Publications by authors named "Angela R Ozburn"

20 Publications

  • Page 1 of 1

Ethanol-Related Behaviors in Mouse Lines Selectively Bred for Drinking to Intoxication.

Brain Sci 2021 Feb 4;11(2). Epub 2021 Feb 4.

Department of Behavioral Neuroscience, Oregon Health & Science University, and VA Portland Health Care System, Portland, OR 97239, USA.

Alcohol use disorder (AUD) is a devastating psychiatric disorder that has significant wide-reaching effects on individuals and society. Selectively bred mouse lines are an effective means of exploring the genetic and neuronal mechanisms underlying AUD and such studies are translationally important for identifying treatment options. Here, we report on behavioral characterization of two replicate lines of mice that drink to intoxication, the High Drinking in the Dark (HDID)-1 and -2 mice, which have been selectively bred (20+ generations) for the primary phenotype of reaching high blood alcohol levels (BALs) during the drinking in the dark (DID) task, a binge-like drinking assay. Along with their genetically heterogenous progenitor line, Hs/Npt, we tested these mice on: DID and drinking in the light (DIL); temporal drinking patterns; ethanol sensitivity, through loss of righting reflex (LORR); and operant self-administration, including fixed ratio (FR1), fixed ratio 3:1 (FR3), extinction/reinstatement, and progressive ratio (PR). All mice consumed more ethanol during the dark than the light and both HDID lines consumed more ethanol than Hs/Npt during DIL and DID. In the dark, we found that the HDID lines achieved high blood alcohol levels early into a drinking session, suggesting that they exhibit front loading like drinking behavior in the absence of the chronicity usually required for such behavior. Surprisingly, HDID-1 (female and male) and HDID-2 (male) mice were more sensitive to the intoxicating effects of ethanol during the dark (as determined by LORR), while Hs/Npt (female and male) and HDID-2 (female) mice appeared less sensitive. We observed lower HDID-1 ethanol intake compared to either HDID-2 or Hs/Npt during operant ethanol self-administration. There were no genotype differences for either progressive ratio responding, or cue-induced ethanol reinstatement, though the latter is complicated by a lack of extinguished responding behavior. Taken together, these findings suggest that genes affecting one AUD-related behavior do not necessarily affect other AUD-related behaviors. Moreover, these findings highlight that alcohol-related behaviors can also differ between lines selectively bred for the same phenotype, and even between sexes within those same line.
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http://dx.doi.org/10.3390/brainsci11020189DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7915226PMC
February 2021

The impact of Drinking in the Dark (DID) procedural manipulations on ethanol intake in High Drinking in the Dark (HDID) mice.

Alcohol 2021 Jun 6;93:45-56. Epub 2021 Feb 6.

Portland Alcohol Research Center, Department of Behavioral Neuroscience at Oregon Health & Science University and Veterans Administration Portland Health Care System, Portland, OR, United States.

The High Drinking in the Dark mouse lines (HDID-1 and HDID-2) were selectively bred to achieve high blood ethanol concentrations (BECs) in the Drinking in the Dark (DID) task, a widely used model of binge-like intake of 20% ethanol. There are several components that differentiate DID from other animal models of ethanol intake: time of day of testing, length of ethanol access, single-bottle access, and individual housing. Here, we sought to determine how some of these individual factors contribute to the high ethanol intake observed in HDID mice. HDID-1, HDID-2, and non-selected HS/NPT mice were tested in a series of DID experiments where one of the following factors was manipulated: length of ethanol access, fluid choice, number of ethanol bottles, and housing condition. We observed that 1) HDID mice achieve intoxicating BECs in DID, even when they are group-housed; 2) HDID mice continue to show elevated ethanol intake relative to HS/NPT mice during an extended access session, but this is most apparent during the first 4 h of access; and 3) offering a water choice during DID prevents elevated intake in the HDID-1 mice, but not necessarily in HDID-2 mice. Together, these results suggest that the lack of choice in the DID paradigm, together with the length of ethanol access, are important factors contributing to elevated ethanol intake in the HDID mice. These results further suggest important differences between the HDID lines in response to procedural manipulations of housing condition and ethanol bottle number in the DID paradigm, highlighting the distinct characteristics that each of these lines possess, despite being selectively bred for the same phenotype.
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http://dx.doi.org/10.1016/j.alcohol.2021.02.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8113115PMC
June 2021

Effects of Tacrolimus and Other Immune Targeting Compounds on Binge-Like Ethanol Drinking in High Drinking in the Dark Mice.

Neurosci Insights 2020 25;15:2633105520975412. Epub 2020 Nov 25.

Portland Alcohol Research Center, Department of Behavioral Neuroscience at Oregon Health and Science University and VA Portland Health Care System, Portland, OR, USA.

