RESUMO
Amino acid substitutions in the kinase domain of the human CSF1R gene are associated with autosomal dominant adult-onset leukoencephalopathy with axonal spheroids and pigmented glia (ALSP). To model the human disease, we created a disease-associated mutation (pGlu631Lys; E631K) in the mouse Csf1r locus. Homozygous mutation (Csf1rE631K/E631K) phenocopied the Csf1r knockout, with prenatal mortality or severe postnatal growth retardation and hydrocephalus. Heterozygous mutation delayed the postnatal expansion of tissue macrophage populations in most organs. Bone marrow cells from Csf1rE631K/+mice were resistant to CSF1 stimulation in vitro, and Csf1rE631K/+ mice were unresponsive to administration of a CSF1-Fc fusion protein, which expanded tissue macrophage populations in controls. In the brain, microglial cell numbers and dendritic arborisation were reduced in Csf1rE631K/+ mice, as in patients with ALSP. The microglial phenotype is the opposite of microgliosis observed in Csf1r+/- mice. However, we found no evidence of brain pathology or impacts on motor function in aged Csf1rE631K/+ mice. We conclude that heterozygous disease-associated CSF1R mutations compromise CSF1R signalling. We speculate that leukoencephalopathy associated with dominant human CSF1R mutations requires an environmental trigger and/or epistatic interaction with common neurodegenerative disease-associated alleles.
Assuntos
Leucoencefalopatias , Doenças Neurodegenerativas , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos , Animais , Humanos , Leucoencefalopatias/genética , Leucoencefalopatias/patologia , Camundongos , Mutação/genética , Doenças Neurodegenerativas/patologia , Neuroglia , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/genéticaRESUMO
Macrophages regulate metabolic homeostasis in health and disease. Macrophage colony-stimulating factor (CSF1)-dependent macrophages contribute to homeostatic control of the size of the liver. This study aimed to determine the systemic metabolic consequences of elevating circulating CSF1. Acute administration of a CSF1-Fc fusion protein to mice led to monocytosis, increased resident tissue macrophages in the liver and all major organs, and liver growth. These effects were associated with increased hepatic glucose uptake and extensive mobilization of body fat. The impacts of CSF1 on macrophage abundance, liver size, and body composition were rapidly reversed to restore homeostasis. The effects of CSF1 on metabolism were independent of several known endocrine regulators and did not impact the physiological fasting response. Analysis using implantable telemetry in metabolic cages revealed progressively reduced body temperature and physical activity with no change in diurnal food intake. These results demonstrate the existence of a dynamic equilibrium between CSF1, the mononuclear phagocyte system, and control of liver-to-body weight ratio, which in turn controls systemic metabolic homeostasis. This novel macrophage regulatory axis has the potential to promote fat mobilization, without changes in appetence, which may have novel implications for managing metabolic syndrome.NEW & NOTEWORTHY CSF1 administration expands tissue macrophages, which transforms systemic metabolism. CSF1 drives fat mobilization and glucose uptake to support liver growth. The effects of CSF1 are independent of normal hormonal metabolic regulation. The effects of CSF1 are rapidly reversible, restoring homeostatic body composition. CSF1-dependent macrophages and liver size are coupled in a dynamic equilibrium.
