1 4161 121 MECP2 REGULATES ETHANOL SENSITIVITY AND INTAKE. WE HAVE INVESTIGATED THE EXPRESSION OF CHROMATIN-REGULATING GENES IN THE PREFRONTAL CORTEX AND IN THE SHELL SUBDIVISION OF THE NUCLEUS ACCUMBENS DURING PROTRACTED WITHDRAWAL IN MICE WITH INCREASED ETHANOL DRINKING AFTER CHRONIC INTERMITTENT ETHANOL (CIE) VAPOR EXPOSURE AND IN MICE WITH A HISTORY OF NON-DEPENDENT DRINKING. WE OBSERVED THAT THE METHYL-CPG BINDING PROTEIN 2 (MECP2) WAS ONE OF THE FEW CHROMATIN-REGULATING GENES TO BE DIFFERENTIALLY REGULATED BY A HISTORY OF DEPENDENCE. AS MECP2 HAS THE POTENTIAL OF ACTING AS A BROAD GENE REGULATOR, WE INVESTIGATED SENSITIVITY TO ETHANOL AND ETHANOL DRINKING IN MECP2(308/) (Y) MICE, WHICH HARBOR A TRUNCATED MECP2 ALLELE BUT HAVE A MILDER PHENOTYPE THAN MECP2 NULL MICE. WE OBSERVED THAT MECP2(308/) (Y) MICE WERE MORE SENSITIVE TO ETHANOL'S STIMULATORY AND SEDATIVE EFFECTS THAN WILD-TYPE (WT) MICE, DRANK LESS ETHANOL IN A LIMITED ACCESS 2 BOTTLE CHOICE PARADIGM AND DID NOT SHOW INCREASED DRINKING AFTER INDUCTION OF DEPENDENCE WITH EXPOSURE TO CIE VAPORS. ALCOHOL METABOLISM DID NOT DIFFER IN MECP2(308/) (Y) AND WT MICE. ADDITIONALLY, MECP2(308/) (Y) MICE DID NOT DIFFER FROM WT MICE IN ETHANOL PREFERENCE IN A 24-HOUR PARADIGM NOR IN THEIR INTAKE OF GRADED SOLUTIONS OF SACCHARIN OR QUININE, SUGGESTING THAT THE MECP2(308/) (Y) MUTATION DID NOT ALTER TASTE FUNCTION. LASTLY, USING THE GENE SET ENRICHMENT ANALYSIS ALGORITHM, WE FOUND A SIGNIFICANT OVERLAP IN THE GENES REGULATED BY ALCOHOL AND BY MECP2. TOGETHER, THESE RESULTS SUGGEST THAT MECP2 CONTRIBUTES TO THE REGULATION OF ETHANOL SENSITIVITY AND DRINKING. 2014 2 2156 43 EPIGENETIC MECHANISMS ARE INVOLVED IN THE REGULATION OF ETHANOL CONSUMPTION IN MICE. BACKGROUND: REPEATED ALCOHOL EXPOSURE IS KNOWN TO INCREASE SUBSEQUENT ETHANOL CONSUMPTION IN MICE. HOWEVER, THE UNDERLYING MECHANISMS HAVE NOT BEEN FULLY ELUCIDATED. ONE POSTULATED MECHANISM INVOLVES EPIGENETIC MODIFICATIONS, INCLUDING HISTONE MODIFICATIONS AND DNA METHYLATION OF RELEVANT GENES SUCH AS NR2B OR BDNF. METHODS: TO INVESTIGATE THE ROLE OF EPIGENETIC MECHANISMS IN THE DEVELOPMENT OF ALCOHOL DRINKING BEHAVIOR, AN ESTABLISHED CHRONIC INTERMITTENT ETHANOL EXPOSURE REINFORCED ETHANOL DRINKING MOUSE MODEL WITH VAPOR INHALATION OVER TWO 9-DAY TREATMENT REGIMENS WAS USED. THE DNA METHYLTRANSFERASE INHIBITOR, 5-AZACYTIDINE OR THE HISTONE DEACETYLASE INHIBITOR, TRICHOSTATIN A WAS ADMINISTERED (INTRAPERITONEALLY) TO C57BL/6 MICE 30 MIN BEFORE DAILY EXPOSURE TO CHRONIC INTERMITTENT ETHANOL. CHANGES IN ETHANOL CONSUMPTION WERE MEASURED USING THE 2-BOTTLE CHOICE TEST. RESULTS: THE RESULTS INDICATED THAT SYSTEMIC ADMINISTRATION OF TRICHOSTATIN A (2.5 MICROG/G) FACILITATED CHRONIC INTERMITTENT ETHANOL-INDUCED ETHANOL DRINKING, BUT SYSTEMIC ADMINISTRATION OF 5-AZACYTIDINE (2 MICROG/G) DID NOT CAUSE THE SAME EFFECT. HOWEVER, WHEN 5-AZACYTIDINE WAS ADMINISTERED BY INTRACEREBROVENTRICULAR INJECTION, IT FACILITATED CHRONIC INTERMITTENT ETHANOL-INDUCED ETHANOL DRINKING. FURTHERMORE, THE INCREASED DRINKING CAUSED BY CHRONIC INTERMITTENT ETHANOL WAS PREVENTED BY INJECTION OF A METHYL DONOR, S-ADENOSYL-L-METHIONINE. TO PROVIDE EVIDENCE THAT CHRONIC INTERMITTENT ETHANOL- OR TRICHOSTATIN A-INDUCED DNA DEMETHYLATION AND HISTONE MODIFICATIONS OF THE NR2B PROMOTER MAY UNDERLIE THE ALTERED ETHANOL CONSUMPTION, WE EXAMINED EPIGENETIC MODIFICATIONS AND NR2B EXPRESSION IN THE PREFRONTAL CORTEX OF THESE MICE. CHRONIC INTERMITTENT ETHANOL OR TRICHOSTATIN A DECREASED DNA METHYLATION AND INCREASED HISTONE ACETYLATION IN THE NR2B GENE PROMOTER, AS WELL AS MRNA LEVELS OF NR2B IN THESE MICE. CONCLUSIONS: TAKEN TOGETHER, THESE RESULTS INDICATE THAT EPIGENETIC MODIFICATIONS ARE INVOLVED IN REGULATING ETHANOL DRINKING BEHAVIOR, PARTIALLY THROUGH ALTERING NR2B EXPRESSION. 2014 3 5177 47 PREFRONTAL CORTEX EXPRESSION OF CHROMATIN MODIFIER GENES IN MALE WSP AND WSR MICE CHANGES ACROSS ETHANOL DEPENDENCE, WITHDRAWAL, AND ABSTINENCE. ALCOHOL-USE DISORDER (AUD) IS A RELAPSING DISORDER ASSOCIATED WITH EXCESSIVE ETHANOL CONSUMPTION. RECENT STUDIES SUPPORT THE INVOLVEMENT OF EPIGENETIC MECHANISMS IN THE DEVELOPMENT OF AUD. STUDIES CARRIED OUT SO FAR HAVE FOCUSED ON A FEW SPECIFIC EPIGENETIC MODIFICATIONS. THE GOAL OF THIS PROJECT WAS TO INVESTIGATE GENE EXPRESSION CHANGES OF EPIGENETIC REGULATORS THAT MEDIATE A BROAD ARRAY OF CHROMATIN MODIFICATIONS AFTER CHRONIC ALCOHOL EXPOSURE, CHRONIC ALCOHOL EXPOSURE FOLLOWED BY 8 H WITHDRAWAL, AND CHRONIC ALCOHOL EXPOSURE FOLLOWED BY 21 DAYS OF ABSTINENCE IN WITHDRAWAL-RESISTANT (WSR) AND WITHDRAWAL SEIZURE-PRONE (WSP) SELECTED MOUSE LINES. WE FOUND THAT CHRONIC VAPOR EXPOSURE TO HIGHLY INTOXICATING LEVELS OF ETHANOL ALTERS THE EXPRESSION OF SEVERAL CHROMATIN REMODELING GENES MEASURED BY QUANTITATIVE PCR ARRAY ANALYSES. THE IDENTIFIED EFFECTS WERE INDEPENDENT OF SELECTED LINES, WHICH, HOWEVER, DISPLAYED BASELINE DIFFERENCES IN EPIGENETIC GENE EXPRESSION. WE