Brain-Region-Specific Genes Form the Major Pathways Featuring Their Basic Functional Role: Their Implication in Animal Chronic Stress Model.

The analysis of RNA-Sec data from murine bulk tissue samples taken from five brain regions associated with behavior and stress response was conducted. The focus was on the most contrasting brain region-specific genes (BRSG) sets in terms of their expression rates. These BRSGs are identified as genes with a distinct outlying (high) expression rate in a specific region compared to others used in the study. The analysis suggested that BRSG sets form non-randomly connected compact gene networks, which correspond to the major neuron-mediated functional processes or pathways in each brain region. The number of BRSGs and the connection rate were found to depend on the heterogeneity and coordinated firing rate of neuron types in each brain region. The most connected pathways, along with the highest BRSG number, were observed in the Striatum, referred to as Medium Spiny Neurons (MSNs), which make up 95% of neurons and exhibit synchronous firing upon dopamine influx. However, the Ventral Tegmental Area/Medial Raphe Nucleus (VTA/MRN) regions, although primarily composed of monoaminergic neurons, do not fire synchronously, leading to a smaller BRSG number. The Hippocampus (HPC) region, on the other hand, displays significant neuronal heterogeneity, with glutamatergic neurons being the most numerous and synchronized. Interestingly, the two monoaminergic regions involved in the study displayed a common BRSG subnetwork architecture, emphasizing their proximity in terms of axonal throughput specifics and high-energy metabolism rates. This finding suggests the concerted evolution of monoaminergic neurons, leading to unique adaptations at the genic repertoire scale. With BRSG sets, we were able to highlight the contrasting features of the three groups: control, depressive, and aggressive mice in the animal chronic stress model. Specifically, we observed a decrease in serotonergic turnover in both the depressed and aggressive groups, while dopaminergic emission was high in both groups. There was also a notable absence of dopaminoceptive receptors on the postsynaptic membranes in the striatum in the depressed group. Additionally, we confirmed that neurogenesis BRSGs are specific to HPC, with the aggressive group showing attenuated neurogenesis rates compared to the control/depressive groups. We also confirmed that immune-competent cells like microglia and astrocytes play a crucial role in depressed phenotypes, including mitophagy-related gene Prkcd. Based on this analysis, we propose the use of BRSG sets as a suitable framework for evaluating case-control group-wise assessments of specific brain region gene pathway responses.

Maturation of cortical input to dorsal raphe nucleus increases behavioral persistence in mice.

The ability to persist toward a desired objective is a fundamental aspect of behavioral control whose impairment is implicated in several behavioral disorders. One of the prominent features of behavioral persistence is that its maturation occurs relatively late in development. This is presumed to echo the developmental time course of a corresponding circuit within late-maturing parts of the brain, such as the prefrontal cortex, but the specific identity of the responsible circuits is unknown. Here, we used a genetic approach to describe the maturation of the projection from layer 5 neurons of the neocortex to the dorsal raphe nucleus in mice. Using optogenetic-assisted circuit mapping, we show that this projection undergoes a dramatic increase in synaptic potency between postnatal weeks 3 and 8, corresponding to the transition from juvenile to adult. We then show that this period corresponds to an increase in the behavioral persistence that mice exhibit in a foraging task. Finally, we used a genetic targeting strategy that primarily affected neurons in the medial prefrontal cortex, to selectively ablate this pathway in adulthood and show that mice revert to a behavioral phenotype similar to juveniles. These results suggest that frontal cortical to dorsal raphe input is a critical anatomical and functional substrate of the development and manifestation of behavioral persistence.

The avBNSTGABA-VTA and avBNSTGABA-DRN pathways are respectively involved in the regulation of anxiety-like behaviors in parkinsonian rats.

