RESUMEN
Oligodendrocyte pathology is increasingly implicated in neurodegenerative diseases as oligodendrocytes both myelinate and provide metabolic support to axons. In multiple sclerosis (MS), demyelination in the central nervous system thus leads to neurodegeneration, but the severity of MS between patients is very variable. Disability does not correlate well with the extent of demyelination1, which suggests that other factors contribute to this variability. One such factor may be oligodendrocyte heterogeneity. Not all oligodendrocytes are the same-those from the mouse spinal cord inherently produce longer myelin sheaths than those from the cortex2, and single-cell analysis of the mouse central nervous system identified further differences3,4. However, the extent of human oligodendrocyte heterogeneity and its possible contribution to MS pathology remain unknown. Here we performed single-nucleus RNA sequencing from white matter areas of post-mortem human brain from patients with MS and from unaffected controls. We identified subclusters of oligodendroglia in control human white matter, some with similarities to mouse, and defined new markers for these cell states. Notably, some subclusters were underrepresented in MS tissue, whereas others were more prevalent. These differences in mature oligodendrocyte subclusters may indicate different functional states of oligodendrocytes in MS lesions. We found similar changes in normal-appearing white matter, showing that MS is a more diffuse disease than its focal demyelination suggests. Our findings of an altered oligodendroglial heterogeneity in MS may be important for understanding disease progression and developing therapeutic approaches.
Asunto(s)
Encéfalo/metabolismo , Encéfalo/patología , Esclerosis Múltiple/patología , Oligodendroglía/patología , Adulto , Anciano , Anciano de 80 o más Años , Animales , Autopsia , Biomarcadores , Estudios de Casos y Controles , Progresión de la Enfermedad , Femenino , Perfilación de la Expresión Génica , Regulación de la Expresión Génica/genética , Humanos , Masculino , Ratones , Persona de Mediana Edad , Esclerosis Múltiple/genética , Vaina de Mielina/genética , Vaina de Mielina/metabolismo , Vaina de Mielina/patología , Oligodendroglía/metabolismo , Remielinización/genética , Análisis de Secuencia de ARN , Transcripción Genética , Sustancia Blanca/citología , Sustancia Blanca/metabolismo , Sustancia Blanca/patologíaRESUMEN
Astrocytes are involved in non-cell-autonomous pathogenic cascades in amyotrophic lateral sclerosis (ALS); however, their role is still debated. We show that astrocytic NF-κB activation drives microglial proliferation and leukocyte infiltration in the SOD1 (G93A) ALS model. This response prolongs the presymptomatic phase, delaying muscle denervation and decreasing disease burden, but turns detrimental in the symptomatic phase, accelerating disease progression. The transition corresponds to a shift in the microglial phenotype showing two effects that can be dissociated by temporally controlling NF-κB activation. While NF-κB activation in astrocytes induced a Wnt-dependent microglial proliferation in the presymptomatic phase with neuroprotective effects on motoneurons, in later stage, astrocyte NF-κB-dependent microglial activation caused an accelerated disease progression. Notably, suppression of the early microglial response by CB2R agonists had acute detrimental effects. These data identify astrocytes as important regulators of microglia expansion and immune response. Therefore, stage-dependent microglia modulation may be an effective therapeutic strategy in ALS.
Asunto(s)
Esclerosis Amiotrófica Lateral/inmunología , Astrocitos/inmunología , FN-kappa B/inmunología , Esclerosis Amiotrófica Lateral/genética , Esclerosis Amiotrófica Lateral/patología , Esclerosis Amiotrófica Lateral/terapia , Animales , Astrocitos/patología , Modelos Animales de Enfermedad , Ratones , Ratones Transgénicos , Microglía/inmunología , Microglía/patología , Neuronas Motoras/inmunología , Neuronas Motoras/patología , FN-kappa B/genética , Receptor Cannabinoide CB2/agonistas , Receptor Cannabinoide CB2/genética , Receptor Cannabinoide CB2/inmunología , Superóxido Dismutasa/genética , Superóxido Dismutasa/inmunología , Superóxido Dismutasa-1/genética , Superóxido Dismutasa-1/inmunologíaRESUMEN
Immune-inflammatory activation impacts extracellular vesicles (EVs), including their miRNA cargo. There is evidence for changes in the EV miRNome in inflammation-associated neuropsychiatric disorders. This mouse study investigated: (1) effects of systemic lipopolysaccharide (LPS) and chronic social stress (CSS) on plasma EV miRNome; and (2) physiological, transcriptional, and behavioural effects of peripheral or central delivered LPS-activated EVs in recipient mice. LPS or CSS effects on the plasma EV miRNome were assessed by using microRNA sequencing. Recipient mice received plasma EVs isolated from LPS-treated or SAL-treated donor mice or vehicle only, either intravenously or into the nucleus accumbens (NAc), on three consecutive days. Bodyweight, spleen or NAc transcriptome and reward (sucrose) motivation were assessed. LPS and CSS increased the expression of 122 and decreased expression of 20 plasma EV miRNAs, respectively. Peripheral LPS-EVs reduced bodyweight, and both LPS-EVs and SAL-EVs increased spleen expression of immune-relevant genes. NAc-infused LPS-EVs increased the expression of 10 immune-inflammatory genes. Whereas motivation increased similarly across test days in all groups, the effect of test days was more pronounced in mice that received peripheral or central LPS-EVs compared with other groups. This study provides causal evidence that increased EV levels impact physiological and behavioural processes and are of potential relevance to neuropsychiatric disorders.
