Epigenetic signatures in antidepressant treatment response: a methylome-wide association study in the EMC trial.

Although the currently available antidepressants are well established in the treatment of the major depressive disorder (MDD), there is strong variability in the response of individual patients. Reliable predictors to guide treatment decisions before or in an early stage of treatment are needed. DNA-methylation has been proven a useful biomarker in different clinical conditions, but its importance for mechanisms of antidepressant response has not yet been determined. 80 MDD patients were selected out of >500 participants from the Early Medication Change (EMC) cohort with available genetic material based on their antidepressant response after four weeks and stratified into clear responders and age- and sex-matched non-responders (N = 40, each). Early improvement after two weeks was analyzed as a secondary outcome. DNA-methylation was determined using the Illumina EPIC BeadChip. Epigenome-wide association studies were performed and differentially methylated regions (DMRs) identified using the comb-p algorithm. Enrichment was tested for hallmark gene-sets and in genome-wide association studies of depression and antidepressant response. No epigenome-wide significant differentially methylated positions were found for treatment response or early improvement. Twenty DMRs were associated with response; the strongest in an enhancer region in SORBS2, which has been related to cardiovascular diseases and type II diabetes. Another DMR was located in CYP2C18, a gene previously linked to antidepressant response. Results pointed towards differential methylation in genes associated with cardiac function, neuroticism, and depression. Linking differential methylation to antidepressant treatment response is an emerging topic and represents a step towards personalized medicine, potentially facilitating the prediction of patients' response before treatment.

Delineation of molecular pathway activities of the chronic antidepressant treatment response suggests important roles for glutamatergic and ubiquitin-proteasome systems.

The aim of this study was to identify molecular pathways related to antidepressant response. We administered paroxetine to the DBA/2J mice for 28 days. Following the treatment, the mice were grouped into responders or non-responders depending on the time they spent immobile in the forced swim test. Hippocampal metabolomics and proteomics analyses revealed that chronic paroxetine treatment affects glutamate-related metabolite and protein levels differentially in the two groups. We found significant differences in the expression of N-methyl-d-aspartate receptor and neuronal nitric oxide synthase proteins between the two groups, without any significant alterations in the respective transcript levels. In addition, we found that chronic paroxetine treatment altered the levels of proteins associated with the ubiquitin-proteasome system (UPS). The soluble guanylate cyclase-ß1, proteasome subunit α type-2 and ubiquitination levels were also affected in peripheral blood mononuclear cells from antidepressant responder and non-responder patients suffering from major depressive disorder. We submit that the glutamatergic system and UPS have a crucial role in the antidepressant treatment response in both mice and humans.

Dioxins, PCBs, chlorinated pesticides and brominated flame retardants in free-range chicken eggs from peri-urban areas in Arusha, Tanzania: Levels and implications for human health.

The environment in the northern part of Tanzania is influenced by rapid population growth, and increased urbanization. Urban agriculture is common and of economic value for low income families. In Arusha, many households sell eggs from free-ranging backyard chicken. In 2011, 159 eggs from different households in five different locations in Arusha were collected, homogenized, pooled into 28 composite samples and analyzed for a wide selection of POPs. Levels of POPs varied widely within and between the locations. The levels of dieldrin and ΣDDT ranged between 2 and 98,791 and 2 and 324ng/g lipid weight (lw), respectively. EU MRLs of 0.02mg/kg dieldrin for eggs were exceeded in 4/28 samples. PCBs, HCHs, chlordanes, toxaphenes and endosulfanes were found at lower frequency and levels. Brominated flame retardants (BFRs), e.g polybrominated diphenylethers (PBDEs), hexabromocyclododecane (HBCD) and 1,2-bis(2,4,6-tribromphenoxy)ethane (BTBPE) were present in 100%, 60% and 46% of the composite samples, respectively. Octa-and deca-BDEs were the dominating PBDEs and BDE 209 levels ranged between

SLC6A15, a novel stress vulnerability candidate, modulates anxiety and depressive-like behavior: involvement of the glutamatergic system.

