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A strong bidirectional link between metabolic and psychiatric disorders exists; yet the molecular basis underlying this interaction remains unresolved. Here we explored the role of the brown fat issue (BAT) as modulatory interface, focusing on the involvement of uncoupling protein 1 (UCP-1), a key metabolic regulator highly expressed in BAT, in the control of emotional behavior.Male and female constitutive UCP-1 knockout (KO) mice were used to investigate the consequences of UCP-1 deficiency on anxiety-related and depression-related behaviors under mild thermogenic (23°C) and thermoneutral (29°C) conditions.UCP-1 KO mice displayed a selective enhancement of anxiety-related behavior exclusively under thermogenic conditions, but not at thermoneutrality. Neural and endocrine stress mediators were not affected in UCP-1 KO mice, which showed an activation of the integrated stress response alongside enhanced Fibroblast-growth factor-21 (FGF-21) levels. However, viral-mediated overexpression of FGF-21 did not phenocopy the behavioral alterations of UCP-1 KO mice and blocking FGF-21 activity did not rescue the anxiogenic phenotype of UCP-1 KO mice. No effects of surgical removal of the intrascapular BAT on anxiety-like behavior or FGF-21 levels were observed in either UCP-1 KO or WT mice.We provide evidence for a novel role of UCP-1 in the regulation of emotions that manifests as inhibitory constraint on anxiety-related behavior, exclusively under thermogenic conditions. We propose this function of UCP-1 to be independent of its activity in the BAT and likely mediated through a central role of UCP-1 in brain regions with converging involvement in energy and emotional control.

Psychostimulants interacting with the dopamine transporter (DAT) can be used illicitly or for the treatment of specific neuropsychiatric disorders. However, they can also pro-duce severe and persistent adverse events. Often, their pharmacological properties in vitro do not fully correlate to their pharmacological profile in vivo. Here, we investigated the pharmacological effects of enantiomers of pyrovalerone, α-pyrrolidinovalerophenone, and 3,4-methylenedioxypyrovalerone as compared to the traditional psychostimulants cocaine and methylphenidate, using a variety of in vitro, computational, and in vivo approaches. We found that in vitro drug-binding kinetics at DAT correlate with the time-course of in vivo psychostimulant action in mice. In particular, a slow dissociation (i.e., slow koff) of S-enantiomers of pyrovalerone analogs from DAT predicts their more persistent in vivo effects when compared to cocaine and methylphenidate. Overall, our findings highlight the critical importance of drug-binding kinetics at DAT for deter-mining the in vivo profile of effects produced by psychostimulant drugs Psychostimulants interacting with the dopamine transporter (DAT) can be used illicitly or for the treatment of specific neuropsychiatric disorders. However, they can also produce severe and persistent adverse events. Often, their pharmacological properties in vitro do not fully correlate to their pharmacological profile in vivo. Here, we investigated the pharmacological effects of enantiomers of pyrovalerone, α-pyrrolidinovalerophenone, and 3,4-methylenedioxypyrovalerone as compared to the traditional psychostimulants cocaine and methylphenidate, using a variety of in vitro, computational, and in vivo approaches. We found that in vitro drug-binding kinetics at DAT correlate with the time-course of in vivo psychostimulant action in mice. In particular, a slow dissociation (i.e., slow koff) of S-enantiomers of pyrovalerone analogs from DAT predicts their more persistent in vivo effects when compared to cocaine and methylphenidate. Overall, our findings highlight the critical importance of drug-binding kinetics at DAT for determining the in vivo profile of effects produced by psychostimulant drugs.

Epidemiological studies indicate a bidirectional association between metabolic disturbances, including obesity and related pathological states, and mood disorders, most prominently major depression. However, the biological mechanisms mediating the comorbid relationship between the deranged metabolic and mood states remain incompletely understood. Here, we tested the hypothesis that the enhanced activation of brown fat tissue (BAT), known to beneficially regulate obesity and accompanying dysfunctional metabolic states, is also paralleled by an alteration of affective behaviour. We used upstream stimulatory factor 1 (USF-1) knock-out (KO) mice as a genetic model of constitutively activated BAT and positive cardiometabolic traits and found a reduction of depression-like and anxiety-like behaviours associated with USF-1 deficiency. Surgical removal of interscapular BAT did not impact the behavioural phenotype of USF-1 KO mice. Further, the absence of USF-1 did not lead to alterations of adult hippocampal neural progenitor cell proliferation, differentiation, or survival. RNA-seq analysis characterised the molecular signature of USF-1 deficiency in the hippocampus and revealed a significant increase in the expression of several members of the X-linked lymphocyte-regulated (xlr) genes, including xlr3b and xlr4b. Xlr genes are the mouse orthologues of the human FAM9 gene family and are implicated in the regulation of dendritic branching, dendritic spine number and morphology. The transcriptional changes were associated with morphological alterations in hippocampal neurons, manifested in reduced dendritic length and complexity in USF-1 KO mice. Collectively these data suggest that the metabolic regulator USF-1 is involved in the control of affective behaviour in mice and that this modulation of mood states is unrelated to USF-1-dependent BAT activation, but reflected in structural changes in the brain.