Asunto(s)
Vesículas Extracelulares/genética , Perfilación de la Expresión Génica/métodos , Lipopolisacáridos/efectos adversos , MicroARNs/genética , Estrés Psicológico/genética , Animales , Vesículas Extracelulares/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Masculino , Ratones , Tamaño de los Órganos/efectos de los fármacos , Proyectos Piloto , Análisis de Secuencia de ARN , Bazo/efectos de los fármacos , Bazo/metabolismo , Estrés Psicológico/psicologíaRESUMEN
Multiple sclerosis (MS) is a chronic inflammatory autoimmune disease of the central nervous system (CNS). Its first clinical presentation (clinically isolated syndrome, CIS) is often followed by the development of relapsing-remitting MS (RRMS). The periphery-to-CNS transmission of inflammatory molecules is a major pathophysiological pathway in MS. This could include signalling via extracellular vesicle (EV) microRNAs (miRNAs). In this study, we investigated the serum EV miRNome in CIS and RRMS patients and matched controls, with the aims to identify MS stage-specific differentially expressed miRNAs and investigate their biomarker potential and pathophysiological relevance. miRNA sequencing was conducted on serum EVs from CIS-remission, RRMS-relapse, and viral inflammatory CNS disorder patients, as well as from healthy and hospitalized controls. Differential expression analysis was conducted, followed by predictive power and target-pathway analysis. A moderate number of dysregulated serum EV miRNAs were identified in CIS-remission and RRMS-relapse patients, especially relative to healthy controls. Some of these miRNAs were also differentially expressed between the two MS stages and had biomarker potential for patient-control and CIS-RRMS separations. For the mRNA targets of the RRMS-relapse-specific EV miRNAs, biological processes inherent to MS pathophysiology were identified using in silico analysis. Study findings demonstrate that specific serum EV miRNAs have MS stage-specific biomarker potential and contribute to the identification of potential targets for novel, efficacious therapies.
Asunto(s)
Vesículas Extracelulares/metabolismo , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , MicroARNs/sangre , Esclerosis Múltiple Recurrente-Remitente/sangre , Adolescente , Adulto , Anciano , Biomarcadores/sangre , Femenino , Humanos , Masculino , Persona de Mediana EdadRESUMEN
Genetic variants in the triggering receptor expressed on myeloid cells 2 (TREM2) increase the risk for several neurodegenerative diseases including Alzheimer's disease and frontotemporal dementia (FTD). Homozygous TREM2 missense mutations, such as p.T66M, lead to the FTD-like syndrome, but how they cause pathology is unknown. Using CRISPR/Cas9 genome editing, we generated a knock-in mouse model for the disease-associated Trem2 p.T66M mutation. Consistent with a loss-of-function mutation, we observe an intracellular accumulation of immature mutant Trem2 and reduced generation of soluble Trem2 similar to patients with the homozygous p.T66M mutation. Trem2 p.T66M knock-in mice show delayed resolution of inflammation upon in vivo lipopolysaccharide stimulation and cultured macrophages display significantly reduced phagocytic activity. Immunohistochemistry together with in vivo TSPO small animal positron emission tomography (µPET) demonstrates an age-dependent reduction in microglial activity. Surprisingly, perfusion magnetic resonance imaging and FDG-µPET imaging reveal a significant reduction in cerebral blood flow and brain glucose metabolism. Thus, we demonstrate that a TREM2 loss-of-function mutation causes brain-wide metabolic alterations pointing toward a possible function of microglia in regulating brain glucose metabolism.
Asunto(s)
Encéfalo/patología , Demencia Frontotemporal/patología , Glucosa/metabolismo , Glicoproteínas de Membrana/genética , Microglía/fisiología , Mutación Missense , Perfusión , Receptores Inmunológicos/genética , Animales , Modelos Animales de Enfermedad , Técnicas de Sustitución del Gen , Humanos , Inmunohistoquímica , Imagen por Resonancia Magnética , Ratones , Proteínas Mutantes/genética , Tomografía de Emisión de PositronesRESUMEN
Maternal immune activation (mIA) in rodents is rapidly emerging as a key model for neurodevelopmental disorders such as autism spectrum disorder (ASD) and schizophrenia. Here, we optimise a mIA model in rats, aiming to address certain limitations of current work in this field. Specifically, the lack of clear evidence for methodology chosen, identification of successful induction of mIA in the dams and investigation of male offspring only. We focus on gestational and early juvenile changes in offspring following mIA, as detailed information on these critical early developmental time points is sparse. Following strain (Wistar, Lister Hooded, Sprague Dawley) comparison and selection, and polyriboinosinic-polyribocytidylic acid (poly I:C) dose selection (2.5-15â¯mg/kg single or once daily for 5â¯days), mIA was induced in pregnant Wistar rats with 10â¯mg/kg poly I:C i.p. on gestational day (GD) 15. Early morphometric analysis was conducted in male and female offspring at GD21 and postnatal day (PD) 21, eight dams for each treatment at each time point were used, 32 in total. Subsequent microglia analysis was conducted at PD21 in a small group of offspring. Poly I:C at 10â¯mg/kg i.p. induced a robust, but variable, plasma IL-6 response 3â¯h post-injection and reduced body weight at 6â¯h and 24â¯h post-injection in two separate cohorts of Wistar rats at GD15. Plasma IL-6 was not elevated at PD21 in offspring or dams. Poly I:C-induced mIA did not affect litter numbers, but resulted in PD21 pup, and GD21 placenta growth restriction. Poly I:C significantly increased microglial activation at PD21 in male hippocampi. We have identified 10â¯mg/kg poly I:C i.p on GD15 as a robust experimental approach for inducing mIA in Wistar rats and used this to identify early neurodevelopmental changes. This work provides a framework to study the developmental trajectory of disease-relevant, sex-specific phenotypic changes in rats.