The anteroventral bed nucleus of stria terminalis (avBNST) is a key brain region which involves negative emotional states, such as anxiety. The most neurons in the avBNST are GABAergic, and it sends GABAergic projections to the ventral tegmental area (VTA) and the dorsal raphe nucleus (DRN), respectively. The VTA and DRN contain dopaminergic and serotonergic cell groups in the midbrain which regulate anxiety-like behaviors. However, it is unclear the role of GABAergic projections from the avBNST to the VTA and the DRN in the regulation of anxiety-like behaviors, particularly in Parkinson's disease (PD)-related anxiety. In the present study, unilateral 6-hydroxydopamine (6-OHDA) lesions of the substantia nigra pars compacta in rats induced anxiety-like behaviors, and decreased level of dopamine (DA) in the basolateral amygdala (BLA). Chemogenetic activation of avBNSTGABA-VTA or avBNSTGABA-DRN pathway induced anxiety-like behaviors and decreased DA or 5-HT release in the BLA in sham and 6-OHDA rats, while inhibition of avBNSTGABA-VTA or avBNSTGABA-DRN pathway produced anxiolytic-like effects and increased level of DA or 5-HT in the BLA. These findings suggest that avBNST inhibitory projections directly regulate dopaminergic neurons in the VTA and serotonergic neurons in the DRN, and the avBNSTGABA-VTA and avBNSTGABA-DRN pathways respectively exert impacts on PD-related anxiety-like behaviors.

LC-derived excitatory synaptic transmission to dorsal raphe serotonin neurons is inhibited by activation of alpha2-adrenergic receptors.

In the central nervous system, noradrenaline transmission controls the degree to which we are awake, alert, and attentive. Aberrant noradrenaline transmission is associated with pathological forms of hyper- and hypo-arousal that present in numerous neuropsychiatric disorders often associated with dysfunction in serotonin transmission. In vivo, noradrenaline regulates the release of serotonin because noradrenergic input drives the serotonin neurons to fire action potentials via activation of excitatory α1-adrenergic receptors (α1-AR). Despite the critical influence of noradrenaline on the activity of dorsal raphe serotonin neurons, the source of noradrenergic afferents has not been resolved and the presynaptic mechanisms that regulate noradrenaline-dependent synaptic transmission have not been described. Using an acute brain slice preparation from male and female mice and electrophysiological recordings from dorsal raphe serotonin neurons, we found that selective optogenetic activation of locus coeruleus terminals in the dorsal raphe was sufficient to produce an α1-AR-mediated excitatory postsynaptic current (α1-AR-EPSC). Activation of inhibitory α2-adrenergic receptors (α2-AR) with UK-14,304 eliminated the α1-AR-EPSC via presynaptic inhibition of noradrenaline release, likely via inhibition of voltage-gated calcium channels. In a subset of serotonin neurons, activation of postsynaptic α2-AR produced an outward current through activation of GIRK potassium conductance. Further, in vivo activation of α2-AR by systemic administration of clonidine reduced the expression of c-fos in the dorsal raphe serotonin neurons, indicating reduced neural activity. Thus, α2-AR are critical regulators of serotonin neuron excitability.

Aggression modulator: Understanding the multifaceted role of the dorsal raphe nucleus.

Aggressive behavior is instinctively driven behavior that helps animals to survive and reproduce and is closely related to multiple behavioral and physiological processes. The dorsal raphe nucleus (DRN) is an evolutionarily conserved midbrain structure that regulates aggressive behavior by integrating diverse brain inputs. The DRN consists predominantly of serotonergic (5-HT:5-hydroxytryptamine) neurons and decreased 5-HT activity was classically thought to increase aggression. However, recent studies challenge this 5-HT deficiency model, revealing a more complex role for the DRN 5-HT system in aggression. Furthermore, emerging evidence has shown that non-5-HT populations in the DRN and specific neural circuits contribute to the escalation of aggressive behavior. This review argues that the DRN serves as a multifaceted modulator of aggression, acting not only via 5-HT but also via other neurotransmitters and neural pathways, as well as different subsets of 5-HT neurons. In addition, we discuss the contribution of DRN neurons in the behavioral and physiological aspects implicated in aggressive behavior, such as arousal, reward, and impulsivity, to further our understanding of DRN-mediated aggression modulation.