Immune activation is one of the most common complications during pregnancy, predominantly evoked by viral infections. Nevertheless, how immune activation affects mother–offspring relationships postpartum remains unknown. Here, by using the polyinosinic-polycytidylic acid (Poly I:C) model of gestational infection we show that viral-like immune activation at mid-gestation persistently changes hypothalamic neurocircuit parameters in mouse dams and, consequently, is adverse to parenting behavior. Poly I:C-exposed dams favor non-pup-directed exploratory behavior at the expense of pup retrieval. These behavioral deficits are underlain by dendrite pruning and lesser immediate early gene activation in Galanin (Gal)+ neurons with dam-specific transcriptional signatures that reside in the medial preoptic area (mPOA). Reduced activation of an exclusively inhibitory contingent of these distal-projecting Gal+ neurons allows for increased feed-forward inhibition onto putative dopaminergic neurons in the ventral tegmental area (VTA) in Poly I:C-exposed dams. Notably, destabilized VTA output specifically accompanies post-pup retrieval epochs. We suggest that gestational immunogenic insults bias both threat processing and reward perception, manifesting as disfavored infant caregiving.

First-generation vaccines against SARS-CoV-2 do not provide adequate immune protection. Therefore, we engineered a divalent gene construct combining the receptor-binding domain (RBD) of the spike protein and the immunodominant region of the viral nucleocapsid. This fusion protein was produced in either E. coli or a recombinant baculovirus system. Subsequently, the fusion protein was mixed with adjuvant and administered to mice in a prime-booster mode. Mice (72%) produced an IgG response against both proteins (titer: 10−4–10−5) 14 days after the first booster injection, which was increased to 100% by a second booster. Comparable IgG responses were detected against the delta, gamma and omicron variants of the RBD region. Durability testing revealed IgGs beyond 90 days. In addition, cytolytic effector cell molecules were increased in lymphocytes isolated from peripheral blood. Ex vivo stimulation of T cells by nucleocapsid and RBD peptides showed antigen-specific upregulation of CD44 among the CD4+ and CD8+ T cells of vaccinated mice. No side effect was documented in the central nervous system. Cumulatively, these data represent a proof-of-principle approach alternative to existing mRNA vaccination strategies.

The signal transducer and activator of transcription 3 (STAT3) signalling pathway is activated through phosphorylation by Janus kinases in response to a diverse set of immunogenic and non-immunogenic triggers. Several distinct lines of evidence propose an intricate involvement of STAT3 in neural function relevant to behaviour in health and disease. However, in part due to the pleiotropic effects resulting from its DNA binding activity and the consequent regulation of expression of a variety of genes with context-dependent cellular consequences, the precise nature of STAT3 involvement in the neural mechanisms underlying psychopathology remains incompletely understood. Here, we focused on the midbrain serotonergic system, a central hub for the regulation of emotions, to examine the relevance of STAT3 signalling for emotional behaviour in mice by selectively knocking down raphe STAT3 expression using germline genetic (STAT3 KO) and viral-mediated approaches. Mice lacking serotonergic STAT3 presented with reduced negative behavioural reactivity and a blunted response to the sensitising effects of amphetamine, alongside alterations in midbrain neuronal firing activity of serotonergic neurons and transcriptional control of gene networks relevant for neuropsychiatric disorders. Viral knockdown of dorsal raphe (DR) STAT3 phenocopied the behavioural alterations of STAT3 KO mice, excluding a developmentally determined effect and suggesting that disruption of STAT3 signalling in the DR of adult mice is sufficient for the manifestation of behavioural traits relevant to psychopathology. Collectively, these results suggest DR STAT3 as a molecular gate for the control of behavioural reactivity, constituting a mechanistic link between the upstream activators of STAT3, serotonergic neurotransmission and psychopathology.

Prenatal exposure to infectious or inflammatory insults is increasingly recognized in the etiology of neuropsychiatric diseases, including schizophrenia, autism, depression and bipolar disorder. New discoveries highlight that maternal immune activation can lead to pathological effects on brain and behavior in multiple generations. This review describes the transgenerational consequences of maternal immune activation in shaping brain and behavior anomalies and disease risk across generations. We discuss potential underlying mechanisms of transmission, by which prenatal immune activation can mediate generation-spanning changes in brain development and functions and how external influences could further determine the specificity of the phenotype across generations. The identification of the underlying mechanisms appears relevant to infection-related neuropsychiatric illnesses independently of existing diagnostic classifications and may help identifying complex patterns of generation-spanning transmission beyond genetic inheritance. The herein described principles emphasize the importance of considering ancestral infectious histories in clinical research aiming at developing new preventive treatment strategies against infection-related neurodevelopmental disorders and mental illnesses.

