Midbrain projection to the basolateral amygdala encodes anxiety-like but not depression-like behaviors.

Anxiety disorders are complex diseases, and often co-occur with depression. It is as yet unclear if a common neural circuit controls anxiety-related behaviors in both anxiety-alone and comorbid conditions. Here, utilizing the chronic social defeat stress (CSDS) paradigm that induces singular or combined anxiety- and depressive-like phenotypes in mice, we show that a ventral tegmental area (VTA) dopamine circuit projecting to the basolateral amygdala (BLA) selectively controls anxiety- but not depression-like behaviors. Using circuit-dissecting ex vivo electrophysiology and in vivo fiber photometry approaches, we establish that expression of anxiety-like, but not depressive-like, phenotypes are negatively correlated with VTA → BLA dopamine neuron activity. Further, our optogenetic studies demonstrate a causal link between such neuronal activity and anxiety-like behaviors. Overall, these data establish a functional role for VTA → BLA dopamine neurons in bi-directionally controlling anxiety-related behaviors not only in anxiety-alone, but also in anxiety-depressive comorbid conditions in mice.

Mitochondrial localization and moderated activity are key to murine erythroid enucleation.

Mammalian red blood cells (RBCs), which primarily contain hemoglobin, exemplify an elaborate maturation process, with the terminal steps of RBC generation involving extensive cellular remodeling. This encompasses alterations of cellular content through distinct stages of erythroblast maturation that result in the expulsion of the nucleus (enucleation) followed by the loss of mitochondria and all other organelles and a transition to anaerobic glycolysis. Whether there is any link between erythroid removal of the nucleus and the function of any other organelle, including mitochondria, remains unknown. Here we demonstrate that mitochondria are key to nuclear clearance. Using live and confocal microscopy and high-throughput single-cell imaging, we show that before nuclear polarization, mitochondria progressively move toward one side of maturing erythroblasts and aggregate near the nucleus as it extrudes from the cell, a prerequisite for enucleation to proceed. Although we found active mitochondrial respiration is required for nuclear expulsion, levels of mitochondrial activity identify distinct functional subpopulations, because terminally maturing erythroblasts with low relative to high mitochondrial membrane potential are at a later stage of maturation, contain greatly condensed nuclei with reduced open chromatin-associated acetylation histone marks, and exhibit higher enucleation rates. Lastly, to our surprise, we found that late-stage erythroblasts sustain mitochondrial metabolism and subsequent enucleation, primarily through pyruvate but independent of in situ glycolysis. These findings demonstrate the critical but unanticipated functions of mitochondria during the erythroblast enucleation process. They are also relevant to the in vitro production of RBCs as well as to disorders of the erythroid lineage.

Wilm's tumor 1 promotes memory flexibility.

Under physiological conditions, strength and persistence of memory must be regulated in order to produce behavioral flexibility. In fact, impairments in memory flexibility are associated with pathologies such as post-traumatic stress disorder or autism; however, the underlying mechanisms that enable memory flexibility are still poorly understood. Here, we identify transcriptional repressor Wilm's Tumor 1 (WT1) as a critical synaptic plasticity regulator that decreases memory strength, promoting memory flexibility. WT1 is activated in the hippocampus following induction of long-term potentiation (LTP) or learning. WT1 knockdown enhances CA1 neuronal excitability, LTP and long-term memory whereas its overexpression weakens memory retention. Moreover, forebrain WT1-deficient mice show deficits in both reversal, sequential learning tasks and contextual fear extinction, exhibiting impaired memory flexibility. We conclude that WT1 limits memory strength or promotes memory weakening, thus enabling memory flexibility, a process that is critical for learning from new experiences.

α1- and ß3-Adrenergic Receptor-Mediated Mesolimbic Homeostatic Plasticity Confers Resilience to Social Stress in Susceptible Mice.