REPORTED DYSREGULATION IN THE EXPRESSION OF GENES INVOLVED IN HISTONE ACETYLATION, DEACETYLATION, LYSINE AND ARGININE METHYLATION AND UBIQUITINATIONHYLATION DURING CHRONIC ETHANOL EXPOSURE AND WITHDRAWAL, BUT NOT AFTER 21 DAYS OF ABSTINENCE. ETHANOL-INDUCED CHANGES ARE CONSISTENT WITH DECREASED HISTONE ACETYLATION AND WITH DECREASED DEPOSITION OF THE PERMISSIVE UBIQUITINATION MARK H2BK120UB, ASSOCIATED WITH REDUCED TRANSCRIPTION. ON THE OTHER HAND, ETHANOL-INDUCED CHANGES IN THE EXPRESSION OF GENES INVOLVED IN HISTONE LYSINE METHYLATION ARE CONSISTENT WITH INCREASED TRANSCRIPTION. THE NET RESULT OF THESE MODIFICATIONS ON GENE EXPRESSION IS LIKELY TO DEPEND ON THE COMBINATION OF THE SPECIFIC HISTONE TAIL MODIFICATIONS PRESENT AT A GIVEN TIME ON A GIVEN PROMOTER. SINCE ALCOHOL DOES NOT MODULATE GENE EXPRESSION UNIDIRECTIONALLY, IT IS NOT SURPRISING THAT ALCOHOL DOES NOT UNIDIRECTIONALLY ALTER CHROMATIN STRUCTURE TOWARD A CLOSED OR OPEN STATE, AS SUGGESTED BY THE RESULTS OF THIS STUDY. 2017 4 4931 36 PATERNAL ALCOHOL EXPOSURE REDUCES ALCOHOL DRINKING AND INCREASES BEHAVIORAL SENSITIVITY TO ALCOHOL SELECTIVELY IN MALE OFFSPRING. ALCOHOL USE DISORDER (AUD) IS HERITABLE, BUT THE GENETIC BASIS FOR THIS DISEASE REMAINS POORLY UNDERSTOOD. ALTHOUGH NUMEROUS GENE VARIANTS HAVE BEEN ASSOCIATED WITH AUD, THESE VARIANTS ACCOUNT FOR ONLY A SMALL FRACTION OF THE TOTAL RISK. THE IDEA OF INHERITANCE OF ACQUIRED CHARACTERISTICS, I.E. "EPIGENETIC INHERITANCE," IS RE-EMERGING AS A PROVEN ADJUNCT TO TRADITIONAL MODES OF GENETIC INHERITANCE. WE HYPOTHESIZED THAT ALCOHOL DRINKING AND NEUROBIOLOGICAL SENSITIVITY TO ALCOHOL ARE INFLUENCED BY ANCESTRAL ALCOHOL EXPOSURE. TO TEST THIS HYPOTHESIS, WE EXPOSED MALE MICE TO CHRONIC VAPOR ETHANOL OR CONTROL CONDITIONS, MATED THEM TO ETHANOL-NAIVE FEMALES, AND TESTED ADULT OFFSPRING FOR ETHANOL DRINKING, ETHANOL-INDUCED BEHAVIORS, GENE EXPRESSION, AND DNA METHYLATION. WE FOUND THAT ETHANOL-SIRED MALE OFFSPRING HAD REDUCED ETHANOL PREFERENCE AND CONSUMPTION, ENHANCED SENSITIVITY TO THE ANXIOLYTIC AND MOTOR-ENHANCING EFFECTS OF ETHANOL, AND INCREASED BDNF EXPRESSION IN THE VENTRAL TEGMENTAL AREA (VTA) COMPARED TO CONTROL-SIRED MALE OFFSPRING. THERE WERE NO DIFFERENCES AMONG ETHANOL- AND CONTROL-SIRED FEMALE OFFSPRING ON THESE ASSAYS. ETHANOL EXPOSURE ALSO DECREASED DNA METHYLATION AT THE BDNFAEPROMOTER OF SIRE'S GERM CELLS AND HYPOMETHYLATION WAS MAINTAINED IN THE VTA OF BOTH MALE AND FEMALE ETHANOL-SIRED OFFSPRING. OUR FINDINGS SHOW THAT PATERNAL ALCOHOL EXPOSURE IS A PREVIOUSLY UNRECOGNIZED REGULATOR OF ALCOHOL DRINKING AND BEHAVIORAL SENSITIVITY TO ALCOHOL IN MALE, BUT NOT FEMALE, OFFSPRING. PATERNAL ALCOHOL EXPOSURE ALSO INDUCES EPIGENETIC ALTERATIONS (DNA HYPOMETHYLATION) AND GENE EXPRESSION CHANGES THAT PERSIST IN THE VTA OF OFFSPRING. THESE RESULTS PROVIDE NEW INSIGHT INTO THE INHERITANCE AND DEVELOPMENT OF ALCOHOL DRINKING BEHAVIORS. 2014 5 6801 45 [EPIGENETIC MECHANISMS AND ALCOHOL USE DISORDERS: A POTENTIAL THERAPEUTIC TARGET]. ALCOHOL USE DISORDER IS A DEVASTATING ILLNESS WITH A PROFOUND HEALTH IMPACT, AND ITS DEVELOPMENT IS DEPENDENT ON BOTH GENETIC AND ENVIRONMENTAL FACTORS. THIS DISEASE OCCURS OVER TIME AND REQUIRES CHANGES IN BRAIN GENE EXPRESSION. THERE IS CONVERGING EVIDENCE SUGGESTING THAT THE EPIGENETIC PROCESSES MAY PLAY A ROLE IN THE ALCOHOL-INDUCED GENE REGULATIONS AND BEHAVIOR SUCH AS THE INTERVENTION OF DNA METHYLATION AND HISTONE ACETYLATION. HISTONE ACETYLATION, LIKE HISTONE METHYLATION, IS A HIGHLY DYNAMIC PROCESS REGULATED BY TWO CLASSES OF ENZYMES: HISTONE ACETYLTRANSFERASES AND HISTONE DEACETYLASES (HDACS). TO DATE, 18 HUMAN HDAC ISOFORMS HAVE BEEN CHARACTERIZED, AND BASED ON THEIR SEQUENCE HOMOLOGIES AND COFACTOR DEPENDENCIES, THEY HAVE BEEN PHYLOGENETICALLY CATEGORIZED INTO 4 MAIN CLASSES: CLASSES I, II (A AND B), III, AND IV. IN THE BRAIN, EXPRESSION OF THE DIFFERENT CLASSES OF HDACS VARIES BETWEEN CELL TYPES AND ALSO IN THEIR SUBCELLULAR LOCALIZATION (NUCLEUS AND/OR CYTOSOL). FURTHERMORE, WE RECENTLY SHOWED THAT A SINGLE ETHANOL EXPOSURE INHIBITS HDAC ACTIVITY AND INCREASES BOTH H3 AND H4 HISTONE ACETYLATION WITHIN THE AMYGDALA OF RATS. IN THE BRAIN OF ALCOHOLIC PATIENTS, ETHANOL HAS BEEN SHOWN TO INDUCE HISTONE-RELATED AND DNA METHYLATION EPIGENETIC CHANGES IN SEVERAL REWARD REGIONS INVOLVED IN REWARD PROCESSES SUCH AS HIPPOCAMPUS, PREFRONTAL CORTEX, AND AMYGDALA. WE RECENTLY DEMONSTRATED ALTERATION OF HISTONE H3 ACETYLATION LEVELS IN SEVERAL BRAIN REGIONS FROM THE REWARD CIRCUIT OF RATS MADE DEPENDENT TO ALCOHOL AFTER CHRONIC AND INTERMITTENT EXPOSURE TO ETHANOL VAPOR. IN NEURONAL CELL LINE CULTURE, ETHANOL WAS SHOWN TO INDUCE HDAC EXPRESSION. IN MOUSE AND RAT BRAIN, NUMEROUS STUDIES REPORTED EPIGENETIC ALTERATIONS FOLLOWING ETHANOL EXPOSURE. WE ALSO DEMONSTRATED THAT BOTH THE EXPRESSION OF GENES AND THE ACTIVITY OF ENZYMES INVOLVED IN EPIGENETIC MECHANISMS ARE CHANGED AFTER REPEATED ADMINISTRATIONS OF