Asunto(s)
Inmunidad Activa/fisiología , Activación de Linfocitos/inmunología , Efectos Tardíos de la Exposición Prenatal/inmunología , Animales , Conducta Animal/fisiología , Citocinas/inmunología , Modelos Animales de Enfermedad , Femenino , Hipocampo/efectos de los fármacos , Inmunidad Activa/inmunología , Interleucina-6/metabolismo , Activación de Linfocitos/fisiología , Masculino , Modelos Animales , Actividad Motora/efectos de los fármacos , Trastornos del Neurodesarrollo , Placenta/metabolismo , Poli I-C/farmacología , Embarazo , Ratas , Ratas Wistar , Esquizofrenia/inmunología , Linfocitos T/inmunologíaRESUMEN
Aberrant epileptic activity is detectable at early disease stages in Alzheimer's disease (AD) patients and in AD mouse models. Here, we investigated in young ArcticAß mice whether AD-like pathology renders neuronal networks more susceptible to the development of acquired epilepsy induced by unilateral intrahippocampal injection of kainic acid (IHK). In this temporal lobe epilepsy model, IHK induces a status epilepticus followed after two weeks by spontaneous recurrent seizures (SRS). ArcticAß mice exhibited more severe status epilepticus and early onset of SRS. This hyperexcitable phenotype was characterized in CA1 neurons by decreased synaptic strength, increased kainic acid-induced LTP and reduced frequency of spontaneous inhibitory currents. However, no difference in neurodegeneration, neuroinflammation, axonal reorganization or adult neurogenesis was observed in ArcticAß mice compared to wild-type littermates following IHK-induced epileptogenesis. Neuropeptide Y (NPY) expression was reduced at baseline and its IHK-induced elevation in mossy fibres and granule cells was attenuated. However, although this alteration might underlie premature seizure onset, neutralization of soluble Aß species by intracerebroventricular Aß-specific antibody application mitigated the hyperexcitable phenotype of ArcticAß mice and prevented early SRS onset. Therefore, the development of seizures at early stages of AD is mediated primarily by Aß species causing widespread changes in synaptic function.
Asunto(s)
Enfermedad de Alzheimer , Péptidos beta-Amiloides , Epilepsia del Lóbulo Temporal , Hipocampo , Plasticidad Neuronal , Convulsiones , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/fisiopatología , Péptidos beta-Amiloides/metabolismo , Animales , Región CA1 Hipocampal/metabolismo , Región CA1 Hipocampal/fisiopatología , Giro Dentado/metabolismo , Giro Dentado/fisiopatología , Modelos Animales de Enfermedad , Electroencefalografía , Epilepsia del Lóbulo Temporal/metabolismo , Epilepsia del Lóbulo Temporal/fisiopatología , Agonistas de Aminoácidos Excitadores/farmacología , Hipocampo/metabolismo , Hipocampo/fisiopatología , Ácido Kaínico/farmacología , Masculino , Ratones , Ratones Transgénicos , Plasticidad Neuronal/fisiología , Convulsiones/metabolismo , Convulsiones/fisiopatología , Estado Epiléptico/metabolismo , Estado Epiléptico/fisiopatologíaRESUMEN
Amyloid-ß (Aß) deposition in the cerebral vasculature is accompanied by remodeling which has a profound influence on vascular integrity and function. In the current study we have quantitatively assessed the age-dependent changes of the cortical vasculature in the arcAß model of cerebral amyloidosis. To estimate the density of the cortical microvasculature in vivo, we used contrast-enhanced magnetic resonance microangiography (CE-µMRA). Three-dimensional gradient echo datasets with 60 µm isotropic resolution were acquired in 4- and 24-month-old arcAß mice and compared with wild-type (wt) control mice of the same age before and after administration of superparamagnetic iron oxide nanoparticles. After segmentation of the cortical vasculature from difference images, an automated algorithm was applied for assessing the number and size distribution of intracortical vessels. With CE-µMRA, cerebral arteries and veins with a diameter of less than the nominal pixel resolution (60 µm) can be visualized. A significant age-dependent reduction in the number of functional intracortical microvessels (radii of 20-80 µm) has been observed in 24-month-old arcAß mice compared with age-matched wt mice, whereas there was no difference between transgenic and wt mice of 4 months of age. Immunohistochemistry demonstrated strong fibrinogen and Aß deposition in small- and medium-sized vessels, but not in large cerebral arteries, of 24-month-old arcAß mice. The reduced density of transcortical vessels may thus be attributed to impaired perfusion and vascular occlusion caused by deposition of Aß and fibrin. The study demonstrated that remodeling of the cerebrovasculature can be monitored noninvasively with CE-µMRA in mice.
