Chapter Nine - Genes and Alcohol Consumption: Studies with Mutant Mice

https://doi.org/10.1016/bs.irn.2016.02.014Get rights and content

Abstract

In this chapter, we review the effects of global null mutant and overexpressing transgenic mouse lines on voluntary self-administration of alcohol. We examine approximately 200 publications pertaining to the effects of 155 mouse genes on alcohol consumption in different drinking models. The targeted genes vary in function and include neurotransmitter, ion channel, neuroimmune, and neuropeptide signaling systems. The alcohol self-administration models include operant conditioning, two- and four-bottle choice continuous and intermittent access, drinking in the dark limited access, chronic intermittent ethanol, and scheduled high alcohol consumption tests. Comparisons of different drinking models using the same mutant mice are potentially the most informative, and we will highlight those examples. More mutants have been tested for continuous two-bottle choice consumption than any other test; of the 137 mouse genes examined using this model, 97 (72%) altered drinking in at least one sex. Overall, the effects of genetic manipulations on alcohol drinking often depend on the sex of the mice, alcohol concentration and time of access, genetic background, as well as the drinking test.

Introduction

Alcohol use disorder (AUD) is a multifactorial disease, and its risk factors are determined by the interplay of genetic and environmental factors, combined with neuroadaptations following acute and repeated alcohol exposure. Alcohol targets ion channels and signaling cascades, producing intoxication, anxiolysis, and a sense of reward. After prolonged, repeated exposure, alcohol-induced changes in gene expression and synaptic function are thought to contribute to the development tolerance, sensitization, and compulsive consumption and drug seeking.

More than 100 genes have been shown to affect alcohol consumption and other alcohol-related behaviors in mouse models (Crabbe, Phillips, Harris, Arends, & Koob, 2006). Excessive alcohol consumption is a common model of addictive behavior, and animal models of voluntary self-administration are valuable for profiling genetic determinants of AUD (Green & Grahame, 2008). The preference to drink alcohol is a reliable measure that depends upon mouse genotype and has been consistent across laboratories despite variations of the drinking protocol used. Preclinical models, in conjunction with human genetic studies, may expose overlapping target genes and identify the most relevant drinking models and biological systems associated with AUD.

In this review, we focus on a single phenotype, voluntary alcohol self-administration, and summarize the global genetic manipulations in mice published to date on this behavior. Most of the studies used two-bottle choice (2BC) tests, where mice had a choice between water and ethanol and access was usually measured in continuous 24-h periods. In some cases, 2BC access to ethanol was intermittent (eg, every other day), which typically (Hwa et al., 2011, Melendez, 2011, Rosenwasser et al., 2013), but not always (Crabbe, Harkness, Spence, Huang, & Metten, 2012), results in higher ethanol intake compared to continuous access. A few studies used four-bottle choice (4BC) access, where mice have simultaneous access to water and three different concentrations of ethanol. Because rodents distribute their drinking across the circadian cycle and because limited access to ethanol tends to increase intake, restricted access during the dark cycle is often used to study periods of high consumption and to model binge-like drinking in humans (Thiele & Navarro, 2014). In the classic mouse drinking in the dark (DID) test, drinking session times begin a few hours after the start of the dark cycle and usually last 2–4 h over a few days. High levels of ethanol drinking and pharmacologically relevant blood ethanol concentrations (BECs) are achieved using this model (Thiele & Navarro, 2014). The scheduled high access consumption (SHAC) test uses fluid restriction to promote drinking of a low ethanol concentration (Finn et al., 2005). This chronic drinking model can also produce high BECs. Fluid access is first restricted and then gradually relaxed until the effects of fluid limitation are minimized. In operant self-administration tests, mice are trained to self-administer quantities of ethanol that produce moderate to high BECs. Removal of access to alcohol followed by restored access transiently increases consumption in dependent mice, and the effects of mutant genes on this alcohol deprivation effect (ADE; Rodd et al., 2004, Vengeliene et al., 2014), a model of relapse drinking, are also presented. As shown in the tables throughout the chapter, the effects of some mutants can depend on the drinking test used as well as the ethanol concentration, time of access, genetic background, and sex of the mice (Vanderlinden, Saba, Bennett, Hoffman, & Tabakoff, 2015).

