Convergent functional genomics of anxiety disorders: translational identification of genes, biomarkers, pathways and mechanisms

Anxiety disorders are prevalent and disabling yet understudied from a genetic standpoint, compared with other major psychiatric disorders such as bipolar disorder and schizophrenia. The fact that they are more common, diverse and perceived as embedded in normal life may explain this relative oversight. In addition, as for other psychiatric disorders, there are technical challenges related to the identification and validation of candidate genes and peripheral biomarkers. Human studies, particularly genetic ones, are susceptible to the issue of being underpowered, because of genetic heterogeneity, the effect of variable environmental exposure on gene expression, and difficulty of accrual of large, well phenotyped cohorts. Animal model gene expression studies, in a genetically homogeneous and experimentally tractable setting, can avoid artifacts and provide sensitivity of detection. Subsequent translational integration of the animal model datasets with human genetic and gene expression datasets can ensure cross-validatory power and specificity for illness. We have used a pharmacogenomic mouse model (involving treatments with an anxiogenic drug—yohimbine, and an anti-anxiety drug—diazepam) as a discovery engine for identification of anxiety candidate genes as well as potential blood biomarkers. Gene expression changes in key brain regions for anxiety (prefrontal cortex, amygdala and hippocampus) and blood were analyzed using a convergent functional genomics (CFG) approach, which integrates our new data with published human and animal model data, as a translational strategy of cross-matching and prioritizing findings. Our work identifies top candidate genes (such as FOS, GABBR1, NR4A2, DRD1, ADORA2A, QKI, RGS2, PTGDS, HSPA1B, DYNLL2, CCKBR and DBP), brain–blood biomarkers (such as FOS, QKI and HSPA1B), pathways (such as cAMP signaling) and mechanisms for anxiety disorders—notably signal transduction and reactivity to environment, with a prominent role for the hippocampus. Overall, this work complements our previous similar work (on bipolar mood disorders and schizophrenia) conducted over the last decade. It concludes our programmatic first pass mapping of the genomic landscape of the triad of major psychiatric disorder domains using CFG, and permitted us to uncover the significant genetic overlap between anxiety and these other major psychiatric disorders, notably the under-appreciated overlap with schizophrenia. PDE10A, TAC1 and other genes uncovered by our work provide a molecular basis for the frequently observed clinical co-morbidity and interdependence between anxiety and other major psychiatric disorders, and suggest schizo-anxiety as a possible new nosological domain

Convergent functional genomic studies of omega-3 fatty acids in stress reactivity, bipolar disorder and alcoholism

Omega-3 fatty acids have been proposed as an adjuvant treatment option in psychiatric disorders. Given their other health benefits and their relative lack of toxicity, teratogenicity and side effects, they may be particularly useful in children and in females of child-bearing age, especially during pregnancy and postpartum. A comprehensive mechanistic understanding of their effects is needed. Here we report translational studies demonstrating the phenotypic normalization and gene expression effects of dietary omega-3 fatty acids, specifically docosahexaenoic acid (DHA), in a stress-reactive knockout mouse model of bipolar disorder and co-morbid alcoholism, using a bioinformatic convergent functional genomics approach integrating animal model and human data to prioritize disease-relevant genes. Additionally, to validate at a behavioral level the novel observed effects on decreasing alcohol consumption, we also tested the effects of DHA in an independent animal model, alcohol-preferring (P) rats, a well-established animal model of alcoholism. Our studies uncover sex differences, brain region-specific effects and blood biomarkers that may underpin the effects of DHA. Of note, DHA modulates some of the same genes targeted by current psychotropic medications, as well as increases myelin-related gene expression. Myelin-related gene expression decrease is a common, if nonspecific, denominator of neuropsychiatric disorders. In conclusion, our work supports the potential utility of omega-3 fatty acids, specifically DHA, for a spectrum of psychiatric disorders such as stress disorders, bipolar disorder, alcoholism and beyond

The Necessity of α4* Nicotinic Receptors in Nicotine-Driven Behaviors: Dissociation Between Reinforcing and Motor Effects of Nicotine

Here we utilize a mouse line with a targeted deletion of the α4 subunit (α4−/− mice), to investigate the role of α4* nAChRs in reinforcing and locomotor effects of nicotine. Within a conditioned place preference paradigm, both α4−/− mice and wild-type (WT) littermates showed a similar place preference to nicotine (0.5 mg/kg i.p.) conditioning. When assessed for operant intravenous self-administration of nicotine (0.05 mg/kg/infusion), α4−/− mice did not differ from their WT littermates in self-administration behavior. To further examine a modulatory role for α4* nAChRs in the reinforcing effects of nicotine, a transgenic mouse with a point mutation of the α4 subunit (α4-S248F) that renders increased sensitivity to low dose nicotine, was assessed for nicotine self-administration over a range of doses. At higher doses examined (0.05 and 0.07 mg/kg/infusion) there was no difference in intravenous nicotine self-administration; however, when mice were offered a lower dose of nicotine (0.03 mg/kg/infusion), α4-S248F mice showed greater nicotine intake than controls. Acute administration of 0.5 mg/kg nicotine caused significant locomotor depression in WT mice but α4−/− mice instead showed significant hyperactivity. Following chronic, intermittent administration of this dose of nicotine only WT mice displayed significant tolerance. Analogous experiments utilizing administration of the nicotinic antagonist mecamylamine in WT mice confirmed a dissociation between the putative nicotinic receptor subtypes required for mediating psychomotor and reinforcing effects of nicotine. These data demonstrate a necessary role for α4* nAChRs in the locomotor depressant effect of nicotine but not the reinforcing effects that support ongoing self-administration of nicotine

The hypocretin/orexin system mediates the extinction of fear memories

Anxiety disorders are often associated with an inability to extinguish learned fear responses. The hypocretin/orexin system is involved in the regulation of emotional states and could also participate in the consolidation and extinction of aversive memories. Using hypocretin receptor-1 and hypocretin receptor-2 antagonists, hypocretin-1 and hypocretin-2 peptides, and hypocretin receptor-1 knockout mice, we investigated the role of the hypocretin system in cue- and context-dependent fear conditioning and extinction. Hypocretins were crucial for the consolidation of fear conditioning, and this effect was mainly observed in memories with a high emotional component. Notably, after the acquisition of fear memory, hypocretin receptor-1 blockade facilitated fear extinction, whereas hypocretin-1 administration impaired this extinction process. The extinction-facilitating effects of the hypocretin receptor-1 antagonist SB334867 were associated with increased expression of cFos in the basolateral amygdala and the infralimbic cortex. Intra-amygdala, but neither intra-infralimbic prefrontal cortex nor intra-dorsohippocampal infusion of SB334867 enhanced fear extinction. These results reveal a key role for hypocretins in the extinction of aversive memories and suggest that hypocretin receptor-1 blockade could represent a novel therapeutic target for the treatment of diseases associated with inappropriate retention of fear, such as post-traumatic stress disorder and phobias.This work was supported by the Instituto de Salud Carlos III grants, #PI13/00042 and #RD12/0028/0023 (RTA-RETICS), by the Spanish Ministry of Science #SAF2011-29864, the/nCatalan Government (SGR2009-00731), and by the Catalan Institution for Research and Advanced Studies (ICREA Academia program). África Flores is a recipient of a predoctoral fellowship from the Spanish Ministry of Education