Major depression is a multifactorial disease, involving both environmental and genetic risk factors. Recently, SLC6A15 - a neutral amino acid transporter mainly expressed in neurons - was proposed as a new candidate gene for major depression and stress vulnerability. Risk allele carriers for a single nucleotide polymorphism (SNP) in a SLC6A15 regulatory region display altered hippocampal volume, glutamate levels, and hypothalamus-pituitary-adrenal axis activity, all markers associated with major depression. Despite this genetic link between SLC6A15 and depression, its functional role with regard to the development and maintenance of depressive disorder is still unclear. The aim of the current study was therefore to characterize the role of mouse slc6a15 in modulating brain function and behavior, especially in relation to stress as a key risk factor for the development of mood disorders. We investigated the effects of slc6a15 manipulation using two mouse models, a conventional slc6a15 knock-out mouse line (SLC-KO) and a virus-mediated hippocampal slc6a15 overexpression (SLC-OE) model. Mice were tested under basal conditions and following chronic social stress. We found that SLC-KO animals displayed a similar behavioral profile to wild-type littermates (SLC-WT) under basal conditions. Interestingly, following chronic social stress SLC-KO animals showed lower levels of anxiety- and depressive-like behavior compared to stressed WT littermates. In support of these findings, SLC-OE animals displayed increased anxiety-like behavior already under basal condition. We also provide evidence that GluR1 expression in the dentate gyrus, but not GluR2 or NR1, are regulated by slc6a15 expression, and may contribute to the difference in stress responsiveness observed between SLC-KO and SLC-WT animals. Taken together, our data demonstrate that slc6a15 plays a role in modulating emotional behavior, possibly mediated by its impact on glutamatergic neurotransmission.

Deciphering the spatio-temporal expression and stress regulation of Fam107B, the paralog of the resilience-promoting protein DRR1 in the mouse brain.

Understanding the molecular mechanisms that promote stress resilience might open up new therapeutic avenues to prevent stress-related disorders. We recently characterized a stress and glucocorticoid-regulated gene, down-regulated in renal cell carcinoma - DRR1 (Fam107A). DRR1 is expressed in the mouse brain; it is up-regulated by stress and glucocorticoids and modulates neuronal actin dynamics. In the adult mouse, DRR1 was shown to facilitate specific behaviors which might be protective against some of the deleterious consequences of stress exposure: in the hippocampal CA3 region, DRR1 improved cognitive performance whereas in the septum, it specifically increased social behavior. Therefore DRR1 was suggested as a candidate protein promoting stress-resilience. Fam107B (family with sequence similarity 107, member B) is the unique paralog of DRR1, and both share high sequence similarities, predicted glucocorticoid response elements, heat-shock induction and tumor suppressor properties. So far, the role of Fam107B in the central nervous system was not studied. The aim of the present investigation, therefore, was to analyze whether Fam107B and DRR1 display comparable mRNA expression patterns in the brain and whether both are modulated by stress and glucocorticoids. Spatio-temporal mapping of Fam107B mRNA expression in the embryonic and adult mouse brain, by means of in situ hybridization, showed that Fam107B was expressed during embryogenesis and in the adulthood, with particularly high and specific expression in the forming telencephalon suggestive of an involvement in corticogenesis. In the adult mouse, expression was restricted to neurogenic niches, like the dentate gyrus. In contrast to DRR1, Fam107B mRNA expression failed to be modulated by glucocorticoids and social stress in the adult mouse. In summary, Fam107B and DRR1 show different spatio-temporal expression patterns in the central nervous system, suggesting at least partially different functional roles in the brain, and where the glucocorticoid receptor (GR)-induced regulation appears to be a unique property of DRR1.

Time-dependent metabolomic profiling of Ketamine drug action reveals hippocampal pathway alterations and biomarker candidates.

Ketamine, an N-methyl-D-aspartate receptor (NMDAR) antagonist, has fast-acting antidepressant activities and is used for major depressive disorder (MDD) patients who show treatment resistance towards drugs of the selective serotonin reuptake inhibitor (SSRI) type. In order to better understand Ketamine's mode of action, a prerequisite for improved drug development efforts, a detailed understanding of the molecular events elicited by the drug is mandatory. In the present study we have carried out a time-dependent hippocampal metabolite profiling analysis of mice treated with Ketamine. After a single injection of Ketamine, our metabolomics data indicate time-dependent metabolite level alterations starting already after 2 h reflecting the fast antidepressant effect of the drug. In silico pathway analyses revealed that several hippocampal pathways including glycolysis/gluconeogenesis, pentose phosphate pathway and citrate cycle are affected, apparent by changes not only in metabolite levels but also connected metabolite level ratios. The results show that a single injection of Ketamine has an impact on the major energy metabolism pathways. Furthermore, seven of the identified metabolites qualify as biomarkers for the Ketamine drug response.

