Others have shown that dopamine receptors regulate the migration of GABAergic cortical interneurons (cINs) to the developing cortex. Given the strong expression of Drd1, the gene encoding the D1 dopamine receptor (D1R) in the developing cortex, we examined here the role of D1R in the cortical migration of interneurons born in the medial ganglionic eminence (MGE). Embryos of transgenic mice expressing cytoplasmic GFP under the control of the Drd1 promoter exhibited strong GFP expression in cells located in the deep cortical layers, including the subplate, and in the marginal zone. In co-culture experiments aimed at characterizing the effect of selective Drd1 ablation either in interneurons or in cortical plate cells on the migratory behavior of interneurons, we identified a prominent pro-migratory non-cell autonomous effect of Drd1 ablation in the cortical substrate. To assess whether Drd1 ablation in cortical cells could influence the final interneuron distribution in vivo, we analyzed the cortical distribution of parvalbumin and somatostatin positive interneurons in the cortex of Drd1-CKO (Drd1-/- cortical cells, Drd1+/+ interneurons) mice. Wild type parvalbumin and somatostatin interneurons exhibited slight but significant density changes and alterations of latero-dorsal distribution compatible with the pro-migratory effect of Drd1-/- cortical cells. In Drd1-KO animals (Drd1-/- cortical cells and Drd1-/- interneurons), the distribution alterations of parvalbumin and somatostatin interneurons were reminiscent of those in Drd1-CKO mutants. We thus propose that D1R regulates in the cortex the motility and distribution of MGE-derived cINs by preponderant non-cell-autonomous mechanism.

Immediate early genes are widely used markers of neuronal activation, but their function in neurons is not well understood. We focused on the role of Egr1 in the nucleus accumbens core (NAc-c) in the long-lasting behavioral effects of cocaine, using an AAV expressing short hairpin RNA (Egr1-shRNA). Egr1 knockdown did not alter acute cocaine locomotor effects or conditioned place preference. In contrast, shEgr1 markedly decreased the locomotor sensitization induced by repeated cocaine administration. Because EGR1 is a transcription factor, we explored the transcriptomic alterations using RNAseq completed by RT-qPCR and protein studies. Egr1 knockdown modified the expression of numerous genes. Analysis of the upregulated genes revealed indirect activation of astrocytes and microglia evidenced by immunohistofluorescence, but shEgr1-induced dampening of cocaine sensitization was unaffected by minocycline, a microglia inhibitor. Proteasome genes were upregulated by shEgr1, possibly contributing to its functional consequences. Downregulated genes included potential EGR1 targets and comprised many genes characteristic of striatal neurons, including those coding signaling proteins (DARPP-32, CDK5 activator p35), glutamate ionotropic (NMDA NR1/2B, AMPA GluA1-3) and metabotropic (mGluR1/5) receptors, and postsynaptic proteins (PSD-95). We confirmed these alterations at the protein level and found decreased cocaine-induced phospho-Ser845-GluA1. Thus our study shows the broad transcriptional consequences of silencing Egr1 in neurons. It provides a mechanism by which Egr1 knockdown in the NAc-c can alter cocaine-induced locomotor sensitization, through downregulation of many genes including key components of glutamate neurotransmission. This broad role of EGR1 in regulating transcription provides clues about its function and role in learning, memory, and synaptic plasticity.

Many psychiatric diseases have been associated with serotonin (5-HT) neuron dysfunction. The firing of 5-HT neurons is known to be under 5-HT1A receptor -mediated autoinhibition, but functional consequences of coexpressed receptors are unknown. Using co-immunoprecipitation, BRET, confocal, and super-resolution microscopy in hippocampal and 5-HT neurons, we present evidence that 5-HT1A and 5-HT2B receptors can form heterodimers and co-cluster at the plasma membrane of dendrites. Selective agonist stimulation of coexpressed 5-HT1A and 5-HT2B receptors prevents 5-HT1A receptor internalization and increases 5-HT2B receptor membrane clustering. Current clamp recordings of 5-HT neurons revealed that 5-HT1A receptor stimulation of acute slices from mice lacking 5-HT2B receptors in 5-HT neurons increased their firing activity trough Ca2+-activated potassium channel inhibition compared to 5-HT neurons from control mice. This work supports the hypothesis that the relative expression of 5-HT1A and 5-HT2B receptors tunes the neuronal excitability of serotonergic neurons through potassium channel regulation.