High Drinking in the Dark (HDID-1) mice represent a unique genetic risk model of binge-like drinking and a novel means of screening potential pharmacotherapies to treat alcohol use disorders (AUDs). We tested the effects of tacrolimus (0, 0.5, 1, and 2 mg/kg), sirolimus (0, 5, 10, and 20 mg/kg), palmitoylethanolamide (PEA; 0, 75, 150, and 225 mg/kg), and secukinumab (0, 5, 20, and 60 mg/kg) on binge-like ethanol intake (2-day, "Drinking in the Dark" [DID]) and blood alcohol levels (BALs) in HDID-1 mice. Tacrolimus reduced ethanol intake and BALs. Tacrolimus had no effect on water intake, but reduced saccharin intake. There was no effect of sirolimus, PEA, or secukinumab on ethanol intake or BALs. These results compare and contrast with previous work addressing these compounds or their targeted mechanisms of action on ethanol drinking, highlighting the importance of screening a wide range of models and genotypes to inform the role of neuroimmune signaling in AUDs.
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http://dx.doi.org/10.1177/2633105520975412DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7705291PMC
November 2020

Valproate reverses mania-like behaviors in mice via preferential targeting of HDAC2.

Mol Psychiatry 2020 Nov 24. Epub 2020 Nov 24.

Translational Neuroscience Program, Department of Psychiatry, University of Pittsburgh School of Medicine, Pittsburgh, PA, 15219, USA.

Valproate (VPA) has been used in the treatment of bipolar disorder since the 1990s. However, the therapeutic targets of VPA have remained elusive. Here we employ a preclinical model to identify the therapeutic targets of VPA. We find compounds that inhibit histone deacetylase proteins (HDACs) are effective in normalizing manic-like behavior, and that class I HDACs (e.g., HDAC1 and HDAC2) are most important in this response. Using an RNAi approach, we find that HDAC2, but not HDAC1, inhibition in the ventral tegmental area (VTA) is sufficient to normalize behavior. Furthermore, HDAC2 overexpression in the VTA prevents the actions of VPA. We used RNA sequencing in both mice and human induced pluripotent stem cells (iPSCs) derived from bipolar patients to further identify important molecular targets. Together, these studies identify HDAC2 and downstream targets for the development of novel therapeutics for bipolar mania.
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http://dx.doi.org/10.1038/s41380-020-00958-2DOI Listing
November 2020

Effects of chemogenetic manipulation of the nucleus accumbens core in male C57BL/6J mice.

Alcohol 2021 03 4;91:21-27. Epub 2020 Nov 4.

Portland Veterans Affairs Medical Center, Research and Development Service, 3710 SW U.S. Veterans Hospital Road, Portland, OR, 97239, United States; Oregon Health and Science University, Department of Behavioral Neuroscience, 3181 SW Sam Jackson Park Road, Portland, OR, 97239, United States. Electronic address:

Binge drinking is a widespread public health concern with limited effective treatment options. To better select pharmaceutical targets, it is imperative to expand our knowledge of the underlying neural mechanisms involved in binge drinking. Our previous experiments in C57BL/6J female mice found that increasing activity in the nucleus accumbens (NAc) core using excitatory Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) reduced binge-like drinking. These results differed from what has been found in males; however, it is unclear whether differences in experimental procedures or sex underlie these discrepancies. We matched the conditions used in our female study and asked whether bidirectional manipulation of NAc core activity has different effects on binge-like drinking in males. Male C57BL/6J mice were stereotaxically injected with AAV2 hSyn-HA hM3Dq (excitatory), -hM4Di (inhibitory), or -eGFP bilaterally into the NAc core. We tested the effects of altering NAc activity on binge-like ethanol intake using Drinking in the Dark (DID). During the first week, mice were pre-treated with vehicle to establish baseline ethanol intake. In week 2, mice were treated with 1 mg/kg CNO prior to DID to determine the effects of DREADD-induced changes in NAc core activity on ethanol intake. Decreasing activity via CNO/hM4Di significantly decreased binge-like drinking in male mice relative to eGFP and hM4Di groups. We also measured intake of sucrose, quinine, and water after CNO treatment and found that increasing NAc core activity via CNO/hM3Dq increased quinine intake, and increased water intake over time. We did not observe significant differences in the GFP or hM4Di groups. This work suggests there exist apparent sex-related differences in NAc core contributions to binge-like alcohol drinking, thus demonstrating the need for inclusion of both sexes in future work.
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http://dx.doi.org/10.1016/j.alcohol.2020.10.005DOI Listing
March 2021

Targeting the Glucocorticoid Receptor Reduces Binge-Like Drinking in High Drinking in the Dark (HDID-1) Mice.

Alcohol Clin Exp Res 2020 05 27;44(5):1025-1036. Epub 2020 Mar 27.