Experimental investigation of early postnatal brain development in infant mice (postnatal day 10) is challenging because of the rapid expansion of their brain volume and the fragility of cranial bones. Stereotaxic adaptors that are commercially available usually evoke significant compression forces, thus limiting the accuracy of micromanipulations. Here, we took advantage of recent progress in 3D printing for the cost-effective production of stereotaxic adaptors for infant mice. Our designs offer gentle head positioning, tailored incisor placement and high precision, while also integrating evaporator and tubing components for gaseous anaesthesia. Thus, post-procedural lethality is minimized even upon time-consuming surgeries. Viral microinjections through glass capillaries are shown as proof of principle to experimentally validate targeting accuracy in mice aged 6-10 days. The exact engineering workflow and printing protocol are disseminated for open-label use. Overall, we present technical details and experimental data on a stereotaxic device that can help neurobiologists working at a particularly complex developmental stage in mice.

Recent and rapidly developing movements relating to the increasing awareness and reports of gender bias, discrimination, and abuse have reached the academic environments. The consideration that negative attitudes toward women and abuse of power creates a hostile environment for female scientists, facilitating sexual harassment and driving women out of science, can be easily related to. Rationally inaccessible gender biases are not only evident at the level of the researchers, but are also paralleled by a corresponding imbalance at the level of the research subjects. Here, we focus on the maternal immune activation (MIA) animal model to illustrate exemplarily the current state of ex-/inclusion of female research subjects and the consideration of sex as biological variable in the basic neurosciences. We demonstrate a strong sex disparity with a major emphasis on male animals in studies examining behavioral and neurochemical alterations in MIA offspring. We put forward the hypothesis that this neglect of female subjects in basic research may stem from a hard-wired sex/gender bias, which may also be reflected in a similar attitude toward female scientists. We suggest exploring the possibility that by dismantling sex bias and male dominance in basic research one would get an additional handle on favorably modifying the perception and appreciation for women in science.

The serotonin transporter (SERT, Slc6a4) plays an important role in the regulation of serotonergic neurotransmission and its aberrant expression has been linked to several psychiatric conditions. While SERT density has been proven to be amenable to in vivo quantitative evaluation by positron emission tomography (PET) in humans, this approach is in its infancy for rodents. Here we set out to evaluate the feasibility of using small-animal PET employing [11C]DASB ([11C]-3-amino-4-(2-dimethylaminomethyl-phenylsulfanyl)-benzonitrile) as a radiotracer to measure SERT density in designated areas of the mouse brain. Using Slc6a4 +/+ , Slc6a4 +/− , and Slc6a4 −/− mice as a genetic model of different SERT expression levels, we showed the feasibility of SERT imaging in the mouse brain with [11C]DASB-PET. The PET analysis was complemented by an evaluation of SERT protein expression using western blot, which revealed a highly significant correlation between in vivo and ex vivo measurements. [11C]DASB-PET was then applied to the examination of dynamic changes of SERT levels in different brain areas in the chronic corticosterone mouse model of chronic stress. The observed significant reduction in SERT density in corticosterone-treated mice was independently validated by and correlated with western blot analysis. This is the first demonstration of a quantitative in vivo evaluation of SERT density in subregions of the mouse brain using [11C]DASB-PET. The evidenced decrease in SERT density in response to chronic corticosterone treatment adds a new dimension to the complex involvement of SERT in the pathophysiology of stress-induced mental illnesses.

Acute intermittent porphyria (AIP) is an inborn error of heme biosynthesis due to the deficiency of hydroxymethylbilane synthase (HMBS) activity. Human AIP heterozygotes have episodic acute neurovisceral attacks that typically start after puberty, whereas patients with homozygous dominant AIP (HD-AIP) have early-onset chronic neurological impairment, including ataxia and psychomotor retardation. To investigate the dramatically different manifestations, knock-in mice with human HD-AIP missense mutations c.500G>A (p.Arg167Glu) or c.518_519GC>AG (p.Arg173Glu), designated R167Q or R173Q mice, respectively, were generated and compared with the previously established T1/T2 mice with ~30% residual HMBS activity and the heterozygous AIP phenotype. Homozygous R173Q mice were embryonic lethal, while R167Q homozygous mice (R167Q(+/+)) had ~5% of normal HMBS activity, constitutively elevated plasma and urinary 5-aminolevulinic acid (ALA) and porphobilinogen (PBG), profound early-onset ataxia, delayed motor development and markedly impaired rotarod performance. Central nervous system (CNS) histology was grossly intact, but CNS myelination was delayed and overall myelin volume was decreased. Heme concentrations in liver and brain were similar to those of T1/T2 mice. Notably, ALA and PBG concentrations in the cerebral spinal fluid and CNS regions were markedly elevated in R167Q(+/+) mice compared with T1/T2 mice. When the T1/T2 mice were administered phenobarbital, ALA and PBG markedly accumulated in their liver and plasma, but not in the CNS, indicating that ALA and PBG do not readily cross the blood–brain barrier. Taken together, these studies suggest that the severe HD-AIP neurological phenotype results from decreased myelination and the accumulation of locally produced neurotoxic porphyrin precursors within the CNS.