BACKGROUND: Homeostatic plasticity in mesolimbic dopamine (DA) neurons plays an essential role in mediating resilience to social stress. Recent evidence implicates an association between stress resilience and projections from the locus coeruleus (LC) to the ventral tegmental area (VTA) (LC→VTA) DA system. However, the precise circuitry and molecular mechanisms of the homeostatic plasticity in mesolimbic DA neurons mediated by the LC→VTA circuitry, and its role in conferring resilience to social defeat stress, have not been described. METHODS: In a well-established chronic social defeat stress model of depression, using projection-specific electrophysiological recordings and optogenetic, pharmacological, and molecular profiling techniques, we investigated the functional role and molecular basis of an LC→VTA circuit in conferring resilience to social defeat stress. RESULTS: We found that LC neurons projecting to the VTA exhibit enhanced firing activity in resilient, but not susceptible, mice. Optogenetically mimicking this firing adaptation in susceptible mice reverses their depression-related behaviors, and induces reversal of cellular hyperactivity and homeostatic plasticity in VTA DA neurons projecting to the nucleus accumbens. Circuit-specific molecular profiling studies reveal that α1- and ß3-adrenergic receptors are highly expressed in VTA→nucleus accumbens DA neurons. Pharmacologically activating these receptors induces similar proresilient effects at the ion channel and cellular and behavioral levels, whereas antagonizing these receptors blocks the proresilient effect of optogenetic activation of LC→VTA circuit neurons in susceptible mice. CONCLUSIONS: These findings reveal a key role of the LC→VTA circuit in mediating homeostatic plasticity in stress resilience and reveal α1- and ß3-adrenergic receptors as new molecular targets for therapeutically promoting resilience.

Sex Differences in the Neuroadaptations of Reward-related Circuits in Response to Subchronic Variable Stress.

Women are twice as likely to be diagnosed with major depressive disorder. However, fewer studies in rodent models of depression have used female animals, leading to a relative lack of understanding of the female brain's response to stress, especially at a neural circuit level. In this study, we utilized a 6-day subchronic variable stress (SCVS) mouse model and measured novelty suppressed feeding as behavioral criteria to evaluate susceptibility to SCVS in male and female mice. First, we showed that SCVS induced a decrease in latency to eat (susceptible phenotype) in female mice, but not in males (resilient phenotype). After determining behavioral phenotypes, we investigated the firing activities of dopamine (DA) neurons in the ventral tegmental area (VTA), as well as the neurons that project from lateral habenula (LHb) to the VTA and from locus coeruleus (LC) to the VTA. Utilizing retrograding lumafluor fluorescent tracers and electrophysiology techniques, we performed cell type- and circuit-specific measures of neuronal firing rates. Our data show that SCVS significantly increased the firing rate of LHb-VTA circuit neurons in female mice when compared to that of their female controls, an effect that was absent in SCVS-exposed males. Interestingly, SCVS did not induce significant firing alterations in VTA DA neurons and LC-VTA circuit neurons in either female mice or male mice when compared to their stress-naïve controls. Overall, our data show sex differences in the LHb-VTA circuit responses to SCVS, and implicates a potential role of this projection in mediating vulnerability of female mice to stress-induced depression.

Quantitative Whole-mount Immunofluorescence Analysis of Cardiac Progenitor Populations in Mouse Embryos.

The use of ever-advancing imaging techniques has contributed broadly to our increased understanding of embryonic development. Pre-implantation development and organogenesis are two areas of research that have benefitted greatly from these advances, due to the high quality of data that can be obtained directly from imaging pre-implantation embryos or ex vivo organs. While pre-implantation embryos have yielded data with especially high spatial resolution, later stages have been less amenable to three-dimensional reconstruction. Obtaining high-quality 3D or volumetric data for known embryonic structures in combination with fate mapping or genetic lineage tracing will allow for a more comprehensive analysis of the morphogenetic events taking place during embryogenesis. This protocol describes a whole-mount immunofluorescence approach that allows for the labeling, visualization, and quantification of progenitor cell populations within the developing cardiac crescent, a key structure formed during heart development. The approach is designed in such a way that both cell- and tissue-level information can be obtained. Using confocal microscopy and image processing, this protocol allows for three-dimensional spatial reconstruction of the cardiac crescent, thereby providing the ability to analyze the localization and organization of specific progenitor populations during this critical phase of heart development. Importantly, the use of reference antibodies allows for successive masking of the cardiac crescent and subsequent quantitative measurements of areas within the crescent. This protocol will not only enable a detailed examination of early heart development, but with adaptations should be applicable to most organ systems in the gastrula to early somite stage mouse embryo.

Critical Role of Histone Turnover in Neuronal Transcription and Plasticity.