Department of Behavioral Neuroscience, Portland Alcohol Research Center, Oregon Health & Science University, Portland, Oregon.

Background: Chronic alcohol exposure can alter glucocorticoid receptor (GR) function in some brain areas that promotes escalated and compulsive-like alcohol intake. GR antagonism can prevent dependence-induced escalation in drinking, but very little is known about the role of GR in regulating high-risk nondependent alcohol intake. Here, we investigate the role of GR in regulating binge-like drinking and aversive responses to alcohol in the High Drinking in the Dark (HDID-1) mice, which have been selectively bred for high blood ethanol (EtOH) concentrations (BECs) in the Drinking in the Dark (DID) test, and in their founder line, the HS/NPT.

Methods: In separate experiments, male and female HDID-1 mice were administered one of several compounds that inhibited GR or its negative regulator, FKBP51 (mifepristone [12.5, 25, 50, 100 mg/kg], CORT113176 [20, 40, 80 mg/kg], and SAFit2 [10, 20, 40 mg/kg]) during a 2-day DID task. EtOH consumption and BECs were measured. EtOH conditioned taste and place aversion (CTA and CPA, respectively) were measured in separate HDID-1 mice after mifepristone administration to assess GR's role in regulating the conditioned aversive effects of EtOH. Lastly, HS/NPT mice were administered CORT113176 during DID to assess whether dissimilar effects from those of HDID-1 would be observed, which could suggest that selective breeding had altered sensitivity to the effects of GR antagonism on binge-like drinking.

Results: GR antagonism (with both mifepristone and CORT113176) selectively reduced binge-like EtOH intake and BECs in the HDID-1 mice, while inhibition of FKBP51 did not alter intake or BECs. In contrast, GR antagonism had no effect on EtOH intake or BECs in the HS/NPT mice. Although HDID-1 mice exhibit attenuated EtOH CTA, mifepristone administration did not enhance the aversive effects of EtOH in either a CTA or CPA task.

Conclusion: These data suggest that the selection process increased sensitivity to GR antagonism on EtOH intake in the HDID-1 mice, and support a role for the GR as a genetic risk factor for high-risk alcohol intake.
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http://dx.doi.org/10.1111/acer.14318DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7211124PMC
May 2020

Chronic Chemogenetic Stimulation of the Nucleus Accumbens Produces Lasting Reductions in Binge Drinking and Ameliorates Alcohol-Related Morphological and Transcriptional Changes.

Brain Sci 2020 Feb 18;10(2). Epub 2020 Feb 18.

Department of Behavioral Neuroscience, Oregon Health & Science University, Portland, OR 97239, USA.

Binge drinking is a dangerous pattern of behavior. We tested whether chronically manipulating nucleus accumbens (NAc) activity (via clozapine-N-oxide (CNO) and Designer Receptors Exclusively Activated by Designer Drugs (DREADD)) could produce lasting effects on ethanol binge-like drinking in mice selectively bred to drink to intoxication. We found chronically increasing NAc activity (4 weeks, via CNO and the excitatory DREADD, hM3Dq) decreased binge-like drinking, but did not observe CNO-induced changes in drinking with the inhibitory DREADD, hM4Di. The CNO/hM3Dq-induced reduction in ethanol drinking persisted for at least one week, suggesting adaptive neuroplasticity via transcriptional and epigenetic mechanisms. Therefore, we defined this plasticity at the morphological and transcriptomic levels. We found that chronic binge drinking (6 weeks) altered neuronal morphology in the NAc, an effect that was ameliorated with CNO/hM3Dq. Moreover, we detected significant changes in expression of several plasticity-related genes with binge drinking that were ameliorated with CNO treatment (e.g., ). Lastly, we found that LMK235, an HDAC4/5 inhibitor, reduced binge-like drinking. Thus, we were able to target specific molecular pathways using pharmacology to mimic the behavioral effects of DREADDs.
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http://dx.doi.org/10.3390/brainsci10020109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7071376PMC
February 2020

Effects of Pharmacologically Targeting Neuroimmune Pathways on Alcohol Drinking in Mice Selectively Bred to Drink to Intoxication.

Alcohol Clin Exp Res 2020 02 13;44(2):553-566. Epub 2020 Jan 13.

Department of Behavioral Neuroscience, Portland Alcohol Research Center, VA Portland Health Care System, Oregon Health & Science University, Portland, Oregon.

Background: Rodent models of high alcohol drinking offer opportunities to better understand factors for alcohol use disorders (AUD) and test potential treatments. Selective breeding was carried out to create 2 unique High Drinking in the Dark (HDID-1, HDID-2) mouse lines that represent models of genetic risk for binge-like drinking. A number of studies have indicated that neuroimmune genes are important for regulation of alcohol drinking. We tested whether compounds shown to reduce drinking in other models also reduce alcohol intake in these unique genetic lines.