Levels and patterns of persistent organic pollutants (POPs) in tilapia (Oreochromis sp.) from four different lakes in Tanzania: geographical differences and implications for human health.

In Tanzania fish is one of the most important protein sources for the rapidly increasing population. Wild fish is threatened by overfishing and pollution from agriculture, industries, mining, household effluents and vector control. To monitor possible implications for public health, the geographical differences of the occurrence and levels of persistent organic pollutants (POPs) in tilapia fish (Oreochromis sp.) from four different Tanzanian lakes were investigated in 2011. Concentrations of organochlorine pesticides (OCPs), polychlorinated biphenyls (PCBs), polybrominated diphenyls (PBDEs) and hexabromocyclododecane (HBCDD) were determined in pooled samples of tilapia muscle from Lake (L) Victoria, L. Tanganyika, L. Nyasa (also called L. Malawi) and L. Babati in Tanzania in 2011. Levels of Σ-DDTs (274 ng/g lipid weight (lw)) and sum of 7 indicator PCBs (Σ-7PCBs) (17 ng/g lw) were significantly higher in tilapia from L. Tanganyika compared to the other lakes. The highest levels of Σ-endosulfan (94 ng/g lw) were detected in tilapia from L. Victoria. Toxaphenes were detected in low levels in fish from L. Tanganyika and L. Babati. Results revealed a geographic difference in the use of DDT and endosulfan between L. Victoria and L. Tanganyika. Low ratios of DDE/DDT in tilapia from L. Tanganyika indicated an on-going use of DDT in the area. Median levels of ΣBDEs, including BDE-209, were highest in L. Victoria (19.4 ng/g lw) and BDE-209 was present in 68% of the samples from this lake. The presence of BDE-209 indicates increasing influence of imported products from heavy industrialized countries. The measured POP levels in the studied tilapia were all below MRLs of EU or were lower than recommended levels, and thus the fish is considered as safe for human consumption. They may, however, pose a risk to the fish species and threaten biodiversity.

Convergent animal and human evidence suggests the activin/inhibin pathway to be involved in antidepressant response.

Despite the overt need for improved treatment modalities in depression, efforts to develop conceptually novel antidepressants have been relatively unsuccessful so far. Here we present a translational approach combining results from hypothesis-free animal experiments with data from a genetic association study in depression. Comparing genes regulated by chronic paroxetine treatment in the mouse hippocampus with genes showing nominally significant association with antidepressant treatment response in two pharmacogenetic studies, the activin pathway was the only one to show this dual pattern of association and therefore selected as a candidate. We examined the regulation of activin A and activin receptor type IA mRNA following antidepressant treatment. We investigated the effects of stereotaxic infusion of activin into the hippocampus and the amygdala in a behavioural model of depression. To analyse whether variants in genes in the activin signalling pathway predict antidepressant treatment response, we performed a human genetic association study. Significant changes in the expression of genes in the activin signalling pathway were observed following 1 and 4 weeks of treatment. Injection of activin A into the hippocampus exerts acute antidepressant-like effects. Polymorphisms in the betaglycan gene, a co-receptor mediating functional antagonism of activin signalling, significantly predict treatment outcome in our system-wide pharmacogenetics study in depression. We provide convergent evidence from mouse and human data that genes in the activin signalling pathway are promising novel candidates involved in the neurobiogical mechanisms underlying antidepressant mechanisms of action. Further, our data suggest this pathway to be a target for more rapid-acting antidepressants in the future.

Cervicothoracic intradural arachnoid cyst misdiagnosed as motor neuron disease.

Recognizing syndromes which mimic ALS is crucial both to avoid giving this diagnosis erroneously and since there may be appropriate treatments. We report a 63-year-old woman diagnosed with possible ALS five years ago based on upper and lower motor neuron signs with typical electrophysiology and normal cranial MRI. At reassessment, spinal MRI revealed a cervicothoracic cyst with cord compression that was successfully treated neurosurgically. Histopathology confirmed an arachnoid origin as suspected from MRI. Spinal cysts may mimic ALS and need to be thoroughly excluded by appropriate imaging.

A novel chronic social stress paradigm in female mice.