Asunto(s)
Péptidos beta-Amiloides/genética , Circulación Cerebrovascular/genética , Medios de Contraste , Proteínas del Citoesqueleto/genética , Angiografía por Resonancia Magnética/métodos , Microvasos/fisiología , Proteínas del Tejido Nervioso/genética , Péptidos beta-Amiloides/metabolismo , Animales , Angiopatía Amiloide Cerebral/genética , Angiopatía Amiloide Cerebral/metabolismo , Angiopatía Amiloide Cerebral/fisiopatología , Proteínas del Citoesqueleto/metabolismo , Femenino , Fibrinógeno/genética , Fibrinógeno/metabolismo , Humanos , Masculino , Ratones , Ratones Transgénicos , Microvasos/diagnóstico por imagen , Proteínas del Tejido Nervioso/metabolismo , RadiografíaRESUMEN
The synaptic Ras/Rap-GTPase-activating protein (SynGAP1) plays a unique role in regulating specific downstream intracellular events in response to N-methyl-D-aspartate receptor (NMDAR) activation. Constitutive heterozygous loss of SynGAP1 disrupts NMDAR-mediated physiological and behavioral processes, but the disruptions might be of developmental origin. Therefore, the precise role of SynGAP1 in the adult brain, including its relative functional significance within specific brain regions, remains unexplored. The present study constitutes the first attempt in achieving adult hippocampal-specific SynGAP1 knockout using the Cre/loxP approach. Here, we report that this manipulation led to a significant numerical increase in both small and large GluA1 and NR1 immunoreactive clusters, many of which were non-opposed to presynaptic terminals. In parallel, the observed marked decline in the amplitude of spontaneous excitatory currents (sEPSCs) and inter-event intervals supported the impression that SynGAP1 loss might facilitate the accumulation of extrasynaptic glutamatergic receptors. In addition, SynGAP1-mediated signaling appears to be critical for the proper integration and survival of newborn neurons. The manipulation impaired reversal learning in the probe test of the water maze and induced a delay-dependent impairment in spatial recognition memory. It did not significantly affect anxiety or reference memory acquisition but induced a substantial elevation in spontaneous locomotor activity in the open field test. Thus, the present study demonstrates the functional significance of SynGAP1 signaling in the adult brain by capturing several changes that are dependent on NMDAR and hippocampal integrity.
Asunto(s)
Hipocampo/citología , Discapacidades para el Aprendizaje/genética , Neuronas/fisiología , Transmisión Sináptica/genética , Proteínas Activadoras de ras GTPasa/deficiencia , Análisis de Varianza , Animales , Reacción de Prevención/fisiología , Proteínas de Dominio Doblecortina , Regulación de la Expresión Génica/genética , Vectores Genéticos/genética , Proteínas Fluorescentes Verdes/genética , Hipocampo/metabolismo , Integrasas/genética , Integrasas/metabolismo , Aprendizaje por Laberinto/fisiología , Potenciales de la Membrana/genética , Trastornos de la Memoria/genética , Ratones , Ratones Transgénicos , Proteínas Asociadas a Microtúbulos/metabolismo , Actividad Motora/genética , Neuronas/citología , Neuronas/efectos de los fármacos , Neuropéptidos/metabolismo , Técnicas de Placa-Clamp , Tiempo de Reacción/genética , Receptores AMPA/genética , Receptores AMPA/metabolismo , Receptores de N-Metil-D-Aspartato/genética , Receptores de N-Metil-D-Aspartato/metabolismo , Conducta Espacial/fisiología , Transmisión Sináptica/efectos de los fármacos , Transducción Genética , Proteínas Activadoras de ras GTPasa/metabolismoRESUMEN
Cellular senescence is the consequence of repetitive exposures to oxidative stress, perturbed energy homeostasis, accumulation of damaged proteins and lesions in their nucleic acids. Whereas mitotic cells are equipped with efficient cell replacement strategies; postmitotic neurons have--with a few exceptions--no mechanism to substitute dysfunctional cells within a complex neuronal network. Here we propose a potential strategy by which aging neurons contend against abnormal accumulation of damaged/misfolded proteins. The suggested mechanism involves the formation of 'budding-like' extrusions and their subsequent clearance by glia. This hypothesis emerged from our previous investigations of the aged hippocampus revealing layer-specific accumulations of Reelin, a glycoprotein with fundamental roles during brain development and adult synaptic plasticity. We showed that Reelin deposits constitute a conserved neuropathological feature of aging, which is significantly accelerated in adult wild-type mice prenatally exposed to a viral-like infection. Here, we employed two- and three-dimensional immunoelectron microscopy to elucidate their morphological properties, localization and origin in immune challenged vs. control mice. In controls, Reelin-positive deposits were dispersed in the neuropil, some being engulfed by glia. In immune challenged mice, however, significantly more Reelin-immunoreactive deposits were associated with neuritic swellings containing mitochondria, vacuoles and cellular debris, pointing to their intracellular origin and suggesting that 'budding-like' neuronal extrusions of misfolded proteins and glial clearance may represent a protective strategy to counteract aging-associated impairments in proteosomal/lysosomal degradation. Neurons exposed to chronic neuroinflammation with increased levels of misfolded/damaged proteins, however, may fail to combat intraneuronal protein accumulations, a process probably underlying neuronal dysfunction and degeneration during aging.