Behavioral and Neurophysiological Implications of Pathological Human Tau Expression in Serotonin Neurons.

Alzheimer's disease (AD) is a progressive degenerative disorder that results in a severe loss of brain cells and irreversible cognitive decline. Memory problems are the most recognized symptoms of AD. However, approximately 90% of patients diagnosed with AD suffer from behavioral symptoms, including mood changes and social impairment years before cognitive dysfunction. Recent evidence indicates that the dorsal raphe nucleus (DRN) is among the initial regions that show tau pathology, which is a hallmark feature of AD. The DRN harbors serotonin (5-HT) neurons, which are critically involved in mood, social, and cognitive regulation. Serotonergic impairment early in the disease process may contribute to behavioral symptoms in AD. However, the mechanisms underlying vulnerability and contribution of the 5-HT system to AD progression remain unknown. Here, we performed behavioral and electrophysiological characterizations in mice expressing a phosphorylation-prone form of human tau (hTauP301L) in 5-HT neurons. We found that pathological tau expression in 5-HT neurons induces anxiety-like behavior and alterations in stress-coping strategies in female and male mice. Female mice also exhibited social disinhibition and mild cognitive impairment in response to 5-HT neuron-specific hTauP301L expression. Behavioral alterations were accompanied by disrupted 5-HT neuron physiology in female and male hTauP301L expressing mice with exacerbated excitability disruption in females only. These data provide mechanistic insights into the brain systems and symptoms impaired early in AD progression, which is critical for disease intervention.

Estrogen signaling in the dorsal raphe regulates binge-like drinking in mice.

Estrogens promote binge alcohol drinking and contribute to sex differences in alcohol use disorder. However, the mechanisms are largely unknown. This study aims to test if estrogens act on 5-hydroxytryptamine neurons in the dorsal raphe nucleus (5-HTDRN) to promote binge drinking. We found that female mice drank more alcohol than male mice in chronic drinking in the dark (DID) tests. This sex difference was associated with distinct alterations in mRNA expression of estrogen receptor α (ERα) and 5-HT-related genes in the DRN, suggesting a potential role of estrogen/ERs/5-HT signaling. In supporting this view, 5-HTDRN neurons from naïve male mice had lower baseline firing activity but higher sensitivity to alcohol-induced excitation compared to 5-HTDRN neurons from naïve female mice. Notably, this higher sensitivity was blunted by 17ß-estradiol treatment in males, indicating an estrogen-dependent mechanism. We further showed that both ERα and ERß are expressed in 5-HTDRN neurons, whereas ERα agonist depolarizes and ERß agonist hyperpolarizes 5-HTDRN neurons. Notably, both treatments blocked the stimulatory effects of alcohol on 5-HTDRN neurons in males, even though they have antagonistic effects on the activity dynamics. These results suggest that ERs' inhibitory effects on ethanol-induced burst firing of 5-HTDRN neurons may contribute to higher levels of binge drinking in females. Consistently, chemogenetic activation of ERα- or ERß-expressing neurons in the DRN reduced binge alcohol drinking. These results support a model in which estrogens act on ERα/ß to prevent alcohol-induced activation of 5-HTDRN neurons, which in return leads to higher binge alcohol drinking.

Neonatal treatment with para-chlorophenylalanine (pCPA) induces adolescent hyperactivity associated with changes in the paraventricular nucleus Crh and Trh expressions.