Assuntos
Fator Estimulador de Colônias de Macrófagos , Macrófagos , Animais , Camundongos , Fator Estimulador de Colônias de Macrófagos/farmacologia , Fator Estimulador de Colônias de Macrófagos/metabolismo , Macrófagos/metabolismo , Metabolismo dos Carboidratos , Glucose/metabolismo , LipídeosRESUMO
Homozygous mutation of the Csf1r locus (Csf1rko) in mice, rats and humans leads to multiple postnatal developmental abnormalities. To enable analysis of the mechanisms underlying the phenotypic impacts of Csf1r mutation, we bred a rat Csf1rko allele to the inbred dark agouti (DA) genetic background and to a Csf1r-mApple reporter transgene. The Csf1rko led to almost complete loss of embryonic macrophages and ablation of most adult tissue macrophage populations. We extended previous analysis of the Csf1rko phenotype to early postnatal development to reveal impacts on musculoskeletal development and proliferation and morphogenesis in multiple organs. Expression profiling of 3-week old wild-type (WT) and Csf1rko livers identified 2760 differentially expressed genes associated with the loss of macrophages, severe hypoplasia, delayed hepatocyte maturation, disrupted lipid metabolism and the IGF1/IGF binding protein system. Older Csf1rko rats developed severe hepatic steatosis. Consistent with the developmental delay in the liver Csf1rko rats had greatly-reduced circulating IGF1. Transfer of WT bone marrow (BM) cells at weaning without conditioning repopulated resident macrophages in all organs, including microglia in the brain, and reversed the mutant phenotypes enabling long term survival and fertility. WT BM transfer restored osteoclasts, eliminated osteopetrosis, restored bone marrow cellularity and architecture and reversed granulocytosis and B cell deficiency. Csf1rko rats had an elevated circulating CSF1 concentration which was rapidly reduced to WT levels following BM transfer. However, CD43hi non-classical monocytes, absent in the Csf1rko, were not rescued and bone marrow progenitors remained unresponsive to CSF1. The results demonstrate that the Csf1rko phenotype is autonomous to BM-derived cells and indicate that BM contains a progenitor of tissue macrophages distinct from hematopoietic stem cells. The model provides a unique system in which to define the pathways of development of resident tissue macrophages and their local and systemic roles in growth and organ maturation.
Assuntos
Fígado Gorduroso/genética , Macrófagos/metabolismo , Anormalidades Musculoesqueléticas/genética , Desenvolvimento Musculoesquelético/genética , Osteopetrose/genética , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/genética , Animais , Medula Óssea/metabolismo , Medula Óssea/patologia , Transplante de Medula Óssea , Modelos Animais de Doenças , Embrião de Mamíferos , Fígado Gorduroso/metabolismo , Fígado Gorduroso/patologia , Fígado Gorduroso/terapia , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Técnicas de Inativação de Genes , Genes Reporter , Humanos , Proteínas de Ligação a Fator de Crescimento Semelhante a Insulina/deficiência , Proteínas de Ligação a Fator de Crescimento Semelhante a Insulina/genética , Fator de Crescimento Insulin-Like I/deficiência , Fator de Crescimento Insulin-Like I/genética , Metabolismo dos Lipídeos , Fígado/metabolismo , Fígado/patologia , Macrófagos/patologia , Masculino , Anormalidades Musculoesqueléticas/metabolismo , Anormalidades Musculoesqueléticas/patologia , Anormalidades Musculoesqueléticas/terapia , Osteopetrose/metabolismo , Osteopetrose/patologia , Osteopetrose/terapia , Ratos , Ratos Transgênicos , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/deficiênciaRESUMO
The laboratory rat continues to be the model of choice for many studies of physiology, behavior, and complex human diseases. Cells of the mononuclear phagocyte system (MPS; monocytes, macrophages, and dendritic cells) are abundant residents in every tissue in the body and regulate postnatal development, homeostasis, and innate and acquired immunity. Recruitment and proliferation of MPS cells is an essential component of both initiation and resolution of inflammation. The large majority of current knowledge of MPS biology is derived from studies of inbred mice, but advances in technology and resources have eliminated many of the advantages of the mouse as a model. In this article, we review the tools available and the current state of knowledge of development, homeostasis, regulation, and diversity within the MPS of the rat.