Asunto(s)
Moléculas de Adhesión Celular Neuronal/metabolismo , Senescencia Celular/fisiología , Proteínas de la Matriz Extracelular/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Neuronas/metabolismo , Pliegue de Proteína , Serina Endopeptidasas/metabolismo , Animales , Moléculas de Adhesión Celular Neuronal/química , Proteínas de la Matriz Extracelular/química , Femenino , Hipocampo/fisiopatología , Masculino , Ratones , Ratones Endogámicos C57BL , Microscopía Electrónica de Rastreo , Microscopía Inmunoelectrónica , Proteínas del Tejido Nervioso/química , Neuroglía/metabolismo , Neuroglía/ultraestructura , Neuronas/ultraestructura , Poli I-C/farmacología , Embarazo , Efectos Tardíos de la Exposición Prenatal/inmunología , Proteína Reelina , Serina Endopeptidasas/químicaRESUMEN
BACKGROUND: Alzheimer's disease (AD) is the most prevalent form of age-related dementia, and its effect on society increases exponentially as the population ages. Accumulating evidence suggests that neuroinflammation, mediated by the brain's innate immune system, contributes to AD neuropathology and exacerbates the course of the disease. However, there is no experimental evidence for a causal link between systemic inflammation or neuroinflammation and the onset of the disease. METHODS: The viral mimic, polyriboinosinic-polyribocytidilic acid (PolyI:C) was used to stimulate the immune system of experimental animals. Wild-type (WT) and transgenic mice were exposed to this cytokine inducer prenatally (gestation day (GD)17) and/or in adulthood. Behavioral, immunological, immunohistochemical, and biochemical analyses of AD-associated neuropathologic changes were performed during aging. RESULTS: We found that a systemic immune challenge during late gestation predisposes WT mice to develop AD-like neuropathology during the course of aging. They display chronic elevation of inflammatory cytokines, an increase in the levels of hippocampal amyloid precursor protein (APP) and its proteolytic fragments, altered Tau phosphorylation, and mis-sorting to somatodendritic compartments, and significant impairments in working memory in old age. If this prenatal infection is followed by a second immune challenge in adulthood, the phenotype is strongly exacerbated, and mimics AD-like neuropathologic changes. These include deposition of APP and its proteolytic fragments, along with Tau aggregation, microglia activation and reactive gliosis. Whereas Aß peptides were not significantly enriched in extracellular deposits of double immune-challenged WT mice at 15 months, they dramatically increased in age-matched immune-challenged transgenic AD mice, precisely around the inflammation-induced accumulations of APP and its proteolytic fragments, in striking similarity to the post-mortem findings in human patients with AD. CONCLUSION: Chronic inflammatory conditions induce age-associated development of an AD-like phenotype in WT mice, including the induction of APP accumulations, which represent a seed for deposition of aggregation-prone peptides. The PolyI:C mouse model therefore provides a unique tool to investigate the molecular mechanisms underlying the earliest pathophysiological changes preceding fibrillary Aß plaque deposition and neurofibrillary tangle formations in a physiological context of aging. Based on the similarity between the changes in immune-challenged mice and the development of AD in humans, we suggest that systemic infections represent a major risk factor for the development of AD.
Asunto(s)
Enfermedad de Alzheimer/patología , Encéfalo/patología , Efectos Tardíos de la Exposición Prenatal/patología , Anciano de 80 o más Años , Enfermedad de Alzheimer/inmunología , Animales , Encéfalo/inmunología , Modelos Animales de Enfermedad , Femenino , Humanos , Masculino , Aprendizaje por Laberinto/fisiología , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Transgénicos , Poli I-C/toxicidad , Embarazo , Efectos Tardíos de la Exposición Prenatal/inducido químicamente , Efectos Tardíos de la Exposición Prenatal/inmunologíaRESUMEN
The main olfactory epithelium consists of 4 major cell types: sensory neurons, supporting cells, microvillar cells, and basal progenitor cells. Several populations of microvillar olfactory cells have been described, whose properties are not yet fully understood. In this study, we aimed to clarify the classification of microvillar cells by introducing a specific marker, CD73. Furthermore, we investigated the turnover of CD73-microvillar cells during adult life. Using direct and indirect immunofluorescence in adult main olfactory epithelium, we first demonstrate that ecto-5'-nucleotidase (CD73) is a reliable marker for microvillar cells reported previously to express phospholipase C ß2 (PLC ß2) along with type 3 IP(3) receptors (IP(3)R3) and transient receptor potential channels 6 (TRPC6), as well as for cells labeled by transgenic expression of tauGFP driven by the IP(3)R3 promoter. The ubiquitous CD73 immunoreactivity in the microvilli of these 2 cell populations indicates that they correspond to the same cell type (CD73-microvillar cell), endowed with a signal transduction cascade mobilizing Ca(++) from intracellular stores. These microvillar cells respond to odors, possess a basal process, and do not degenerate after bulbectomy, suggesting that they contribute to cellular homeostasis in the olfactory epithelium. Next, we examined whether CD73-microvillar cells undergo turnover in the adult olfactory epithelium. By combining CD73 immunofluorescence and BrdU pulse labeling, we show delayed BrdU incorporation in a small fraction of CD73-positive microvillar cells, which persists for several weeks after BrdU administration. These findings indicate that CD73-microvillar cells likely differentiate from proliferating progenitor cells and have a slow turnover despite their apical position in the olfactory epithelium. These combined properties are unique among olfactory cells, in line with the possibility that they might regulate cellular homeostasis driven by extracellular ATP and adenosine.