Turnover and exchange of nucleosomal histones and their variants, a process long believed to be static in post-replicative cells, remains largely unexplored in brain. Here, we describe a novel mechanistic role for HIRA (histone cell cycle regulator) and proteasomal degradation-associated histone dynamics in the regulation of activity-dependent transcription, synaptic connectivity, and behavior. We uncover a dramatic developmental profile of nucleosome occupancy across the lifespan of both rodents and humans, with the histone variant H3.3 accumulating to near-saturating levels throughout the neuronal genome by mid-adolescence. Despite such accumulation, H3.3-containing nucleosomes remain highly dynamic-in a modification-independent manner-to control neuronal- and glial-specific gene expression patterns throughout life. Manipulating H3.3 dynamics in both embryonic and adult neurons confirmed its essential role in neuronal plasticity and cognition. Our findings establish histone turnover as a critical and previously undocumented regulator of cell type-specific transcription and plasticity in mammalian brain.

Cocoa extracts reduce oligomerization of amyloid-ß: implications for cognitive improvement in Alzheimer's disease.

BACKGROUND: Alzheimer's disease (AD) is the most common age-related neurodegenerative disorder, characterized by pathological aggregates of amyloid peptide-ß (Aß) and tau protein. Currently available therapies mediate AD symptoms without modifying disease progression. Polyphenol-rich diets are reported to reduce the risk for AD. OBJECTIVE: In the present study, we investigated the AD disease-modifying effects of cocoa, a rich source of flavanols, which are a class of polyphenols. We hypothesized that cocoa extracts interfere with amyloid-ß oligomerization to prevent synaptic deficits. METHODS: We tested the effects of three different cocoa extracts, viz. Natural, Dutched, and Lavado extracts, on Aß42 and Aß40 oligomerization, using photo-induced cross-linking of unmodified proteins technique. To assess the effects of cocoa extracts on synaptic function, we measured long term potentiation in mouse brain hippocampal slices exposed to oligomeric Aß. RESULTS: Our results indicate that cocoa extracts are effective in preventing the oligomerization of Aß, with Lavado extract being most effective. Lavado extract, but not Dutched extract, was effective in restoring the long term potentiation response reduced by oligomeric Aß. CONCLUSION: Our findings indicate that cocoa extracts have multiple disease-modifying properties in AD and present a promising route of therapeutic and/or preventative initiatives.

Receptor heteromerization expands the repertoire of cannabinoid signaling in rodent neurons.

A fundamental question in G protein coupled receptor biology is how a single ligand acting at a specific receptor is able to induce a range of signaling that results in a variety of physiological responses. We focused on Type 1 cannabinoid receptor (CB1R) as a model GPCR involved in a variety of processes spanning from analgesia and euphoria to neuronal development, survival and differentiation. We examined receptor dimerization as a possible mechanism underlying expanded signaling responses by a single ligand and focused on interactions between CB1R and delta opioid receptor (DOR). Using co-immunoprecipitation assays as well as analysis of changes in receptor subcellular localization upon co-expression, we show that CB1R and DOR form receptor heteromers. We find that heteromerization affects receptor signaling since the potency of the CB1R ligand to stimulate G-protein activity is increased in the absence of DOR, suggesting that the decrease in CB1R activity in the presence of DOR could, at least in part, be due to heteromerization. We also find that the decrease in activity is associated with enhanced PLC-dependent recruitment of arrestin3 to the CB1R-DOR complex, suggesting that interaction with DOR enhances arrestin-mediated CB1R desensitization. Additionally, presence of DOR facilitates signaling via a new CB1R-mediated anti-apoptotic pathway leading to enhanced neuronal survival. Taken together, these results support a role for CB1R-DOR heteromerization in diversification of endocannabinoid signaling and highlight the importance of heteromer-directed signal trafficking in enhancing the repertoire of GPCR signaling.

Synaptic stimulation of mTOR is mediated by Wnt signaling and regulation of glycogen synthetase kinase-3.