ETHANOL IN MICE SENSITIZED TO THE MOTOR STIMULANT EFFECT OF ETHANOL (A MODEL OF DRUG-INDUCED NEUROPLASTICITY). NUMEROUS STUDIES HAVE SHOWN THAT HDAC INHIBITORS ARE ABLE TO COUNTER ETHANOL-INDUCED BEHAVIORS AND THE ETHANOL-INDUCED CHANGES IN THE LEVELS OF HDAC AND/OR LEVELS OF ACETYLATED HDAC. FOR EXAMPLE, TRICHOSTATIN A (TSA) TREATMENT CAUSED THE REVERSAL OF ETHANOL-INDUCED TOLERANCE, ANXIETY, AND ETHANOL DRINKING BY INHIBITING HDAC ACTIVITY, THEREBY INCREASING HISTONE ACETYLATION IN THE AMYGDALA OF RATS. ANOTHER STUDY DEMONSTRATED THAT TSA PREVENTED THE DEVELOPMENT OF ETHANOL WITHDRAWAL INDUCED ANXIETY IN RATS BY RESCUING DEFICITS IN HISTONE ACETYLATION INDUCED BY INCREASED HDAC ACTIVITY IN THE AMYGDALA. WE HAVE DEMONSTRATED THAT TREATMENT WITH THE HDAC INHIBITOR SODIUM BUTYRATE BLOCKS BOTH THE DEVELOPMENT AND THE EXPRESSION OF ETHANOL-INDUCED BEHAVIORAL SENSITIZATION IN MICE. IN THIS CONTEXT, CONVERGING EVIDENCE INDICATES THAT HDAC INHIBITORS COULD BE USEFUL IN COUNTERACTING ETHANOL-INDUCED GENE REGULATIONS VIA EPIGENETIC MECHANISMS, THAT IS, HDAC INHIBITORS COULD AFFECT DIFFERENT ACETYLATION SITES AND MAY ALSO ALTER THE EXPRESSION OF DIFFERENT GENES THAT COULD IN TURN COUNTERACT THE EFFECT OF ETHANOL. RECENT WORK IN RODENTS HAS SHOWN THAT SYSTEMIC ADMINISTRATION OF PAN HDAC CLASS I AND II INHIBITORS, TSA AND N-HYDROXY-N-PHENYL-OCTANEDIAMIDE [SUBEROYLANILIDE HYDROXAMIC ACID] (SAHA), AND OF THE MORE SELECTIVE INHIBITOR (MAINLY HDAC1 AND HDAC9) MS-275, DECREASE BINGE-LIKE ALCOHOL DRINKING IN MICE. SAHA SELECTIVELY REDUCED ETHANOL OPERANT SELF-ADMINISTRATION AND SEEKING IN RATS. OUR PREVIOUS STUDY REVEALED THAT MS-275 STRONGLY DECREASED OPERANT ETHANOL SELF-ADMINISTRATION IN ALCOHOL-DEPENDENT RATS WHEN ADMINISTERED 30 MINUTES BEFORE THE SESSION AT THE SECOND DAY OF INJECTION. WE ALSO DEMONSTRATED THAT INTRA-CEREBRO-VENTRICULAR INFUSION OF MS-275 INCREASES ACETYLATION OF HISTONE 4 WITHIN THE NUCLEUS ACCUMBENS AND THE DORSOLATERAL STRIATUM, ASSOCIATED TO A DECREASE IN ETHANOL SELF-ADMINISTRATION BY ABOUT 75%. MS-275 ALSO DIMINISHED BOTH THE MOTIVATION TO CONSUME ETHANOL (25% DECREASE), RELAPSE (BY ABOUT 50%) AND POSTPONED REACQUISITION AFTER ABSTINENCE. BOTH LITERATURE AND SEVERAL OF OUR STUDIES STRONGLY SUPPORT THE POTENTIAL THERAPEUTIC INTEREST OF TARGETING EPIGENETIC MECHANISMS IN EXCESSIVE ALCOHOL DRINKING AND STRENGTHEN THEINTEREST OF FOCUSING ON SPECIFIC ISOFORMS OF HISTONE DEACETYLASES. 2017 6 5445 44 REPEATED VAPOR ETHANOL EXPOSURE INDUCES TRANSIENT HISTONE MODIFICATIONS IN THE BRAIN THAT ARE MODIFIED BY GENOTYPE AND BRAIN REGION. BACKGROUND: EMERGING RESEARCH IMPLICATES ETHANOL (ETOH)-INDUCED EPIGENETIC MODIFICATIONS IN REGULATING GENE EXPRESSION AND ETOH CONSUMPTION. HOWEVER, CONSENSUS ON SPECIFIC EPIGENETIC MODIFICATIONS INDUCED BY ETOH HAS NOT YET EMERGED, MAKING IT CHALLENGING TO IDENTIFY MECHANISMS AND DEVELOP TARGETED TREATMENTS. WE HYPOTHESIZED THAT CHRONIC INTERMITTENT ETOH (CIE) INDUCES PERSISTENT CHANGES IN HISTONE MODIFICATIONS ACROSS THE CEREBRAL CORTEX (CCX), NUCLEUS ACCUMBENS (NAC), AND PREFRONTAL CORTEX (PFC), AND THAT THESE HISTONE MODIFICATIONS ARE ALTERED IN A KNOCK-IN MOUSE STRAIN WITH ALTERED SENSITIVITY TO ETOH. METHODS: C57BL/6J (B6) MICE AND ALPHA1SHLA KNOCKIN MICE ON A B6 BACKGROUND WERE EXPOSED TO 16 H OF VAPOR ETOH OR ROOM AIR FOLLOWED BY 8 H OF ROOM AIR FOR 4 CONSECUTIVE DAYS AND SACRIFICED AT MULTIPLE TIME POINTS UP TO 72 H FOLLOWING EXPOSURE. HISTONE MODIFICATIONS WERE ASSESSED USING WESTERN BLOT AND DOT BLOT. RT-QPCR WAS USED TO STUDY EXPRESSION OF CHROMATIN MODIFYING ENZYMES IN NAC AND PFC. RESULTS: IN NAC, CIE SIGNIFICANTLY INCREASED ACETYLATION OF HISTONE SUBUNIT H3 AT LYSINE 9 (H3K9AC) BUT NOT LYSINE 14 (H3K14AC) OR LYSINE 27 (H3K27AC). IN PFC, CIE SIGNIFICANTLY INCREASED H3K9AC BUT NOT H3K14 OR H3K27AC. THERE WERE NO SIGNIFICANT CHANGES AT 8 OR 72 H AFTER ETOH EXPOSURE IN EITHER NAC OR PFC. CIE WAS ALSO ASSOCIATED WITH INCREASED EXPRESSION OF KAT2B, KAT5, AND TET1 IN NAC BUT NOT PFC. IN CCX, CIE HAD A SIGNIFICANT EFFECT ON LEVELS OF H3K18AC; THERE WAS ALSO A SIGNIFICANT EFFECT OF THE ALPHA1SHLA MUTATION ON LEVELS OF H3K27ME3, H3K14AC, AND H3K18AC AS WELL AS A TREND FOR H3S10PK14AC. CONCLUSIONS: THE ETOH-INDUCED HISTONE MODIFICATIONS OBSERVED WERE TRANSIENT AND VARIED SIGNIFICANTLY BETWEEN BRAIN REGIONS. A GENETIC MUTATION THAT ALTERED SENSITIVITY TO ETOH WAS ASSOCIATED WITH ALTERED INDUCTION OF HISTONE MODIFICATIONS DURING CIE. THESE RESULTS HAVE IMPLICATIONS FOR STUDYING ETOH-INDUCED HISTONE MODIFICATIONS AND ETOH SENSITIVITY. 