Regulator of G-protein Signaling 2 (RGS2) is a key regulator of G-protein-coupled signaling pathways involved in fear and anxiety. Data from rodent models and genetic analysis of anxiety-related traits and disorders in humans suggest down-regulation of RGS2 expression to be a risk factor for anxiety. Here we investigated, whether genetic variation in microRNAs mediating posttranscriptional down-regulation of RGS2 may be a risk factor for anxiety as well. 75 microRNAs predicted to regulate RGS2 were identified by four bioinformatic algorithms and validated experimentally by luciferase reporter gene assays. Specificity was confirmed for six microRNAs (hsa-miR-1271-5p, hsa-miR-22-3p, hsa-miR-3591-3p, hsa-miR-377-3p, hsa-miR-4717-5p, hsa-miR-96-5p) by disrupting their seed sequence at the 3' untranslated region of RGS2. Hsa-miR-4717-5p showed the most robust effect on RGS2 and regulated two other candidate genes of anxiety disorders (CNR1 and IKBKE) as well. Two SNPs (rs150925, rs161427) within and 1,000 bp upstream of the hostgene of hsa-miR-4717-5p (MIR4717) show a minor allele frequency greater than 0.05. Both were in high linkage disequilibrium (r(2) = 1, D' = 1) and both major (G) alleles showed a trend for association with panic disorder with comorbid agoraphobia in one of two patient/control samples (combined n(patients) = 497). Dimensional anxiety traits, as described by Anxiety Sensitivity Index (ASI) and Agoraphobic Cognitions Questionnaire (ACQ) were significantly higher among carriers of both major (G) alleles in a combined patient/control sample (n(combined) = 831). Taken together, data indicate that MIR4717 regulates human RGS2 and contributes to the genetic risk towards anxiety-related traits.

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Anxiety disorders result from a complex interplay of genetic and environmental factors such as stress. On the level of cellular signaling, regulator of G protein signaling 2 (Rgs2) has been implicated in human and rodent anxiety. However, there is limited knowledge about the role of Rgs2 in fear learning and reactivity to stress. In this study, Rgs2 mice showed increased fear learning, male mice displayed increased contextual and cued fear learning, while females showed selectively enhanced cued fear learning. Male Rgs2 mice displayed increased long-term-contextual fear memory, but increased cued fear extinction. Learning in spatial non-aversive paradigms was also increased in Rgs2 mice. Female, but not male mice show increased spatial learning in the Barnes maze, while male mice showed enhanced place preference in the IntelliCage, rendering enhanced cognitive function non-specific for aversive stimuli. Consistent with the previous results, Rgs2 deletion resulted in increased innate anxiety, including neophobic behavior expressed as hypolocomotion, in three different tests based on the approach-avoidance conflict. Acute electric foot shock stress provoked hypolocomotion in several exploration-based tests, suggesting fear generalization in both genotypes. Rgs2 deletion was associated with reduced monoaminergic neurotransmitter levels in the hippocampus and prefrontal cortex and disturbed corresponding GPCR expression of the adrenergic, serotonergic, dopaminergic and neuropeptide Y system. Taken together, Rgs2 deletion promotes improved cognitive function as well as increased anxiety-like behavior, but has no effect on acute stress reactivity. These effects may be related to the observed disruption of the monoaminergic systems.