The proper maturation of emotional and sensory circuits requires fine-tuning of serotonin (5-HT) level during early postnatal development. Consistently, dysfunctions of the serotonergic system have been associated with neurodevelopmental psychiatric diseases, including autism spectrum disorders (ASD). However, the mechanisms underlying the developmental effects of 5-HT remain partially unknown, one obstacle being the action of 5-HT on different cell types. Here, we focused on microglia, which play a role in brain wiring refinement, and we investigated whether the control of these cells by 5-HT is relevant for neurodevelopment and spontaneous behaviors in mice. Since the main 5-HT sensor in microglia is the 5-HT2B receptor subtype, we prevented 5-HT signaling specifically in microglia by conditional invalidation of the Htr2b gene in these cells. We observed that abrogating the serotonergic control of microglia during early postnatal development affects the phagolysosomal compartment of these cells and their proximity to dendritic spines and perturbs neuronal circuits maturation. Furthermore, this early ablation of microglial 5-HT2B receptors leads to adult hyperactivity in a novel environment and behavioral defects in sociability and flexibility. Importantly, we show that these behavioral alterations result from a developmental effect, since they are not observed when microglial Htr2b invalidation is induced later, at P30 onward. Thus, a primary alteration of 5-HT sensing in microglia, during a critical time window between birth and P30, is sufficient to impair social and flexibility skills. This link between 5-HT and microglia may explain the association between serotonergic dysfunctions and behavioral traits like impaired sociability and inadaptability to novelty, which are prominent in psychiatric disorders such as ASD.

Approximately 15 million babies are born prematurely every year and many will face lifetime motor and/or cognitive deficits. Children born prematurely are at higher risk of developing perinatal brain lesions, especially white matter injuries (WMI). Evidence in humans and rodents demonstrates that systemic inflammation-induced neuroinflammation, including microglial and astrocyte reactivity, is the prominent processes of WMI associated with preterm birth. Thus, a new challenge in the field of perinatal brain injuries is to develop new neuroprotective strategies to target neuroinflammation to prevent WMI. Serotonin (5-HT) and its receptors play an important role in inflammation, and emerging evidence indicates that 5-HT may regulate brain inflammation by the modulation of microglial reactivity and astrocyte functions. The present study is based on a mouse model of WMI induced by intraperitoneal (i.p.) injections of IL-1β during the first 5 days of life. In this model, certain key lesions of preterm brain injuries can be summarized by (i) systemic inflammation, (ii) pro-inflammatory microglial and astrocyte activation, and (iii) inhibition of oligodendrocyte maturation, leading to hypomyelination. We demonstrate that Htr7 mRNA (coding for the HTR7/5-HT7 receptor) is significantly overexpressed in the anterior cortex of IL-1β-exposed animals, suggesting it as a potential therapeutic target. LP-211 is a specific high-affinity HTR7 agonist that crosses the blood–brain barrier (BBB). When co-injected with IL-1β, LP-211 treatment prevented glial reactivity, the down-regulation of myelin-associated proteins, and the apparition of anxiety-like phenotypes. Thus, HTR7 may represent an innovative therapeutic target to protect the developing brain from preterm brain injuries.

AbstractMany psychiatric diseases including depression, schizophrenia and anxiety have been associated with serotonin (5-HT) neuron dysfunction. Pacemaker-like firing of raphe 5-HT neurons was proposed to be under unique 5-HT1Areceptor-mediated autoinhibition. We previously showed that 5-HT2Breceptors were expressed by 5-HT neurons together with 5-HT1Areceptors. However, functional consequences on 5-HT neurons of putative interaction between these receptors are unknown. Using co-immunoprecipitation, BRET, confocal and super-resolution microscopy in hippocampal and 5-HT neurons, we present converging evidence that 5-HT1Aand 5-HT2Breceptors can form heterodimers and co-cluster at the surface of dendrites. 5-HT2Breceptor clusters were redistributed upon 5-HT1Areceptor expression supporting functional interactions between the two receptors. Furthermore, 5-HT2Breceptor expression prevented agonist-induced internalization of 5-HT1Areceptors, whereas 5-HT1Areceptors mimicked the clustering effect of 5-HT2Breceptor stimulation on its surface expression. The functional impact of this interactionin-vivowas assessed by recording 5-HT neuron excitability from mice lacking 5-HT2Breceptors in 5-HT neurons. Upon 5-HT1Areceptor stimulation, the firing activity of 5-HT neurons was increased in the absence of 5-HT2Breceptors and decreased in their presence through regulation of SK channels, thus demonstrating functional output of this interaction in controlling 5-HT neuron firing activity.

