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BACKGROUND AND PURPOSE: Stroke is a major cause of chronic neurological disability. There is considerable interest in understanding how acute transcriptome changes evolve into subacute and chronic patterns that facilitate or limit spontaneous recovery. Here we mapped longitudinal changes in gene expression at multiple time points after stroke in mice out to 6 months. METHODS: Adult C57BL/6 mice were subjected to transient middle cerebral artery occlusion. Longitudinal transcriptome levels were measured at 10 time points after stroke from acute to recovery phases of ischemic stroke. Localization and the number of mononuclear phagocytes were determined in the postischemic brain. Whole-mount brain imaging was performed in asplenic mice receiving GFP+ (green fluorescent protein)-tagged splenocytes. RESULTS: Sustained stroke-induced mRNA abundance changes were observed in both hemispheres with 2989 ipsilateral and 822 contralateral genes significantly perturbed. In the hemisphere ipsilateral to the infarct, genes associated with immune functions were strongly affected, including temporally overlapping innate and adaptive immunity and macrophage M1 and M2 phenotype-related genes. The strong immune gene activation was accompanied by the sustained infiltration of peripheral immune cells at acute, subacute, and recovery stages of stroke. The infiltrated immune cells were found in the infarcted area but also in remote regions at 2 months after stroke. CONCLUSIONS: The study identifies that immune components are the predominant molecular signatures and they may propagate or continuously respond to brain injury in the subacute to chronic phase after central nervous system injury. The study suggests a potential immune-based strategy to modify injury progression and tissue remodeling in ischemic stroke, even months after the initiating event.
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Isquemia Encefálica/diagnóstico por imagen , Isquemia Encefálica/inmunología , Movimiento Celular/fisiología , Inmunidad Celular/fisiología , Recuperación de la Función/fisiología , Transcripción Genética/fisiología , Animales , Isquemia Encefálica/genética , Células Cultivadas , Femenino , Masculino , Ratones , Ratones Endogámicos C57BLRESUMEN
Vascular endothelial growth factor (VEGF) treatment during pilocarpine-induced status epilepticus (SE) causes sustained preservation of behavioral function in rats in the absence of enduring neuroprotection (Nicoletti et al., 2010), suggesting the possibility that other cells or mechanisms could be involved in the beneficial effects of VEGF during SE. Astrocytes have been reported to contribute to epileptiform discharges in the hippocampus (Tian et al., 2005; Kang et al., 1998) and to express VEGF receptors (Krum & Rosenstein, 2002). We report here that VEGF treatment significantly alters multiple astrocyte parameters. This study investigated astrocyte morphology one month after SE in animals treated with VEGF or inactivated VEGF control protein during SE. Individual GFAP-immunostained astrocytes from CA1 and dentate gyrus hilus were traced and morphologically quantified, and both somatic and process structures were analyzed. VEGF treatment during SE significantly prevented post-SE increases in number of branch intersections, process length, and node count. Furthermore, analysis of distance to nearest neighboring astrocytes revealed that VEGF treatment significantly increased inter-astrocyte distance. Overall, VEGF treatment during SE did not significantly alter the shape of the astrocytes, but did prevent SE-induced changes in branching complexity, size, and spatial patterning. Because astrocyte morphology may be related to astrocyte function, it is possible that VEGF's enduring effects on astrocyte morphology may impact the functioning of the post-seizure hippocampus.
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Astrocitos/patología , Convulsiones/tratamiento farmacológico , Convulsiones/patología , Estado Epiléptico/tratamiento farmacológico , Estado Epiléptico/patología , Factor A de Crecimiento Endotelial Vascular/uso terapéutico , Animales , Astrocitos/efectos de los fármacos , Células Cultivadas , Giro Dentado/efectos de los fármacos , Giro Dentado/patología , Hipocampo/efectos de los fármacos , Hipocampo/patología , Masculino , Pilocarpina/efectos adversos , Ratas , Ratas Sprague-Dawley , Resultado del Tratamiento , Factor A de Crecimiento Endotelial Vascular/farmacologíaRESUMEN
Conditional mutagenesis is becoming a method of choice for studying gene function, but constructing conditional alleles is often laborious, limited by target gene structure, and at times, prone to incomplete conditional ablation. To address these issues, we developed a technology termed conditionals by inversion (COIN). Before activation, COINs contain an inverted module (COIN module) that lies inertly within the antisense strand of a resident gene. When inverted into the sense strand by a site-specific recombinase, the COIN module causes termination of the target gene's transcription and simultaneously provides a reporter for tracking this event. COIN modules can be inserted into natural introns (intronic COINs) or directly into coding exons as part of an artificial intron (exonic COINs), greatly simplifying allele design and increasing flexibility over previous conditional KO approaches. Detailed analysis of over 20 COIN alleles establishes the reliability of the method and its broad applicability to any gene, regardless of exon-intron structure. Our extensive testing provides rules that help ensure success of this approach and also explains why other currently available conditional approaches often fail to function optimally. Finally, the ability to split exons using the COIN's artificial intron opens up engineering modalities for the generation of multifunctional alleles.