Methods: We report tests of gabapentin, tesaglitazar, fenofibrate, caffeic acid phenethyl ester (CAPE), ibrutinib, and rolipram. Although these compounds have different mechanisms of action, they have all been shown to reduce inflammatory responses. We evaluated effects of these compounds on alcohol intake. In order to facilitate comparison with previously published findings for some compounds, we employed similar schedules that were previously used for that compound.

Results: Gabapentin increased ethanol (EtOH) binge-like alcohol drinking in female HDID-1 and HS/NPT mice. Tesaglitazar and fenofibrate did not alter 2-bottle choice (2BC) drinking in male HDID-1 or HS/NPT mice. However, tesaglitazar had no effect on DID EtOH intake but reduced blood alcohol levels (BAL), and fenofibrate increased DID intake with no effects on BAL. CAPE had no effect on EtOH intake. Ibrutinib reduced intake in female HDID-1 in initial testing, but did not reduce intake in a second week of testing. Rolipram reduced DID intake and BALs in male and female HDID-1, HDID-2, and HS/NPT mice.

Conclusions: A number of compounds shown to reduce EtOH drinking in other models, and genotypes are not effective in HDID mice or their genetically heterogeneous founders, HS/NPT. The most promising compound was the PDE4 inhibitor, rolipram. These results highlight the importance of assessing generalizability when rigorously testing compounds for therapeutic development.
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http://dx.doi.org/10.1111/acer.14269DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7018604PMC
February 2020

Ethanol Conditioned Taste Aversion in High Drinking in the Dark Mice.

Brain Sci 2019 Jan 1;9(1). Epub 2019 Jan 1.

Portland Alcohol Research Center, Department of Behavioral Neuroscience, Oregon Health & Science University and VA Portland Health Care System, Portland, OR 97239, USA.

Two independent lines of High Drinking in the Dark (HDID-1, HDID-2) mice have been bred to reach high blood alcohol levels after a short period of binge-like ethanol drinking. Male mice of both lines were shown to have reduced sensitivity to develop a taste aversion to a novel flavor conditioned by ethanol injections as compared with their unselected HS/NPT founder stock. We have subsequently developed inbred variants of each line. The current experiments established that reduced ethanol-conditioned taste aversion is also seen in the inbred variants, in both males and females. In other experiments, we asked whether HDID mice would ingest sufficient doses of ethanol to lead to a conditioned taste aversion upon retest. Different manipulations were used to elevate consumption of ethanol on initial exposure. Access to increased ethanol concentrations, to multiple tubes of ethanol, and fluid restriction to increase thirst motivation all enhanced initial drinking of ethanol. Each condition led to reduced intake the next day, consistent with a mild conditioned taste aversion. These experiments support the conclusion that one reason contributing to the willingness of HDID mice to drink to the point of intoxication is a genetic insensitivity to the aversive effects of ethanol.
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http://dx.doi.org/10.3390/brainsci9010002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6356868PMC
January 2019

Pharmacogenetic Manipulation of the Nucleus Accumbens Alters Binge-Like Alcohol Drinking in Mice.

Alcohol Clin Exp Res 2018 05 18;42(5):879-888. Epub 2018 Apr 18.

Research and Development Service, Portland Veterans Affairs Medical Center, Portland, Oregon.

Background: Chronic alcohol intake leads to long-lasting changes in reward- and stress-related neuronal circuitry. The nucleus accumbens (NAc) is an integral component of this circuitry. Here, we investigate the effects of DREADDs (Designer Receptors Exclusively Activated by Designer Drugs) on neuronal activity in the NAc and binge-like drinking.

Methods: C57BL/6J mice were stereotaxically injected with AAV2 hSyn-HA hM3Dq, -hM4Di, or -eGFP bilaterally into NAc [core + shell, core or shell]. We measured clozapine-n-oxide (CNO)-induced changes in NAc activity and assessed binge-like ethanol (EtOH) or tastant/fluid intake in a limited access Drinking in the Dark (DID) schedule.

Results: We found that CNO increased NAc firing in hM3Dq positive cells and decreased firing in hM4Di cells, confirming the efficacy of these channels to alter neuronal activity both spatially and temporally. Increasing NAc core + shell activity decreased binge-like drinking without altering intake of other tastants. Increasing activity specifically in the NAc core reduced binge-like drinking, and decreasing activity in the NAc core increased drinking. Manipulation of NAc shell activity did not alter DID. Thus, we find that increasing activity in the entire NAc, or just the NAc core is sufficient to decrease binge drinking.