Disruption of the brain serotoninergic (5-HT) system during development induces long-lasting changes in molecular profile, cytoarchitecture, and function of neurons, impacting behavioral regulation throughout life. In male and female rats, we investigate the effect of neonatal tryptophan hydroxylase (TPH) inhibition by using para-chlorophenylalanine (pCPA) on the expression of 5-HTergic system components and neuropeptides related to adolescent social play behavior regulation. We observed sex-dependent 5-HT levels decrease after pCPA-treatment in the dorsal raphe nucleus (DRN) at 17 and 35 days. Neonatal pCPA-treatment increased playing, social and locomotory behaviors assessed in adolescent rats of both sexes. The pCPA-treated rats demonstrated decreased Crh (17 days) and increased Trh (35 days) expression in the hypothalamic paraventricular nucleus (PVN). There was sex dimorphism in Htr2c (17 days) and VGF (35 days) in the prefrontal cortex, with the females expressing higher levels of it than males. Our results indicate that neonatal pCPA-treatment results in a long-lasting and sex-dependent DRN 5-HT synthesis changes, decreased Crh, and increased Trh expression in the PVN, resulting in a hyperactivity-like phenotype during adolescence. The present work demonstrates that the impairment of TPH function leads to neurobehavioral disorders related to hyperactivity and impulsivity, such as attention deficit hyperactivity disorder (ADHD).

Whole Brain Mapping of Orexin Receptor mRNA Expression Visualized by Branched In Situ Hybridization Chain Reaction.

Orexins, which are produced within neurons of the lateral hypothalamic area, play a pivotal role in the regulation of various behaviors, including sleep/wakefulness, reward behavior, and energy metabolism, via orexin receptor type 1 (OX1R) and type 2 (OX2R). Despite the advanced understanding of orexinergic regulation of behavior at the circuit level, the precise distribution of orexin receptors in the brain remains unknown. Here, we develop a new branched in situ hybridization chain reaction (bHCR) technique to visualize multiple target mRNAs in a semiquantitative manner, combined with immunohistochemistry, which provided comprehensive distribution of orexin receptor mRNA and neuron subtypes expressing orexin receptors in mouse brains. Only a limited number of cells expressing both Ox1r and Ox2r were observed in specific brain regions, such as the dorsal raphe nucleus and ventromedial hypothalamic nucleus. In many brain regions, Ox1r-expressing cells and Ox2r-expressing cells belong to different cell types, such as glutamatergic and GABAergic neurons. Moreover, our findings demonstrated considerable heterogeneity in Ox1r- or Ox2r-expressing populations of serotonergic, dopaminergic, noradrenergic, cholinergic, and histaminergic neurons. The majority of orexin neurons did not express orexin receptors. This study provides valuable insights into the mechanism underlying the physiological and behavioral regulation mediated by the orexin system, as well as the development of therapeutic agents targeting orexin receptors.

Protective effects of SSRI, Citalopram in mutant APP and mutant Tau expressed dorsal raphe neurons in Alzheimer's disease.

Depression is among the most common neuropsychiatric comorbidities in Alzheimer's disease (AD) and other Tauopathies. Apart from its anti-depressive and anxiolytic effects, selective serotonin reuptake inhibitor (SSRI) treatment also offers intracellular modifications that may help to improve neurogenesis, reduce amyloid burden & Tau pathologies, and neuroinflammation in AD. Despite its multifaceted impact in the brain, the exact physiological and molecular mechanism by which SSRIs such as Citalopram improve neurogenesis and synaptogenesis in dementia is poorly understood. In the current study, we investigated the protective role of SSRI, Citalopram, in serotonergic, medullary raphe neurons (RN46A-B14). RN46A-B14 cells were transfected with wild-type and mutant APP and Tau cDNAs for 24 h and then treated with 20 µM Cit for 24 h. We then assessed mRNA and protein levels of pTau, total Tau, serotonin related proteins such as TPH2, SERT, and 5HTR1a, synaptic proteins and the cytoskeletal structure. We also assessed cell survival, mitochondrial respiration and mitochondrial morphology. The mutant APP and Tau transfected cells showed increased levels of serotonin related proteins and mRNA, while the mRNA and protein levels of synaptic proteins were downregulated. Citalopram treatment significantly reduced pathologically pTau level along with the serotonin related protein levels. On the other hand, there was a significant increase in the mRNA and protein levels of synaptic genes and cytoskeletal structure in the treated groups. Further, Citalopram also improved cell survival, mitochondrial respiration and mitochondrial morphology in the treated cells that express mAPP and mTau. Taken together these findings suggest Citalopram could not only be a promising therapeutic drug for treating patients with depression, but also for AD patients.