There are important considerations regarding genetic engineering methods, including the potential alteration of genes other than the mutated gene and the influence of the background strain carrying the genotype. C57BL/6J (B6) mice are a high alcohol-drinking strain and, as the tables in this chapter demonstrate, occupy a central role in voluntary drinking studies. This review focuses on global homozygous knockouts, although a few studies used hypofunctional or overexpressing transgenic lines. Strategies to reduce confounding and compensatory effects of null mutations include the use of knockin mice and brain regional or cell-specific knockouts, and a few of these studies are noted. In this review, we do not debate the genetic engineering methods used; instead, our aim is to provide a summary of homozygous null or overexpressing mutants and their role (or lack thereof) in alcohol consumption in mice. Current mouse gene and protein names from Uniprot (http://www.uniprot.org) are listed in the tables, which in some cases differ from the nomenclature used in the published studies. We first review the effects of mutant neurotransmitter receptor subunits on alcohol drinking in mice and then examine mutations in other ion channel receptors, cannabinoid and opioid receptors, neuropeptides, kinases/enzymes, and immune-related genes.

Section snippets

γ-Aminobutyric Acid

Alcohol potentiates γ-aminobutyric acid type A (GABAA) receptor-mediated responses and enhances inhibitory neurotransmission, and some of the top candidate genes implicated in alcohol consumption code for specific GABAA receptor subunits (Trudell, Messing, Mayfield, & Harris, 2014). Deletion of the α1 subunit decreased ethanol consumption in operant and 2BC tests (Blednov et al., 2003, June et al., 2007), and knockdown of α5 reduced drinking in male (Boehm, Ponomarev, et al., 2004) but not

Cannabinioids and Opioids

The endocannabinoid system is involved in brain reward signaling and drug-seeking behavior (Panagis, Mackey, & Vlachou, 2014). In several studies of different genetic backgrounds, male and female cannabinoid 1 receptor (CB1R) knockout mice showed reduced ethanol intake and/or preference for ethanol than wild-type mice (Hungund et al., 2003, Lallemand and de Witte, 2005, Naassila et al., 2004, Poncelet et al., 2003, Racz et al., 2003, Thanos, Dimitrakakis, et al., 2005, Vinod, Yalamanchili, et

Immune-Related Genes

The interplay between brain, behavior, and immune responses in the etiology and progression of drug abuse is a current area of interest in addiction research (http://www.arcr.niaaa.nih.gov/arcr/arcr372/toc37_2.htm). The neuroimmune system, encompassing innate immune signaling within the peripheral and central nervous systems, is important in the pathophysiology and potential treatment of alcohol abuse and dependence (Crews and Vetreno, 2015, Mayfield et al., 2013, Robinson et al., 2014).

Ion Channels

Some of the rapid-onset actions of alcohol are likely mediated by direct action on ion channels (Howard et al., 2014, Trudell et al., 2014). In addition to the prominent and well-studied neurotransmitter systems in alcohol dependence discussed previously (eg, GABA, glutamate), other ion channels that have been implicated in alcohol intake and preference are described below.

Protein Kinases

In addition to direct effects on ion channels, ethanol indirectly modulates channel function via phosphorylation and other posttranslational processing mechanisms (Trudell et al., 2014). Mice lacking protein kinase C type ɛ (PKCɛ) drank less ethanol than wild type, and this effect has been observed in different drinking tests across multiple labs (Besheer et al., 2006, Choi et al., 2002, Hodge et al., 1999, Olive et al., 2005, Olive et al., 2000, Wallace et al., 2007; Table 9). Selective

Enzymes

Table 10 shows the effects of other assorted enzymes on ethanol drinking in mice. In particular, aldehyde dehydrogenase (ALDH) is one of the few known genes to affect risk of developing AUD in humans (Chen, Ferreira, Gross, & Mochly-Rosen, 2014). ALDH2 plays a major role in the detoxification of ethanol-derived acetaldehyde, and inhibition of ALDH is the mechanism of action of disulfiram, an FDA-approved drug for AUD. A mutation in ALDH2 produces an enzyme incapable of metabolizing ethanol,

Neuropeptides/Hormones

Table 11 summarizes the effects of deletion or overexpression of classical or putative neuropeptides and hormones and their receptors on voluntary ethanol administration in mice. Of the genes represented here, the corticotropin-releasing factor/urocortin family and other stress-related neuromodulators are promising for future studies of genetic determinants of AUD (Schank, Ryabinin, Giardino, Ciccocioppo, & Heilig, 2012).

Other Gene Targets

Table 12 represents an assortment of genes that did not specifically fit into the previous categories. These genes are associated with synaptic function, development, circadian regulation, and other cellular regulatory functions but are not discussed individually herein. One of the most pronounced phenotypes among mutant mice is the almost complete blockade of alcohol consumption in mice lacking any one of the three taste genes (Gnat3, Tas1r3, and Trpm5) (Blednov et al., 2008).