2015 7 4150 38 MECHANISTIC INSIGHTS INTO EPIGENETIC MODULATION OF ETHANOL CONSUMPTION. THERE IS GROWING EVIDENCE THAT SMALL-MOLECULE INHIBITORS OF EPIGENETIC MODULATORS, SUCH AS HISTONE DEACETYLASES (HDAC) AND DNA METHYLTRANSFERASES (DNMT), CAN REDUCE VOLUNTARY ETHANOL CONSUMPTION IN ANIMAL MODELS, BUT MOLECULAR AND CELLULAR PROCESSES UNDERLYING THIS BEHAVIORAL EFFECT ARE POORLY UNDERSTOOD. WE USED C57BL/6J MALE MICE TO INVESTIGATE THE EFFECTS OF TWO FDA-APPROVED DRUGS, DECITABINE (A DNMT INHIBITOR) AND SAHA (AN HDAC INHIBITOR), ON ETHANOL CONSUMPTION USING TWO TESTS: BINGE-LIKE DRINKING IN THE DARK (DID) AND CHRONIC INTERMITTENT EVERY OTHER DAY (EOD) DRINKING. DECITABINE BUT NOT SAHA REDUCED ETHANOL CONSUMPTION IN BOTH TESTS. WE FURTHER INVESTIGATED DECITABINE'S EFFECTS ON THE BRAIN'S REWARD PATHWAY BY GENE EXPRESSION PROFILING IN THE VENTRAL TEGMENTAL AREA (VTA), USING RNA SEQUENCING AND ELECTROPHYSIOLOGICAL RECORDINGS FROM VTA DOPAMINERGIC NEURONS. DECITABINE-INDUCED DECREASES IN EOD DRINKING WERE ASSOCIATED WITH GLOBAL CHANGES IN GENE EXPRESSION, IMPLICATING REGULATION OF CEREBRAL BLOOD FLOW, EXTRACELLULAR MATRIX ORGANIZATION, AND NEUROIMMUNE FUNCTIONS IN DECITABINE ACTIONS. IN ADDITION, AN IN VIVO ADMINISTRATION OF DECITABINE SHORTENED ETHANOL-INDUCED EXCITATION OF VTA DOPAMINERGIC NEURONS IN VITRO, SUGGESTING THAT DECITABINE REDUCES ETHANOL DRINKING VIA CHANGES IN THE REWARD PATHWAY. TAKEN TOGETHER, OUR DATA SUGGEST A CONTRIBUTION OF BOTH NEURONAL AND NON-NEURONAL MECHANISMS IN THE VTA IN THE REGULATION OF ETHANOL CONSUMPTION. DECITABINE AND OTHER EPIGENETIC COMPOUNDS HAVE BEEN APPROVED FOR CANCER TREATMENT, AND UNDERSTANDING THEIR MECHANISMS OF ACTIONS IN THE BRAIN MAY ASSIST IN REPURPOSING THESE DRUGS AND DEVELOPING NOVEL THERAPIES FOR CENTRAL DISORDERS, INCLUDING DRUG ADDICTION. 2017 8 894 29 CHRONIC ETHANOL FEEDING ALTERS HEPATOCYTE MEMORY WHICH IS NOT ALTERED BY ACUTE FEEDING. BACKGROUND: GENE EXPRESSION CHANGES IN THE LIVER AFTER ACUTE BINGE DRINKING MAY DIFFER FROM THE CHANGES SEEN IN CHRONIC ETHANOL FEEDING IN THE RAT. THE CHANGES IN GENE EXPRESSION AFTER CHRONIC ETHANOL FEEDING MAY SENSITIZE THE LIVER TO ALCOHOL-INDUCED LIVER DAMAGE, WHICH IS NOT SEEN AFTER ACUTE BINGE DRINKING. METHODS: TO TEST THIS HYPOTHESIS, GENE MICROARRAY ANALYSIS WAS PERFORMED ON THE LIVERS OF RATS (N = 3) FED AN ACUTE BINGE DOSE OF ETHANOL (6 G/KG BODY WT) AND KILLED AT 3 AND 12 HOURS AFTER ETHANOL BY GAVAGE. THE GENE MICROARRAYS WERE COMPARED WITH THOSE MADE ON THE LIVER OF RATS FROM A PREVIOUS STUDY, IN WHICH THE RATS WERE FED ETHANOL BY INTRAGASTRIC TUBE FOR 1 MONTH (36% OF CALORIES DERIVED FROM ETHANOL). RESULTS: MICROARRAY ANALYSIS DATA VARIED BETWEEN THE ACUTE AND CHRONIC MODELS IN SEVERAL IMPORTANT RESPECTS. GROWTH FACTORS INCREASED MAINLY IN THE CHRONIC ALCOHOL FED RAT. CHANGES IN ENZYMES INVOLVED IN OXIDATIVE STRESS WERE NOTED ONLY WITH CHRONIC ETHANOL FEEDING. GENE EXPRESSION OF FAT METABOLISM WAS INCREASED ONLY WITH CHRONIC ETHANOL FEEDING. MOST IMPORTANTLY, EPIGENETIC RELATED ENZYMES AND ACETYLATION AND METHYLATION OF HISTONES CHANGED ONLY AFTER CHRONIC ETHANOL FEEDING. CONCLUSIONS: THE RESULTS SUPPORT THE CONCEPT THAT CHRONIC ETHANOL INGESTION INDUCES ALTERED GENE EXPRESSION AS A RESULT OF CHANGES IN EPIGENETIC MECHANISMS, WHERE ACETYLATION AND METHYLATION OF HISTONES WERE ALTERED. 2009 9 6177 35 THE HISTONE METHYLTRANSFERASE G9A MEDIATES STRESS-REGULATED ALCOHOL DRINKING. THE EPIGENETIC ENZYME G9A IS A HISTONE METHYLTRANSFERASE THAT DIMETHYLATES LYSINE 9 ON HISTONE H3 (H3K9ME2), AND IN THE ADULT NUCLEUS ACCUMBENS (NAC), G9A REGULATES MULTIPLE BEHAVIORS ASSOCIATED WITH SUBSTANCE USE DISORDER. WE SHOW HERE THAT CHRONIC INTERMITTENT ETHANOL (CIE) EXPOSURE IN MALE MICE REDUCED BOTH G9A AND H3K9ME2 LEVELS IN THE ADULT NAC, BUT NOT DORSAL STRIATUM. VIRAL-MEDIATED REDUCTION OF G9A IN THE NAC HAD NO EFFECTS ON BASELINE VOLITIONAL ETHANOL DRINKING OR ESCALATED ALCOHOL DRINKING PRODUCED BY CIE EXPOSURE; HOWEVER, NAC G9A WAS REQUIRED FOR STRESS-REGULATED CHANGES IN ETHANOL DRINKING, INCLUDING POTENTIATED ALCOHOL DRINKING PRODUCED BY ACTIVATION OF THE KAPPA-OPIOID RECEPTOR. IN ADDITION, WE OBSERVED THAT CHRONIC SYSTEMIC ADMINISTRATION OF A G9A INHIBITOR, UNC0642, ALSO BLOCKED STRESS-POTENTIATED ALCOHOL DRINKING. TOGETHER, OUR FINDINGS SUGGEST THAT CHRONIC ALCOHOL USE, SIMILAR TO OTHER ABUSED SUBSTANCES, PRODUCES A NAC-SELECTIVE REDUCTION IN G9A LEVELS THAT SERVES TO LIMIT STRESS-REGULATED ALCOHOL DRINKING. MOREOVER, OUR FINDINGS SUGGEST THAT PHARMACOLOGICAL INHIBITION OF G9A MIGHT PROVIDE A NOVEL THERAPEUTIC APPROACH TO TREAT STRESS-INDUCED ALCOHOL DRINKING, WHICH IS A MAJOR TRIGGER OF RELAPSE IN INDIVIDUALS SUFFERING FROM AUD. 2022 10 5485 37 REVERSAL OF COCAINE-CONDITIONED PLACE PREFERENCE THROUGH METHYL SUPPLEMENTATION IN MICE: ALTERING GLOBAL DNA METHYLATION IN THE PREFRONTAL CORTEX. ANALYSIS OF GLOBAL METHYLATION IN CELLS HAS REVEALED CORRELATIONS BETWEEN OVERALL DNA METHYLATION STATUS AND SOME BIOLOGICAL STATES. RECENT STUDIES SUGGEST THAT EPIGENETIC REGULATION THROUGH DNA METHYLATION COULD BE RESPONSIBLE FOR NEUROADAPTATIONS INDUCED BY ADDICTIVE DRUGS. HOWEVER, THERE IS NO INVESTIGATION TO DETERMINE GLOBAL DNA METHYLATION STATUS FOLLOWING REPEATED EXPOSURE TO ADDICTIVE DRUGS. USING MICE CONDITIONED PLACE PREFERENCE (CPP) PROCEDURE, WE MEASURED