Characterization of dopaminergic projections from the ventral tegmental area and the dorsal raphe nucleus to the orbital frontal cortex.

The orbitofrontal cortex (OFC) is a key node in the cortico-limbic-striatal circuitry that influences decision-making guided by the relative value of outcomes. Midbrain dopamine from either the ventral tegmental area (VTA) or the dorsal raphe nucleus (DRN) has the potential to modulate OFC neurons; however, it is unknown at what concentrations these terminals release dopamine. Male and female adult dopamine transporter (DAT)IRES-Cre-tdTomato mice were injected with AAV2/8-EF1a-DIO-eYFP into either the DRN or the VTA or the retrograde label cholera toxin B (CTB) 488 in the medial or lateral OFC. We quantified co-expression of CTB 488 or enhanced yellow fluorescent protein (eYFP) with tdTomato fluorescence in VTA or DRN and eYFP fibre density in the medial or lateral OFC. Both VTA and DRN dopamine neurons project to either the medial OFC or the lateral OFC, with greater expression of fibres in the medial OFC. Using fast-scan cyclic voltammetry, we detected optogenetically evoked dopamine from channelrhodopsin 2 (ChR2)-expressing VTA or DRN dopamine terminals in either the medial OFC or the lateral OFC. We assessed if optical stimulation of dopamine from the VTA or the DRN onto the medial OFC could alter layer V pyramidal neuronal firing; however, we did not observe a change in firing at stimulation parameters that evoked dopamine release from either projection even though bath application of dopamine with the monoamine transporter inhibitor, nomifensine, decreased firing. In summary, dopaminergic neurons from the VTA or the DRN project to the OFC and release submicromolar dopamine in the medial and lateral OFC.

Enhancing HIF-1α-P2X2 signaling in dorsal raphe serotonergic neurons promotes psychological resilience.

Major depressive disorder (MDD) is a devastating condition. Although progress has been made in the past seven decades, patients with MDD continue to receive an inadequate treatment, primarily due to the late onset of first-line antidepressant drugs and to their acute withdrawal symptoms. Resilience is the ability to rebound from adversity in a healthy manner and many people have psychological resilience. Revealing the mechanisms and identifying methods promoting resilience will hopefully lead to more effective prevention strategies and treatments for depression. In this study, we found that intermittent hypobaric hypoxia training (IHHT), a method for training pilots and mountaineers, enhanced psychological resilience in adult mice. IHHT produced a sustained antidepressant-like effect in mouse models of depression by inducing long-term (up to 3 months after this treatment) overexpression of hypoxia-inducible factor (HIF)-1α in the dorsal raphe nucleus (DRN) of adult mice. Moreover, DRN-infusion of cobalt chloride, which mimics hypoxia increasing HIF-1α expression, triggered a rapid and long-lasting antidepressant-like effect. Down-regulation of HIF-1α in the DRN serotonergic (DRN5-HT) neurons attenuated the effects of IHHT. HIF-1α translationally regulated the expression of P2X2, and conditionally knocking out P2rx2 (encodes P2X2 receptors) in DRN5-HT neurons, in turn, attenuated the sustained antidepressant-like effect of IHHT, but not its acute effect. In line with these results, a single sub-anesthetic dose of ketamine enhanced HIF-1α-P2X2 signaling, which is essential for its rapid and long-lasting antidepressant-like effect. Notably, we found that P2X2 protein levels were significantly lower in the DRN of patients with MDD than that of control subjects. Together, these findings elucidate the molecular mechanism underlying IHHT promoting psychological resilience and highlight enhancing HIF-1α-P2X2 signaling in DRN5-HT neurons as a potential avenue for screening novel therapeutic treatments for MDD.

Dorsal raphe stimulation relays a reward signal to the ventral tegmental area via GluN2C NMDA receptors.