Concluding Remarks

Mouse models of voluntary ethanol administration have been instrumental for profiling putative behavioral and genetic determinants in human alcoholics, who exhibit excessive consumption as a hallmark of the disease. The impact of more than 150 genes on alcohol consumption has been evaluated by construction of mutant mice. The global knockout strategy has been used extensively in addiction research to link proteins with behavior, and most studies presented in this chapter used this approach.

Medication Development

FDA-approved drugs for AUD have provided only modest benefit and are not routinely prescribed, and so the search continues for more effective drugs. Identifying existing drugs that could be repurposed to treat AUD is a current goal for researchers and, if successful, would fast track therapeutic options for the disease. A strategy for prioritizing relevant genes from a large list of potential targets is to examine the preclinical evidence in combination with genetic association studies in human

Future Directions

While the study of individual genes is informative, combining gene network and systems biology approaches to identify interrelated networks and pathways is critical in the future treatment of AUD. Because complex trait diseases involve coordinated expression changes in multiple gene families, examining gene clusters is an important research direction, as supported by the INIA studies showing that coexpression patterns can distinguish gene modules related to alcohol consumption in animal models (

Acknowledgments

The authors acknowledge funding from NIAAA grants AA006399, AA013520, AA020926, and AA012404. The authors have no conflicts of interest with this material. The views expressed herein are solely those of the authors and do not necessarily represent those of the funding agencies.

References (264)

  • S.L. Boehm et al.

    Over-expression of the fyn-kinase gene reduces hypnotic sensitivity to ethanol in mice

    Neuroscience Letters

    (2004)
  • S.L. Boehm et al.

    gamma-Aminobutyric acid A receptor subunit mutant mice: New perspectives on alcohol actions

    Biochemical Pharmacology

    (2004)
  • J.A. Bouwknecht et al.

    Ethanol intake is not elevated in male 5-HT(1B) receptor knockout mice

    European Journal of Pharmacology

    (2000)
  • J.M. Boyce-Rustay et al.

    Dopamine D3 receptor knockout mice and the motivational effects of ethanol

    Pharmacology, Biochemistry, and Behavior

    (2003)
  • Z.B. Bulwa et al.

    Increased consumption of ethanol and sugar water in mice lacking the dopamine D2 long receptor

    Alcohol

    (2011)
  • H.K. Caldwell et al.

    The acute intoxicating effects of ethanol are not dependent on the vasopressin 1a or 1b receptors

    Neuropeptides

    (2006)
  • P. Charbogne et al.

    15 years of genetic approaches in vivo for addiction research: Opioid receptor and peptide gene knockout in mouse models of drug abuse

    Neuropharmacology

    (2014)
  • J. Chen et al.

    Altered glutamatergic neurotransmission in the striatum regulates ethanol sensitivity and intake in mice lacking ENT1

    Behavioural Brain Research

    (2010)
  • K. Chu et al.

    Dependence-induced increases in ethanol self-administration in mice are blocked by the CRF1 receptor antagonist antalarmin and by CRF1 receptor knockout

    Pharmacology, Biochemistry, and Behavior

    (2007)
  • M.S. Cowen et al.

    Neurobehavioral effects of alcohol in AMPA receptor subunit (GluR1) deficient mice

    Neuropharmacology

    (2003)
  • A. Dawson et al.

    The beta2 nicotinic acetylcholine receptor subunit differentially influences ethanol behavioral effects in the mouse

    Alcohol

    (2013)
  • M. El-Ghundi et al.

    Disruption of dopamine D1 receptor gene expression attenuates alcohol-seeking behavior

    European Journal of Pharmacology

    (1998)
  • M.A. Enoch

    The role of GABA(A) receptors in the development of alcoholism

    Pharmacology, Biochemistry, and Behavior

    (2008)
  • F. Faber et al.

    Lack of angiotensin II conversion to angiotensin III increases water but not alcohol consumption in aminopeptidase A-deficient mice

    Regulatory Peptides

    (2006)
  • S.P. Farris et al.

    Applying the new genomics to alcohol dependence

    Alcohol

    (2015)
  • L.B. Ferguson et al.

    PPAR agonists regulate brain gene expression: Relationship to their effects on ethanol consumption

    Neuropharmacology

    (2014)
  • X. Gallego et al.

    Transgenic over expression of nicotinic receptor alpha 5, alpha 3, and beta 4 subunit genes reduces ethanol intake in mice

    Alcohol

    (2012)
  • G. Gorini et al.