GLOBAL DNA METHYLATION LEVEL IN THE NUCLEUS ACCUMBENS (NAC) AND THE PREFRONTAL CORTEX (PFC) ASSOCIATED WITH DRUG REWARDING EFFECTS. WE FOUND THAT COCAINE-, BUT NOT MORPHINE- OR FOOD-CPP TRAINING DECREASED GLOBAL DNA METHYLATION IN THE PFC. CHRONIC TREATMENT WITH METHIONINE, A METHYL DONOR, FOR 25 CONSECUTIVE DAYS PRIOR TO AND DURING CPP TRAINING INHIBITED THE ESTABLISHMENT OF COCAINE, BUT NOT MORPHINE OR FOOD CPP. WE ALSO FOUND THAT BOTH MRNA AND PROTEIN LEVEL OF DNMT (DNA METHYTRANSFERASE) 3B IN THE PFC WERE DOWNREGULATED FOLLOWING THE ESTABLISHMENT OF COCAINE CPP, AND THE DOWNREGULATION COULD BE REVERSED BY REPEATED ADMINISTRATION OF METHIONINE. OUR STUDY INDICATES A CRUCIAL ROLE OF GLOBAL PFC DNA HYPOMETHYLATION IN THE REWARDING EFFECTS OF COCAINE. REVERSAL OF GLOBAL DNA HYPOMETHYLATION COULD SIGNIFICANTLY ATTENUATE THE REWARDING EFFECTS INDUCED BY COCAINE. OUR RESULTS SUGGEST THAT METHIONINE MAY HAVE BECOME A POTENTIAL THERAPEUTIC TARGET TO TREAT COCAINE ADDICTION. 2012 11 6175 30 THE HISTONE DEACETYLASE INHIBITOR SUBEROYLANILIDE HYDROXAMIC ACID (SAHA) ALLEVIATES DEPRESSION-LIKE BEHAVIOR AND NORMALIZES EPIGENETIC CHANGES IN THE HIPPOCAMPUS DURING ETHANOL WITHDRAWAL. WITHDRAWAL FROM CHRONIC ALCOHOL DRINKING CAN CAUSE DEPRESSION, LEADING TO AN INABILITY TO FUNCTION IN DAILY LIFE AND AN INCREASED RISK FOR RELAPSE TO HARMFUL DRINKING. UNDERSTANDING THE CAUSES OF ALCOHOL WITHDRAWAL-RELATED DEPRESSION MAY LEAD TO NEW THERAPEUTIC TARGETS FOR TREATMENT. EPIGENETIC FACTORS HAVE RECENTLY EMERGED AS IMPORTANT CONTRIBUTORS TO BOTH DEPRESSION AND ALCOHOL USE DISORDER (AUD). SPECIFICALLY, ACETYLATION OF THE N-TERMINAL TAILS OF HISTONE PROTEINS THAT PACKAGE DNA INTO NUCLEOSOMES IS ALTERED IN STRESS-INDUCED MODELS OF DEPRESSION AND DURING ALCOHOL WITHDRAWAL. THE GOAL OF THIS STUDY WAS TO EXAMINE DEPRESSION-LIKE BEHAVIOR DURING ALCOHOL WITHDRAWAL AND ASSOCIATED CHANGES IN HISTONE ACETYLATION AND EXPRESSION OF HISTONE DEACETYLASE 2 (HDAC2) IN THE HIPPOCAMPUS, A BRAIN REGION CRITICAL FOR MOOD REGULATION AND DEPRESSION. MALE SPRAGUE-DAWLEY RATS WERE TREATED WITH THE LIEBER-DECARLI ETHANOL LIQUID DIET FOR 15 DAYS AND THEN UNDERWENT WITHDRAWAL. RATS WERE TREATED WITH THE HDAC INHIBITOR, SUBEROYLANILIDE HYDROXAMIC ACID (SAHA), DURING WITHDRAWAL AND WERE TESTED FOR DEPRESSION-LIKE BEHAVIOR. IN A SEPARATE GROUP OF RATS, THE HIPPOCAMPUS WAS ANALYZED FOR MRNA AND PROTEIN EXPRESSION OF HDAC2 AND LEVELS OF HISTONE H3 LYSINE 9 ACETYLATION (H3K9AC) DURING CHRONIC ETHANOL EXPOSURE AND WITHDRAWAL. RATS UNDERGOING ETHANOL WITHDRAWAL EXHIBITED DEPRESSION-LIKE BEHAVIOR AND HAD INCREASED HDAC2 AND DECREASED H3K9AC LEVELS IN SPECIFIC STRUCTURES OF THE HIPPOCAMPUS. TREATMENT WITH SAHA DURING WITHDRAWAL AMELIORATED DEPRESSION-LIKE BEHAVIOR AND NORMALIZED CHANGES IN HIPPOCAMPAL HDAC2 AND H3K9AC LEVELS. THESE RESULTS DEMONSTRATE THAT ETHANOL WITHDRAWAL CAUSES AN ALTERED EPIGENETIC STATE IN THE HIPPOCAMPUS. TREATMENT WITH AN HDAC INHIBITOR CAN CORRECT THIS STATE AND ALLEVIATE DEPRESSION-LIKE SYMPTOMS DEVELOPED DURING WITHDRAWAL. TARGETING HISTONE ACETYLATION MAY BE A NOVEL STRATEGY TO REDUCE ETHANOL WITHDRAWAL-INDUCED DEPRESSION. 2019 12 1731 40 DYSREGULATION OF THE HISTONE DEMETHYLASE KDM6B IN ALCOHOL DEPENDENCE IS ASSOCIATED WITH EPIGENETIC REGULATION OF INFLAMMATORY SIGNALING PATHWAYS. EPIGENETIC ENZYMES OVERSEE LONG-TERM CHANGES IN GENE EXPRESSION BY INTEGRATING GENETIC AND ENVIRONMENTAL CUES. WHILE THERE ARE HUNDREDS OF ENZYMES THAT CONTROL HISTONE AND DNA MODIFICATIONS, THEIR POTENTIAL ROLES IN SUBSTANCE ABUSE AND ALCOHOL DEPENDENCE REMAIN UNDEREXPLORED. A FEW RECENT STUDIES HAVE SUGGESTED THAT EPIGENETIC PROCESSES COULD UNDERLIE TRANSCRIPTOMIC AND BEHAVIORAL HALLMARKS OF ALCOHOL ADDICTION. IN THE PRESENT STUDY, WE SOUGHT TO IDENTIFY EPIGENETIC ENZYMES IN THE BRAIN THAT ARE DYSREGULATED DURING PROTRACTED ABSTINENCE AS A CONSEQUENCE OF CHRONIC AND INTERMITTENT ALCOHOL EXPOSURE. THROUGH QUANTITATIVE MRNA EXPRESSION ANALYSIS OF OVER 100 EPIGENETIC ENZYMES, WE IDENTIFIED 11 THAT ARE SIGNIFICANTLY ALTERED IN ALCOHOL-DEPENDENT RATS COMPARED WITH CONTROLS. FOLLOW-UP STUDIES OF ONE OF THESE ENZYMES, THE HISTONE DEMETHYLASE KDM6B, SHOWED THAT THIS ENZYME EXHIBITS REGION-SPECIFIC DYSREGULATION IN THE PREFRONTAL CORTEX AND NUCLEUS ACCUMBENS OF ALCOHOL-DEPENDENT RATS. KDM6B WAS ALSO UPREGULATED IN THE HUMAN ALCOHOLIC BRAIN. UPREGULATION OF KDM6B PROTEIN IN ALCOHOL-DEPENDENT RATS WAS ACCOMPANIED BY A DECREASE OF TRIMETHYLATION LEVELS AT HISTONE H3, LYSINE 27 (H3K27ME3), CONSISTENT WITH THE KNOWN DEMETHYLASE SPECIFICITY OF KDM6B. SUBSEQUENT EPIGENETIC (CHROMATIN IMMUNOPRECIPITATION [CHIP]-SEQUENCING) ANALYSIS SHOWED THAT ALCOHOL-INDUCED CHANGES IN H3K27ME3 WERE SIGNIFICANTLY ENRICHED AT GENES IN THE IL-6 SIGNALING PATHWAY, CONSISTENT WITH THE WELL-CHARACTERIZED ROLE OF KDM6B IN MODULATION OF INFLAMMATORY RESPONSES. KNOCKDOWN OF KDM6B IN CULTURED MICROGLIAL CELLS DIMINISHED IL-6 INDUCTION IN RESPONSE TO AN INFLAMMATORY STIMULUS. OUR FINDINGS IMPLICATE A NOVEL KDM6B-MEDIATED EPIGENETIC SIGNALING PATHWAY INTEGRATED WITH INFLAMMATORY SIGNALING PATHWAYS THAT ARE KNOWN TO UNDERLIE THE DEVELOPMENT OF ALCOHOL ADDICTION. 