Conclusions: We conclude that the reduction in EtOH drinking is not due to general malaise, altered perception of taste, or reduced calorie-seeking. Furthermore, we provide the first evidence for bidirectional control of NAc core and binge-like drinking. These findings could have promising implications for treatment.
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http://dx.doi.org/10.1111/acer.13626DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6034712PMC
May 2018

Genome-Wide Expression Profiles Drive Discovery of Novel Compounds that Reduce Binge Drinking in Mice.

Neuropsychopharmacology 2018 05 18;43(6):1257-1266. Epub 2017 Dec 18.

The Waggoner Center for Alcohol and Addiction Research, The University of Texas at Austin, Austin, TX, USA.

Transcriptome-based drug discovery has identified new treatments for some complex diseases, but has not been applied to alcohol use disorder (AUD) or other psychiatric diseases, where there is a critical need for improved pharmacotherapies. High Drinking in the Dark (HDID-1) mice are a genetic model of AUD risk that have been selectively bred (from the HS/Npt line) to achieve intoxicating blood alcohol levels (BALs) after binge-like drinking. We compared brain gene expression of HDID-1 and HS/Npt mice, to determine a molecular signature for genetic risk for high intensity, binge-like drinking. Using multiple computational methods, we queried LINCS-L1000 (Library of Integrated Network-Based Cellular Signatures), a database containing gene expression signatures of thousands of compounds, to predict candidate drugs with the greatest potential to decrease alcohol consumption. Our analyses predicted novel compounds for testing, many with anti-inflammatory properties, providing further support for a neuroimmune mechanism of excessive alcohol drinking. We validated the top 2 candidates in vivo as a proof-of-concept. Terreic acid (a Bruton's tyrosine kinase inhibitor) and pergolide (a dopamine and serotonin receptor agonist) robustly reduced alcohol intake and BALs in HDID-1 mice, providing the first evidence for transcriptome-based drug discovery to target an addiction trait. Effective drug treatments for many psychiatric diseases are lacking, and the emerging tools and approaches outlined here offer researchers studying complex diseases renewed opportunities to discover new or repurpose existing compounds and expedite treatment options.
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http://dx.doi.org/10.1038/npp.2017.301DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5916369PMC
May 2018

NPAS2 Regulation of Anxiety-Like Behavior and GABAA Receptors.

Front Mol Neurosci 2017 3;10:360. Epub 2017 Nov 3.

Department of Psychiatry, University of Pittsburgh Medical Center, Pittsburgh, PA, United States.

Abnormal circadian rhythms and circadian genes are strongly associated with several psychiatric disorders. Neuronal PAS Domain Protein 2 (NPAS2) is a core component of the molecular clock that acts as a transcription factor and is highly expressed in reward- and stress-related brain regions such as the striatum. However, the mechanism by which NPAS2 is involved in mood-related behaviors is still unclear. We measured anxiety-like behaviors in mice with a global null mutation in (Npas2 null mutant mice) and found that Npas2 null mutant mice exhibit less anxiety-like behavior than their wild-type (WT) littermates (in elevated plus maze, light/dark box and open field assay). We assessed the effects of acute or chronic stress on striatal expression, and found that both stressors increased levels of . Moreover, knockdown of in the ventral striatum resulted in a similar reduction of anxiety-like behaviors as seen in the Npas2 null mutant mouse. Additionally, we identified genes as transcriptional targets of NPAS2, found that Npas2 null mutant mice exhibit reduced sensitivity to the GABAa positive allosteric modulator, diazepam and that knockdown of reduced expression and response to diazepam in the ventral striatum. These results: (1) implicate in the response to stress and the development of anxiety; and (2) provide functional evidence for the regulation of GABAergic neurotransmission by NPAS2 in the ventral striatum.
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http://dx.doi.org/10.3389/fnmol.2017.00360DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5675889PMC
November 2017

High Drinking in the Dark (HDID) mice are sensitive to the effects of some clinically relevant drugs to reduce binge-like drinking.

Pharmacol Biochem Behav 2017 09 5;160:55-62. Epub 2017 Aug 5.

Portland Alcohol Research Center, Department of Behavioral Neuroscience, Oregon Health & Science University, and VA Portland Health Care System, Portland, OR 97239, USA.

Background: There is a serious public health need for better understanding of alcohol use disorder disease mechanisms and for improved treatments. At this writing, only three drugs are approved by the Food and Drug Administration as medications to treat alcohol use disorders - disulfiram, naltrexone, and acamprosate. Binge drinking is a form of abusive alcohol drinking defined by the NIAAA as a drinking to blood alcohol levels (BALs)>0.08% during a period of approximately 2h. To model genetic risk for binge-like drinking, we have used selective breeding to create a unique animal model, High Drinking in the Dark (HDID) mice. Behavioral characterization of HDID mice has revealed that HDID mice exhibit behavioral impairment after drinking, withdrawal after a single binge-drinking session, and escalate their intake in response to induction of successive cycles of dependence. Notably, HDID mice do not exhibit altered tastant preference or alcohol clearance rates. We therefore asked whether drugs of known clinical relevance could modulate binge-like ethanol drinking in HDID mice, reasoning that this characterization of HDID responses should inform future use of this genetic animal model for screening and development of novel potential therapeutics.