The defense against pathogens is mediated by innate and adaptive immune mechanisms that act in the periphery and in the central nervous system. In periphery, serotonin (5-HT) is synthesized in the gastrointestinal tract and the enteric nerves, and is stored in particularly high abundance in platelet granules. In the CNS, serotonin is synthesized by serotonergic neurons that innervate many brain areas and is stored in synaptic vesicles. Serotonin can regulate inflammation and immunity by acting at serotonin receptors that are differentially expressed on immune cells. Serotonin acts as a potent chemoattractant, recruiting innate immune cells to sites of inflammation, alters the production and release of cytokines and modulates immune cell activation/proliferation. In addition, serotonin has been widely involved in neuropsychiatric diseases, major depressive disorders, or autism spectrum disorders, and in neurodegenerative diseases, including Alzheimer’s disease or amyotrophic lateral sclerosis. Independent evidence suggests the contribution of inflammatory mediators in these pathologies. Indeed, peripheral infections as well as chronic diseases of the central nervous system cause activation of microglia, the resident macrophages of the brain. In this review, we summarize evidence that serotonin via 5-HT2B receptors regulates inflammation, either developmentally, acutely, or during neurodegenerative diseases, and in turn influences the course of these diseases through immune cell modulation.

5 HT receptors expressed throughout the human body are targets for established therapeutics and various drugs in development. Their diversity of structure and function reflects the important role 5-HT receptors play in physiologic and pathophysiological processes. The present review offers a framework for the official receptor nomenclature and a detailed understanding of each of the 14 5-HT receptor subtypes, their roles in the systems of the body, and, where appropriate, the (potential) utility of therapeutics targeting these receptors. Significance Statement-This review provides a comprehensive account of the classification and function of 5-hydroxytryptamine receptors, including how they are targeted for therapeutic benefit.

Microglial cells, the brain resident macrophages, participate to brain development and function and help maintaining its homeostasis. To play these roles, they need to detect and adapt to modifications of their environment, including changes in the activity of neurons. The neuromodulators serotonin, dopamine, norepinephrine, acetylcholine and histamine are synthesized and released by specialized neurons to coordinate the activity of other neurons in different regions. In this review, we summarize the current evidence obtained in vitro or in vivo that neuromodulators act on microglia. On the short term, they can modify their motility, morphology and phagocytic activity; on the mid-long term they can modulate their transition between different immune activation states. Lastly, we review some recent data suggesting that these regulations of microglia by neuromodulators are involved in vivo in some aspects of central nervous system development, function and homeostasis.

Severe peripheral infections induce an adaptive sickness behavior and an innate immune reaction in various organs including the brain. On the long term, persistent alteration of microglia, the brain innate immune cells, is associated with an increased risk of psychiatric disorders. It is thus critical to identify genes and mechanisms controlling the intensity and duration of the neuroinflammation induced by peripheral immune challenges. We tested the hypothesis that the 5-HT2B receptor, the main serotonin receptor expressed by microglia, might represent a valuable candidate. First, we observed that Htr2b-/- mice, knock-out for the 5-HT2B receptor gene, developed, when exposed to a peripheral lipopolysaccharide (LPS) challenge, a stronger weight loss compared to wild-type mice; in addition, comparison of inflammatory markers in brain, 4 and 24 hr after LPS injection, showed that Htr2b deficiency leads to a prolonged neuroinflammation. Second, to assess the specific contribution of the microglial 5-HT2B receptor, we investigated the response to LPS of conditional knock-out mice invalidated for Htr2b in microglia only. We found that deletion of Htr2b in microglia since birth is sufficient to cause enhanced weight loss and increased neuroinflammatory response upon LPS injection at adult stage. In contrast, mice deleted for microglial Htr2b in adulthood responded normally to LPS, revealing a neonatal developmental effect. These results highlight the role of microglia in the response to a peripheral immune challenge and suggest the existence of a developmental, neonatal period, during which instruction of microglia through 5-HT2B receptors is necessary to prevent microglia overreactivity in adulthood.