Assuntos
Imunidade Adaptativa , Modelos Animais de Doenças , Imunidade Inata , Sistema Fagocitário Mononuclear/imunologia , Ratos , Animais , Genoma , Homeostase/imunologia , Inflamação/imunologia , Fator Estimulador de Colônias de Macrófagos/genética , Camundongos , Mutação , FenótipoRESUMO
Mutations in the human CSF1R gene have been associated with dominant and recessive forms of neurodegenerative disease. Here we describe the impacts of Csf1r mutation in the rat on development of the brain. Diffusion imaging indicated small reductions in major fiber tracts that may be associated in part with ventricular enlargement. RNA-seq profiling revealed a set of 105 microglial markers depleted in all brain regions of the Csf1rko rats. There was no evidence of region or sex-specific expression of microglia-associated transcripts. Other than the microglial signature, Csf1rko had no effect on any neuronal or region-specific transcript cluster. Expression of markers of oligodendrocytes, astrocytes, dopaminergic neurons and Purkinje cells was minimally affected. However, there were defects in dendritic arborization of doublecortin-positive neurogenic precursors and expression of poly-sialylated neural cell adhesion molecule (PS-NCAM) in the dentate gyrus of the hippocampus. Heterozygous Csf1rko rats had no detectable brain phenotype. We conclude that most brain developmental processes occur normally in the absence of microglia and that CSF1R haploinsufficiency is unlikely to cause leukoencephalopathy.
Assuntos
Microglia , Doenças Neurodegenerativas/genética , Neurogênese/fisiologia , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/deficiência , Animais , Modelos Animais de Doenças , Feminino , Técnicas de Inativação de Genes , Humanos , Masculino , Mutação , Ratos , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/genéticaRESUMO
Repeated cycles of binge-like alcohol consumption and abstinence change the activity of several neurotransmitter systems. Some of these changes are consolidated following prolonged alcohol use and are thought to play an important role in the development of dependence. We have previously shown that systemic administration of the dual beta-adrenergic antagonist and 5-HT1A/1B partial agonist pindolol selectively reduces long-term but not short-term binge-like consumption of ethanol and alters excitatory postsynaptic currents in basolateral amygdala (BLA) principal neurons. The aim of this study was to investigate the effects of pindolol microinfusions in the BLA on long-term ethanol intake using the drinking-in-the-dark paradigm in mice. We also microinfused RU24969 (5-HT1A/1B receptor partial agonist) and CGP12177 (ß1/2 adrenergic antagonist) following long-term ethanol intake and determined the densities of 5-HT1A/1B receptors and ß1/2 adrenergic in the BLA following short-term (4 weeks) and long-term ethanol (12 weeks) consumption. We show that intra-BLA infusion of pindolol (1000 pmol/0.5 µl), RU24969 (0.3 and 3 pmol/0.5 µl) and CGP12177 (500 pmol/0.5 µl) produce robust decreases in long-term ethanol consumption. Additionally, we identified reduced ß1/2 adrenergic receptor expression and no change in 5-HT1A/1B receptor density in the BLA of long-term ethanol-consuming mice. Collectively, our data highlight the effects of pindolol on voluntary, binge-like ethanol consumption behavior following long-term intake.
Assuntos
Antagonistas de Receptores Adrenérgicos beta 2/farmacologia , Complexo Nuclear Basolateral da Amígdala/efeitos dos fármacos , Depressores do Sistema Nervoso Central/administração & dosagem , Etanol/administração & dosagem , Pindolol/farmacologia , Antagonistas da Serotonina/farmacologia , Agonistas do Receptor de Serotonina/farmacologia , Animais , Complexo Nuclear Basolateral da Amígdala/metabolismo , Depressores do Sistema Nervoso Central/farmacologia , Agonismo Parcial de Drogas , Etanol/farmacologia , Humanos , Indóis/farmacologia , Camundongos , Norepinefrina/metabolismo , Propanolaminas/farmacologia , Receptor 5-HT1A de Serotonina/efeitos dos fármacos , Receptor 5-HT1A de Serotonina/metabolismo , Receptor 5-HT1B de Serotonina/efeitos dos fármacos , Receptor 5-HT1B de Serotonina/metabolismo , Receptores Adrenérgicos beta 1/efeitos dos fármacos , Receptores Adrenérgicos beta 1/metabolismo , Receptores Adrenérgicos beta 2/efeitos dos fármacos , Receptores Adrenérgicos beta 2/metabolismo , Serotonina/metabolismoRESUMO