Methods: We tested the efficacy of acamprosate and naltrexone to reduce binge-like drinking in HDID mice. Additionally, we tested the GABA receptor agonist, baclofen, based on recent pre-clinical and clinical studies demonstrating that it reduces alcohol drinking. We elected not to include disulfiram due to its more limited clinical usage. Mice were tested after acute doses of drugs in the limited-access Drinking in the Dark (DID) paradigm.

Results: HDID mice were sensitive to the effects of acamprosate and baclofen, but not naltrexone. Both drugs reduced binge-like drinking. However, naltrexone failed to reduce drinking in HDID mice. Thus, HDID mice may represent a useful model for screening novel compounds.
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http://dx.doi.org/10.1016/j.pbb.2017.08.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5603423PMC
September 2017

Functional Implications of the CLOCK 3111T/C Single-Nucleotide Polymorphism.

Front Psychiatry 2016 21;7:67. Epub 2016 Apr 21.

Department of Psychiatry and Translational Neuroscience Program, University of Pittsburgh School of Medicine , Pittsburgh, PA , USA.

Circadian rhythm disruptions are prominently associated with bipolar disorder (BD). Circadian rhythms are regulated by the molecular clock, a family of proteins that function together in a transcriptional-translational feedback loop. The CLOCK protein is a key transcription factor of this feedback loop, and previous studies have found that manipulations of the Clock gene are sufficient to produce manic-like behavior in mice (1). The CLOCK 3111T/C single-nucleotide polymorphism (SNP; rs1801260) is a genetic variation of the human CLOCK gene that is significantly associated with increased frequency of manic episodes in BD patients (2). The 3111T/C SNP is located in the 3'-untranslated region of the CLOCK gene. In this study, we sought to examine the functional implications of the human CLOCK 3111T/C SNP by transfecting a mammalian cell line (mouse embryonic fibroblasts isolated from Clock(-/-) knockout mice) with pcDNA plasmids containing the human CLOCK gene with either the T or C SNP at position 3111. We then measured circadian gene expression over a 24-h time period. We found that the CLOCK3111C SNP resulted in higher mRNA levels than the CLOCK 3111T SNP. Furthermore, we found that Per2, a transcriptional target of CLOCK, was also more highly expressed with CLOCK 3111C expression, indicating that the 3'-UTR SNP affects the expression, function, and stability of CLOCK mRNA.
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http://dx.doi.org/10.3389/fpsyt.2016.00067DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4838618PMC
May 2016

Commonalities and Distinctions Among Mechanisms of Addiction to Alcohol and Other Drugs.

Alcohol Clin Exp Res 2015 Oct;39(10):1863-77

Research & Development Service, Portland VA Medical Center, Portland, Oregon.

Background: Alcohol abuse is comorbid with abuse of many other drugs, some with similar pharmacology and others quite different. This leads to the hypothesis of an underlying, unitary dysfunctional neurobiological basis for substance abuse risk and consequences.

Methods: In this review, we discuss commonalities and distinctions of addiction to alcohol and other drugs. We focus on recent advances in preclinical studies using rodent models of drug self-administration.

Results: While there are specific behavioral and molecular manifestations common to alcohol, psychostimulant, opioid, and nicotine dependence, attempts to propose a unifying theory of the addictions inevitably face details where distinctions are found among classes of drugs.

Conclusions: For alcohol, versus other drugs of abuse, we discuss and compare advances in: (i) neurocircuitry important for the different stages of drug dependence; (ii) transcriptomics and genetical genomics; and (iii) enduring effects, noting in particular the contributions of behavioral genetics and animal models.
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http://dx.doi.org/10.1111/acer.12810DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4594192PMC
October 2015

Circadian clock genes: effects on dopamine, reward and addiction.

Alcohol 2015 Jun 8;49(4):341-9. Epub 2015 Jan 8.

University of Pittsburgh School of Medicine, Department of Psychiatry, 450 Technology Drive, Suite 223, Pittsburgh, PA 15219, USA. Electronic address:

Addiction is a widespread public health issue with social and economic ramifications. Substance abuse disorders are often accompanied by disruptions in circadian rhythms including sleep/wake cycles, which can exacerbate symptoms of addiction and dependence. Additionally, genetic disturbance of circadian molecular mechanisms can predispose some individuals to substance abuse disorders. In this review, we will discuss how circadian genes can regulate midbrain dopaminergic activity and subsequently, drug intake and reward. We will also suggest future directions for research on circadian genes and drugs of abuse.
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http://dx.doi.org/10.1016/j.alcohol.2014.09.034DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4457686PMC
June 2015

Direct regulation of diurnal Drd3 expression and cocaine reward by NPAS2.