Major depression is one of the most prevalent stress-related psychiatric diseases. Next to environmental influences such as chronic social stress, gender is among the strongest risk factors for major depression, with women having a twice as high risk to develop the disease compared to men. While there is abundant literature on the effects of chronic social stress in male rodents, there is a serious lack of information on gender-specific effects. Especially in mice, which due to the wide availability of transgenic lines offer a unique opportunity to study gene x environment interactions, there is no existing model of chronic social stress that is applicable to both sexes. We here describe the effects of chronic social stress based on the disruption of the social network in a group-housed situation in female mice, a model that was recently described and validated for male mice. In this model, the group composition of the mice is changed twice per week for a period of 7 weeks, covering the adolescent and early adulthood period. We observed that housing in an unpredictable social environment resulted in chronic stress in female mice. The observed effects, which included increased adrenal weight, decreased thymus weight, increased corticosterone levels, and increased anxiety-like behavior, were very similar to the described effects of this paradigm in male mice. In addition, we observed a distinct expression of stress system-related genes in female mice following chronic stress exposure. Our results validate this model as a suitable approach to study chronic social stress in female mice and open up the opportunity to use this model with transgenic or knockout mouse lines.

High susceptibility to chronic social stress is associated with a depression-like phenotype.

Chronic stress is a key risk factor for a variety of diseases, including depression. There is a large degree of individual variation in the ability to recover successfully from a chronic stress exposure, but the determinants of this individual stress susceptibility are still poorly understood. We recently developed a novel mouse paradigm for chronic social stress during adolescence, which closely mimics the human condition of chronic social stress in respect to construct, face and predictive validity. By applying this chronic stress model to a large number of animals we aimed at identifying individuals that are either resilient or vulnerable to the persistent effects of chronic social stress exposure. Animals showing markedly elevated basal corticosterone levels 5 weeks following the end of the stress paradigm were considered "vulnerable", whereas individuals recovering quickly and being indistinguishable from controls were classified as "resilient". Stress vulnerability was associated with an increased level of corticotropin-releasing hormone in the paraventricular nucleus, decreased hippocampal mineralocorticoid receptor expression as well as increased anxiety- and depression-like behavior compared to resilient and control animals. In summary, we show that by using a large cohort of animals it is possible to select individuals that are vulnerable or resilient to the lasting effects of chronic social stress. The vulnerable phenotype mimics many aspects of stress-related human affective disorders and this may be used as a novel approach to study depression in an animal model, ultimately contributing to a better understanding and treatment of stress-related disorders.

A single episode of restraint stress regulates central corticotrophin- releasing hormone receptor expression and binding in specific areas of the mouse brain.

The importance of restraining stress-induced activation of the hypothalamic-pituitary-adrenocortical (HPA) system within tolerable limits requires efficient mechanisms for feedback inhibition. Recently, central corticotrophin-releasing hormone (CRH) receptor type 1 (CRHR1) has been shown to mediate HPA system feedback inhibition. To date, most of the data regarding stress-associated expression changes of CRHR1 and CRHR2 mRNA and their ligand CRH have been generated in rats. Taken considerable species differences into consideration, and with the growing importance of transgenic mice, a systematic analysis of the time course of expression changes of CRH and its two receptors in the mouse brain is needed to provide more insight into the regulation of the HPA system, both under physiological and pathophysiological conditions in this species. We analysed in detail the time course of expression changes of CRH, CRHR1 and CRHR2 mRNA after of restraint stress in mice in stress-relevant brain regions (paraventricular nucleus, hippocampus, neocortex). We could show a rapid, strong and long-lasting decrease in cortical and hippocampal CRHR1 mRNA expression after stress, whereas CRHR2 mRNA increased in the same neuroanatomical areas. In situ hybridisation analyses could be further confirmed at the protein level by CRH receptor autoradiography with changes in CRH binding that persisted even 7 days after a single episode of restraint stress. Our observation that stress has opposing effects on CRHR1 and CRHR2 neuronal systems supports the idea that regulation of the relative contribution of the two CRH receptors to brain CRH pathways may be essential in coordinating physiological responses to stress. We further hypothesise that the sustained alteration of CRH receptor expression and binding after a single episode of stress could mediate the long-term effects of stress on neuroendocrine function and emotional regulation.

Chronic social stress during adolescence in mice alters fat distribution in late life: prevention by antidepressant treatment.