Alcohol dependence is a debilitating disorder with current therapies displaying limited efficacy and/or compliance. Consequently, there is a critical need for improved pharmacotherapeutic strategies to manage alcohol use disorders (AUDs). Previous studies have shown that the development of alcohol dependence involves repeated cycles of binge-like ethanol intake and abstinence. Therefore, we used a model of binge-ethanol consumption (drinking-in-the-dark) in mice to test the effects of compounds known to modify the activity of neurotransmitters implicated in alcohol addiction. From this, we have identified the FDA-approved antihypertensive drug pindolol, as a potential candidate for the management of AUDs. We show that the efficacy of pindolol to reduce ethanol consumption is enhanced following long-term (12 weeks) binge-ethanol intake, compared with short-term (4 weeks) intake. Furthermore, pindolol had no effect on locomotor activity or consumption of the natural reward sucrose. Because pindolol acts as a dual beta-adrenergic antagonist and 5-HT1A/1B partial agonist, we examined its effect on spontaneous synaptic activity in the basolateral amygdala (BLA), a brain region densely innervated by serotonin and norepinephrine-containing fibres. Pindolol increased spontaneous excitatory post-synaptic current frequency of BLA principal neurons from long-term ethanol-consuming mice but not naïve mice. Additionally, this effect was blocked by the 5-HT1A/1B receptor antagonist methiothepin, suggesting that altered serotonergic activity in the BLA may contribute to the efficacy of pindolol to reduce ethanol intake following long-term exposure. Although further mechanistic investigations are required, this study demonstrates the potential of pindolol as a new treatment option for AUDs that can be fast-tracked into human clinical studies.
Assuntos
Anti-Hipertensivos/farmacologia , Comportamento Animal/efeitos dos fármacos , Consumo Excessivo de Bebidas Alcoólicas/tratamento farmacológico , Etanol/administração & dosagem , Pindolol/farmacologia , Animais , Modelos Animais de Doenças , Masculino , Camundongos , Camundongos Endogâmicos C57BL , TempoRESUMO
Macrophages contribute to many aspects of development and homeostasis, innate and acquired immunity, immunopathology, and tissue repair. Every tissue contains an abundant resident macrophage population. Inflammatory stimuli promote the recruitment of monocytes from the blood and their adaptation promotes the removal of the stimulus and subsequent restoration of normal tissue architecture. Dysregulation of this response leads to chronic inflammation and tissue injury. In many tissues, their differentiation and survival are dependent on the colony stimulating factor 1 receptor (CSF1R) signalling axis, which is highly conserved across all vertebrates. Complete loss of either CSF1R or its cognate ligands, colony stimulating factor 1 (CSF1), and interleukin 34 (IL-34), results in the loss of many tissue-resident macrophage populations. This provides a useful paradigm to study macrophages.There are many tools used to visualize tissue-resident macrophages and their precursors, monocytes, in mice and humans. Particularly in mice there are genetic tools available to delete, enhance and manipulate monocytes and macrophages and their gene products to gain insight into phenotype and function. The laboratory rat has many advantages as an experimental model for the understanding of human disease, but the analytical resources are currently more limited than in mice. Here, we describe available genetic models, antibodies, and immunohistochemistry (IHC) methods that may be used to visualize tissue-resident macrophages in rats.
Assuntos
Fator Estimulador de Colônias de Macrófagos , Macrófagos , Humanos , Ratos , Camundongos , Animais , Imuno-Histoquímica , Monócitos , Aclimatação , Receptores Proteína Tirosina QuinasesRESUMO
Macrophage and osteoclast proliferation, differentiation and survival are regulated by colony-stimulating factor-1 receptor (CSF1R) signaling. Osteopetrosis associated with Csf1 and Csf1r mutations has been attributed to the loss of osteoclasts and deficiency in bone resorption. Here we demonstrate that homozygous Csf1r mutation in rat leads to delayed postnatal skeletal ossification associated with substantial loss of osteal macrophages (osteomacs) in addition to osteoclasts. Osteosclerosis and site-specific skeletal abnormalities were reversed by intraperitoneal transfer of wild-type bone marrow cells (BMT) at weaning. Following BMT, IBA1+ macrophages were detected before TRAP+ osteoclasts at sites of ossification restoration. These observations extend evidence that osteomacs independently contribute to bone anabolism and are required for normal postnatal bone growth and morphogenesis.