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Alelos , Silenciador del Gen , Ingeniería Genética/métodos , Mutagénesis Insercional/métodos , Inversión de Secuencia/genética , ADN Nucleotidiltransferasas/metabolismoRESUMEN
The artemin-GFRα3 signaling pathway has been implicated in various painful conditions including migraine, cold allodynia, hyperalgesia, inflammatory bone pain, and mouse knees contain GFRα3-immunoreactive nerve endings. We developed high affinity mouse (REGN1967) and human (REGN5069) GFRα3-blocking monoclonal antibodies and, following in vivo evaluations in mouse models of chronic joint pain (osteoarthritic-like and inflammatory), conducted a first-in-human phase 1 pharmacokinetics (PK) and safety trial of REGN5069 (NCT03645746) in healthy volunteers, and a phase 2 randomized placebo-controlled efficacy and safety trial of REGN5069 (NCT03956550) in patients with knee osteoarthritis (OA) pain. In three commonly used mouse models of chronic joint pain (destabilization of the medial meniscus, intra-articular monoiodoacetate, or Complete Freund's Adjuvant), REGN1967 and REGN5069 attenuated evoked behaviors including tactile allodynia and thermal hyperalgesia without discernably impacting joint pathology or inflammation, prompting us to further evaluate REGN5069 in humans. In the phase 1 study in healthy subjects, the safety profiles of single doses of REGN5069 up to 3000 mg (intravenous) or 600 mg (subcutaneous) were comparable to placebo; PK were consistent with a monoclonal antibody exhibiting target-mediated disposition. In the phase 2 study in patients with OA knee pain, two doses of REGN5069 (100 mg or 1000 mg intravenous every 4 weeks) for 8 weeks failed to achieve the 12-week primary and secondary efficacy endpoints relative to placebo. In addition to possible differences in GFRα3 biology between mice and humans, we highlight here differences in experimental parameters that could have contributed to a different profile of efficacy in mouse models versus human OA pain. Additional research is required to more fully evaluate any potential role of GFRα3 in human pain.
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This study investigated hyperreligiosity in persons with partial epilepsy by exploring a relationship between aura symptoms and spirituality. It was reasoned that patients with high frequencies of auras that are suggestive of metaphysical phenomena, termed numinous-like auras, would report increased spirituality of an unconventional form, both during their seizures and generally. Numinous-like auras included: dreaminess/feeling of detachment, autoscopy, derealization, depersonalization, time speed alterations, bodily distortions, and pleasure. A high-frequency aura group, low-frequency aura group, and nonseizure reference group were compared on the Expressions of Spirituality-Revised. The High group had significantly greater Experiential/Phenomenological Dimension and Paranormal Beliefs factor scores than the Low group, and significantly greater Experiential/Phenomenological Dimension factor scores than the reference group. There were no differences between the Low group and the reference group. In addition, there were no differences among the three groups on traditional measures of religiosity. The results provide preliminary evidence that epilepsy patients with frequent numinous-like auras have greater ictal and interictal spirituality of an experiential, personalized, and atypical form, which may be distinct from traditional, culturally based religiosity. This form of spirituality may be better described by the term cosmic spirituality than hyperreligiosity. It is speculated that this spirituality is due to an overactivation and subsequent potentiation of the limbic system, with frequent numinous-like auras indicating sufficient activation for this process to occur. It is likely that numinous-like experiences foster cosmic spirituality in a number of circumstances, including seizures, psychosis, near-death experiences, psychedelic drug use, high-elevation exposure, and also normal conditions.