2021 13 2347 33 EPIGENETIC REGULATION OF MIR-124 UNDER ETHANOL DEPENDENCE AND WITHDRAWAL. WITHDRAWAL FROM CHRONIC ALCOHOL CAUSE THE PERSISTENT MOLECULAR ALTERATION, SUCH AS CHANGES IN THE RELEASE OF NEUROTRANSMITTER AND GENE EXPRESSION. THE ALTERATIONS ARE THOUGHT TO INCREASE IN THE RISK OF RELAPSE. RECENT STUDIES SUGGEST THAT THE GENE EXPRESSION REGULATED BY HISTONE ACETYLATION MAY PLAY AN IMPORTANT ROLE IN THE DEPENDENCE OF ABUSED DRUGS, INCLUDING OF ETHANOL. FURTHERMORE, MIRNA, ANOTHER REGULATOR OF GENE EXPRESSION, ARE ALSO IMPORTANT MOLECULES FOR THE DEPENDENCE. HOWEVER, CHANGES IN THE MOLECULES UNDER ETHANOL WITHDRAWAL AND ITS RELATIONSHIP ARE POORLY UNDERSTOOD. IN THE PRESENT STUDY, WE INVESTIGATED THE EXPRESSION OF ACETYLATED HISTONE H3 AND MIR-124 IN MOUSE BRAIN AT 3 DAYS AFTER ETHANOL WITHDRAWAL. 6-WEEK AGES OF C57BL/6J MICE WERE TREATED WITH LIQUID DIET CONTAINING ETHANOL FOR 10 DAYS. USING THE ESCALATING ETHANOL DOSAGE SCHEDULE, THE MICE WERE FED THE ETHANOL DIET AS FOLLOWS: 1ST DAY: 1 W/V%: 2ND AND 3RD DAY: 3 W/V%; 4TH AND 5TH DAY: 4 W/V% AND FROM THE 6TH TO 10TH DAY: 5 W/V% ETHANOL DIET, RESPECTIVELY. THE PAIR-FED CONTROL MICE WERE GIVEN THE SAME VOLUME OF ETHANOL-FREE LIQUID DIET WITH GLUCOSE SUBSTITUTED IN ISOCALORIC QUANTITIES FOR ETHANOL. AFTER FEEDING ALCOHOL LIQUID DIET, THE MICE SHOWED SEVERE WITHDRAWAL SIGNS. THE EXPRESSION OF ACETYLATED HISTONE H3 WAS SIGNIFICANTLY DECREASED IN LIMBIC FOREBRAIN AT 3 DAYS AFTER WITHDRAWAL. WE FOUND THAT MIR-124 ALSO DECREASED IN THE LIMBIC FOREBRAIN. IT HAS BEEN REPORTED THAT CDC42 REGULATES NEURONAL DEVELOPMENT AS A TARGET OF MIR-124. WE FOUND THAT CDC42 PROTEIN MARKEDLY INCREASED IN BOTH BRAIN REGIONS AT 3 DAYS AFTER WITHDRAWAL. OUR FINDINGS SUGGEST THAT CHANGES IN THE EXPRESSION OF MIR-124 VIA HISTONE ACETYLATION LEADS TO CHANGE THE CDC42 EXPRESSION UNDER ETHANOL WITHDRAWAL. 2012 14 4173 31 MELATONIN INDUCES HISTONE HYPERACETYLATION IN THE RAT BRAIN. WE HAVE REPORTED THAT MELATONIN INDUCES HISTONE HYPERACETYLATION IN MOUSE NEURAL STEM CELLS, SUGGESTING AN EPIGENETIC ROLE FOR THIS PLEIOTROPIC HORMONE. TO SUPPORT SUCH A ROLE, IT IS NECESSARY TO DEMONSTRATE THAT MELATONIN PRODUCES SIMILAR EFFECTS IN VIVO. HISTONE ACETYLATION, FOLLOWING CHRONIC TREATMENT WITH MELATONIN (4MUG/ML IN DRINKING WATER FOR 17 DAYS), WAS EXAMINED BY WESTERN BLOTTING IN SELECTED RAT BRAIN REGIONS. MELATONIN INDUCED SIGNIFICANT INCREASES IN HISTONE H3 AND HISTONE H4 ACETYLATION IN THE HIPPOCAMPUS. HISTONE H4 WAS ALSO HYPERACETYLATED IN THE STRIATUM, BUT THERE WERE NO SIGNIFICANT CHANGES IN HISTONE H3 ACETYLATION IN THIS BRAIN REGION. NO SIGNIFICANT CHANGES IN THE ACETYLATION OF EITHER HISTONE H3 OR H4 WERE OBSERVED IN THE MIDBRAIN AND CEREBELLUM. AN EXAMINATION OF KINASE ACTIVATION, WHICH MAY BE RELATED TO THESE CHANGES, REVEALED THAT MELATONIN TREATMENT INCREASED THE LEVELS OF PHOSPHO-ERK (EXTRACELLULAR SIGNAL-REGULATED KINASE) IN THE HIPPOCAMPUS AND STRIATUM, BUT PHOSPHO-AKT (PROTEIN KINASE B) LEVELS WERE UNCHANGED. THESE FINDINGS SUGGEST THAT CHROMATIN REMODELING AND ASSOCIATED CHANGES IN THE EPIGENETIC REGULATION OF GENE EXPRESSION UNDERLIE THE MULTIPLE PHYSIOLOGICAL EFFECTS OF MELATONIN. 2013 15 1086 35 COCAINE ADMINISTRATION AND ITS WITHDRAWAL ENHANCE THE EXPRESSION OF GENES ENCODING HISTONE-MODIFYING ENZYMES AND HISTONE ACETYLATION IN THE RAT PREFRONTAL CORTEX. CHRONIC EXPOSURE TO COCAINE, CRAVING, AND RELAPSE ARE ATTRIBUTED TO LONG-LASTING CHANGES IN GENE EXPRESSION ARISING THROUGH EPIGENETIC AND TRANSCRIPTIONAL MECHANISMS. ALTHOUGH SEVERAL BRAIN REGIONS ARE INVOLVED IN THESE PROCESSES, THE PREFRONTAL CORTEX SEEMS TO PLAY A CRUCIAL ROLE NOT ONLY IN MOTIVATION AND DECISION-MAKING BUT ALSO IN EXTINCTION AND SEEKING BEHAVIOR. IN THIS STUDY, WE APPLIED COCAINE SELF-ADMINISTRATION AND EXTINCTION TRAINING PROCEDURES IN RATS WITH A YOKED TRIAD TO DETERMINE DIFFERENTIALLY EXPRESSED GENES IN PREFRONTAL CORTEX. MICROARRAY ANALYSIS SHOWED SIGNIFICANT UPREGULATION OF SEVERAL GENES ENCODING HISTONE MODIFICATION ENZYMES DURING EARLY EXTINCTION TRAINING. SUBSEQUENT REAL-TIME PCR TESTING OF THESE GENES FOLLOWING COCAINE SELF-ADMINISTRATION OR EARLY (THIRD DAY) AND LATE (TENTH DAY) EXTINCTION REVEALED ELEVATED LEVELS OF THEIR TRANSCRIPTS. INTERESTINGLY, WE FOUND THE ENRICHMENT OF BRD1 MESSENGER RNA IN RATS SELF-ADMINISTERING COCAINE THAT LASTED UNTIL EXTINCTION TRAINING DURING COCAINE WITHDRAWAL WITH CONCOMITANT INCREASED ACETYLATION OF H3K9 AND H4K8. HOWEVER, DESPITE ELEVATED LEVELS OF METHYL- AND DEMETHYLTRANSFERASE-ENCODED TRANSCRIPTS, NO CHANGES IN GLOBAL DI- AND TRI-METHYLATION OF HISTONE H3 AT LYSINE 4, 9, 27, AND 79 WERE OBSERVED. SURPRISINGLY, AT THE END OF EXTINCTION TRAINING (10 DAYS OF COCAINE WITHDRAWAL), MOST OF THE ANALYZED GENES IN THE RATS ACTIVELY AND PASSIVELY ADMINISTERING COCAINE RETURNED TO THE CONTROL LEVEL. TOGETHER, THE ALTERATIONS IDENTIFIED IN THE RAT PREFRONTAL CORTEX MAY SUGGEST ENHANCED CHROMATIN REMODELING AND TRANSCRIPTIONAL ACTIVITY INDUCED BY EARLY COCAINE ABSTINENCE; HOWEVER, TO KNOW WHETHER THEY ARE BENEFICIAL OR NOT FOR THE EXTINCTION OF DRUG-SEEKING BEHAVIOR, FURTHER IN VIVO EVALUATION IS REQUIRED. 