RESUMO
Resident and recruited macrophages control the development and proliferation of the liver. We have previously shown in multiple species that treatment with a macrophage colony stimulating factor (CSF1)-Fc fusion protein initiated hepatocyte proliferation and promoted repair in models of acute hepatic injury in mice. Here, we investigated the impact of CSF1-Fc on resolution of advanced fibrosis and liver regeneration, using a non-resolving toxin-induced model of chronic liver injury and fibrosis in C57BL/6J mice. Co-administration of CSF1-Fc with exposure to thioacetamide (TAA) exacerbated inflammation consistent with monocyte contributions to initiation of pathology. After removal of TAA, either acute or chronic CSF1-Fc treatment promoted liver growth, prevented progression and promoted resolution of fibrosis. Acute CSF1-Fc treatment was also anti-fibrotic and pro-regenerative in a model of partial hepatectomy in mice with established fibrosis. The beneficial impacts of CSF1-Fc treatment were associated with monocyte-macrophage recruitment and increased expression of remodelling enzymes and growth factors. These studies indicate that CSF1-dependent macrophages contribute to both initiation and resolution of fibrotic injury and that CSF1-Fc has therapeutic potential in human liver disease.
Assuntos
Hepatopatias , Fator Estimulador de Colônias de Macrófagos , Animais , Fibrose , Fígado/metabolismo , Hepatopatias/patologia , Fator Estimulador de Colônias de Macrófagos/metabolismo , Fator Estimulador de Colônias de Macrófagos/farmacologia , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BLRESUMO
While the dire cardiometabolic consequences of the hypercaloric modern 'Western' diet are well known, there is not much information on the health impact of a high sucrose diet not inducing weight gain. Here, we tested the hypothesis that rats reared with intermittent binge access to sucrose in addition to normal chow would develop an inflammatory response in brain. To test this hypothesis, we undertook serial PET/MRI scans with the TSPO ligand [18F]DPA714 in a group of (n=9) rats at baseline and again after voluntarily consuming 5% sucrose solution three days a week for three months. Compared to a control group fed with normal chow (n=9), the sucrose rats indeed showed widespread increases in the availability of cerebral binding sites for the microglial marker, despite normal weight gain compared to the control diet group. Subsequent immunofluorescence staining of the brains confirmed the PET findings, showing a widespread 20% increase in the abundance of IBA-1-positive microglia with characteristic 'semi-activated' morphology in the binge sucrose rats, which had 23% lower density of microglial endpoints and 25% lower mean process length compared to microglia in the control rats with ordinary feeding. GFAP immunofluorescence showed no difference in astroglial coverage in the sucrose rats, except for a slight reduction in hypothalamus. The binge sucrose diet-induced neuroinflammation was associated with a significant elevation of white blood cell counts. Taking these results together, we find that long-term intake of sucrose in a binge paradigm, similar in sucrose content to the contemporary Western diet, triggered a low-grade systemic and central inflammation in non-obese rats. The molecular mechanism of this phenomenon remains to be established.