Asunto(s)
5'-Nucleotidasa/metabolismo , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Microvellosidades/metabolismo , Mucosa Olfatoria/metabolismo , Animales , Bromodesoxiuridina/química , Calcio/metabolismo , Receptores de Inositol 1,4,5-Trifosfato/genética , Ratones , Ratones Transgénicos , Fosfolipasa C beta/metabolismo , Regiones Promotoras Genéticas , Transducción de Señal , Olfato/fisiología , Canales Catiónicos TRPC/metabolismo , Canal Catiónico TRPC6RESUMEN
In addition to the fundamental role of the extracellular glycoprotein Reelin in neuronal development and adult synaptic plasticity, alterations in Reelin-mediated signaling have been suggested to contribute to neuronal dysfunction associated with Alzheimer's disease (AD). In vitro data revealed a biochemical link between Reelin-mediated signaling, Tau phosphorylation, and amyloid precursor protein (APP) processing. To directly address the role of Reelin in amyloid-beta plaque and Tau pathology in vivo, we crossed heterozygous Reelin knock-out mice (reeler) with transgenic AD mice to investigate the temporal and spatial AD-like neuropathology. We demonstrate that a reduction in Reelin expression results in enhanced amyloidogenic APP processing, as indicated by the precocious production of amyloid-beta peptides, the significant increase in number and size of amyloid-beta plaques, as well as age-related aggravation of plaque pathology in double mutant compared with single AD mutant mice of both sexes. Numerous amyloid-beta plaques accumulate in the hippocampal formation and neocortex of double mutants, precisely in layers with strongest Reelin expression and highest accumulation of Reelin plaques in aged wild-type mice. Moreover, concentric accumulations of phosphorylated Tau-positive neurons around amyloid-beta plaques were evident in 15-month-old double versus single mutant mice. Silver stainings indicated the presence of neurofibrillary tangles, selectively associated with amyloid-beta plaques and dystrophic neurites in the entorhinal cortex and hippocampus. Our findings suggest that age-related Reelin aggregation and concomitant reduction in Reelin-mediated signaling play a proximal role in synaptic dysfunction associated with amyloid-beta deposition, sufficient to enhance Tau phosphorylation and tangle formation in the hippocampal formation in aged Reelin-deficient transgenic AD mice.
Asunto(s)
Enfermedad de Alzheimer , Péptidos beta-Amiloides/metabolismo , Moléculas de Adhesión Celular Neuronal/metabolismo , Proteínas de la Matriz Extracelular/metabolismo , Regulación de la Expresión Génica/genética , Proteínas del Tejido Nervioso/metabolismo , Placa Amiloide/patología , Serina Endopeptidasas/metabolismo , Proteínas tau/metabolismo , Factores de Edad , Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/patología , Precursor de Proteína beta-Amiloide/genética , Animales , Moléculas de Adhesión Celular Neuronal/genética , Modelos Animales de Enfermedad , Proteínas de la Matriz Extracelular/genética , Hipocampo/metabolismo , Hipocampo/patología , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Mutación/genética , Proteínas del Tejido Nervioso/genética , Ovillos Neurofibrilares/patología , Proteína Reelina , Serina Endopeptidasas/genética , alfa-Sinucleína/metabolismoRESUMEN
Cajal-Retzius cells play a crucial role during ontogeny in regulating cortical lamination via release of reelin. In adult brain, they comprise small calretinin-positive interneurons located in the marginal zone of the cerebral cortex and in the hippocampal fissure. Alterations of reelin signaling or expression have been involved in major neurological disorders, and they underlie granule cell dispersion (GCD) in mesial temporal lobe epilepsy (TLE). Here, we investigated in a mouse model of TLE the contribution of Cajal-Retzius cells to reelin production in epileptic hippocampus and the molecular mechanisms underlying GCD. Following unilateral intrahippocampal Kainic acid injection in adult mice to induce an epileptic focus, we observed that Cajal-Retzius cells gradually became strongly immunopositive for reelin, due to intracellular accumulation. This phenotype resembled the morphology of Cajal-Retzius cells in reeler Orleans (reln (orl/orl) ) mice, which express a secretion-deficient 310-kDa reelin fragment. The possibility that GCD might result from abnormal reelin processing in Cajal-Retzius cells, leading to a lack of reelin secretion, was confirmed by KA injection in reln (orl/+) mice, which induced severe GCD. Furthermore, Western blot analysis in KA-treated wildtype mice revealed increased production of â¼300-kDa reelin fragments, confirming abnormal proteolytic processing. This effect was not seen upon treatment with Botulinum neurotoxin E (BoNT/E), which prevents GCD in KA-lesioned hippocampus by chronic blockade of synaptic transmission. Furthermore, BoNT/E blocked upregulation of TrkB in Cajal-Retzius cells, suggesting that production of truncated reelin in KA-treated hippocampus is activity-dependent and regulated by BDNF. Altogether, these data reveal that GCD results from abnormal reelin processing in Cajal-Retzius cells under the control of BDNF. Our findings highlight the critical role played by Cajal-Retzius cells for hippocampal neuronal reorganization in TLE.
Asunto(s)
Factor Neurotrófico Derivado del Encéfalo/metabolismo , Moléculas de Adhesión Celular Neuronal/metabolismo , Epilepsia del Lóbulo Temporal/fisiopatología , Proteínas de la Matriz Extracelular/metabolismo , Hipocampo/patología , Proteínas del Tejido Nervioso/metabolismo , Serina Endopeptidasas/metabolismo , Animales , Toxinas Botulínicas/farmacología , Región CA1 Hipocampal/metabolismo , Región CA1 Hipocampal/patología , Región CA3 Hipocampal/metabolismo , Región CA3 Hipocampal/patología , Calbindina 2 , Recuento de Células , Giro Dentado/metabolismo , Giro Dentado/patología , Modelos Animales de Enfermedad , Epilepsia del Lóbulo Temporal/inducido químicamente , Epilepsia del Lóbulo Temporal/metabolismo , Agonistas de Aminoácidos Excitadores/efectos adversos , Hipocampo/metabolismo , Interneuronas/efectos de los fármacos , Interneuronas/metabolismo , Ácido Kaínico/efectos adversos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes Neurológicos , Ratones Transgénicos , Receptor trkB/metabolismo , Proteína Reelina , Proteína G de Unión al Calcio S100/metabolismo , Transmisión Sináptica/efectos de los fármacosRESUMEN
The brain-specific Ras/Rap-GTPase activating protein (SynGAP) is a prime candidate linking N-methyl-d-aspartate receptors to the regulation of the ERK/MAP kinase signalling cascade, suggested to be essential for experience-dependent synaptic plasticity. Here, we evaluated the behavioural phenotype of SynGAP heterozygous knockout mice (SG(+/-)), expressing roughly half the normal levels of SynGAP. In the cognitive domain, SG(+/-) mice demonstrated severe working and reference memory deficits in the radial arm maze task, a mild impairment early in the transfer test of the water maze task, and a deficiency in spontaneous alternation in an elevated T-maze. In the non-cognitive domain, SG(+/-) mice were hyperactive in the open field and appeared less anxious in the elevated plus maze test. In contrast, object recognition memory performance was not impaired in SG(+/-) mice. The reduction in SynGAP thus resulted in multiple behavioural traits suggestive of aberrant cognitive and non-cognitive processes normally mediated by the hippocampus. Immunohistochemical evaluation further revealed a significant reduction in calbindin-positive interneurons in the hippocampus and doublecortin-positive neurons in the dentate gyrus of adult SG(+/-) mice. Heterozygous constitutive deletion of SynGAP is therefore associated with notable behavioural as well as morphological phenotypes indicative of hippocampal dysfunction. Any suggestion of a possible causal link between them however remains a matter for further investigation.
