Bioenergetic-related gene expression in the hippocampus predicts internalizing vs. externalizing behavior in an animal model of temperament

Externalizing and internalizing behavioral tendencies underlie many psychiatric and substance use disorders. These tendencies are associated with differences in temperament that emerge early in development via the interplay of genetic and environmental factors. To better understand the neurobiology of temperament, we have selectively bred rats for generations to produce two lines with highly divergent behavior: bred Low Responders (bLRs) are highly inhibited and anxious in novel environments, whereas bred High Responders (bHRs) are highly exploratory, sensation-seeking, and prone to drug-seeking behavior. Recently, we delineated these heritable differences by intercrossing bHRs and bLRs (F₀-F₁-F₂) to produce a heterogeneous F₂ sample with well-characterized lineage and behavior (exploratory locomotion, anxiety-like behavior, Pavlovian conditioning). The identified genetic loci encompassed variants that could influence behavior via many mechanisms, including proximal effects on gene expression. Here we measured gene expression in male and female F₀s (<i>n</i> = 12 bHRs, 12 bLRs) and in a large sample of heterogeneous F₂s (<i>n</i> = 250) using hippocampal RNA-Seq. This enabled triangulation of behavior with both genetic and functional genomic data to implicate specific genes and biological pathways. Our results show that bHR/bLR differential gene expression is robust, surpassing sex differences in expression, and predicts expression associated with F₂ behavior. In F₀ and F₂ samples, gene sets related to growth/proliferation are upregulated with bHR-like behavior, whereas gene sets related to mitochondrial function, oxidative stress, and microglial activation are upregulated with bLR-like behavior. Integrating our F₂ RNA-Seq data with previously-collected whole genome sequencing data identified genes with hippocampal expression correlated with proximal genetic variation (<i>cis</i>-expression quantitative trait loci or <i>cis</i>-eQTLs). These <i>cis</i>-eQTLs successfully predict bHR/bLR differential gene expression based on F₀ genotype. Sixteen of these genes are associated with <i>cis</i>-eQTLs colocalized within loci we previously linked to behavior and are strong candidates for mediating the influence of genetic variation on behavioral temperament. Eight of these genes are related to bioenergetics. Convergence between our study and others targeting similar behavioral traits revealed five more genes consistently related to temperament. Overall, our results implicate hippocampal bioenergetic regulation of oxidative stress, microglial activation, and growth-related processes in shaping behavioral temperament, thereby modulating vulnerability to psychiatric and addictive disorders.