Assuntos
Encéfalo/patologia , Dieta/efeitos adversos , Sacarose Alimentar/efeitos adversos , Inflamação/patologia , Obesidade/patologia , Animais , Astrócitos/patologia , Gliose/sangue , Gliose/complicações , Gliose/patologia , Inflamação/sangue , Inflamação/complicações , Fígado/patologia , Masculino , Microglia/patologia , Obesidade/sangue , Obesidade/complicações , Ratos , Ratos WistarRESUMO
A subpopulation of raphe 5-HT neurons expresses the vesicular glutamate transporter VGLUT3 with the co-release of glutamate and serotonin proposed to play a pivotal role in encoding reward- and anxiety-related behaviors. Serotonin axons are identifiable by immunolabeling of either serotonin (5-HT) or the plasma membrane 5-HT transporter (SERT), with SERT labeling demonstrated to be only partially overlapping with 5-HT staining. Studies investigating the colocalization or segregation of VGLUT3 within SERT or 5-HT immunolabeled boutons have led to inconsistent results. Therefore, we combined immunohistochemistry, high resolution confocal imaging, and 3D-reconstruction techniques to map and quantify the distribution of VGLUT3 immunoreactive boutons within 5-HT vs. SERT-positive axons in various regions of the mouse forebrain, including the prefrontal cortex, nucleus accumbens core and shell, bed nucleus of the stria terminalis, dorsal striatum, lateral septum, basolateral and central amygdala, and hippocampus. Our results demonstrate that about 90% of 5-HT boutons are colocalized with SERT in almost all the brain regions studied, which therefore reveals that VGLUT3 and SERT do not segregate. However, in the posterior part of the NAC shell, we confirmed the presence of a subtype of 5-HT immunoreactive axons that lack the SERT. Interestingly, about 90% of the 5-HT/VGLUT3 boutons were labeled for the SERT in this region, suggesting that VGLUT3 is preferentially located in SERT immunoreactive 5-HT boutons. This work demonstrates that VGLUT3 and SERT cannot be used as specific markers to classify the different subtypes of 5-HT axons.
RESUMO
Long-term binge alcohol consumption alters the signaling of numerous neurotransmitters in the brain including noradrenaline (NE) and serotonin (5-HT). Alterations in the signaling of these neuronal pathways result in dysfunctional emotional states like anxiety and depression which are typically seen during alcohol withdrawal. Interestingly, studies have demonstrated that the development of alcohol-induced negative affective states is linked to disrupted neurogenesis in the dentate gyrus (DG) region of the hippocampus in alcohol-dependent animals. We have previously shown that modulation of NE and 5-HT activity by pharmacological targeting of ß-adrenoreceptors (ß-ARs) and 5-HT1A/1B receptors with pindolol reduces consumption in long-term alcohol-consuming mice. Since these receptors are also involved in emotional homeostasis and hippocampal neurogenesis, we investigated the effects of pindolol administration on emotional and neurogenic deficits in mice consuming long-term alcohol (18 weeks). We report that acute administration of pindolol (32 mg/kg) reduces anxiety-like behavior in mice at 24 h withdrawal in the marble-burying test (MBT) and the elevated plus-maze (EPM). We also show that chronic (2 weeks) pindolol treatment (32 mg/kg/day) attenuates alcohol-induced impairments in the density of immature neurons (DCX+) but not newborn cells (BrdU+) in the hippocampal DG. Pindolol treatment also restores the normal proportion of newborn proliferating cells (BrdU+/Ki67+/DCX-), newborn proliferating immature neurons (BrdU+/Ki67+/DCX+) and newborn non-proliferating immature neurons (BrdU+/Ki67-/DCX+) following long-term alcohol intake. These results suggest that pindolol, through its unique pharmacology may rescue some but not all deficits of long-term alcohol abuse on the brain, adding further value to its properties as a strong pharmaceutical option for alcohol use disorders (AUDs).