The defense against pathogens is mediated by innate and adaptive immune mechanisms that act in periphery and central nervous system (CNS). Outside the CNS, serotonin is found in gastrointestinal tract and enteric nerves, in hematopoietic stem cells, and in particularly high abundance in platelets. Serotonin regulates inflammation and immunity by acting on serotonin receptors that are differentially expressed on immune cells, both in rodents and humans. Serotonin acts as a potent chemoattractant, recruiting innate immune cells to sites of inflammation. Serotonin also alters the production and release of cytokines and cell activation/proliferation. Some immune cells, including mast cells and T lymphocytes, have the capacity to synthesize and release serotonin, expanding the range of tissues for serotonin signaling.

Membrane receptors often form complexes with other membrane proteins that directly interact with different effectors of the signal transduction machinery. G-protein-coupled receptors (GPCRs) were for long time considered as single pharmacological entities. However, evidence for oligomerization appeared for various classes and subtypes of GPCRs. This review focuses on metabotropic serotonin (5-hydroxytryptamine, 5-HT) receptors, which belong to the rhodopsin-like class A of GPCRs, and will summarize the convergent evidence that homo- and hetero-dimers containing 5-HT receptors exist in transfected cells and in-vivo. We will show that complexes involving 5-HT receptors may acquire new signal transduction pathways and new physiological roles. In some cases, these complexes participate in disease-specific deregulations, that can be differentially affected by various drugs. Hence, selecting receptor complex-specific responses of these heterodimers may constitute an emerging strategy likely to improve beneficial therapeutic effects. (C) 2019 Elsevier B.V. and Societe Francaise de Biochimie et Biologie Moleculaire (SFBBM). All rights reserved.

Microglial cells are resident innate immune cells of the brain that constantly scan their environment with their long processes and, upon disruption of homeostasis, undergo rapid morphological changes. For example, a laser lesion induces in a few minutes an oriented growth of microglial processes, also called "directional motility", toward the site of injury. A similar effect can be obtained by delivering locally ATP or serotonin (5-hydroxytryptamine [5-HT]). In this article, we describe a protocol to induce a directional growth of microglial processes toward a local application of ATP or 5-HT in acute brain slices of young and adult mice and to image this attraction over time by multiphoton microscopy. A simple method of quantification with free and open-source image analysis software is proposed. A challenge that still characterizes acute brain slices is the limited time, decreasing with age, during which the cells remain in a physiological state. This protocol, thus, highlights some technical improvements (medium, air-liquid interface chamber, imaging chamber with a double perfusion) aimed at optimizing the viability of microglial cells over several hours, especially in slices from adult mice.