Biol Psychiatry 2015 Mar 13;77(5):425-433. Epub 2014 Aug 13.

Department of Psychiatry and Translational Neuroscience Program, University of Pittsburgh School of Medicine, Pittsburgh, PA 15219.

Background: Circadian gene disruptions are associated with the development of psychiatric disorders, including addiction. However, the mechanisms by which circadian genes regulate reward remain poorly understood.

Methods: We used mice with a mutation in Npas2 and adeno-associated virus-short hairpin RNA mediated knockdown of Npas2 and Clock in the nucleus accumbens (NAc). We performed conditioned place preference assays. We utilized cell sorting quantitative real-time polymerase chain reaction, and chromatin immunoprecipitation followed by deep sequencing.

Results: Npas2 mutants exhibit decreased sensitivity to cocaine reward, which is recapitulated with a knockdown of neuronal PAS domain protein 2 (NPAS2) specifically in the NAc, demonstrating the importance of NPAS2 in this region. Interestingly, reducing circadian locomotor output cycles kaput (CLOCK) (a homologue of NPAS2) in the NAc had no effect, suggesting an important distinction in NPAS2 and CLOCK function. Furthermore, we found that NPAS2 expression is restricted to Drd1 expressing neurons while CLOCK is ubiquitous. Moreover, NPAS2 and CLOCK have distinct temporal patterns of DNA binding, and we identified novel and unique binding sites for each protein. We identified the Drd3 dopamine receptor as a direct transcriptional target of NPAS2 and found that NPAS2 knockdown in the NAc disrupts its diurnal rhythm in expression. Chronic cocaine treatment likewise disrupts the normal rhythm in Npas2 and Drd3 expression in the NAc, which may underlie behavioral plasticity in response to cocaine.

Conclusions: Together, these findings identify an important role for the circadian protein, NPAS2, in the NAc in the regulation of dopamine receptor expression and drug reward.
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http://dx.doi.org/10.1016/j.biopsych.2014.07.030DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4315729PMC
March 2015

Chronic self-administration of alcohol results in elevated ΔFosB: comparison of hybrid mice with distinct drinking patterns.

BMC Neurosci 2012 Oct 29;13:130. Epub 2012 Oct 29.

Waggoner Center for Alcoholism and Addiction Research, Institute for Neuroscience, University of Texas at Austin, Austin, TX 78712, USA.

Background: The inability to reduce or regulate alcohol intake is a hallmark symptom for alcohol use disorders. Research on novel behavioral and genetic models of experience-induced changes in drinking will further our knowledge on alcohol use disorders. Distinct alcohol self-administration behaviors were previously observed when comparing two F1 hybrid strains of mice: C57BL/6J x NZB/B1NJ (BxN) show reduced alcohol preference after experience with high concentrations of alcohol and periods of abstinence while C57BL/6J x FVB/NJ (BxF) show sustained alcohol preference. These phenotypes are interesting because these hybrids demonstrate the occurrence of genetic additivity (BxN) and overdominance (BxF) in ethanol intake in an experience dependent manner. Specifically, BxF exhibit sustained alcohol preference and BxN exhibit reduced alcohol preference after experience with high ethanol concentrations; however, experience with low ethanol concentrations produce sustained alcohol preference for both hybrids. In the present study, we tested the hypothesis that these phenotypes are represented by differential production of the inducible transcription factor, ΔFosB, in reward, aversion, and stress related brain regions.

Results: Changes in neuronal plasticity (as measured by ΔFosB levels) were experience dependent, as well as brain region and genotype specific, further supporting that neuronal circuitry underlies motivational aspects of ethanol consumption. BxN mice exhibiting reduced alcohol preference had lower ΔFosB levels in the Edinger-Westphal nucleus than mice exhibiting sustained alcohol preference, and increased ΔFosB levels in central medial amygdala as compared with control mice. BxN mice showing sustained alcohol preference exhibited higher ΔFosB levels in the ventral tegmental area, Edinger-Westphal nucleus, and amygdala (central and lateral divisions). Moreover, in BxN mice ΔFosB levels in the Edinger-Westphal nucleus and ventral tegmental regions significantly positively correlated with ethanol preference and intake. Additionally, hierarchical clustering analysis revealed that many ethanol-naïve mice with overall low ΔFosB levels are in a cluster, whereas many mice displaying sustained alcohol preference with overall high ΔFosB levels are in a cluster together.