RESUMO
Repeated episodes of binge-like alcohol consumption produce anxiety, depression and various deleterious effects including alterations in neurogenesis. While the involvement of the serotonin receptor 1 A (5-HT1A) in the regulation of anxiety-like behavior and neurogenesis is well documented, its contribution to alcohol withdrawal-induced anxiety and alcohol-induced deficits in neurogenesis is less documented. Using the Drinking-In-the-Dark (DID) paradigm to model chronic long-term (12 weeks) binge-like voluntary alcohol consumption in mice, we show that the selective partial activation of 5-HT1A receptors by tandospirone (3 mg/kg) prevents alcohol withdrawal-induced anxiety in a battery of behavioral tests (marble burying, elevated-plus-maze, open-field), which is accompanied by a robust decrease in binge-like ethanol intake (1 and 3 mg/kg). Furthermore, using triple immunolabelling of proliferation and neuronal differentiation markers, we show that long-term DID elicits profound deficits in neurogenesis and neuronal fate specification in the dorsal hippocampus that are entirely reversed by a 2-week chronic treatment with the 5-HT1A partial agonist tandospirone (3 mg/kg/day). Together, our results confirm previous observations that 5-HT1A receptors play a pivotal role in alcohol drinking behavior and the associated emotional and neurogenic impairments, and suggest that 5-HT1A partial agonists represent a promising treatment strategy for alcohol abuse.
Assuntos
Consumo de Bebidas Alcoólicas/efeitos adversos , Ansiedade/induzido quimicamente , Transtorno Depressivo/induzido quimicamente , Emoções/efeitos dos fármacos , Doenças do Sistema Nervoso/induzido quimicamente , Receptor 5-HT1A de Serotonina/metabolismo , Animais , Ansiedade/metabolismo , Ansiedade/patologia , Transtorno Depressivo/metabolismo , Transtorno Depressivo/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Doenças do Sistema Nervoso/metabolismo , Doenças do Sistema Nervoso/patologiaRESUMO
Serotonin neurons arise from the brainstem raphe nuclei and send their projections throughout the brain to release 5-HT which acts as a modulator of several neuronal populations. Previous electron microscopy studies in rats have morphologically determined the distribution of 5-HT release sites (boutons) in certain brain regions and have shown that 5-HT containing boutons form synaptic contacts that are either symmetric or asymmetric. In addition, 5-HT boutons can form synaptic triads with the pre- and postsynaptic specializations of either symmetrical or asymmetrical synapses. However, due to the labor intensive processing of serial sections required by electron microscopy, little is known about the neurochemical properties or the quantitative distribution of 5-HT triads within whole brain or discrete subregions. Therefore, we used a semi-automated approach that combines immunohistochemistry and high-resolution confocal microscopy to label serotonin transporter (SERT) immunoreactive axons and reconstruct in 3D their distribution within limbic brain regions. We also used antibodies against key pre- (synaptophysin) and postsynaptic components of excitatory (PSD95) or inhibitory (gephyrin) synapses to (1) identify putative 5-HTergic boutons within SERT immunoreactive axons and, (2) quantify their close apposition to neurochemical excitatory or inhibitory synapses. We provide a 5-HTergic axon density map and have determined the ratio of synaptic triads consisting of a 5-HT bouton in close proximity to either neurochemical excitatory or inhibitory synapses within different limbic brain areas. The ability to model and map changes in 5-HTergic axonal density and the formation of triadic connectivity within whole brain regions using this rapid and quantitative approach offers new possibilities for studying neuroplastic changes in the 5-HTergic pathway.
