intake. Sazetidine-A [6-(5(((S)-azetidine-2-yl)methoxy)pyridine-3-yl)hex-5-yn-1-ol] is a novel compound that potently and selectively desensitizes alpha4beta2 nicotinic receptors with only modest receptor activation. OBJECTIVES: The goal of the present study was to examine the effects of sazetidine-A on alcohol and nicotine self-administration in alcohol-preferring (P) rats. METHODS: P rats were given the choice of water or alcohol. Once stable baselines were established, the acute (0, 0.1, 0.3, 1, and 3 mg/kg, s.c.) and chronic (3 mg/kg for 10 days) effects of sazetidine-A on alcohol intake were assessed. Naltrexone (2.5 mg/kg) served as a positive control. The effect of sazetidine-A (3 mg/kg) and naltrexone (4 mg/kg) on saccharin (0.2%) preference was also assessed. In addition, the acute effects of sazetidine-A (3 mg/kg) and naltrexone (4 mg/kg) on alcohol intake after alcohol deprivation were evaluated. In another experiment, the effects of sazetidine-A (0, 1, or 3 mg/kg) on i.v. nicotine self-administration in P and NP rats were assessed. RESULTS: Sazetidine-A caused a dose-dependent reduction in alcohol intake. Chronic sazetidine-A also effectively reduced alcohol intake until the seventh day of treatment, when partial tolerance appeared to develop. In the post-deprivation study, sazetidine-A significantly reduced alcohol intake and preference. Sazetidine-A at 3 mg/kg significantly reduced nicotine self-administration in both lines. CONCLUSIONS: Sazetidine-A significantly reduced alcohol and nicotine intake in P rats that self-administer higher levels of both drugs. Sazetidine-A may hold promise for the treatment of alcohol and nicotine addiction.

Genomewide SNP screen to detect quantitative trait loci for alcohol preference in the high alcohol preferring and low alcohol preferring mice.

BACKGROUND: The high and low alcohol preferring (HAP1 and LAP1) mouse lines were selectively bred for differences in alcohol intake. The HAP1 and LAP1 mice are essentially noninbred lines that originated from the outbred colony of HS/Ibg mice, a heterogeneous stock developed from intercrossing 8 inbred strains of mice. METHODS: A total of 867 informative SNPs were genotyped in 989 HAP1 x LAP1 F2, 68 F1s, 14 parents (6 LAP1, 8 HAP1), as well as the 8 inbred strains of mice crossed to generate the HS/Ibg colony. Multipoint genome wide analyses were performed to simultaneously detect linked QTLs and also fine map these regions using the ancestral haplotypes. RESULTS: QTL analysis detected significant evidence of association on 4 chromosomes: 1, 3, 5, and 9. The region on chromosome 9 was previously found linked in a subset of these F2 animals using a whole genome microsatellite screen. CONCLUSIONS: We have detected strong evidence of association to multiple chromosomal regions in the mouse. Several of these regions include candidate genes previously associated with alcohol dependence in humans or other animal models.

Effects of concurrent access to multiple ethanol concentrations and repeated deprivations on alcohol intake of high-alcohol-drinking (HAD) rats.

High-alcohol-drinking rats, given access to 10% ethanol, expressed an alcohol deprivation effect (ADE) only after multiple deprivations. In alcohol-preferring (P) rats, concurrent access to multiple ethanol concentrations combined with repeated cycles of EtOH access and deprivation produced excessive ethanol drinking. The current study was undertaken to examine the effects of repeated alcohol deprivations with concurrent access to multiple concentrations of ethanol on ethanol intake of HAD replicate lines of rats. HAD-1 and HAD-2 rats received access to 10, 20 and 30% (v/v) ethanol for 6 weeks. Rats from each replicate line were assigned to: (1) a non-deprived group; (2) a group initially deprived of ethanol for 2 weeks; or (3) a group initially deprived for 8 weeks. Following the restoration of the ethanol solutions, cycle of 2 weeks of ethanol exposure and 2 weeks of alcohol deprivation was repeated three times for a total of four deprivations. Following the initial ethanol deprivation period, deprived groups significantly increased ethanol intakes during the initial 24-hour re-exposure period. Multiple deprivations increased ethanol intakes, shifted preference to higher ethanol concentrations and prolonged the duration of the elevated ethanol intakes for up to 5 days. In addition, repeated deprivations increased ethanol intake in the first 2-hour re-exposure period as high as 5-7 g/kg (which are equivalent to amounts consumed in 24 hours by HAD rats), and produced blood ethanol levels in excess of 150 mg%. The results indicate that HAD rats exhibit 'loss-of-control' of alcohol drinking with repeated deprivations when multiple ethanol concentrations are available.

Acute ethanol effects on local cerebral glucose utilization in select central nervous system regions of adolescent alcohol-preferring (P) and alcohol-nonpreferring (NP) rats.

BACKGROUND: Alcohol abuse among adolescents is a major health and developmental problem. The 2-[(14)C]deoxyglucose (2-DG) technique allows for the in vivo quantification of local cerebral glucose utilization (LCGU) as a measure of functional neuronal activity. METHODS: Local cerebral glucose utilization rates were examined after acute ethanol administration within selected brain regions of adolescent alcohol-preferring (P) and -nonpreferring (NP) rats. Postnatal day 45 male P and NP rats were injected with saline or 1.0 g/kg ethanol, i.p., 10 minutes prior to an intravenous bolus of [(14)C]-2-deoxyglucose (125 microCi/kg). Image densities were determined using quantitative autoradiography and LCGU values calculated. RESULTS: Acute ethanol injection significantly decreased LCGU rates in select brain regions including the olfactory tubercles, the frontal cortex (Fr), and subregions of the posterior hippocampus (pCA1 and pCA3). Acute ethanol had no significant effects on LCGU rates in any region of the adolescent NP rats. Significant basal LCGU rate differences were apparent between the rat lines in a nearly global fashion with adolescent P rats having much higher basal LCGU rates compared with adolescent NP rats. CONCLUSIONS: These findings suggest that the adolescent P and NP rats are less sensitive to the effects of acute ethanol than their adult counterparts. The adolescent P rat is relatively more sensitive to the initial effects of acute ethanol in select brain regions as compared with the adolescent NP rat. Additionally, the innate hyper-excited state of the adolescent P central nervous system is a likely factor in the development of their high alcohol drinking behaviors.