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Trastornos Disociativos/psicología , Epilepsias Parciales/psicología , Religión y Medicina , Religión y Psicología , Espiritualidad , Adulto , Femenino , Humanos , Masculino , Persona de Mediana Edad , Inventario de Personalidad/estadística & datos numéricos , Psicometría , Adulto JovenRESUMEN
Vascular endothelial growth factor (VEGF) is a vascular growth factor more recently recognized as a neurotrophic factor (for review, see Storkebaum E, Lambrechts D, Carmeliet P. BioEssays 2004;26:943-54). We previously reported that endogenous VEGF protein is dramatically upregulated after pilocarpine-induced status epilepticus in the rat, and that intra-hippocampal infusions of recombinant human VEGF significantly protected against the loss of hippocampal CA1 neurons in this model (Nicoletti JN, Shah SK, McCloskey DP, et al. Neuroscience 2008;151:232-41). We hypothesized that we would see a preservation of cognitive and emotional functioning with VEGF treatment accompanying the neuroprotection previously observed in this paradigm. Using the Morris water maze to evaluate learning and memory, and the light-dark task to assess anxiety, we found a selective profile of preservation. Specifically, VEGF completely preserved normal anxiety functioning and partially but significantly protected learning and memory after status epilepticus. To determine whether the ability of VEGF to attenuate behavioral deficits was accompanied by sustained preservation of hippocampal neurons, we stereologically estimated CA1 pyramidal neuron densities 4 weeks after status epilepticus. At this time point, we found no significant difference in neuronal densities between VEGF- and control-treated status epilepticus animals, suggesting that VEGF could have protected hippocampal functioning independent of its neuroprotective effect.
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Síntomas Conductuales/tratamiento farmacológico , Síntomas Conductuales/etiología , Estado Epiléptico/complicaciones , Factor A de Crecimiento Endotelial Vascular/uso terapéutico , Adaptación Ocular/efectos de los fármacos , Análisis de Varianza , Animales , Síntomas Conductuales/patología , Modelos Animales de Enfermedad , Hipocampo/patología , Humanos , Locomoción/efectos de los fármacos , Masculino , Aprendizaje por Laberinto/efectos de los fármacos , Pilocarpina , Células Piramidales/efectos de los fármacos , Células Piramidales/metabolismo , Ratas , Ratas Sprague-Dawley , Proteínas Recombinantes/uso terapéutico , Estado Epiléptico/inducido químicamenteRESUMEN
Administration of dextran sodium sulfate (DSS) to rodents at varying concentrations and exposure times is commonly used to model human inflammatory bowel disease (IBD). Currently, the criteria used to assess IBD-like pathology seldom include surrogate measures of visceral pain. Thus, we sought to standardize the model and then identify surrogate measures to assess effects on visceral pain. We used various 4% DSS protocols and evaluated effects on weight loss, colon pathology, biochemistry, RNA signature, and open field behavior. We then tested the therapeutic potential of NPY Y1 and/or Y2 receptor inhibition for the treatment of IBD pathology using this expanded panel of outcome measures. DSS caused weight loss and colon shrinkage, increased colon NPY and inflammatory cytokine expression, altered behaviors in the open field and induced a distinct gene metasignature that significantly overlapped with that of human IBD patients. Inhibition of Y1 and/or Y2 receptors failed to improve gross colon pathology. Y1 antagonism significantly attenuated colon inflammatory cytokine expression without altering pain-associated behaviors while Y2 antagonism significantly inhibited pain-associated behaviors in spite of a limited effect on inflammatory markers. A protocol using 7 days of 4% DSS most closely modeled human IBD pathology. In this model, rearing behavior potentially represents a tool for evaluating visceral pain/discomfort that may be pharmacologically dissociable from other features of pathology. The finding that two different NPY receptor antagonists exhibited different efficacy profiles highlights the benefit of including a variety of outcome measures in IBD efficacy studies to most fully evaluate the therapeutic potential of experimental treatments.
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Colitis/tratamiento farmacológico , Sulfato de Dextran/toxicidad , Modelos Animales de Enfermedad , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Receptores de Neuropéptido Y/antagonistas & inhibidores , Animales , Arginina/análogos & derivados , Arginina/farmacología , Benzazepinas/farmacología , Peso Corporal , Colitis/inducido químicamente , Colitis/metabolismo , Colitis/patología , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos C57BLRESUMEN
One challenge of neuroscience educators is to make accessible to students as many aspects of brain structure and function as possible. The anatomy and function of the cerebrovasculature is among many topics of neuroscience that are underrepresented in undergraduate neuroscience education. Recognizing this deficit, we evaluated methods to produce archival tissue specimens of the cerebrovasculature and the "neurovascular unit" for instruction and demonstration in the teaching lab. An additional goal of this project was to identify the costs of each method as well as to determine which method(s) could be adapted into lab exercises, where students participate in the tissue preparation, staining, etc. In the present report, we detail several methods for demonstrating the cerebrovasculature and suggest that including this material can be a valuable addition to more traditional anatomy/physiology demonstrations and exercises.