The persistent or "late" phase of long-term potentiation (L-LTP), which requires protein synthesis, can be induced by relatively intense synaptic activity. The ability of such strong synaptic protocols to engage the translational machinery and produce plasticity-related proteins, while weaker protocols activate only posttranslational processes and transient potentiation (early LTP; E-LTP), is not understood. Among the major translation control pathways in neurons, the stimulation of mammalian target of rapamycin (mTOR) is a key event in the induction of L-LTP. We report that mTOR is tonically suppressed in rat hippocampus under resting conditions, a consequence of the basal activity of glycogen synthetase kinase 3 (GSK3). This suppression could be overcome by weak synaptic stimulation in the presence of the ß-adrenergic agonist isoproterenol, a combination that induced L-LTP, and activation of mTOR coincided with the Akt-mediated phosphorylation of GSK3. Surprisingly, while isoproterenol alone elevated Akt activity, it failed to increase GSK3 phosphorylation or mTOR signaling, showing that Akt was uncoupled from these effectors in the absence of synaptic stimulation. With the addition of weak stimulation, Akt signaled to GSK3 and mTOR, a gating effect that was mediated by voltage-dependent Ca(2+) channels and the Wnt pathway. mTOR could be stimulated by pharmacological inhibition, enabling weak HFS to induce L-LTP. These results establish GSK3 as an integrator of Akt and Wnt signals and suggest that overcoming GSK3-mediated suppression of mTOR is a key event in the induction of L-LTP by synaptic activity.

Reliability and validity of the Greek version of the Derogatis Psychiatric Rating Scale (DPRS).

BACKGROUND: The aim of this paper is to report on the Greek version of the Derogatis Psychiatric Rating Scale (DPRS) and its validity and reliability when administered to a Greek sample. The DPRS is a clinician-rated measure of psychiatric symptoms with nine basic and eight complementary symptom dimensions and a Global Pathology Index. METHOD: The DPRS was translated into Greek and administered to 161 psychiatric patients and healthy volunteers. Its validity was assessed against the Semistructured Clinical Interview (SCID) and the Symptoms Checklist-90-Revised (SCL-90-R). The inter-rater reliability and test-retest reliability were also evaluated for each symptom dimension. RESULTS: All the symptom dimensions of the DPRS, except euphoria, have concurrent validity with the SCID on the relevant symptom dimensions diagnoses set by the interview. At the cut-off point 2/3 the DPRS has a sensitivity of 98% and a specificity of 74% in indicating active psychiatric patients. The symptom dimensions of the DPRS also have strong correlations with the relevant dimensions of SCL-90-R. The range for inter-rater reliability was from 0.48 to 0.88, the range for test-retest reliability was from 0.25 to 0.85, and the internal consistency was 0.90. CONCLUSION: The Greek version of the DPRS has acceptable psychometric properties, rendering it a useful screening instrument of global psychopathology as well as a multidimensional measure of some basic psychiatric symptoms.

Matricide by person with bipolar disorder and dependent overcompliant personality.

Matricide is an infrequent form of homicide. This paper is to present a case of matricide with typical characteristics of the act but interesting particularities as well. The perpetrator was a 43-year-old man, respected member of his community, with over compliant characteristics, eagerness in serving people and caring his parents, good social adaptation before and after the crime. He abandoned his family and work in order to better serve his old, disabled but over demanding mother who frequently insulted and humiliated him. Suddenly he came to a state of "mental confusion" and strangled her. After the crime, the perpetrator manifested the symptoms of a bipolar disorder and also received the diagnosis of dependant personality disorder. Years later, he presented again a crisis of escalating aggressive urge for which he was hospitalized. Many people and associations of his hometown actively demanded the minimal possible punishment for him. The case is discussed especially concerning: a) hypotheses about the aetiopathogeny of the act, b) the constant support provided to the perpetrator by his family and social environment.

Translation and validation of the Depression Outcomes Module (DOM) in Greece.

In Greece, as in other countries, major depressive disorder is underdiagnosed. Its severity, implications and outcomes are often not adequately evaluated. The Depression Outcomes Module (DOM) was developed in order to meet the need for a global assessment of this disorder. The objective of the current study was to estimate the psychometric properties of DOM in a Greek population presenting depressive symptoms. The DOM was translated into Greek. Patients were examined twice (baseline and follow-up assessment). The psychometric properties of DOM were calculated. Subjects were 83 psychiatric inpatients and outpatients presenting depressive symptoms. The measures used were DOM, Structured Clinical Interview for DSM III-R (SCID) and Hamilton Rating Scale for Depression (Ham-D). The results were: (a) baseline assessment: test-retest reliability k = 0.90, internal consistency 0.93, sensitivity 97%, specificity 90%; (b) follow up assessment: test-retest reliability k = 0.89, sensitivity 81% and specificity 67%. Recovery from depression detected by DOM at the follow-up was significantly correlated both with pharmacotherapy and with a combination of pharmacotherapy and supportive psychotherapy. It was concluded that the Greek version of DOM is a comprehensive, useful instrument for diagnosing, assessing depression and evaluating its outcomes.