Learned safety is a fear inhibitory mechanism, which regulates fear responses, promotes episodes of safety and generates positive affective states. Despite its potential as experimental model for several psychiatric illnesses, including post-traumatic stress disorder and depression, the molecular mechanisms of learned safety remain poorly understood, We here investigated the molecular mediators of learned safety, focusing on the characterization of miRNA expression in the basolateral amygdala (BLA). Comparing levels of 22 miRNAs in learned safety and learned fear trained mice, six safety-related miRNAs, including three members of the miR-132/-212 family, were identified. A gain-of-function approach based upon in-vivo transfection of a specific miRNA mimic, and miR-132/212 knock-out mice as loss-of-function tool were used in order to determine the relevance of miR-132 for learned safety at the behavioral and the neuronal functional levels. Using a designated bioinformatic approach, PTEN and GAT1 were identified as potential novel miR-132 target genes and further experimentally validated. We here firstly provide evidence for a regulation of amygdala miRNA expression in learned safety and propose miR-132 as signature molecule to be considered in future preclinical and translational approaches testing the transdiagnostic relevance of learned safety as intermediate phenotype in fear and stress-related disorders.

Acute intermittent porphyria (AIP) is an autosomal dominant inborn error of heme biosynthesis due to a pathogenic mutation in the Hmbs gene, resulting in half-normal activity of hydroxymethylbilane synthase. Factors that induce hepatic heme biosynthesis induce episodic attacks in heterozygous patients. The clinical presentation of acute attacks involves the signature neurovisceral pain and may include psychiatric symptoms. Here we used a knock-in mouse line that is biallelic for the Hmbs c.500G > A (p.R167Q) mutation with ~ 5% of normal hydroxymethylbilane synthase activity to unravel the consequences of severe HMBS deficiency on affective behavior and brain physiology. Hmbs knock-in mice (KI mice) model the rare homozygous dominant form of AIP and were used as tool to elucidate the hitherto unknown pathophysiology of the behavioral manifestations of the disease and its neural underpinnings. Extensive behavioral analyses revealed a selective depression-like phenotype in Hmbs KI mice; transcriptomic and immunohistochemical analyses demonstrated aberrant myelination. The uncovered compromised mitochondrial function in the hippocampus of knock-in mice and its ensuing neurogenic and neuroplastic deficits lead us to propose a mechanistic role for disrupted mitochondrial energy production in the pathogenesis of the behavioral consequences of severe HMBS deficiency and its neuropathological sequelae in the brain.

Mice lacking the epidermal growth factor receptor (EGFR) develop an early postnatal degeneration of the frontal cortex and olfactory bulbs and show increased cortical astrocyte apoptosis. The poor health and early lethality of EGFR-/- mice prevented the analysis of mechanisms responsible for the neurodegeneration and function of the EGFR in the adult brain. Here, we show that postnatal EGFR-deficient neural stem cells are impaired in their self-renewal potential and lack clonal expansion capacity in vitro. Mice lacking the EGFR in the brain (EGFRΔbrain ) show low penetrance of cortical degeneration compared to EGFR-/- mice despite genetic recombination of the conditional allele. Adult EGFRΔ mice establish a proper blood-brain barrier and perform reactive astrogliosis in response to mechanical and infectious brain injury, but are more sensitive to Kainic acid-induced epileptic seizures. EGFR-deficient cortical astrocytes, but not midbrain astrocytes, have reduced expression of glutamate transporters Glt1 and Glast, and show reduced glutamate uptake in vitro, illustrating an excitotoxic mechanism to explain the hypersensitivity to Kainic acid and region-specific neurodegeneration observed in EGFR-deficient brains.

Maternal immune activation (MIA) models that are based on administration of the viral mimetic, poly(I:C), are widely used as experimental tools to study neuronal and behavioral dysfunctions in relation to immune-mediated neurodevelopmental disorders and mental illnesses. Evidence from investigations in non-pregnant rodents suggests that different poly(I:C) products can vary in terms of their immunogenicity, even if they are obtained from the same vendor. The present study aimed at extending these findings to pregnant mice, while also controlling various poly(I:C) products for potential contamination with lipopolysaccharide (LPS). We found significant variability between different batches of poly(I:C) potassium salt obtained from the same vendor (Sigma-Aldrich) in terms of the relative amount of dsRNA fragments in the high molecular weight range (1000-6000 nucleotides long) and with regards to their effects on maternal thermoregulation and immune responses in maternal plasma, placenta and fetal brain. Batches of poly(I:C) potassium salt containing larger amounts of high molecular weight fragments induced more extensive effects on thermoregulation and immune responses compared to batches with minimal amounts of high molecular weight fragments. Consistent with these findings, poly(I:C) enriched for high molecular weight dsRNA (HMW) caused larger maternal and placental immune responses compared to low molecular weight (LMW) poly(I:C). These variable effects were unrelated to possible LPS contamination. Finally, we found marked variability between different batches of the poly(I:C) potassium salt in terms of their effects on spontaneous abortion rates. This batch-to-batch variability was confirmed by three independent research groups using distinct poly(I:C) administration protocols in mice. Taken together, the present data confirm that different poly(I:C) products can induce varying immune responses and can differentially affect maternal physiology and pregnancy outcomes. It is therefore pivotal that researchers working with poly(I:C)-based MIA models ascertain and consider the precise molecular composition and immunogenicity of the product in use. We recommend the establishment of reference databases that combine phenotype data with empirically acquired quality information, which can aid the design, implementation and interpretation of poly(I:C)-based MIA models.