RESUMEN
In addition to its potent effects on vasculature, it has become clear that vascular endothelial growth factor (VEGF) has effects on both neurons and glia, and recent studies suggest that it can be neuroprotective. To determine potential mechanisms underlying this neuroprotection, recombinant human VEGF was bath applied to adult rat hippocampal slices, and both extracellular and intracellular recordings were used to examine intrinsic properties and synaptic responses of hippocampal principal neurons. Initial studies in area CA1 showed that VEGF significantly reduced the amplitude of responses elicited by Schaffer collateral stimulation, without influencing membrane properties. Similar effects occurred in CA3 pyramidal cells and dentate gyrus granule cells when their major glutamatergic afferents were stimulated. Because VEGF expression is increased after seizures, effects of VEGF were also examined in rats with recurrent spontaneous seizures. VEGF reduced spontaneous discharges in slices from these rats but had surprisingly little effect on epileptiform discharges produced by disinhibition of slices from control rats. These results demonstrate a previously unknown effect of VEGF on neuronal activity and also demonstrate a remarkable potency in the epileptic brain. Based on this, we suggest that VEGF or VEGF-related targets could provide useful endpoints to direct novel therapeutic strategies for epilepsy.
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Hipocampo/fisiopatología , Fármacos Neuroprotectores/metabolismo , Convulsiones/fisiopatología , Transmisión Sináptica , Factor A de Crecimiento Endotelial Vascular/metabolismo , Potenciales de Acción , Animales , Enfermedad Crónica , Convulsivantes , Relación Dosis-Respuesta a Droga , Potenciales Evocados , Técnicas In Vitro , Masculino , Inhibición Neural , Neuronas , Fármacos Neuroprotectores/administración & dosificación , Fármacos Neuroprotectores/farmacología , Células Piramidales/efectos de los fármacos , Células Piramidales/fisiología , Ratas , Ratas Sprague-Dawley , Convulsiones/inducido químicamente , Sinapsis , Transmisión Sináptica/efectos de los fármacos , Factores de Tiempo , Factor A de Crecimiento Endotelial Vascular/administración & dosificación , Factor A de Crecimiento Endotelial Vascular/farmacologíaRESUMEN
Nerve growth factor (NGF) plays a role in sympathetic neuron integrity and survival. Brain-derived neurotrophic factor (BDNF) also has trophic effects on sympathetic neurons. We report here the serendipitous finding that co-treatment of hippocampus with BDNF and the NGF antagonist TrkA-Fc leads to perivascular inflammation and marked vasoconstriction. This effect is not observed with either reagent alone or in combination with other control proteins. Because NGF supports sympathetic neuron health, we tested the hypothesis that BDNF combined with sympathetic compromise caused this effect. Superior cervical ganglia were removed bilaterally with concurrent BDNF infusion into hippocampus. Perivascular inflammation was observed at 3 days, but not 12 days post treatment, when sympathetic terminals had receded, suggesting that the presence of these terminals was necessary for inflammation. Since sympathetic dysfunction may lead to compensatory overactivity of norepinephrine (NE) signaling, we co-infused BDNF with NE in the hippocampus and observed perivascular inflammation. In humans, sympathetic overactivity has been reported in a variety of vascular diseases. Some of these diseases, e.g. primary Raynaud's, are not accompanied by serious inflammatory disease whereas others, such as scleroderma and systemic lupus, are. We speculate that BDNF may contribute to the transformation of sympathetic dysfunction to inflammatory disease.