Assuntos
Axônios/metabolismo , Mapeamento Encefálico , Sistema Límbico/citologia , Proteínas da Membrana Plasmática de Transporte de Serotonina/metabolismo , Sinapses/metabolismo , Sinapses/fisiologia , Análise de Variância , Animais , Axônios/ultraestrutura , Proteínas de Transporte/metabolismo , Proteína 4 Homóloga a Disks-Large , Guanilato Quinases/metabolismo , Imageamento Tridimensional , Sistema Límbico/fisiologia , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Microscopia Confocal , Microscopia Eletrônica , Terminações Pré-Sinápticas/metabolismo , Terminações Pré-Sinápticas/ultraestrutura , Serotonina/metabolismo , Sinapses/ultraestrutura , Sinaptofisina/metabolismoRESUMO
Alcohol addiction is a debilitating disorder producing maladaptive changes in the brain, leading drinkers to become more sensitive to stress and anxiety. These changes are key factors contributing to alcohol craving and maintaining a persistent vulnerability to relapse. Serotonin (5-Hydroxytryptamine, 5-HT) is a monoamine neurotransmitter widely expressed in the central nervous system where it plays an important role in the regulation of mood. The serotonin system has been extensively implicated in the regulation of stress and anxiety, as well as the reinforcing properties of all of the major classes of drugs of abuse, including alcohol. Dysregulation within the 5-HT system has been postulated to underlie the negative mood states associated with alcohol use disorders. This review will describe the serotonergic (5-HTergic) neuroplastic changes observed in animal models throughout the alcohol addiction cycle, from prenatal to adulthood exposure. The first section will focus on alcohol-induced 5-HTergic neuroadaptations in offspring prenatally exposed to alcohol and the consequences on the regulation of stress/anxiety. The second section will compare alterations in 5-HT signalling induced by acute or chronic alcohol exposure during adulthood and following alcohol withdrawal, highlighting the impact on the regulation of stress/anxiety signalling pathways. The third section will outline 5-HTergic neuroadaptations observed in various genetically-selected ethanol preferring rat lines. Finally, we will discuss the pharmacological manipulation of the 5-HTergic system on ethanol- and anxiety/stress-related behaviours demonstrated by clinical trials, with an emphasis on current and potential treatments.
RESUMO
BACKGROUND: Our laboratory has previously shown that the smoking-cessation agent varenicline, an agonist/partial agonist of α4ß2*, α3ß4*, α3ß2*, α6ß2* (* indicates the possibility of additional subunits) and α7 subunits of nicotinic acetylcholine receptors (nAChRs), reduces ethanol consumption in rats only after long-term exposure (12 weeks). As compounds having partial agonistic activity on α3ß4* nAChRs were shown to decrease ethanol consumption in rodents, we assessed here the involvement of the ß4 subunit in the effect of varenicline in the reduction of short- and long-term binge-like ethanol drinking in mice. METHODS: We used the well-validated drinking-in-the-dark (DID) paradigm to model chronic binge-like ethanol drinking in ß4-/- and ß4+/+ littermate mice and compare the effect of intraperitoneal injection of varenicline (2mg/kg) on ethanol intake following short- (4 weeks) or long-term (12 weeks) exposure. RESULTS: Drinking pattern and amounts of ethanol intake were similar in ß4-/- and ß4+/+ mice. Interestingly, our results showed that varenicline reduces ethanol consumption following short- and long-term ethanol exposure in the DID. Although the effect of varenicline on the reduction of ethanol consumption was slightly more pronounced in ß4-/- mice than their ß4+/+ littermates no significant differences were observed between genotypes. CONCLUSION: In mice, varenicline reduces binge-like ethanol consumption both after short- and long-term exposure in the DID and this effect is independent of ß4 nAChR subunit.
Assuntos
Consumo Excessivo de Bebidas Alcoólicas/terapia , Proteínas do Tecido Nervoso/metabolismo , Receptores Nicotínicos/metabolismo , Vareniclina/uso terapêutico , Animais , Consumo Excessivo de Bebidas Alcoólicas/metabolismo , Etanol/sangue , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas do Tecido Nervoso/genética , Receptores Nicotínicos/genética , Fatores de TempoRESUMO
Excess sugar consumption has been shown to contribute directly to weight gain, thus contributing to the growing worldwide obesity epidemic. Interestingly, increased sugar consumption has been shown to repeatedly elevate dopamine levels in the nucleus accumbens (NAc), in the mesolimbic reward pathway of the brain similar to many drugs of abuse. We report that varenicline, an FDA-approved nicotinic acetylcholine receptor (nAChR) partial agonist that modulates dopamine in the mesolimbic reward pathway of the brain, significantly reduces sucrose consumption, especially in a long-term consumption paradigm. Similar results were observed with other nAChR drugs, namely mecamylamine and cytisine. Furthermore, we show that long-term sucrose consumption increases α4ß2 * and decreases α6ß2* nAChRs in the nucleus accumbens, a key brain region associated with reward. Taken together, our results suggest that nAChR drugs such as varenicline may represent a novel treatment strategy for reducing sugar consumption.