Dementia is a progressive cognitive diminution impeding with normal daily activities that is constantly on the increase. Currently, the estimated prevalence is 50 million affected people worldwide, a figure expected to triple within the next 30 years. While the pathophysiology of the different types of dementia is complex, likely involving the interplay between multiple genetic and environmental factors, strong evidence points towards an important link between diet and cognitive health. Here we examined the consequences of high-fat, high-sugar Western diet (HFSD)-induced obesity on cognitive performance in the fear conditioning task in mice and explored a possible beneficial effect of 6-shogaol (6S), an active constituent of ginger, in this model. Chronic exposure to HFSD significantly enhanced body weight gain in C57BL/6N mice and this effect was prevented by treatment with 6S. HFSD + vehicle-treated mice presented with a selective deficit in cued fear memory, which was not observed in HFSD + 6S-treated animals. The findings of this study provide first evidence for a beneficial effect of 6S on HFSD-induced obesity and emotional memory deficit in mice.

The highly conserved transcription factor LIM-only 3 (Lmo3) is involved in important neurodevelopmental processes in several brain areas including the amygdala, a central hub for the generation and regulation of emotions. Accordingly, a role for Lmo3 in the behavioral responses to ethanol and in the display of anxiety-like behavior in mice has been demonstrated while the potential involvement of Lmo3 in the control of mood-related behavior has not yet been explored. Using a mouse model of Lmo3 depletion (Lmo3z), we here report that genetic Lmo3 deficiency is associated with altered performance in behavioral paradigms assessing anxiety-like and depression-like traits and additionally accompanied by impairments in learned fear. Importantly, long-term potentiation (LTP) in the basolateral amygdala (BLA), a proposed cellular correlate of fear learning, is impaired in Lmo3z mice. RNA-Seq analysis of BLA tissue and gene set enrichment analysis (GSEA) of differentially expressed genes in Lmo3z mice reveals a significant overlap between genes overexpressed in Lmo3z mice and those enriched in the amygdala of a cohort of patients suffering from major depressive disorder. Consequently, we propose that Lmo3 may play a role in the regulation of gene networks that are relevant to the regulation of emotions. Future work may aid to further explore the role of Lmo3 in the pathophysiology of affective disorders and its genetic foundations.

Gestational infection constitutes a risk factor for the occurrence of psychiatric disorders in the offspring. Activation of the maternal immune system (MIA) with subsequent impact on the development of the fetal brain is considered to form the neurobiological basis for aberrant neural wiring and the psychiatric manifestations later in offspring life. The examination of validated animal models constitutes a premier resource for the investigation of the neural underpinnings. Here we used a mouse model of MIA based upon systemic treatment of pregnant mice with Poly(I:C) (polyriboinosinic-polyribocytidilic acid), for the unbiased and comprehensive analysis of the impact of MIA on adult offspring brain activity, morphometry, connectivity and function by a magnetic resonance imaging (MRI) approach. Overall lower neural activity, smaller brain regions and less effective fiber structure were observed for Poly(I:C) offspring compared to the control group. The corpus callosum was significantly smaller and presented with a disruption in myelin/ fiber structure in the MIA progeny. Subsequent resting-state functional MRI experiments demonstrated a paralleling dysfunctional interhemispheric connectivity. Additionally, while the overall flow of information was intact, cortico-limbic connectivity was hampered and limbic circuits revealed hyperconnectivity in Poly(I:C) offspring. Our study sheds new light on the impact of maternal infection during pregnancy on the offspring brain and identifies aberrant resting-state functional connectivity patterns as possible correlates of the behavioral phenotype with relevance for psychiatric disorders.

Major depressive disorder (MDD) is one of the most debilitating psychiatric diseases, affecting a large percentage of the population worldwide. Currently, the underlying pathomechanisms remain incompletely understood, hampering the development of critically needed alternative therapeutic strategies, which further largely depends on the availability of suitable model systems. Here we used a mouse model of early life stress – a precipitating factor for the development of MDD – featuring infectious stress through maternal immune activation (MIA) by polyinosinic:polycytidilic acid (Poly(I:C)) to examine epigenetic modulations as potential molecular correlates of the alterations in brain structure, function and behavior. We found that in adult female MIA offspring anhedonic behavior was associated with modulations of the global histone acetylation profile in the hippocampus. Morevoer, specific changes at the promoter and in the expression of the serotonin transporter (SERT), critically involved in the etiology of MDD and pharmacological antidepressant treatment were detected. Furthermore, an accompanying reduction in hippocampal levels of histone deacetylase (HDAC) 1 was observed in MIA as compared to control offspring. Based on these results we propose a model in which the long-lasting impact of MIA on depression-like behavior and associated molecular and cellular aberrations in the offspring is brought about by the modulation of epigenetic processes and consequent enduring changes in gene expression. These data provide additional insights into the principles underlying the impact of early infectious stress on the development of MDD and may contribute to the development of new targets for antidepressant therapy., Graphical abstract