BACKGROUND: Glutamate relays a reward signal from the dorsal raphe (DR) to the ventral tegmental area (VTA). However, the role of the different subtypes of N-methyl-D-aspartate (NMDA) receptors is complex and not clearly understood. Therefore, we measured NMDA receptors subunits expression in limbic brain areas. In addition, we studied the effects of VTA down-regulation of GluN2C NMDA receptor on the reward signal that arises from DR electrical stimulation. METHODS: Using qPCR, we identified the relative composition of the different Grin2a-d subunits of the NMDA receptors in several brain areas. Then, we used fluorescent in situ hybridization (FISH) to evaluate the colocalization of Grin2c and tyrosine hydroxylase (TH) mRNA in VTA neurons. To assess the role of GluN2C in brain stimulation reward, we downregulated this receptor using small interfering RNA (siRNA) in rats self-stimulating for electrical pulses delivered to the DR. To delineate further the specific role of GluN2C in relaying the reward signal, we pharmacologically altered the function of VTA NMDA receptors by bilaterally microinjecting the NMDA receptor antagonist PPPA. RESULTS: We identified GluN2C as the most abundant subunit of the NMDA receptor expressed in the VTA. FISH revealed that about 50% of TH-positive neurons colocalize with Grin2c transcript. siRNA manipulation produced a selective down-regulation of the GluN2C protein subunit and a significant reduction in brain stimulation reward. Interestingly, PPPA enhanced brain stimulation reward, but only in rats that received the nonactive RNA sequence. CONCLUSION: The present results suggest that VTA glutamate neurotransmission relays a reward signal initiated by DR stimulation by acting on GluN2C NMDA receptors.

Reduction of p11 in dorsal raphe nucleus serotonergic neurons mediates depression-like behaviors.

The pathology of depression is related to the imbalance of various neurotransmitters. The dorsal raphe nucleus (DRN), the main brain region producing 5-HT, is crucially involved in the pathophysiology of depression. It contains several neuron types, in which GABAergic neurons are activated by stimuli associated with negative experiences and 5-HT neurons are activated by reward signals. However, little is known about its underlying molecular mechanisms. Here, we found that p11, a multifunctional protein associated with depression, was down-regulated by chronic social defeat stress in 5-HTDRN neurons. Knockdown of p11 in DRN induced depression-like behaviors, while its overexpression in 5-HTDRN neurons alleviated depression-like behavior caused by chronic social defeat stress. Further, p11 regulates membrane trafficking of glutamate receptors in 5-HTDRN neurons, suggesting a possible molecular mechanism underlying the participation of p11 in the pathological process of depression. This may facilitate the understanding of the molecular and cellular basis of depression.

The Role of GABA in the Dorsal Striatum-Raphe Nucleus Circuit Regulating Stress Vulnerability in Male Mice with High Levels of Shati/Nat8l.

Depression is a frequent and serious illness, and stress is considered the main risk factor for its onset. First-line antidepressants increase serotonin (5-hydroxytryptamine; 5-HT) levels in the brain. We previously reported that an N-acetyltransferase, Shati/Nat8l, is upregulated in the dorsal striatum (dSTR) of stress-susceptible mice exposed to repeated social defeat stress (RSDS) and that dSTR Shati/Nat8l overexpression in mice (dSTR-Shati OE) induces stress vulnerability and local reduction in 5-HT content. Male mice were used in this study, and we found that dSTR 5-HT content decreased in stress-susceptible but not in resilient mice. Moreover, vulnerability to stress in dSTR-Shati OE mice was suppressed by the activation of serotonergic neurons projecting from the dorsal raphe nucleus (dRN) to the dSTR, followed by upregulation of 5-HT content in the dSTR using designer receptors exclusively activated by designer drugs (DREADD). We evaluated the role of GABA in modulating the serotonergic system in the dRN. Stress-susceptible after RSDS and dSTR-Shati OE mice exhibited an increase in dRN GABA content. Furthermore, dRN GABA content was correlated with stress sensitivity. We found that the blockade of GABA signaling in the dRN suppressed stress susceptibility in dSTR-Shati OE mice. In conclusion, we propose that dSTR 5-HT and dRN GABA, controlled by striatal Shati/Nat8l via the dSTR-dRN neuronal circuitry, critically regulate stress sensitivity. Our study provides insights into the neural processes that underlie stress and suggests that dSTR Shati/Nat8l could be a novel therapeutic target for drugs against depression, allowing direct control of the dRN serotonergic system.

