Genome-wide analysis of the nucleus accumbens identifies DNA methylation signals differentiating low/binge from heavy alcohol drinking

Abstract

Alcohol use disorders encompass a range of drinking levels and behaviors, including low, binge and heavy drinking. In this regard, investigating the neural state of individuals who chronically self-administer lower doses of alcohol may provide insight into mechanisms that prevent the escalation of alcohol use. DNA methylation is one of the epigenetic mechanisms that stabilizes adaptations in gene expression and has been associated with alcohol use. Thus, we investigated DNA methylation, gene expression and the predicted neural effects in the nucleus accumbens of male rhesus macaques categorized as “low” or “binge” drinkers, compared to “alcohol-naïve” and “heavy” drinkers based on drinking patterns during a 12 month alcohol self-administration protocol. Using genome-wide CpG-rich region enrichment and bisulfite sequencing, the methylation levels of 2.6 million CpGs were compared between alcohol naïve (AN), low/binge (L/BD) and heavy/very heavy (H/VHD) drinking subjects (n=24). Through regional clustering analysis, we identified nine significant differential methylation regions (DMRs) that specifically distinguished ANs and L/BDs, and then compared those DMRs among H/VHDs. The DMRs mapped to genes encoding ion channels, receptors, cell adhesion molecules and cAMP, NF-κβ and Wnt signaling pathway proteins. Two of the DMRs, linked to PDE10A and PKD2L2, were also differentially methylated in H/VHDs, suggesting an alcohol-dose independent effect. However two other DMRs, linked to the CCBE1 and FZD5 genes, had L/BD methylation levels that significantly differed from both ANs and H/VHDs. The remaining 5 DMRs also differentiated L/BDs and ANs, however H/VHDs methylation levels were not distinguishable from either of the two groups. Functional validation of two DMRs, linked to FZD5 and PDE10A, support their role in regulating gene expression and exon usage, respectively. In summary, the findings demonstrate that L/BD is associated with unique DNA methylation signatures in the primate NAcc, and identifies synaptic genes that may play a role in preventing the escalation of alcohol use.

Studies using macaque monkeys to address excessive alcohol drinking and stress interactions

“…Using this approach the rhesus monoamine oxidase A (MAO-A) expression in the blood of alcohol-naïve subjects was found to be negatively correlated with subsequent alcohol self-administration (Cervera-Juanes et al, 2015). Further, the methylation of the promoter region of the MAO-A gene was associated with MAO-A expression in blood and in the nucleus accumbens (Cervera-Juanes et al, 2016). Because MAO-A is involved in monoaminergic metabolism (serotonin, dopamine and norepinephrine), discrete epigenetic changes altering the expression of this enzyme has direct implications for homeostatic physiological and behavioral processes involved in the risk for heavy drinking.…”

“…Importantly, like humans, NHPs show wide individual differences in their chronic intake of alcohol over years, with a large proportion voluntarily drinking amounts similar to human alcoholic chronic drinking levels (Baker et al, 2014; Baker et al, 2017; Mendelson and Mello, 1972;). More broadly, macaques and humans share similar genetic and epigenetic composition (Barr, 2013; Cervera-Juanes et al, 2016). For example, a recent study found CRH and serotonin gene polymorphisms in male rhesus macaques predicted a blunted ACTH response to dexamethasone (Ferguson et al, 2012).…”

“…Using this animal model, novel approaches to both genetic (Cervera-Juanes et al, 2016) and MRI connectivity studies can help address highly translational endocrine, neuroanatomical and behavioral measures of the temporal sequence and specific mediators of dysregulation in motivational circuitry associated with severity of AUDs.…”

Epigenetic mediators and consequences of excessive alcohol consumption

“…This can be further understood by examining how the epigenome changes across the three components of AUDs: intoxication, withdrawal, and craving (Koob & Volkow, 2010). In this special issue on epigenetics, Cervera-Juanes, Wilhelm, Park, Grant, & Ferguson (this issue) show that within the nucleus accumbens core, which is involved in neurocircuitry for intoxication and withdrawal, chronic low level drinking results in DNA hypomethylation, and high levels of drinking result in hypermethylation. In the prefrontal cortex, which is involved in neurocircuitry for craving, Hashimoto, Gavin, Wiren, Crabbe, and Guizzetti (this issue) show altered gene expression for proteins needed for histone and DNA epigenetic modification creation and maintenance, and they show that ethanol withdrawal has a larger effect on gene expression than ethanol exposure or abstinence.…”

Prefrontal cortex expression of chromatin modifier genes in male WSP and WSR mice changes across ethanol dependence, withdrawal, and abstinence

“…Although there is an emerging literature on the effects of alcohol on epigenetic mediators (Barbier et al, 2015, 2016; Cervera-Juanes, Wilhelm, Park, Grant, and Ferguson, 2016; Pandey, Ugale, Zhang, Tang, and Prakash, 2008; Ponomarev, Wang, Zhang, Harris, and Mayfield, 2012; Warnault, Darcq, Levine, Barak, and Ron, 2013), no study has systematically and comprehensively examined changes in the expression of genes encoding for chromatin modifying proteins after chronic alcohol exposure, alcohol withdrawal, and protracted abstinence.…”