Serotonin is a neurotransmitter involved in many psychiatric diseases. In humans, a lack of 5HT2B receptors is associated with serotonin-dependent phenotypes, including impulsivity and suicidality. A lack of 5-HT2B receptors in mice eliminates the effects of molecules that directly target serotonergic neurons including amphetamine-derivative serotonin releasers, and selective serotonin reuptake inhibitor antidepressants. In this work, we tested the hypothesis that 5-HT2B receptors directly and positively regulate raphe serotonin neuron activity. By ex-vivo electrophysiological recordings, we report that stimulation by the 5-HT2Breceptor agonist, BW723C86, increased the firing frequency of serotonin Pet1-positive neurons. Viral overexpression of 5-HT2B receptors in these neurons increased their excitability. Furthermore, in-vivo 5-HT2B-receptor stimulation by BW723C86 counteracted 5HT1A autoreceptor-dependent reduction in firing rate and hypothermic response in wildtype mice. By a conditional genetic ablation that eliminates 5-HT2B-receptor expression specifically and exclusively from Pet1-positive serotonin neurons (Htr2b mice), we demonstrated that behavioral and sensitizing effects of MDMA, as well as acute behavioral and chronic neurogenic effects of the antidepressant fluoxetine, require 5-HT2B-receptor expression in serotonergic neurons. In Htr2b mice, dorsal raphe serotonin neurons displayed a lower firing frequency compared to control Htr2b mice as assessed by in-vivo extracellular recordings and a stronger hypothermic effect of 5-HT1A-autoreceptor stimulation was observed. The increase in head twitch response to DOI further confirmed the lower serotonergic tone resulting from the absence of 5-HT2B receptors in serotonin neurons. Together, these observations indicate that the 5-HT2B receptor acts as a direct positive modulator of serotonin Pet1-positive neurons in an opposite way as the known 5-HT1A negative autoreceptor. Introduction Serotonin (5-Hydroxytryptamine, 5-HT) is involved in many psychiatric diseases including depression, addiction, impulsivity or psychosis. The 5-HT neurons that innervate forebrain originate predominantly from the rostral cell group of neurons in the dorsal raphe nucleus (DRN) (Commons, 2016; Okaty et al, 2015). These neurons express the serotonergic markers tryptophan hydroxylase (TPH2), and 5-HT transporter (SERT), and also the negative autoreceptors, 5-HT1A and 5-HT1B receptors, whose expression is restricted to somatodendritic compartments of 5-HT neurons, and to axonal terminals, respectively (Riad et al, 2000). The 5-HT1A autoreceptor activation elicits an outward current carried through G protein-coupled inwardly-rectifying potassium channels (GIRK) of the Kir3 family leading to membrane hyperpolarization and inhibition of 5-HT neuron firing (Aghajanian and Lakoski, 1984). The presence of synaptic vesicles in dendrites of 5-HT neurons led to the suggestion that autoinhibition is mediated via dendritic release of 5-HT, for review see (Andrade et al, 2015). However, activity of 5-HT DRN neurons can also be positively modulated by 5HT2A/2B/2C receptors triggering directly or indirectly inward currents (Boothman et al, 2003; Craven et al, 2001; Kirby et al, 2003; Liu et al, 2000; Quérée et al, 2009). Upon electrical stimulation of leech 5-HT neurons, transmembrane Ca entry through L-type channels first evokes an early dendritic exocytosis; subsequently, the released 5-HT activates dendritic 5HT2 autoreceptors coupled to Gq and phospholipase C, resulting in a positive feedforward loop that maintains sustained exocytosis (Leon-Pinzon et al, 2014). It has thus been proposed that DRN neurons can display responses ranging from inhibition to excitation depending on a balance of functional 5-HT1A and 5-HT2 receptors (Marinelli et al, 2004). However, a direct action of 5-HT2 receptor subtypes at 5-HT neurons has not yet been clearly established. In humans, a loss-of-function polymorphism of 5-HT2B receptors is associated with 5-

Microglia are the predominant immune response cells and professional phagocytes of the central nervous system (CNS) that have been shown to be important for brain development and homeostasis. These cells present a broad spectrum of phenotypes across stages of the lifespan and especially in CNS diseases. Their prevalence in all neurological pathologies makes it pertinent to reexamine their distinct roles during steady-state and disease conditions. A major question in the field is determining whether the clustering and phenotypical transformation of microglial cells are leading causes of pathogenesis, or potentially neuroprotective responses to the onset of disease. The recent explosive growth in our understanding of the origin and homeostasis of microglia, uncovering their roles in shaping of the neural circuitry and synaptic plasticity, allows us to discuss their emerging functions in the contexts of cognitive control and psychiatric disorders. The distinct mesodermal origin and genetic signature of microglia in contrast to other neuroglial cells also make them an interesting target for the development of therapeutics. Here, we review the physiological roles of microglia, their contribution to the effects of environmental risk factors (e.g., maternal infection, early-life stress, dietary imbalance), and their impact on psychiatric disorders initiated during development (e.g., Nasu-Hakola disease (NHD), hereditary diffuse leukoencephaly with spheroids, Rett syndrome, autism spectrum disorders (ASDs), and obsessive-compulsive disorder (OCD)) or adulthood (e.g., alcohol and drug abuse, major depressive disorder (MDD), bipolar disorder (BD), schizophrenia, eating disorders and sleep disorders). Furthermore, we discuss the changes in microglial functions in the context of cognitive aging, and review their implication in neurodegenerative diseases of the aged adult (e.g., Alzheimer’s and Parkinson’s). Taking into account the recent identification of microglia-specific markers, and the availability of compounds that target these cells selectively in vivo, we consider the prospect of disease intervention via the microglial route.