2017 16 3587 39 IMPACT OF TLR4 ON BEHAVIORAL AND COGNITIVE DYSFUNCTIONS ASSOCIATED WITH ALCOHOL-INDUCED NEUROINFLAMMATORY DAMAGE. TOLL-LIKE RECEPTORS (TLRS) PLAY AN IMPORTANT ROLE IN THE INNATE IMMUNE RESPONSE, AND EMERGING EVIDENCE INDICATES THEIR ROLE IN BRAIN INJURY AND NEURODEGENERATION. OUR RECENT RESULTS HAVE DEMONSTRATED THAT ETHANOL IS CAPABLE OF ACTIVATING GLIAL TLR4 RECEPTORS AND THAT THE ELIMINATION OF THESE RECEPTORS IN MICE PROTECTS AGAINST ETHANOL-INDUCED GLIAL ACTIVATION, INDUCTION OF INFLAMMATORY MEDIATORS AND APOPTOSIS. THIS STUDY WAS DESIGNED TO ASSESS WHETHER ETHANOL-INDUCED INFLAMMATORY DAMAGE CAUSES BEHAVIORAL AND COGNITIVE CONSEQUENCES, AND IF BEHAVIORAL ALTERATIONS ARE DEPENDENT OF TLR4 FUNCTIONS. HERE WE SHOW IN MICE DRINKING ALCOHOL FOR 5MONTHS, FOLLOWED BY A 15-DAY WITHDRAWAL PERIOD, THAT ACTIVATION OF THE ASTROGLIAL AND MICROGLIAL CELLS IN FRONTAL CORTEX AND STRIATUM IS MAINTAINED AND THAT THESE EVENTS ARE ASSOCIATED WITH COGNITIVE AND ANXIETY-RELATED BEHAVIORAL IMPAIRMENTS IN WILD-TYPE (WT) MICE, AS DEMONSTRATED BY TESTING THE ANIMALS WITH OBJECT MEMORY RECOGNITION, CONDITIONED TASTE AVERSION AND DARK AND LIGHT BOX ANXIETY TASKS. MICE LACKING TLR4 RECEPTORS ARE PROTECTED AGAINST ETHANOL-INDUCED INFLAMMATORY DAMAGE, AND BEHAVIORAL ASSOCIATED EFFECTS. WE FURTHER ASSESS THE POSSIBILITY OF THE EPIGENETIC MODIFICATIONS PARTICIPATING IN SHORT- OR LONG-TERM BEHAVIORAL EFFECTS ASSOCIATED WITH NEUROINFLAMMATORY DAMAGE. WE SHOW THAT CHRONIC ALCOHOL TREATMENT DECREASES H4 HISTONE ACETYLATION AND HISTONE ACETYLTRANSFERASES ACTIVITY IN FRONTAL CORTEX, STRIATUM AND HIPPOCAMPUS OF WT MICE. ALTERATIONS IN CHROMATIN STRUCTURE WERE NOT OBSERVED IN TLR4(-/-) MICE. THESE RESULTS PROVIDE THE FIRST EVIDENCE OF THE ROLE THAT TLR4 FUNCTIONS PLAY IN THE BEHAVIORAL CONSEQUENCES OF ALCOHOL-INDUCED INFLAMMATORY DAMAGE AND SUGGEST THAT THE EPIGENETIC MODIFICATIONS MEDIATED BY TLR4 COULD CONTRIBUTE TO SHORT- OR LONG-TERM ALCOHOL-INDUCED BEHAVIORAL OR COGNITIVE DYSFUNCTIONS. 2011 17 5791 40 STABLE HISTONE METHYLATION CHANGES AT PROTEOGLYCAN NETWORK GENES FOLLOWING ETHANOL EXPOSURE. ALCOHOL USE DISORDER (AUD) IS A CHRONIC MENTAL ILLNESS IN WHICH PATIENTS OFTEN ACHIEVE PROTRACTED PERIODS OF ABSTINENCE PRIOR TO RELAPSE. EPIGENETIC MECHANISMS MAY PROVIDE AN EXPLANATION FOR THE PERSISTING GENE EXPRESSION CHANGES THAT CAN BE OBSERVED EVEN AFTER LONG PERIODS OF ABSTINENCE AND MAY CONTRIBUTE TO RELAPSE. IN THIS STUDY, WE EXAMINED TWO HISTONE MODIFICATIONS, HISTONE 3 LYSINE 4 TRI-METHYLATION (H3K4ME3) AND HISTONE 3 LYSINE 27 TRI-METHYLATION (H3K27ME3), IN THE PREFRONTAL CORTEX OF WITHDRAWAL SEIZURE RESISTANT (WSR) MICE 21 DAYS AFTER 72 H OF ETHANOL VAPOR EXPOSURE. THESE HISTONE MODIFICATIONS WERE SELECTED BECAUSE THEY ARE ASSOCIATED WITH ACTIVE PROMOTERS (H3K4ME3) AND REPRESSED GENE EXPRESSION IN A EUCHROMATIC ENVIRONMENT (H3K27ME3). WE PERFORMED A GENOME-WIDE ANALYSIS TO IDENTIFY DIFFERENCES IN H3K4ME3 AND H3K27ME3 LEVELS IN POST-ETHANOL EXPOSURE VS. CONTROL MICE BY CHIP-SEQ. WE DETECTED A GLOBAL REDUCTION IN H3K4ME3 PEAKS AND INCREASE IN H3K27ME3 PEAKS IN POST-ETHANOL EXPOSURE MICE COMPARED TO CONTROLS, THESE CHANGES ARE CONSISTENT WITH PERSISTENT REDUCTIONS IN GENE EXPRESSION. PATHWAY ANALYSIS OF GENES DISPLAYING CHANGES IN H3K4ME3 AND H3K27ME3 REVEALED ENRICHMENT FOR GENES INVOLVED IN PROTEOGLYCAN AND CALCIUM SIGNALING PATHWAYS, RESPECTIVELY. MICROARRAY ANALYSIS OF 7,683 GENES AND QPCR ANALYSIS IDENTIFIED EIGHT GENES DISPLAYING CONCORDANT REGULATION OF GENE EXPRESSION AND H3K4ME3/H3K27ME3. WE ALSO COMPARED CHANGES IN H3K4ME3 AND/OR H3K27ME3 FROM OUR STUDY WITH CHANGES IN GENE EXPRESSION IN RESPONSE TO ETHANOL FROM PUBLISHED LITERATURE AND WE FOUND THAT THE EXPRESSION OF 52% OF THE GENES WITH ALTERED H3K4ME3 BINDING AND 40% OF GENES WITH H3K27ME3 DIFFERENCES ARE ALTERED BY ETHANOL EXPOSURE. THE CHROMATIN CHANGES ASSOCIATED WITH THE 21-DAY POST-EXPOSURE PERIOD SUGGEST THAT THIS PERIOD IS A UNIQUE STATE IN THE ADDICTION CYCLE THAT DIFFERS FROM ETHANOL INTOXICATION AND ACUTE WITHDRAWAL. THESE RESULTS PROVIDE INSIGHTS INTO THE ENDURING EFFECTS OF ETHANOL ON PROTEOGLYCAN AND CALCIUM SIGNALING GENES IN THE BRAIN. 