Project description:Here, we identify persistent and substantial variation in ethanol drinking behavior within an inbred mouse strain and utilize this model to identify gene networks influencing such non-genetic variation in ethanol intake. C57BL/6NCrl mice showed persistent inter-individual variation of ethanol intake in a two-bottle choice paradigm over a three week period, ranging from less than 1 g/kg to over 14 g/kg ethanol in an 18h interval. Whole genome microarray expression analysis in nucleus accumbens, prefrontal cortex and ventral tegmental area of individual animals identified gene expression patterns correlated with ethanol intake. Results included several gene networks previously implicated in ethanol behaviors, such as glutamate signaling, BDNF and genes involved in synaptic vesicle function. Additionally, genes functioning in epigenetic chromatin or DNA modifications such as acetylation and/or methylation also had expression patterns correlated with ethanol intake. Our results thus implicate specific brain regional gene networks, including chromatin modification factors, as potentially important mechanisms underlying individual variation in ethanol intake. Voluntary two-bottle choice drinking was performed as described previously (Khisti et al. 2006). Briefly, two bottles containing 10%(w/v) ethanol (Aaper Alcohol and Chemical Co. Shelbyville, KY) or tap water were placed into the home cage at the beginning of the dark cycle. Tube position was varied every two days (L, L, R, R). Drinking sessions lasted 18 hours/day followed by 6 hours access to water only. Mice had four consecutive drinking sessions followed by four days of abstinence repeated four times to give 16 total drinking sessions. Three brain regions were harvested 6 days after the last drinking session: prefrontal cortex (PFC), nucleus accumbens (NAc) and ventral tegmental area (VTA) as previously described (Kerns et al. 2005). Labeled cRNA from individual animals (n=19) was hybridized to a single microarray for each brain region (n=58 total microarrays).

Project description:Lasting behavioral and physiological changes such as abusive consumption, dependence, and withdrawal are characteristic features of alcohol use disorders (AUD). Mechanistically, persistent changes in gene expression are hypothesized to contribute to these brain adaptations leading to ethanol toxicity and abuse. Here we employed repeated chronic intermittent ethanol (CIE) exposure by vapor chamber as a mouse model to simulate the cycles of ethanol exposure and withdrawal commonly seen with AUD. This model has previously been shown to induce progressive ethanol consumption in rodents. Brain regional expression networks contributing to CIE-induced behavioral changes were identified by microarray analysis across five brain regions in the mesolimbic dopamine system and extended amygdala with tissue harvested from 0-120 hours following the last cycle of CIE. Weighted Gene Correlated Network Analysis (WGCNA) was used to identify gene networks over-represented for CIE-induced temporal expression changes across brain regions. Differential gene expression analysis of CIE vs. air-treated controls showed that long-lasting gene regulation occurred 5-days after the final cycle of ethanol exposure only in prefrontal cortex (PFC) and hippocampus. In the majority of brain-regions, however, ethanol regulated gene expression changes occurred only immediately following CIE or within the first 8-hours of removal from ethanol. Bioinformatics analysis of modules identified by WGCNA showed that neuroinflammatory responses were seen across multiple brain regions at early time-points, whereas co-expression modules related to neuroplasticity, chromatin remodeling, and neurodevelopment were seen at later time-points and in specific brain regions (PFC or HPC). In PFC a module containing Bdnf was identified as highly CIE responsive in a biphasic manner, with peak changes at 0 hours and 5 days following CIE, suggesting a possible role in mechanisms underlying long-term molecular and behavioral response to CIE. Strikingly, bioinformatics analysis of this network and several other modules identified Let-7 family microRNAs as potential regulators of gene expression changes induced by CIE. Our results suggest a complex temporal and regional pattern of widespread gene network responses involving neuroinflammatory and neuroplasticity related genes as contributing to physiological and behavioral responses to chronic ethanol. In particular, our identification of a potential role for Let-7 miRNAs and a Bdnf-related expression network in long-lasting expression changes after CIE may lead to future druggable gene target identification for novel intervention in AUD.

Dataset Information

Chronic ethanol exposure results in time and brain-region dependent changes in gene coexpression networks.

Publications

Chronic ethanol exposure produces time- and brain region-dependent changes in gene coexpression networks.

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