The serotonin transporter (SERT, Slc6a4) plays an important role in the regulation of serotonergic neurotransmission and its aberrant expression has been linked to several psychiatric conditions. While SERT density has been proven to be amenable to in vivo quantitative evaluation by positron emission tomography (PET) in humans, this approach is in its infancy for rodents. Here we set out to evaluate the feasibility of using small-animal PET employing [11C]DASB ([11C]-3-amino-4-(2-dimethylaminomethyl-phenylsulfanyl)-benzonitrile) as a radiotracer to measure SERT density in designated areas of the mouse brain. Using Slc6a4+/+, Slc6a4+/−, and Slc6a4−/− mice as a genetic model of different SERT expression levels, we showed the feasibility of SERT imaging in the mouse brain with [11C]DASB-PET. The PET analysis was complemented by an evaluation of SERT protein expression using western blot, which revealed a highly significant correlation between in vivo and ex vivo measurements. [11C]DASB-PET was then applied to the examination of dynamic changes of SERT levels in different brain areas in the chronic corticosterone mouse model of chronic stress. The observed significant reduction in SERT density in corticosterone-treated mice was independently validated by and correlated with western blot analysis. This is the first demonstration of a quantitative in vivo evaluation of SERT density in subregions of the mouse brain using [11C]DASB-PET. The evidenced decrease in SERT density in response to chronic corticosterone treatment adds a new dimension to the complex involvement of SERT in the pathophysiology of stress-induced mental illnesses.

Learned safety is a fear inhibitory mechanism, which regulates fear responses, promotes episodes of safety and generates positive affective states. Despite its potential as experimental model for several psychiatric illnesses, including post-traumatic stress disorder and depression, the molecular mechanisms of learned safety remain poorly understood, We here investigated the molecular mediators of learned safety, focusing on the characterization of miRNA expression in the basolateral amygdala (BLA). Comparing levels of 22 miRNAs in learned safety and learned fear trained mice, six safety-related miRNAs, including three members of the miR-132/-212 family, were identified. A gain-of-function approach based upon in-vivo transfection of a specific miRNA mimic, and miR-132/212 knock-out mice as loss-of-function tool were used in order to determine the relevance of miR-132 for learned safety at the behavioral and the neuronal functional levels. Using a designated bioinformatic approach, PTEN and GAT1 were identified as potential novel miR-132 target genes and further experimentally validated. We here firstly provide evidence for a regulation of amygdala miRNA expression in learned safety and propose miR-132 as signature molecule to be considered in future preclinical and translational approaches testing the transdiagnostic relevance of learned safety as intermediate phenotype in fear and stress-related disorders.

Maternal immune activation (MIA) is a well-established model for the investigation of the deleterious effects of gestational infection on offspring mental health later in life. Hence, MIA represents a critical environmental variable determining brain development and the depending neural and behavioral functions in the progeny. Transgenerational transmission of some of the effects of MIA has been recently reported using the Polyinosinic:polycytidylic acid (Poly (I:C)) MIA model in C57BL/6 (C57) inbred mice. However, little is known about the underlying molecular mechanisms and the possible relevance of the specific genetic make-up of the inbred mouse strain used. Here we set out to characterize the effects of gestational Poly (I:C) treatment in C3H/HeNCrl mice (C3H), focusing on maternal care and offspring depression-like behavior and its intergenerational potential. miRNA expression in the offspring hippocampus in the F1 and F2 generations was examined as possible mechanism contributing to the observed behavioral effects. The impact of MIA on maternal care and its transmission to F1 females was previously observed in C57 mice was also found in C3H mice. Depression-like behavior in the adult offspring in C3H F1 and F2 females differed from reports of the C57 strain in the literature, suggesting a potential modulating role of the genetic background in the Poly(I:C) MIA mouse model. As the pattern of expression of selected candidate miRNAs in the F1 and F2 offspring hippocampus was not conserved between the two generations, it is unlikely to be a direct consequence of altered maternal care, or to be an immediate determinant of offspring emotionality.

The ability to recognize and properly respond to instances of protection from impending danger is critical for preventing chronic stress and anxiety—central symptoms of anxiety and affective disorders afflicting large populations of people. Learned safety encompasses learning processes, which lead to the identification of episodes of security and regulation of fear responses. On the basis of insights into the neural circuitry and molecular mechanisms involved in learned safety in mice and humans, we describe learned safety as a tool for understanding neural mechanisms involved in the pathomechanisms of specific affective disorders. This review summarizes our current knowledge on the neurobiological underpinnings of learned safety and discusses potential applications in basic and translational neurosciences.