Asunto(s)
Conducta Animal/fisiología , Hipocampo/fisiología , Aprendizaje por Laberinto/fisiología , Memoria/fisiología , Proteínas Activadoras de ras GTPasa , Animales , Femenino , Hipocampo/citología , Masculino , Ratones , Ratones Noqueados , Neuronas/citología , Neuronas/metabolismo , Pruebas Neuropsicológicas , Proteínas Activadoras de ras GTPasa/genética , Proteínas Activadoras de ras GTPasa/metabolismoRESUMEN
Despite the broad implications of the cannabinoid type 2 receptor (CB2) in neuroinflammatory processes, a suitable CB2-targeted probe is currently lacking in clinical routine. In this work, we synthesized 15 fluorinated pyridine derivatives and tested their binding affinities toward CB2 and CB1. With a sub-nanomolar affinity (Ki for CB2) of 0.8 nM and a remarkable selectivity factor of >12,000 over CB1, RoSMA-18-d6 exhibited outstanding in vitro performance characteristics and was radiofluorinated with an average radiochemical yield of 10.6 ± 3.8% (n = 16) and molar activities ranging from 52 to 65 GBq/µmol (radiochemical purity > 99%). [18F]RoSMA-18-d6 showed exceptional CB2 attributes as demonstrated by in vitro autoradiography, ex vivo biodistribution, and positron emission tomography (PET). Further, [18F]RoSMA-18-d6 was used to detect CB2 upregulation on postmortem human ALS spinal cord tissues. Overall, these results suggest that [18F]RoSMA-18-d6 is a promising CB2 PET radioligand for clinical translation.
Asunto(s)
Piridinas/farmacología , Radiofármacos/farmacología , Receptor Cannabinoide CB2/metabolismo , Animales , Encéfalo/diagnóstico por imagen , Radioisótopos de Flúor/química , Humanos , Ligandos , Masculino , Simulación del Acoplamiento Molecular , Estructura Molecular , Tomografía de Emisión de Positrones , Piridinas/síntesis química , Piridinas/farmacocinética , Radiofármacos/síntesis química , Radiofármacos/farmacocinética , Ratas Wistar , Médula Espinal/diagnóstico por imagen , Bazo/diagnóstico por imagen , Relación Estructura-Actividad , Tritio/químicaRESUMEN
Besides its critical role during neurodevelopment, the extracellular glycoprotein reelin is also a pivotal regulator of adult synaptic function and plasticity, and altered reelin-mediated signalling has been suggested to contribute to neuronal dysfunction associated with Alzheimer's disease. We have recently discovered, in aged rodents and non-human primates, a pronounced decline in reelin-positive interneurons and concomitant accumulation of reelin in extracellular amyloid-like deposits, both being associated with episodic-like memory impairments. Here, we report that these age-related neuropathological changes in hippocampus, entorhinal and piriform cortices of aged wild-type mice are accompanied by abnormal axonal varicosities and altered expression profiles of calcium-binding proteins in plaque-dense areas, as well as a significant reduction in the number of parvalbumin-positive gamma-aminobutyric acid (GABA)ergic projection neurons in basal forebrain areas, including medial septum (MS), ventral and horizontal diagonal Band of Broca (VDB/HDB) and substantia innominata (SI), compared with young subjects. In addition, a significant reduction in the number of choline acetyltransferase-positive cholinergic projection neurons was evident in the HDB/SI area but not in the MS of aged compared with young wild-type mice. No reelin-deposits were found in these basal forebrain regions. Our findings suggest that the elevated reelin plaque load in the projection areas of afferent subcortical GABAergic and cholinergic neurons including hippocampus, entorhinal and piriform cortices affects the axonal integrity and survival of these neurons, potentially contributing to the cognitive impairments observed in aged wild-type mice.