2018 18 917 38 CHRONIC HIGH-FAT DIET DRIVES POSTNATAL EPIGENETIC REGULATION OF MU-OPIOID RECEPTOR IN THE BRAIN. OPIOID SYSTEM DYSREGULATION HAS BEEN OBSERVED IN BOTH GENETIC AND HIGH-FAT DIET (HFD)-INDUCED MODELS OF OBESITY. AN UNDERSTANDING OF THE MOLECULAR MECHANISMS OF MOR TRANSCRIPTIONAL REGULATION, PARTICULARLY WITHIN AN IN VIVO CONTEXT, IS LACKING. USING A DIET-INDUCED MODEL OF OBESITY (DIO), MICE WERE FED A HIGH-FAT DIET (60% CALORIES FROM FAT) FROM WEANING TO >18 WEEKS OF AGE. COMPARED WITH MICE FED THE CONTROL DIET, DIO MICE HAD A DECREASED PREFERENCE FOR SUCROSE. MOR MRNA EXPRESSION WAS DECREASED IN REWARD-RELATED CIRCUITRY (VENTRAL TEGMENTAL AREA (VTA), NUCLEUS ACCUMBENS (NAC), AND PREFRONTAL CORTEX (PFC)) BUT NOT THE HYPOTHALAMUS, IMPORTANT IN THE HOMEOSTATIC REGULATION OF FEEDING. DNA METHYLATION IS AN EPIGENETIC MODIFICATION THAT LINKS ENVIRONMENTAL EXPOSURES TO ALTERED GENE EXPRESSION. WE FOUND A SIGNIFICANT INCREASE IN DNA METHYLATION IN THE MOR PROMOTER REGION WITHIN THE REWARD-RELATED BRAIN REGIONS. METHYL CPG-BINDING PROTEIN 2 (MECP2) CAN BIND METHYLATED DNA AND REPRESS TRANSCRIPTION, AND DIO MICE SHOWED INCREASED BINDING OF MECP2 TO THE MOR PROMOTER IN REWARD-RELATED REGIONS OF THE BRAIN. FINALLY, USING CHIP ASSAYS WE EXAMINED H3K9 METHYLATION (INACTIVE CHROMATIN) AND H3 ACETYLATION (ACTIVE CHROMATIN) WITHIN THE MOR PROMOTER REGION AND FOUND INCREASED H3K9 METHYLATION AND DECREASED H3 ACETYLATION. THESE DATA ARE THE FIRST TO IDENTIFY DNA METHYLATION, MECP2 RECRUITMENT, AND CHROMATIN REMODELING AS MECHANISMS LEADING TO TRANSCRIPTIONAL REPRESSION OF MOR IN THE BRAINS OF MICE FED A HIGH-FAT DIET. 2011 19 1418 29 DIFFERENCES IN DNA METHYLATION REPROGRAMMING UNDERLIE THE SEXUAL DIMORPHISM OF BEHAVIORAL DISORDER CAUSED BY PRENATAL STRESS IN RATS. PRENATAL STRESS (PS) CAN LEAD TO NEUROENDOCRINE AND EMOTIONAL DISORDERS LATER IN ADOLESCENCE. SEXUAL DIMORPHISM IN THESE NEURODEVELOPMENTAL OUTCOMES HAVE BEEN OBSERVED; HOWEVER, THE UNDERLYING MECHANISMS ARE NOT FULLY UNDERSTOOD. TO ADDRESS THIS ISSUE, WE INVESTIGATED WHETHER THERE ARE SEX DIFFERENCES IN EPIGENETIC REPROGRAMMING IN RATS EXPOSED TO PS. PREGNANT FEMALE RATS WERE SUBJECTED TO CHRONIC RESTRAINT STRESS FROM GESTATIONAL DAY (G)12 TO G18. FROM POSTNATAL DAY (P)38 TO P45, SUBGROUPS OF OFFSPRING INCLUDING BOTH MALES AND FEMALES WERE SUBJECTED TO BEHAVIORAL TESTING AND BRAIN TISSUE SPECIMENS WERE ANALYZED BY DNA PYROSEQUENCING, WESTERN BLOTTING, AND GOLGI STAINING TO ASSESS CHANGES IN METHYLATION PATTERN OF GLUCOCORTICOID RECEPTOR (GR) GENE, EXPRESSION OF DNA METHYLTRANSFERASE (DNMT) AND DNA DEMETHYLASE, AND DENDRITE MORPHOLOGY, RESPECTIVELY. THE DNA METHYLTRANSFERASE INHIBITOR DECITABINE WAS ADMINISTERED TO RATS PRIOR TO PS TO FURTHER EVALUATE THE ROLE OF METHYLATION IN THE SEXUALLY DIMORPHIC EFFECTS OF PS. THE RESULTS SHOWED THAT PS INCREASED ANXIETY-LIKE BEHAVIOR IN OFFSPRING, ESPECIALLY IN FEMALES, WHILE DEPRESSION-LIKE BEHAVIOR WAS INCREASED IN MALE OFFSPRING COMPARED TO CONTROL LITTERMATES. THE METHYLATION PATTERN IN THE PROMOTER REGION OF THE GR GENE DIFFERED BETWEEN MALES AND FEMALES. SEX-SPECIFIC CHANGES IN THE EXPRESSION OF DNMTS (DNMT1 AND DNMT3A) AND DNA DEMETHYLASE (TET METHYLCYTOSINE DIOXYGENASE 2) WERE ALSO OBSERVED. INTERESTINGLY, DECITABINE ALLEVIATED THE BEHAVIORAL DISORDER CAUSED BY PS AND RESTORED DENDRITE DENSITY AND MORPHOLOGY IN FEMALE BUT NOT MALE RATS. THESE FINDINGS SUGGEST THAT DIFFERENT CHANGE PATTERNS OF DNMT AND DEMETHYLASE IN THE TWO SEXES AFTER PS ARE RESPONSIBLE FOR THE SEXUALLY DIMORPHISM, WHICH COULD HAVE IMPLICATIONS FOR THE CLINICAL MANAGEMENT OF STRESS-RELATED DISORDERS. 2020 20 1789 22 EFFECT OF CHRONIC HEROIN AND COCAINE ADMINISTRATION ON GLOBAL DNA METHYLATION IN BRAIN AND LIVER. DRUG ABUSE IS ASSOCIATED WITH EPIGENETIC CHANGES, SUCH AS HISTONE MODIFICATIONS AND DNA METHYLATION. THE PURPOSE OF THE PRESENT STUDY WAS TO EXAMINE THE EFFECT OF CHRONIC COCAINE AND HEROIN ADMINISTRATION ON GLOBAL DNA METHYLATION IN BRAIN AND LIVER. MALE, 8 WEEK OLD, C57BL/6J MICE RECEIVED HEROIN IN A CHRONIC 'INTERMITTENT' ESCALATING DOSE PARADIGM, OR COCAINE IN A CHRONIC ESCALATING DOSE 'BINGE' PARADIGM, WHICH MIMIC THE HUMAN PATTERN OF OPIOID OR COCAINE ABUSE RESPECTIVELY. FOLLOWING SACRIFICE, LIVERS AND BRAINS WERE REMOVED AND DNA WAS EXTRACTED FROM THEM. THE EXTRACTED DNA WAS HYDROLYZED AND 2'-DEOXYCYTIDINE AND 5-METHYL-2'-DEOXYCYTIDINE WERE DETERMINED BY HPLC-UV. THE % 5-METHYL-2'-DEOXYCYTIDINE CONTENT OF DNA WAS SIGNIFICANTLY HIGHER IN THE BRAIN COMPARED TO THE LIVER. THERE WERE NO DIFFERENCES BETWEEN THE CONTROL ANIMALS AND THE COCAINE OR HEROIN TREATED ANIMALS IN NEITHER OF THE TISSUES EXAMINED, WHICH IS SURPRISING SINCE COCAINE ADMINISTRATION INDUCED GROSS MORPHOLOGICAL CHANGES IN THE LIVER. MOREOVER, THERE WAS NO DIFFERENCE IN THE % 5-METHYL-2'-DEOXYCYTIDINE CONTENT OF DNA BETWEEN THE COCAINE AND THE HEROIN TREATED ANIMALS. THE GLOBAL DNA METHYLATION STATUS IN THE BRAIN AND LIVER OF MICE CHRONICALLY TREATED WITH COCAINE OR HEROIN REMAINS UNAFFECTED, BUT THIS FINDING CANNOT EXCLUDE THE EXISTENCE OF ANATOMICAL REGION OR GENE-SPECIFIC METHYLATION DIFFERENCES. THIS IS THE FIRST TIME THAT GLOBAL DNA METHYLATION IN THE LIVER AND WHOLE BRAIN HAS BEEN STUDIED FOLLOWING CHRONIC COCAINE OR HEROIN TREATMENT. 2013