Selection for Drinking in the Dark Alters Brain Gene Coexpression Networks

Abstract

Background Heterogeneous stock (HS/NPT) mice have been used to create lines selectively bred in replicate for High Drinking in the Dark (HDID) (Crabbe et al., 2009; Crabbe et al., 2011b). Both selected lines routinely reach a blood ethanol concentration of 1.00 mg/ml or greater at the end of the day 2 four hour period of access. The mechanisms through which genetic differences influence DID are currently unclear. Therefore, the current study examines the transcriptome, the first stage at which genetic variability affects neurobiology. Rather than focusing solely on differential expression, we also examine changes in the ways that gene transcripts collectively interact with each other, as revealed by changes in coexpression patterns. Methods Naïve mice (N=48/group) were genotyped using the Mouse Universal Genotyping Array, which provided 3683 informative markers. QTL analysis used a marker-by-marker strategy with the threshold for a significant LOD set at 10.6. Gene expression in the ventral striatum was measured using the Illumina Mouse 8.2 array. Differential gene expression and the Weighted Gene Coexpression Network Analysis (WGCNA) were implemented largely as described elsewhere (Iancu et al., 2010; Iancu et al., 2012b). Results Significant quantitative trait loci (QTLs) for elevated blood ethanol concentrations after DID were detected on chromosomes 4, 14 and 16; the latter two were associated with gene poor regions. None of the QTLs overlapped with known QTLs for ethanol preference drinking. Ninety- four transcripts were detected as being differentially expressed in both selected lines vs HS controls; there was no overlap with known preference genes. The WGCNA revealed 2 modules as showing significant effects of both selections on intramodular connectivity. A number of genes known to be associated with ethanol phenotypes (e.g. Gabarg1, Glra2, Grik1, Npy2r and Nts) showed significant changes in connectivity. Discussion We found marked and consistent effects of selection on coexpression patterns; differential expression changes were more modest and less concordant. The QTLs and differentially expressed genes detected here are distinct from the preference phenotype (Mulligan et al., 2006). This is consistent with behavioral data and suggests that the DID and preference phenotypes are markedly different genetically.

Glycine Receptors Containingα2 orα3 Subunits Regulate Specific Ethanol-Mediated Behaviors

“…As mentioned, altered gene coexpression networks were found in mouse lines bred for high levels of DID, and Glra2 was one of the genes showing significant changes in connectivity (Iancu et al, 2013). This study highlights the role of gene networks rather than differential expression of individual transcripts in the propensity for ethanol drinking and further supports the importance of the GlyRa2 subunit in ethanol drinking and preference.…”

“…There were no differences among the mutant and WT mice during the DID test, which is a model of binge-like drinking. However, mouse lines selected for high ethanol consumption during DID demonstrated altered gene coexpression networks (in the absence of ethanol exposure), and Glra2 was one of the genes found to have significant changes in connectivity (Iancu et al, 2013).…”

Understanding the addiction cycle: A complex biology with distinct contributions of genotype vs. sex at each stage

“…In addition, little biological confirmation of the important pathways altered as a consequence of alcohol exposure as identified in array analysis has been reported to date. Instead of analysis of neuroadapation, much transcriptional profiling has focused on inherent genetic differences observed between alcohol-related phenotypes (see Iancu et al, 2013, McBride et al, 2013). Compared with RNA-seq, hybridization arrays have reduced dynamic range but improved sensitivity (see Malone and Oliver, 2011).…”

Adenosinergic regulation of binge-like ethanol drinking and associated locomotor effects in male C57BL/6J mice

“…In addition, a significant portion of this work has been limited to knockout models or operant paradigms. Behavioral and quantitative genetic studies in rodents suggest that binge-like alcohol consumption is a distinct phenotype (Fritz et al, 2014b, Bell et al, 2006, Iancu et al, 2013). Therefore, elucidating its neurochemical/biological underpinnings will be necessary to more completely understand this form of excessive alcohol drinking.…”