Obesity and visceral fat accumulation are key features of the metabolic syndrome that represents one of the main health problems in western societies due to its neurovascular and cardiovascular complications. Epidemiological studies have identified chronic stress exposure as an important risk factor for the development of obesity and metabolic syndrome, but also psychiatric diseases, especially affective disorders. However, it is still unclear if chronic stress has merely transient or potentially lasting effects on body composition. Here, we investigated the effects of chronic social stress during the adolescent period on body fat composition in mice one year after the cessation of the stressor. We found that stress exposure during the adolescent period decreases subcutaneous fat content, without change in visceral fat, and consequently increases the visceral fat/subcutaneous fat ratio in adulthood. Further, we demonstrated that treatment with a selective serotonin reuptake inhibitor (paroxetine) during stress exposure prevented later effects on body fat distribution. These results from a recently validated chronic stress paradigm in mice provide evidence that stressful experiences during adolescence can alter body fat distribution in adulthood, thereby possibly contributing to an increased risk for metabolic diseases. Antidepressant treatment disrupted this effect underlining the link between the stress hormone system, metabolic homeostasis and affective disorders.

Consequences of chronic social stress on behaviour and vasopressin gene expression in the PVN of DBA/2OlaHsd mice--influence of treatment with the CRHR1-antagonist R121919/NBI 30775.

Accumulating evidence suggests that corticotropin-releasing hormone (CRH) neurocircuitry modulate the neuroendocrine and behavioural phenotypes in depression and anxiety. Thus, the administration of the selective CRH-receptor 1 (CRHR1)-antagonist R121919/NBI 30775 has proven its ability to act as an anxiolytic in rats. It is still unclear whether vasopressinergic neuronal circuits, which are known to be involved in the regulation of emotionality, are affected by R121919/NBI 30775. Using DBA/2OlaHsd mice, we investigated the effects of chronic social defeat and concomitant treatment with R121919/NBI 30775 on 1) the behavioural profile in the modified hole board test and 2) in-situ hybridization analysis-based expression of arginine vasopressin (AVP) and CRH mRNA in both the hypothalamic paraventricular nucleus and supraoptic nucleus. The results suggest that chronic social defeat leads to increased avoidance behaviour and reduction in directed exploration, general exploration, and locomotion. Chronic treatment with the CRHR1-antagonist was effective in reversing the directed exploration to control level. The dissection of the antagonist-treated group into responders and non-responders using the parameter time spent on board revealed further positive effects of R121919/NBI 30775 on avoidance behaviour and locomotion. Behavioural changes were accompanied by alterations in AVP gene expression in the paraventricular nucleus. Taken together, the anxiolytic action of the CRHR1 antagonist was found in a subgroup of animals only, and further studies have to be done to clarify the inter-individual biological differences in response patterns to this compound to optimise its application under clinical conditions.

Persistent neuroendocrine and behavioral effects of a novel, etiologically relevant mouse paradigm for chronic social stress during adolescence.

Chronic stress is widely regarded as a key risk factor for a variety of diseases. A large number of paradigms have been used to induce chronic stress in rodents. However, many of these paradigms do not consider the etiology of human stress-associated disorders, where the stressors involved are mostly of social nature and the effects of the stress exposure persist even if the stressor is discontinued. In addition, many chronic stress paradigms are problematic with regard to stress adaptation, continuity, duration and applicability. Here we describe and validate a novel chronic social stress paradigm in male mice during adolescence. We demonstrate persistent effects of chronic social stress after 1 week of rest, including altered adrenal sensitivity, decreased expression of corticosteroid receptors in the hippocampus and increased anxiety. In addition, pharmacological treatments with the antidepressant paroxetine (SSRI) or with the corticotropin-releasing hormone receptor 1 antagonist DMP696 were able to prevent aversive long-term consequences of chronic social stress. In conclusion, this novel chronic stress paradigm results in persistent alterations of hypothalamus-pituitary-adrenal axis function and behavior, which are reversible by pharmacological treatment. Moreover, this paradigm allows to investigate the interaction of genetic susceptibility and environmental risk factors.

Pharmacological validation of a novel home cage activity counter in mice.

Many behavioural tests in rodents are based on the premise that basal locomotor activity of the animals is similar between the tested groups. The measurement of basal home cage activity is therefore an essential parameter, that should be included in all studies which employ tests of anxiety or cognition. Currently available systems for the assessment of home cage locomotion are often complex and expensive. Here we describe and validate a novel, simple and cost-efficient apparatus for the assessment of basic home cage locomotor activity in rodents. Circadian dark-light activity patterns can be reliably obtained with the home cage activity counter. Furthermore, changes in locomotion induced by novelty or pharmacological treatment were reliably and sensitively detected by the apparatus. Thus, the here presented home cage activity counter can be used for the measurement of basal home cage locomotor activity.