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Enfermedades del Sistema Nervioso Autónomo/inmunología , Enfermedades del Sistema Nervioso Autónomo/metabolismo , Edema Encefálico/inmunología , Edema Encefálico/metabolismo , Factor Neurotrófico Derivado del Encéfalo/fisiología , Animales , Enfermedades del Sistema Nervioso Autónomo/patología , Enfermedades del Sistema Nervioso Autónomo/fisiopatología , Edema Encefálico/patología , Factor Neurotrófico Derivado del Encéfalo/administración & dosificación , Hipocampo/metabolismo , Inflamación/metabolismo , Inflamación/fisiopatología , Bombas de Infusión , Neuronas/inmunología , Neuronas/metabolismo , Neuronas/patología , Ratas , Ratas Sprague-DawleyRESUMEN
Acute intrahippocampal infusion of brain-derived neurotrophic factor (BDNF) leads to long-term potentiation (BDNF-LTP) of synaptic transmission at medial perforant path-->granule cell synapses in the rat dentate gyrus. Endogenous BDNF is implicated in the maintenance of high-frequency stimulation-induced LTP (HFS-LTP). However, the relationship between exogenous BDNF-LTP and HFS-LTP is unclear. First, we found that BDNF-LTP, like HFS-LTP, is associated with enhancement in both synaptic strength and granule cell excitability (EPSP-spike coupling). Second, treatment with a competitive NMDA receptor (NMDAR) antagonist blocked HFS-LTP but had no effect on the development or magnitude of BDNF-LTP. Thus, NMDAR activation is not required for the induction or expression of BDNF-LTP. Formation of stable, late phase HFS-LTP requires mRNA synthesis and is coupled to upregulation of the immediate early gene activity-regulated cytoskeleton-associated protein (Arc). Local infusion of the transcription inhibitor actinomycin D (ACD) 1 hr before or immediately before BDNF infusion inhibited BDNF-LTP and upregulation of Arc protein expression. ACD applied 2 hr after BDNF infusion had no effect, defining a critical time window of transcription-dependent synaptic strengthening. Finally, the functional role of BDNF-LTP was assessed in occlusion experiments with HFS-LTP. HFS-LTP was induced, and BDNF was infused at time points corresponding to early phase (1 hr) or late phase (4 hr) HFS-LTP. BDNF applied during the early phase led to normal BDNF-LTP. In contrast, BDNF-LTP was completely occluded during the late phase. The results strongly support a role for BDNF in triggering transcription-dependent, late phase LTP in the intact adult brain.
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Factor Neurotrófico Derivado del Encéfalo/farmacología , Hipocampo/efectos de los fármacos , Potenciación a Largo Plazo/efectos de los fármacos , Proteínas del Tejido Nervioso , Transcripción Genética/efectos de los fármacos , Animales , Factor Neurotrófico Derivado del Encéfalo/farmacocinética , Proteínas del Citoesqueleto , Vías de Administración de Medicamentos , Estimulación Eléctrica/métodos , Antagonistas de Aminoácidos Excitadores/farmacología , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Potenciales Postsinápticos Excitadores/fisiología , Hipocampo/fisiología , Proteínas Inmediatas-Precoces/metabolismo , Potenciación a Largo Plazo/fisiología , Plasticidad Neuronal/efectos de los fármacos , Plasticidad Neuronal/fisiología , Inhibidores de la Síntesis del Ácido Nucleico/farmacología , Ratas , Ratas Sprague-Dawley , Receptores de N-Metil-D-Aspartato/antagonistas & inhibidores , Transmisión Sináptica/efectos de los fármacos , Transmisión Sináptica/fisiología , Distribución Tisular , Transcripción Genética/fisiología , Regulación hacia Arriba/efectos de los fármacosRESUMEN
We have previously demonstrated that brain-derived neurotrophic factor (BDNF) induces persistent neuropeptide Y (NPY) production in cortical cultures in an ERK1/2-dependent manner. In some studies, it was shown that BDNF leads to the downregulation of TrkB receptor and some of its downstream responses, whereas in others it does not. We examined whether the BDNF requirement for induction of persistent NPY production correlates with that for induction of phosphorylation of TrkB and ERK1/2. Continuous 24-h exposure to BDNF led to a 2- to 3-fold increase in NPY production (maximal level). While 1 h of BDNF exposure induced NPY production at a half maximal level, 8 h was required for induction of a maximal level. BDNF-induced NPY production was completely inhibited by co-exposure to TrkB-Fc fusion protein (TrkB extracellular domain fused to Fc) and partially inhibited by TrkB-Fc added 1 h after BDNF; TrkC-Fc did not do so. Activation of TrkB receptor was analyzed at two potential tyrosine phosphorylated sites, the activation loop and the Shc binding. BDNF led to coordinated phosphorylation of the two sites that persisted for 6-8 h, and this was not associated with changes in the content of TrkB protein. The presence of BDNF throughout the 6- to 8-h period was required for the persistent phosphorylation of TrkB and ERK1/2. Thus, continuous BDNF activation of TrkB is required for persistent activation of the ERK1/2 pathway and induction of NPY production. We propose that, within the time frame analyzed in this study, BDNF does not lead to the downregulation of TrkB receptor or of the biological responses leading to NPY production.