Conclusions: By comparing and contrasting two alcohol phenotypes, this study demonstrates that the reward- and stress-related circuits (including the Edinger-Westphal nucleus, ventral tegmental area, amygdala) undergo significant plasticity that manifests as reduced alcohol preference.
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http://dx.doi.org/10.1186/1471-2202-13-130DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3504532PMC
October 2012

Specific role of VTA dopamine neuronal firing rates and morphology in the reversal of anxiety-related, but not depression-related behavior in the ClockΔ19 mouse model of mania.

Neuropsychopharmacology 2011 Jun 23;36(7):1478-88. Epub 2011 Mar 23.

Department of Psychiatry, UT Southwestern Medical Center, Dallas, TX 75390-9070, USA.

Lithium has been used extensively for mood stabilization, and it is particularly efficacious in the treatment of bipolar mania. Like other drugs used in the treatment of psychiatric diseases, it has little effect on the mood of healthy individuals. Our previous studies found that mice with a mutation in the Clock gene (ClockΔ19) have a complete behavioral profile that is very similar to human mania, which can be reversed with chronic lithium treatment. However, the cellular and physiological effects that underlie its targeted therapeutic efficacy remain unknown. Here we find that ClockΔ19 mice have an increase in dopaminergic activity in the ventral tegmental area (VTA), and that lithium treatment selectively reduces the firing rate in the mutant mice with no effect on activity in wild-type mice. Furthermore, lithium treatment reduces nucleus accumbens (NAc) dopamine levels selectively in the mutant mice. The increased dopaminergic activity in the Clock mutants is associated with cell volume changes in dopamine neurons, which are also rescued by lithium treatment. To determine the role of dopaminergic activity and morphological changes in dopamine neurons in manic-like behavior, we manipulated the excitability of these neurons by overexpressing an inwardly rectifying potassium channel subunit (Kir2.1) selectively in the VTA of ClockΔ19 mice and wild-type mice using viral-mediated gene transfer. Introduction of this channel mimics the effects of lithium treatment on the firing rate of dopamine neurons in ClockΔ19 mice and leads to a similar change in dopamine cell volume. Furthermore, reduction of dopaminergic firing rates in ClockΔ19 animals results in a normalization of locomotor- and anxiety-related behavior that is very similar to lithium treatment; however, it is not sufficient to reverse depression-related behavior. These results suggest that abnormalities in dopamine cell firing and associated morphology underlie alterations in anxiety-related behavior in bipolar mania, and that the therapeutic effects of lithium come from a reversal of these abnormal phenotypes.
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http://dx.doi.org/10.1038/npp.2011.33DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3096816PMC
June 2011

The vermicelli and capellini handling tests: simple quantitative measures of dexterous forepaw function in rats and mice.

J Vis Exp 2010 Jul 21(41). Epub 2010 Jul 21.

Institute for Neuroscience, University of Texas at Austin, TX, USA.

Previous characterizations of rodent eating behavior have revealed that they use coordinated forepaw movements to manipulate food pieces. We have extended upon this work to develop a simple quantitative measure of forepaw dexterity that is sensitive to lateralized impairments and age-dependent changes. Rodents learn skillful forepaw and digit movements to manage thin pasta pieces, which they eagerly consume. We have previously described methods for quantifying vermicelli handling in rats and showed that the measures are very sensitive to forelimb impairments resulting from unilateral ischemic lesions, middle cerebral artery occlusions and unilateral striatal dopamine depletion [Allred, R.P., Adkins, D.L., Woodlee, M.T., Husbands, L.C., Maldonado M.A., Kane, J.R., Schallert, T. & Jones, T.A. The Vermicelli Handling Test: a simple quantitative measure of dexterous forepaw function in rats. J. Neurosci. Methods 170, 229-244 (2008)]. Here we present a more detailed protocol for this test in rats and compare it with a newly developed version for mice, the Capellini Handling Test. Rats and mice are videotaped while handling short lengths of uncooked vermicelli or capellini pasta, respectively, with a camera positioned to optimize the view of paw movements. Slow motion video playback allows for the identification of forepaw adjustments, defined as any distinct removal and replacement of the paw, or of any number of digits, on the pasta piece after eating commences. Forepaw adjustments per piece are averaged over trials per each testing session. Repeated testing permits sensitive quantitative analysis of changes in forepaw dexterity over time. Protocols for pre-testing habituation and handling practice, as well as procedures for characterizing atypical handling patterns, are described. Because rats and mice perform the pasta handling tests slightly differently, species-specific differences in administration and scoring of these tests are highlighted. All animal use was in accordance with protocols approved by the University of Texas at Austin Animal Care and Use Committee.
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http://dx.doi.org/10.3791/2076DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3039868PMC
July 2010