Taok1 haploinsufficiency leads to autistic-like behaviors in mice via the dorsal raphe nucleus.

Strong evidence from human genetic studies associates the thousand and one amino acid kinase 1 (TAOK1) gene with autism spectrum disorder (ASD). In this work, we discovered a de novo frameshifting mutation in TAOK1 within a Chinese ASD cohort. We found that Taok1 haploinsufficiency induces autistic-like behaviors in mice. Importantly, we observed a significant enrichment of Taok1 in the dorsal raphe nucleus (DRN). The haploinsufficiency of Taok1 considerably restrained the activation of DRN neurons during social interactions, leading to the aberrant phosphorylation of numerous proteins. Intriguingly, the genetic deletion of Taok1 in VGlut3-positive neurons of DRN resulted in mice exhibiting autistic-like behaviors. Ultimately, reintroducing wild-type Taok1, but not its kinase-dead variant, into the DRN of adult mice effectively mitigated the autistic-like behaviors associated with Taok1 haploinsufficiency. This work suggests that Taok1, through its influence in the DRN, regulates social interaction behaviors, providing critical insights into the etiology of ASD.

Impact of the circadian nuclear receptor REV-ERBα in dorsal raphe 5-HT neurons on social interaction behavior, especially social preference.

Social interaction among conspecifics is essential for maintaining adaptive, cooperative, and social behaviors, along with survival among mammals. The 5-hydroxytryptamine (5-HT) neuronal system is an important neurotransmitter system for regulating social behaviors; however, the circadian role of 5-HT in social interaction behaviors is unclear. To investigate whether the circadian nuclear receptor REV-ERBα, a transcriptional repressor of the rate-limiting enzyme tryptophan hydroxylase 2 (Tph2) gene in 5-HT biosynthesis, may affect social interaction behaviors, we generated a conditional knockout (cKO) mouse by targeting Rev-Erbα in dorsal raphe (DR) 5-HT neurons (5-HTDR-specific REV-ERBα cKO) using the CRISPR/Cas9 gene editing system and assayed social behaviors, including social preference and social recognition, with a three-chamber social interaction test at two circadian time (CT) points, i.e., at dawn (CT00) and dusk (CT12). The genetic ablation of Rev-Erbα in DR 5-HTergic neurons caused impaired social interaction behaviors, particularly social preference but not social recognition, with no difference between the two CT points. This deficit of social preference induced by Rev-Erbα in 5-HTDR-specific mice is functionally associated with real-time elevated neuron activity and 5-HT levels at dusk, as determined by fiber-photometry imaging sensors. Moreover, optogenetic inhibition of DR to nucleus accumbens (NAc) 5-HTergic circuit restored the impairment of social preference in 5-HTDR-specific REV-ERBα cKO mice. These results suggest the significance of the circadian regulation of 5-HT levels by REV-ERBα in regulating social interaction behaviors.

The α1 and γ2 subunit-containing GABAA receptor-mediated inhibitory transmission in the anteroventral bed nucleus of stria terminalis is involved in the regulation of anxiety in rats with substantia nigra lesions.