Metabolic signals modulate hypothalamic-pituitary-adrenal axis activation during maternal separation of the neonatal mouse.

The postnatal development of the mouse is characterised by a period of hypo-responsiveness of the hypothalamic-pituitary-adrenal (HPA) axis to moderate stressors. Maternal separation disinhibits this blockade of the HPA axis, but the mechanism responsible is not clear. The present study examined the influence of metabolic signals on the central and peripheral components of the HPA axis in neonatal mice aged 8 days in absence or presence of the mother. Reductions in plasma glucose and leptin as well as rapid increases in plasma ghrelin were apparent in the neonate 4 h following maternal deprivation and maximal at 8 h. In addition, maternal separation induced an increase of neuropeptide Y (NPY) mRNA expression in the arcuate nucleus, a decrease of corticotrophin-releasing hormone (CRH) mRNA expression in the paraventricular nucleus and a rise in serum corticosterone. Pharmacological manipulation of the metabolic signals attenuated the HPA response to maternal separation. Thus, the rise in plasma corticosterone induced by maternal separation was ameliorated by prevention of reduction in blood glucose or blockade of the ghrelin signalling pathway, as were the hypothalamic changes in NPY and CRH mRNAs. By contrast, leptin treatment did not affect the HPA axis response to maternal separation. Together these results suggest that metabolic signals play an important role in triggering the HPA response of the neonate to maternal separation.

Differential disinhibition of the neonatal hypothalamic- pituitary-adrenal axis in brain-specific CRH receptor 1-knockout mice.

In the adult, corticotropin-releasing hormone (CRH) is the key mediator for the behavioural and neuroendocrine response to stress. It has also been hypothesized that, during postnatal development of the stress system, CRH controls the activity of the HPA axis and mediates the effects of early disturbances, e.g. 24 h of maternal deprivation. In the current study we investigated the function of specific brain corticotropin-releasing hormone receptor type 1 (CRHR1) subpopulations in the control of the HPA axis during postnatal development under basal conditions as well as after 24 h of maternal deprivation. We used two conditional CRHR1-deficient mouse lines which lack this receptor, either specifically in forebrain and limbic structures (Cam-CRHR1) or in all neurons (Nes-CRHR1). Basal circulating corticosterone was increased in Nes-CRHR1 mice compared to controls. Corticosterone response to maternal deprivation was significantly increased in both CRHR1-deficient lines. In the paraventricular nucleus, Cam-CRHR1 animals displayed enhanced CRH and decreased vasopressin expression levels. In contrast, gene expression in Nes-CRHR1 pups was strikingly similar to that in maternally deprived control pups. Furthermore, maternal deprivation resulted in an enhanced response of Cam-CRHR1 pups in the brain, while expression levels in Nes-CRHR1 mouse pups were mostly unchanged. Our results demonstrate that brainstem and/or hypothalamic CRHR1 contribute to the suppression of basal corticosterone secretion in the neonate, while limbic and/or forebrain CRHR1 dampen the activation of the neonatal HPA axis induced by maternal deprivation.

Mutagenesis and knockout models: hypothalamic-pituitary-adrenocortical system.

Hyperactivity of central neuropeptidergic circuits such as the corticotropin-releasing hormone (CRH) and vasopressin (AVP) neuronal systems is thought to play a causal role in the etiology and symptomatology of anxiety disorders. Indeed, there is increasing evidence from basic science that chronic stress-induced perturbation of CRH and AVP neurocircuitries may contribute to abnormal neuronal communication in conditions of pathological anxiety. Anxiety disorders aggregate in families, and accumulating evidence supports the notion that the major source of familial risk is genetic. In this context, refined molecular technologies and the creation of genetically engineered mice have allowed us to specifically target individual genes involved in the regulation of the elements of the CRH (e.g., CRH peptides, CRH-related peptides, their receptors, binding protein). During the past few years, studies performed in such mice have complemented and extended our knowledge. The cumulative evidence makes a strong case implicating dysfunction of CRH-related systems in the pathogenesis of anxiety disorders and depression and leads us beyond the monoaminergic synapse in search of eagerly anticipated strategies to discover and develop better therapies.