Gestational infection is increasingly being recognized for its involvement as causative mechanism in severe developmental brain abnormalities and its contribution to the pathogenesis of psychopathologies later in life. First observations in the widely accepted maternal immune activation (MIA) model based upon the systemic administration of the viral mimetic Polyinosinic:polycytidylic acid (poly(I:C)) have recently suggested a transmission of behavioral and transcriptional traits across generations. Although maternal care behavior (MCB) is known as essential mediator of the transgenerational effects of environmental challenges on offspring brain function and behavior, the possible propagation of alterations of MCB resulting from MIA to following generations has not yet been examined. Here we show that poly(I:C) stimulation at embryonic day 12.5 (E12.5) leads to aberrant MCB and that this effect is transmitted to the female F1 offspring. The transgenerational effects on MCB are paralleled by enhanced depression-like behavior in the second generation F2 offspring with contributions of both maternal and paternal heritages. Examination of offspring hippocampal expression of genes known as targets of MCB and relevant for ensuing non-genetic transmission of altered brain function and behavior revealed transgenerationally conserved and modified expressional patterns in the F1 and F2 generation. Collectively these data firstly demonstrate the transgenerational transmission of the impact of gestational immune activation on the reproductive care behavior of the mother. Behavioral and molecular characteristics of first and second generation offspring suggest transgenerationally imprinted consequences of gestational infection on psychopathological traits related to mood disorders which remain to be examined in future cross-fostering experiments.

The lymphocyte-specific protein tyrosine kinase (Lck), which belongs to the Src kinase-family, is expressed in neurons of the hippocampus, a structure critical for learning and memory. Recent evidence demonstrated a significant downregulation of Lck in Alzheimer's disease. Lck has additionally been proposed to be a risk factor for Alzheimer's disease, thus suggesting the involvement of Lck in memory function. The neuronal role of Lck, however, and its involvement in learning and memory remain largely unexplored. Here, in vitro electrophysiology, confocal microscopy, and molecular, pharmacological, genetic and biochemical techniques were combined with in vivo behavioral approaches to examine the role of Lck in the mouse hippocampus. Specific pharmacological inhibition and genetic silencing indicated the involvement of Lck in the regulation of neuritic outgrowth. In the functional pre-established synaptic networks that were examined electrophysiologically, specific Lck-inhibition also selectively impaired the long-term hippocampal synaptic plasticity without affecting spontaneous excitatory synaptic transmission or short-term synaptic potentiation. The selective inhibition of Lck also significantly altered hippocampus-dependent spatial learning and memory in vivo. These data provide the basis for the functional characterization of brain Lck, describing the importance of Lck as a critical regulator of both neuronal morphology and in vivo long-term memory.

Background. The gastrin-releasing peptide receptor (GRPR) is highly expressed in the limbic system, where it importantly regulates emotional functions and in the suprachiasmatic nucleus, where it is central for the photic resetting of the circadian clock. Mice lacking GRPR presented with deficient light-induced phase shift in activity as well altered emotional learning and amygdala function. The effect of GRPR deletion on depression-like behavior and its molecular signature in the amygdala, however, has not yet been evaluated. Methods. GRPR knock-out mice (GRPR-KO) were tested in the forced-swim test and the sucrose preference test for depression-like behavior. Gene expression in the basolateral nucleus of the amygdala was evaluated by micorarray analysis subsequent to laser-capture microdissection-assisted extraction of mRNA. The expression of selected genes was confirmed by RT-PCR. Results. GRPR-KO mice were found to present with increased depression-like behavior. Microarray analysis revealed down-regulation of several glucocorticoid-responsive genes in the basolateral amygdala. Acute administration of dexamethasone reversed the behavioral phenotype and alterations in gene expression. Discussion. We propose that deletion of GRPR leads to the induction of depression-like behavior which is paralleled by dysregulation of amygdala gene expression, potentially resulting from deficient light-induced corticosterone release in GRPR-KO.

Aberrant serotonergic neurotransmission in the brain is considered at the core of the pathophysiological mechanisms involved in neuropsychiatric disorders. Gene by environment interactions contribute to the development of depression and involve modulation of the availability and functional activity of the serotonin transporter (SERT). Using behavioral and in vivo electrophysiological approaches together with biochemical, molecular-biological and molecular imaging tools we establish Flotillin-1 (Flot1) as a novel protein interacting with SERT and demonstrate its involvement in the response to chronic corticosterone (CORT) treatment. We show that genetic Flot1 depletion augments chronic CORT-induced behavioral despair and describe concomitant alterations in the expression of SERT, activity of serotonergic neurons and alterations of the glucocorticoid receptor transport machinery. Hence, we propose a role for Flot1 as modulatory factor for the depressogenic consequences of chronic CORT exposure and suggest Flotillin-1-dependent regulation of SERT expression and activity of serotonergic neurotransmission at the core of the molecular mechanisms involved.

Background. Learned safety is established by negatively correlating the occurrence of a neutral stimulus and a noxious stimulus, which renders the previously neutral stimulus a ‘safety signal’. While the neurophysiological and molecular mechanisms have been characterized in mice, it is currently not known how the neural substrates involved compare between mice and people. Methods. Here we attempt to adapt the original animal protocol to humans and use functional magnetic resonance imaging to examine neural responses to the conditioned stimulus in safety conditioned and fear conditioned subjects. Diffusion tensor imaging (DTI) was used in a parallel group of subjects as a fi rst approach to delineate the underlying neural circuitry. Results. Learned safety is associated with dampened amygdala and increased dorsolateral prefrontal cortex and caudate responses and paralleled by pupillary constriction. A neural connection between the amygdala and the dorsolateral prefrontal cortex is suggested by DTI. Conclusion. We present a translational bridge between mouse and human models of learned safety in which cellular and molecular insights from animal experiments are extended to the human neural circuitry. This study provides an example of how animal experiments can be used to inform and target human studies, which in turn can corroborate results obtained in experimental animals.