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Factor Neurotrófico Derivado del Encéfalo/metabolismo , Corteza Cerebral/metabolismo , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Neuropéptido Y/biosíntesis , Receptor trkB/metabolismo , Animales , Factor Neurotrófico Derivado del Encéfalo/administración & dosificación , Células Cultivadas , Corteza Cerebral/citología , Esquema de Medicación , Activación Enzimática , Femenino , Feto , Cinética , Sistema de Señalización de MAP Quinasas/fisiología , Fosforilación , Embarazo , Ratas , Ratas Sprague-Dawley , Factores de TiempoRESUMEN
Vascular endothelial growth factor (VEGF) is a vascular growth factor which induces angiogenesis (the development of new blood vessels), vascular permeability, and inflammation. In brain, receptors for VEGF have been localized to vascular endothelium, neurons, and glia. VEGF is upregulated after hypoxic injury to the brain, which can occur during cerebral ischemia or high-altitude edema, and has been implicated in the blood-brain barrier breakdown associated with these conditions. Given its recently-described role as an inflammatory mediator, VEGF could also contribute to the inflammatory responses observed in cerebral ischemia. After seizures, blood-brain barrier breakdown and inflammation is also observed in brain, albeit on a lower scale than that observed after stroke. Recent evidence has suggested a role for inflammation in seizure disorders. We have described striking increases in VEGF protein in both neurons and glia after pilocarpine-induced status epilepticus in the brain. Increases in VEGF could contribute to the blood-brain barrier breakdown and inflammation observed after seizures. However, VEGF has also been shown to be neuroprotective across several experimental paradigms, and hence could potentially protect vulnerable cells from damage associated with seizures. Therefore, the role of VEGF after seizures could be either protective or destructive. Although only further research will determine the exact nature of VEGF's role after seizures, preliminary data indicate that VEGF plays a protective role after seizures.
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Barrera Hematoencefálica/fisiología , Circulación Cerebrovascular/fisiología , Epilepsia/fisiopatología , Factor A de Crecimiento Endotelial Vascular/fisiología , Animales , HumanosRESUMEN
Pleiotrophin (PTN) is an extracellular matrix-associated protein with neurotrophic and neuroprotective effects that is involved in a variety of neurodevelopmental processes. Data regarding the cognitive-behavioral and neuroanatomical phenotype of pleiotrophin knockout (KO) mice is limited. The purpose of this study was to more fully characterize this phenotype, with emphasis on the domains of learning and memory, cognitive-behavioral flexibility, exploratory behavior and anxiety, social behavior, and the neuronal and vascular microstructure of the lateral entorhinal cortex (EC). PTN KOs exhibited cognitive rigidity, heightened anxiety, behavioral reticence in novel contexts and novel social interactions suggestive of neophobia, and lamina-specific decreases in neuronal area and increases in neuronal density in the lateral EC. Initial learning of spatial and other associative tasks, as well as vascular density in the lateral EC, was normal in the KOs. These data suggest that the absence of PTN in vivo is associated with disruption of specific cognitive and affective processes, raising the possibility that further study of PTN KOs might have implications for the study of human disorders with similar features.
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Conducta Animal , Proteínas Portadoras/genética , Citocinas/deficiencia , Citocinas/genética , Neuroanatomía , Animales , Ansiedad/metabolismo , Conducta Animal/fisiología , Proteínas Portadoras/metabolismo , Cognición , Citocinas/metabolismo , Corteza Entorrinal/irrigación sanguínea , Corteza Entorrinal/citología , Corteza Entorrinal/fisiología , Conducta Exploratoria , Aprendizaje por Laberinto , Memoria , Ratones , Ratones Noqueados , Neuronas/citología , Fenotipo , Conducta SocialRESUMEN
BACKGROUND: Infusion of exogenous vascular endothelial growth factor (VEGF) into adult brain at doses above 60 ng/day induces dramatic angiogenesis accompanied by vascular leak and inflammation. Blood vessels formed by this treatment are dilated and tortuous, exhibiting a pathological morphology. Pathological VEGF-induced angiogenesis is preceded by vascular leak and inflammation, which have been proposed to mediate subsequent angiogenesis. METHODS: To test this hypothesis, we infused VEGF into the brains of adult rats to induce pathological angiogenesis. Some of these rats were treated with dexamethasone, a potent anti-inflammatory glucocorticoid, to inhibit inflammation and edema. RESULTS: We demonstrate that inhibition of inflammation by treatment with dexamethasone significantly attenuated VEGF-induced pathological angiogenesis. To present converging evidence that inflammation may be important in this angiogenic process, we also demonstrate that mice genetically deficient in the inflammatory mediator intercellular adhesion molecule-1 have attenuated VEGF-induced angiogenesis. These same mice showed normal amounts of physiological angiogenesis in response to enriched environments, however, suggesting that a generalized reduction in vascular plasticity could not account for their poor angiogenic response to VEGF. CONCLUSIONS: Taken together, the data from these experiments suggest that the inflammation which occurs before or during VEGF-induced pathological brain angiogenesis plays a contributory role in the pathological angiogenic process.