Asunto(s)
Envejecimiento/patología , Moléculas de Adhesión Celular Neuronal/metabolismo , Proteínas de la Matriz Extracelular/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Neuronas/patología , Placa Amiloide/metabolismo , Prosencéfalo/patología , Serina Endopeptidasas/metabolismo , Análisis de Varianza , Animales , Astrocitos/metabolismo , Calbindinas , Recuento de Células , Supervivencia Celular , Colina O-Acetiltransferasa/metabolismo , Femenino , Procesamiento de Imagen Asistido por Computador , Inmunohistoquímica , Masculino , Ratones , Neuronas/metabolismo , Tamaño de los Órganos , Parvalbúminas/metabolismo , Placa Amiloide/patología , Prosencéfalo/metabolismo , Proteína Reelina , Proteína G de Unión al Calcio S100/metabolismo , Ácido gamma-Aminobutírico/metabolismoRESUMEN
The cannabinoid type 2 (CB2) receptor has emerged as a valuable target for therapy and imaging of immune-mediated pathologies. With the aim to find a suitable radiofluorinated analogue of the previously reported CB2 positron emission tomography (PET) radioligand [11C]RSR-056, 38 fluorinated derivatives were synthesized and tested by in vitro binding assays. With a Ki (hCB2) of 6 nM and a selectivity factor of nearly 700 over cannabinoid type 1 receptors, target compound 3 exhibited optimal in vitro properties and was selected for evaluation as a PET radioligand. [18F]3 was obtained in an average radiochemical yield of 11 ± 4% and molar activities between 33 and 114 GBq/µmol. Specific binding of [18F]3 to CB2 was demonstrated by in vitro autoradiography and in vivo PET experiments using the CB2 ligand GW-405â¯833. Metabolite analysis revealed only intact [18F]3 in the rat brain. [18F]3 detected CB2 upregulation in human amyotrophic lateral sclerosis spinal cord tissue and may thus become a candidate for diagnostic use in humans.
Asunto(s)
Encéfalo/metabolismo , Radioisótopos de Flúor/metabolismo , Neuroimagen/métodos , Tomografía de Emisión de Positrones/métodos , Piridinas/química , Radiofármacos/metabolismo , Receptor Cannabinoide CB2/metabolismo , Animales , Encéfalo/diagnóstico por imagen , AMP Cíclico/metabolismo , Radioisótopos de Flúor/química , Hepatocitos/metabolismo , Humanos , Ligandos , Masculino , Ratones , Ratones Endogámicos C57BL , Estructura Molecular , Conformación Proteica , Radioquímica , Radiofármacos/química , Ratas , Ratas Wistar , Receptor Cannabinoide CB2/química , Relación Estructura-ActividadRESUMEN
Prenatal exposure to infections represents a risk factor for the emergence of neuropsychiatric disorders in later life, including schizophrenia and autism. However, it remains essentially unknown whether this association is primarily attributable to prenatal and/or postnatal maternal effects on the offspring. Here, we addressed this issue by dissecting the relative contributions of prenatal inflammatory events and postnatal maternal factors in an animal model of prenatal viral-like infection. Pregnant mice were exposed to the inflammatory agent polyriboinosinic-polyribocytidilic acid (PolyI:C; 5 mg/kg, i.v.) or vehicle treatment on gestation day 9, and offspring born to PolyI:C- and vehicle-treated dams were cross fostered to surrogate rearing mothers that had either experienced inflammatory or sham treatment during pregnancy. We demonstrate that a variety of dopamine- and glutamate-related pharmacological and neuroanatomical disturbances emerge after prenatal immune challenge regardless of whether neonates were raised by vehicle- or PolyI:C-exposed surrogate mothers. However, the adoption of prenatal control animals to immune-challenged surrogate mothers was also sufficient to induce specific pharmacological and neuroanatomical abnormalities in the fostered offspring. Multiple schizophrenia-related dysfunctions emerging after prenatal immune challenge are thus mediated by prenatal but not postnatal maternal effects on the offspring, but immunological stress during pregnancy may affect postpartum maternal factors in such a way that being reared by an immune-challenged surrogate mother can confer risk for distinct forms of psychopathology in adult life.
Asunto(s)
Esquizofrenia/etiología , Amidinas/uso terapéutico , Animales , Modelos Animales de Enfermedad , Conducta Exploratoria , Femenino , Habituación Psicofisiológica , Masculino , Conducta Materna , Ratones , Ratones Endogámicos C57BL , Actividad Motora/efectos de los fármacos , Conducta Paterna , Poli I-C/uso terapéutico , Embarazo , Complicaciones del Embarazo/psicología , Esquizofrenia/inmunología , Psicología del EsquizofrénicoRESUMEN
Maternal infection during pregnancy increases the risk for neurodevelopmental disorders such as schizophrenia and autism in the offspring. This association appears to be critically dependent on the precise prenatal timing. However, the extent to which distinct adult psychopathological and neuropathological traits may be sensitive to the precise times of prenatal immune activation remains to be further characterized. Here, we evaluated in a mouse model of prenatal immune challenge by the viral mimic, polyriboinosinic-polyribocytidilic acid (PolyIC), whether prenatal immune activation in early/middle and late gestation may influence the susceptibility to some of the critical cognitive, pharmacological, and neuroanatomical dysfunctions implicated in schizophrenia and autism. We revealed that PolyIC-induced prenatal immune challenge on gestation day (GD) 9 but not GD17 significantly impaired sensorimotor gating and reduced prefrontal dopamine D1 receptors in adulthood, whereas prenatal immune activation specifically in late gestation impaired working memory, potentiated the locomotor reaction to the NMDA-receptor antagonist dizocilpine, and reduced hippocampal NMDA-receptor subunit 1 expression. On the other hand, potentiation of the locomotor reaction to the dopamine-receptor agonist amphetamine and reduction in Reelin- and Parvalbumin-expressing prefrontal neurons emerged independently of the precise times of prenatal immune challenge. Our findings thus highlight that prenatal immune challenge during early/middle and late fetal development in mice leads to distinct brain and behavioral pathological symptom clusters in adulthood. Further examination and evaluation of in utero immune challenge at different times of gestation may provide important new insight into the neuroimmunological and neuropathological mechanisms underlying the segregation of different symptom clusters in heterogeneous neuropsychiatric disorders such as schizophrenia and autism.