Depression, among other mood disorders, represents one of the most common health problems worldwide, with steadily increasing incidence and major socio-economic consequences. However, since the knowledge about the underlying pathophysiological principles is still very scanty, depression and other mood disorders are currently diagnosed solely on clinical grounds. Currently used treatment modalities would therefore benefit enormously from the development of alternative therapeutic interventions. The implementation of proper animal models is a prerequisite for increasing the understanding of the neurobiological basis of mood disorders and is paving the way for the discovery of novel therapeutic targets. In the past thirty years, since the seminal description of the Forced Swim Test as a system to probe antidepressant activity in rodents, the use of animals to model depression and antidepressant activity has come a long way. In this review we describe some of the most commonly used strategies, ranging from screening procedures, such as the Forced Swim Test and the Tail Suspension Test and animal models, such as those based upon chronic stress procedures, to genetic approaches. Finally we also discuss some of the inherent limitations and caveats that need to be considered when using animals as models for mental disorders in basic research.

Corrigendum: STAT3 controls IL6-dependent regulation of serotonin transporter function and depression-like behaviour

With the advent of proteomics technologies it is possible to simultaneously demonstrate the expression of hundreds of proteins. The information offered by proteomics provides context-based understanding of cellular protein networks and has been proven to be a valuable approach in neuroscience studies. The mouse hippocampus has been a major target of analysis in the search for molecular correlates to neuronal information storage. Although human and rat hippocampal samples have been successfully subjected to proteomic profiling, no elaborate analysis providing the fundamental experimental basis for protein-expression studies in the mouse hippocampus has been carried out as yet. This led us to construct a master map generated from the individual hippocampal proteomes of five different mouse strains. A proteomic approach, based upon 2-DE coupled to MS (MALDI-TOF/TOF) has been chosen in an attempt to establish a comprehensive reference database of proteins expressed in the mouse hippocampus. 469 individual proteins, represented by 1156 spots displaying various functional states of the respective gene products were identified. Proteomic profiling of the hippocampus, a brain region with a pivotal role for neuronal information processing and storage may provide insight into the characteristics of proteins serving this highly sophisticated function.

Individual mouse strains differ significantly in terms of behaviour, cognitive function and long-term potentiation. Hippocampal gene expression profiling of eight different mouse strains points towards strain-specific regulation of genes involved in neuronal information storage. Protein expression with regard to strain- dependent expression of structures related to neuronal information storage has not been investigated yet. Herein, a proteomic approach based on two-dimensional gel electrophoresis coupled with mass spectrometry (MALDI-TOF/TOF) has been chosen to address this question by determining strain-dependent expression of proteins involved in neurotransmission and activity-induced actin remodelling in hippocampal tissue of five mouse strains. Of 31 spots representing 16 different gene products analysed and quantified, N-ethylmaleimide-sensitive fusion protein, N-ethylmaleimide-sensitive factor attachment protein-alpha, actin-like protein 3, profilin and cofilin were expressed in a strain-dependent manner. By treating protein expression as a phenotype, we have shown significant genetic variation in brain protein expression. Further experiments in this direction may provide an indication of the genetic elements that contribute to the phenotypic differences between the selected strains through the expressional level of the translated protein. In view of this, we propose that proteomic analysis enabling to concomitantly survey the expression of a large number of proteins could serve as a valuable tool for genetic and physiological studies of central nervous system function.

Increasing epidemiological and experimental evidence implicates gestational infections as one important factor involved in the pathogenesis of several neuropsychiatric disorders. Corresponding preclinical model systems based upon maternal immune activation (MIA) by treatment of the pregnant female have been developed. These MIA animal model systems have been successfully used in basic and translational research approaches, contributing to the investigation of the underlying pathophysiological mechanisms at the molecular, cellular and behavioral levels. The present article focuses on the application of a specific MIA rodent paradigm, based upon treatment of the gestating dam with the viral mimic polyinosinic-polycytidilic acid (Poly(I:C)), a synthetic analog of double-stranded RNA (dsRNA) which activates the Toll-like receptor 3 (TLR3) pathway. Important advantages and constraints of this animal model will be discussed, specifically in light of gestational infection as one vulnerability factor contributing to the complex etiology of mood and psychotic disorders, which are likely the result of intricate multi-level gene×environment interactions. Improving our currently incomplete understanding of the molecular pathomechanistic principles underlying these disorders is a prerequisite for the development of alternative therapeutic approaches which are critically needed in light of the important drawbacks and limitations of currently available pharmacological treatment options regarding efficacy and side effects. The particular relevance of the Poly(I:C) MIA model for the discovery of novel drug targets for symptomatic and preventive therapeutic strategies in mood and psychotic disorders is highlighted in this review article.