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Circulación Cerebrovascular/efectos de los fármacos , Dexametasona/farmacología , Glucocorticoides/farmacología , Molécula 1 de Adhesión Intercelular/genética , Molécula 1 de Adhesión Intercelular/inmunología , Neovascularización Patológica/tratamiento farmacológico , Factores de Edad , Animales , Encéfalo/irrigación sanguínea , Encéfalo/inmunología , Células Cultivadas , Células Endoteliales/citología , Células Endoteliales/efectos de los fármacos , Humanos , Molécula 1 de Adhesión Intercelular/metabolismo , Masculino , Ratones , Ratones Noqueados , Neovascularización Patológica/inducido químicamente , Neovascularización Patológica/inmunología , Ratas , Ratas Sprague-Dawley , Venas Umbilicales/citología , Factor A de Crecimiento Endotelial Vascular/farmacologíaRESUMEN
Neuromedin U (NMU) has recently been reported to have a role in nociception and inflammation. To clarify the function of the two known NMU receptors, NMU receptor 1 (NMUR1) and NMU receptor 2 (NMUR2), during nociception and inflammation in vivo, we generated mice in which the genes for each receptor were independently deleted. Compared to wild type littermates, mice deficient in NMUR2 showed a reduced thermal nociceptive response in the hot plate, but not in the tail flick, test. In addition, the NMUR2 mutant mice showed a reduced behavioral response and a marked reduction in thermal hyperalgesia following capsaicin injection. NMUR2-deficient mice also showed an impaired pain response during the chronic, but not acute, phase of the formalin test. In contrast, NMUR1-deficient mice did not show any nociceptive differences compared to their wild type littermates in any of the behavioral tests used. We observed the same magnitude of inflammation in both lines of NMU receptor mutant mice compared to their wild type littermates after injection with complete Freund's adjuvant (CFA), suggesting no requirement for either receptor in this response. Thus, the pro-nociceptive effects of NMU in mice appear to be mediated through NMUR2, not NMUR1.
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Proteínas de la Membrana/genética , Proteínas de la Membrana/fisiología , Nociceptores/fisiología , Umbral del Dolor/fisiología , Receptores de Neurotransmisores/genética , Receptores de Neurotransmisores/fisiología , Animales , Conducta Animal , Capsaicina , Femenino , Adyuvante de Freund , Expresión Génica , Calor , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Dimensión del DolorRESUMEN
The results of several studies have contributed to the hypothesis that BDNF promotes seizure activity, particularly in adult hippocampus. To test this hypothesis, BDNF, vehicle (phosphate-buffered saline, PBS), or albumin was infused directly into the hippocampus for 2 weeks using osmotic minipumps. Rats were examined behaviorally, electrophysiologically, and anatomically. An additional group was tested for sensitivity to the convulsant pilocarpine. Spontaneous behavioral seizures were observed in BDNF-infused rats (8/32; 25%) but not in controls (0/20; 0%). In a subset of six animals (three BDNF, three albumin), blind electrophysiological analysis of scalp recordings contralateral to the infused hippocampus demonstrated abnormalities in all BDNF rats; but not controls. Neuronal loss in BDNF-treated rats was not detected relative to PBS- or albumin-treated animals, but immunocytochemical markers showed a pattern of expression in BDNF-treated rats that was similar to rats with experimentally induced seizures. Thus, BDNF-infused rats had increased expression of NPY in hilar neurons of the dentate gyrus relative to control rats. NPY and BDNF expression was increased in the mossy fiber axons of dentate gyrus granule cells relative to controls. The increase in NPY and BDNF expression in BDNF-treated rats was bilateral and occurred throughout the septotemporal axis of the hippocampus. Mossy fiber sprouting occurred in five BDNF-treated rats but no controls. In another group of infused rats that was tested for seizure sensitivity to the convulsant pilocarpine, BDNF-infused rats had a shorter latency to status epilepticus than PBS-infused rats. In addition, the progression from normal behavior to severe seizures was faster in BDNF-treated rats. These data support the hypothesis that intrahippocampal BDNF infusion can facilitate, and potentially initiate, seizure activity in adult hippocampus.