The anteroventral bed nucleus of the stria terminalis (avBNST) is widely acknowledged as a key brain structure that regulates negative emotional states, such as anxiety. At present, it is still unclear whether GABAA receptor-mediated inhibitory transmission in the avBNST is involved in Parkinson's disease (PD)-related anxiety. In this study, unilateral 6-hydroxydopamine (6-OHDA) lesions of the substantia nigra pars compacta (SNc) in rats induced anxiety-like behaviors, increased GABA synthesis and release, and upregulated expression of GABAA receptor subunits in the avBNST, as well as decreased level of dopamine (DA) in the basolateral amygdala (BLA). In both sham and 6-OHDA rats, intra-avBNST injection of GABAA receptor agonist muscimol induced the following changes: (i) anxiolytic-like responses, (ii) inhibition of the firing activity of GABAergic neurons in the avBNST, (iii) excitation of dopaminergic neurons in the ventral tegmental area (VTA) and serotonergic neurons in the dorsal raphe nucleus (DRN), and (iv) increase of DA and 5-HT release in the BLA, whereas antagonist bicuculline induced the opposite effects. Collectively, these findings suggest that degeneration of the nigrostriatal pathway enhances GABAA receptor-mediated inhibitory transmission in the avBNST, which is involved in PD-related anxiety. Further, activation and blockade of avBNST GABAA receptors affect the firing activity of VTA dopaminergic and DRN serotonergic neurons, and then change release of BLA DA and 5-HT, thereby regulating anxiety-like behaviors.

GABAergic miR-34a regulates Dorsal Raphè inhibitory transmission in response to aversive, but not rewarding, stimuli.

The brain employs distinct circuitries to encode positive and negative valence stimuli, and dysfunctions of these neuronal circuits have a key role in the etiopathogenesis of many psychiatric disorders. The Dorsal Raphè Nucleus (DRN) is involved in various behaviors and drives the emotional response to rewarding and aversive experiences. Whether specific subpopulations of neurons within the DRN encode these behaviors with different valence is still unknown. Notably, microRNA expression in the mammalian brain is characterized by tissue and neuronal specificity, suggesting that it might play a role in cell and circuit functionality. However, this specificity has not been fully exploited. Here, we demonstrate that microRNA-34a (miR-34a) is selectively expressed in a subpopulation of GABAergic neurons of the ventrolateral DRN. Moreover, we report that acute exposure to both aversive (restraint stress) and rewarding (chocolate) stimuli reduces GABA release in the DRN, an effect prevented by the inactivation of DRN miR-34a or its genetic deletion in GABAergic neurons in aversive but not rewarding conditions. Finally, miR-34a inhibition selectively reduced passive coping with severe stressors. These data support a role of miR-34a in regulating GABAergic neurotransmitter activity and behavior in a context-dependent manner and suggest that microRNAs could represent a functional signature of specific neuronal subpopulations with valence-specific activity in the brain.

The unfixed light pattern contributes to depressive-like behaviors in male mice.

Light pollution is now associated with an increased incidence of mental disorders in humans, and the unfixed light pattern (ULP) is a common light pollution that occurs in such as rotating shift work. However, how much contribution the ULP has to depression and its potential mechanism are yet unknown. Our study aimed to investigate the effect of the ULP on depressive-like behaviors in mice and to explore the links to the circadian-orexinergic system. Male C57BL/6 J mice were exposed to the ULP by subjecting them to an alternating light pattern every 6 days for 54 days. The tail suspension test (TST) and forced swimming test (FST) were conducted to assess depressive-like behaviors. The rhythm of locomotor activity and the circadian expression of cFOS in the suprachiasmatic nucleus (SCN), clock genes in the liver, and corticosterone (CORT) in serum were detected to observe changes in the circadian system. The circadian expression of orexin-A (OX-A) in the lateral hypothalamus area (LHA) and dorsal raphe nucleus (DRN) and serotonin (5-HT) in the DRN were measured to determine alterations in the orexinergic system. The results showed that mice exposed to the ULP exhibited increased immobility time in the TST and FST. The ULP significantly disrupted the circadian rhythm of locomotor activity, clock genes in the liver, and CORT in the serum. Importantly, when exposed to the ULP, cFOS expression in the SCN showed decreased amplitude. Its projection area, the LHA, had a lower mesor of OX-A expression. OX-A projection to the DRN and 5-HT expression in the DRN were reduced in mesor. Our research suggests that the ULP contributes to depressive-like behaviors in mice, which might be related to the reduced amplitude of circadian oscillation in the SCN and hypoactivity of the orexinergic system. These findings may provide novel insights into rotating shift work-related depression.