    Molecular targets of alcohol action: Translational research for pharmacotherapy development and screening

    Progress in Molecular Biology and Translational Science

    (2011)
  • N.J. Grahame et al.

    Naltrexone and alcohol drinking in mice lacking beta-endorphin by site-directed mutagenesis

    Pharmacology, Biochemistry, and Behavior

    (2000)
  • A.S. Green et al.

    Ethanol drinking in rodents: Is free-choice drinking related to the reinforcing effects of ethanol?

    Alcohol

    (2008)
  • J.E. Grisel et al.

    Ethanol oral self-administration is increased in mutant mice with decreased beta-endorphin expression

    Brain Research

    (1999)
  • R.A. Harris et al.

    Testing the silence of mutations: Transcriptomic and behavioral studies of GABA(A) receptor alpha1 and alpha2 subunit knock-in mice

    Neuroscience Letters

    (2011)
  • R.G. Agrawal et al.

    Bioinformatics analyses reveal age-specific neuroimmune modulation as a target for treatment of high ethanol drinking

    Alcoholism, Clinical and Experimental Research

    (2014)
  • I.E. Bauer et al.

    Serotonergic gene variation in substance use pharmacotherapy: A systematic review

    Pharmacogenomics

    (2015)
  • A. Becker et al.

    Rewarding effects of ethanol and cocaine in mu opioid receptor-deficient mice

    Naunyn-Schmiedeberg's Archives of Pharmacology

    (2002)
  • J. Besheer et al.

    GABAA receptor regulation of voluntary ethanol drinking requires PKCepsilon

    Synapse

    (2006)
  • M.K. Bird et al.

    Metabotropic glutamate 5 receptors regulate sensitivity to ethanol in mice

    The International Journal of Neuropsychopharmacology

    (2008)
  • K. Bjork et al.

    Modulation of voluntary ethanol consumption by beta-arrestin 2

    The FASEB Journal

    (2008)
  • Y.A. Blednov et al.

    Peroxisome proliferator-activated receptors alpha and gamma are linked with alcohol consumption in mice and withdrawal and dependence in humans

    Alcoholism, Clinical and Experimental Research

    (2015)
  • Y.A. Blednov et al.

    Glycine receptors containing alpha2 or alpha3 subunits regulate specific ethanol-mediated behaviors

    The Journal of Pharmacology and Experimental Therapeutics

    (2015)
  • Y.A. Blednov et al.

    GABAA receptors containing rho1 subunits contribute to in vivo effects of ethanol in mice

    PLoS One

    (2014)
  • Y.A. Blednov et al.

    PPAR agonists: I. Role of receptor subunits in alcohol consumption in male and female mice

    Alcoholism, Clinical and Experimental Research

    (2016)
  • Y.A. Blednov et al.

    Loss of ethanol conditioned taste aversion and motor stimulation in knockin mice with ethanol-insensitive alpha2-containing GABA(A) receptors

    The Journal of Pharmacology and Experimental Therapeutics

    (2011)
  • Y.A. Blednov et al.

    Role of endocannabinoids in alcohol consumption and intoxication: Studies of mice lacking fatty acid amide hydrolase

    Neuropsychopharmacology

    (2007)
  • Y.A. Blednov et al.

    Metabotropic glutamate receptor 5 (mGluR5) regulation of ethanol sedation, dependence and consumption: Relationship to acamprosate actions

    The International Journal of Neuropsychopharmacology

    (2008)
  • Y.A. Blednov et al.

    Neuroimmune regulation of alcohol consumption: Behavioral validation of genes obtained from genomic studies

    Addiction Biology

    (2012)
  • Y.A. Blednov et al.

    Potassium channels as targets for ethanol: Studies of G-protein-coupled inwardly rectifying potassium channel 2 (GIRK2) null mutant mice

    The Journal of Pharmacology and Experimental Therapeutics

    (2001)
  • Y.A. Blednov et al.

    GABAA receptor alpha 1 and beta 2 subunit null mutant mice: Behavioral responses to ethanol

    The Journal of Pharmacology and Experimental Therapeutics

    (2003)
  • Y.A. Blednov et al.

    Blockade of the leptin-sensitive pathway markedly reduces alcohol consumption in mice

    Alcoholism, Clinical and Experimental Research

    (2004)
  • Y.A. Blednov et al.

    Mice lacking Gad2 show altered behavioral effects of ethanol, flurazepam and gabaxadol

    Addiction Biology

    (2010)
  • Cited by (0)

    View full text