Asunto(s)
Factor Neurotrófico Derivado del Encéfalo/administración & dosificación , Hipocampo/efectos de los fármacos , Sistema Límbico/fisiopatología , Convulsiones/inducido químicamente , Convulsiones/fisiopatología , Animales , Conducta Animal/efectos de los fármacos , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Giro Dentado/metabolismo , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Relación Dosis-Respuesta a Droga , Vías de Administración de Medicamentos , Electroencefalografía/efectos de los fármacos , Hipocampo/metabolismo , Infusiones Parenterales , Masculino , Fibras Musgosas del Hipocampo/metabolismo , Neuropéptido Y/metabolismo , Pilocarpina , Ratas , Estado Epiléptico/inducido químicamente , Estado Epiléptico/fisiopatologíaRESUMEN
Vascular endothelial growth factor (VEGF) has been shown to induce angiogenesis when infused continuously into adult rat brain tissue. In addition, VEGF has been shown to enhance permeability in brain vasculature. Adult rats were continuously infused with mouse VEGF into neocortex for up to 7 days. We studied the development of VEGF-induced vasculature in rat neocortex and evaluated the temporal expression of a wide variety of markers for inflammation and vascular leak in relation to the angiogenic response using immunohistochemistry and Western blot analysis. We report here that VEGF-mediated inflammation in brain is characterized by upregulation of ICAM-1 and the chemokine MIP-1alpha, as well as a preferential extravasation of monocytes. VEGF causes a dramatic breakdown of the blood-brain barrier, which is characterized by decreased investment of the vasculature with astroglial endfeet. Perivascular cells, in contrast, increase around the newly formed cerebrovasculature. In addition, breakdown of the blood-brain barrier, leukocyte extravasation, and extracellular matrix deposition occur before vascular proliferation. Furthermore, administration of low doses of VEGF induces permeability and inflammation without appreciable vascular proliferation.
Asunto(s)
Encéfalo/irrigación sanguínea , Encéfalo/efectos de los fármacos , Encefalitis/inducido químicamente , Mediadores de Inflamación/farmacología , Neovascularización Fisiológica/efectos de los fármacos , Factor A de Crecimiento Endotelial Vascular/farmacología , Animales , Astrocitos/efectos de los fármacos , Astrocitos/patología , Barrera Hematoencefálica/efectos de los fármacos , Western Blotting , Encéfalo/patología , Permeabilidad Capilar/efectos de los fármacos , División Celular/efectos de los fármacos , Quimiocinas/genética , Relación Dosis-Respuesta a Droga , Vías de Administración de Medicamentos , Encefalitis/metabolismo , Encefalitis/patología , Hibridación in Situ , Mediadores de Inflamación/administración & dosificación , Molécula 1 de Adhesión Intercelular/metabolismo , Antígeno Ki-67/metabolismo , Masculino , Neocórtex/irrigación sanguínea , Neocórtex/efectos de los fármacos , Neocórtex/patología , ARN Mensajero/metabolismo , Ratas , Ratas Sprague-Dawley , Factor A de Crecimiento Endotelial Vascular/administración & dosificaciónRESUMEN
Vascular endothelial growth factor (VEGF) plays a critical role during normal embryonic angiogenesis and also in the pathological angiogenesis that occurs in a number of diseases, including cancer. Initial attempts to block VEGF by using a humanized monoclonal antibody are beginning to show promise in human cancer patients, underscoring the importance of optimizing VEGF blockade. Previous studies have found that one of the most effective ways to block the VEGF-signaling pathway is to prevent VEGF from binding to its normal receptors by administering decoy-soluble receptors. The highest-affinity VEGF blocker described to date is a soluble decoy receptor created by fusing the first three Ig domains of VEGF receptor 1 to an Ig constant region; however, this fusion protein has very poor in vivo pharmacokinetic properties. By determining the requirements to maintain high affinity while extending in vivo half life, we were able to engineer a very potent high-affinity VEGF blocker that has markedly enhanced pharmacokinetic properties. This VEGF-Trap effectively suppresses tumor growth and vascularization in vivo, resulting in stunted and almost completely avascular tumors. VEGF-Trap-mediated blockade may be superior to that achieved by other agents, such as monoclonal antibodies targeted against the VEGF receptor.