RESUMO
Social recognition, the ability to recognize individuals that were previously encountered, requires complex integration of sensory inputs with previous experience. Here, we use a variety of approaches to discern how oxytocin-sensitive neurons in the PFC exert descending control over a circuit mediating social recognition in mice. Using male mice with Cre-recombinase directed to the oxytocin receptor gene (Oxtr), we revealed that oxytocin receptors (OXTRs) are expressed on glutamatergic neurons in the PFC, optogenetic stimulation of which elicited activation of neurons residing in several mesolimbic brain structures. Optogenetic stimulation of axons in the BLA arising from OXTR-expressing neurons in the PFC eliminated the ability to distinguish novel from familiar conspecifics, but remarkably, distinguishing between novel and familiar objects was unaffected. These results suggest that an oxytocin-sensitive PFC to BLA circuit is required for social recognition. The implication is that impaired social memory may manifest from dysregulation of this circuit.SIGNIFICANCE STATEMENT Using mice, we demonstrate that optogenetic activation of the neurons in the PFC that express the oxytocin receptor gene (Oxtr) impairs the ability to distinguish between novel and familiar conspecifics, but the ability to distinguish between novel and familiar objects remains intact. Subjects with autism spectrum disorders (ASDs) have difficulty identifying a person based on remembering facial features; however, ASDs and typical subjects perform similarly when remembering objects. In subjects with ASD, viewing the same face increases neural activity in the PFC, which may be analogous to the optogenetic excitation of oxytocin receptor (OXTR) expressing neurons in the PFC that impairs social recognition in mice. The implication is that overactivation of OXTR-expressing neurons in the PFC may contribute to ASD symptomology.
Assuntos
Ácido Glutâmico/metabolismo , Neurônios/metabolismo , Córtex Pré-Frontal/metabolismo , Receptores de Ocitocina/metabolismo , Reconhecimento Psicológico/fisiologia , Comportamento Social , Animais , Masculino , Camundongos , Camundongos Transgênicos , Optogenética , Receptores de Ocitocina/genéticaRESUMO
This study tested the hypothesis that peripheral immune challenges will produce predictable activation patterns in the rat brain consistent with sympathetic excitation. As part of examining this hypothesis, this study asked whether central activation is dependent on capsaicin-sensitive C-fibres. We induced skin contact sensitivity immune responses with 2,4-dinitrochlorobenzene (DNCB), in the presence or absence of the acute C-fibre toxin capsaicin (8-methyl-N-vanillyl-6-nonenamide) to trigger immune responses with and without diminished activity of C-fibres. Innovative blood-oxygen-level-dependent functional magnetic resonance imaging data revealed that the skin contact sensitivity immune responses induced with DNCB were associated with localized increases in brain neuronal activity in treated rats. This response was diminished by pre-treatment with capsaicin 1 week before scans. In the same animals, we found expression of the immediate early gene c-Fos in sub-regions of the amygdala and hypothalamic sympathetic brain nuclei. Significant increases in c-Fos expression were found in the supraoptic nucleus, central amygdala and medial habenula following immune challenges. Our results support the idea that selective brain regions, some of which are associated with sympathetic function, process or modulate immune function through pathways that are partially dependent on C-fibres. Together with previous studies demonstrating the motor control pathways from brain to immune targets, these findings indicate a central neuroimmune system to monitor host status and coordinate appropriate host responses.
Assuntos
Dermatite de Contato/imunologia , Hipotálamo/imunologia , Fibras Nervosas Amielínicas/imunologia , Pele/inervação , Animais , Biomarcadores/metabolismo , Mapeamento Encefálico/métodos , Capsaicina/farmacologia , Dermatite de Contato/metabolismo , Dermatite de Contato/fisiopatologia , Dinitroclorobenzeno , Modelos Animais de Doenças , Hipotálamo/efeitos dos fármacos , Hipotálamo/metabolismo , Hipotálamo/fisiopatologia , Imageamento por Ressonância Magnética , Masculino , Fibras Nervosas Amielínicas/efeitos dos fármacos , Fibras Nervosas Amielínicas/metabolismo , Vias Neurais/imunologia , Vias Neurais/metabolismo , Proteínas Proto-Oncogênicas c-fos/metabolismo , Ratos Sprague-Dawley , Sistema Nervoso Simpático/imunologia , Sistema Nervoso Simpático/fisiopatologia , Fatores de Tempo , Regulação para CimaRESUMO
Neuroinflammation and neurodegeneration have been observed in the brain in type 1 diabetes (T1D). However, little is known about the mediators of these effects. In T1D mice with 12- and 35-wk duration of diabetes we examined two mechanisms of neurodegeneration, loss of the neuroprotective factors insulin-like growth factor I (IGF-I) and IGF-binding protein-3 (IGFBP-3) and changes in indoleamine 2,3-dioxygenase (IDO) expression in the brain, and compared the response to age-matched controls. Furthermore, levels of matrix metalloproteinase-2 (MMP-2), nucleoside triphosphate diphosphohydrolase-1 (CD39), and ionized calcium-binding adaptor molecule 1 (Iba-1) were utilized to assess inflammatory changes in astrocytes, microglia, and blood vessels. In the diabetic hypothalamus (HYPO), we observed 20% reduction in neuronal soma diameter (P<0.05) and reduced neuronal expression of IGFBP-3 (-32%, P<0.05) and IGF-I (-15%, P<0.05) compared with controls at 35 wk. In diabetic HYPO, MMP-2 expression was increased in astrocytes (46%, P<0.01), and IDO⺠cell density rose by (62%, P<0.05). CD39 expression dropped by 30% (P<0.05) in microglia and blood vessels. With 10 wk of systemic treatment using minocycline, an anti-inflammatory agent that crosses the blood-brain barrier, MMP-2, IDO, and CD39 levels normalized (P<0.05). Our results suggest that increased IDO and early loss of CD39⺠protective cells lead to activation of inflammation in sympathetic centers of the CNS. As a downstream effect, the loss of the neuronal survival factors IGFBP-3 and IGF-I and the neurotoxic products of the kynurenine pathway contribute to the loss of neuronal density observed in the HYPO in T1D.
Assuntos
Diabetes Mellitus Tipo 1/complicações , Neuropatias Diabéticas/metabolismo , Regulação para Baixo , Encefalite/complicações , Hipotálamo/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Sistema Nervoso Simpático/metabolismo , Animais , Anti-Inflamatórios não Esteroides/uso terapêutico , Astrócitos/efeitos dos fármacos , Astrócitos/imunologia , Astrócitos/metabolismo , Astrócitos/patologia , Diabetes Mellitus Tipo 1/tratamento farmacológico , Diabetes Mellitus Tipo 1/fisiopatologia , Neuropatias Diabéticas/imunologia , Neuropatias Diabéticas/patologia , Neuropatias Diabéticas/prevenção & controle , Progressão da Doença , Regulação para Baixo/efeitos dos fármacos , Encefalite/imunologia , Encefalite/metabolismo , Encefalite/prevenção & controle , Hipotálamo/efeitos dos fármacos , Hipotálamo/imunologia , Hipotálamo/patologia , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Mediadores da Inflamação/antagonistas & inibidores , Mediadores da Inflamação/metabolismo , Proteína 3 de Ligação a Fator de Crescimento Semelhante à Insulina/metabolismo , Fator de Crescimento Insulin-Like I/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/metabolismo , Macrófagos/patologia , Masculino , Camundongos Endogâmicos C57BL , Microglia/efeitos dos fármacos , Microglia/imunologia , Microglia/metabolismo , Microglia/patologia , Minociclina/uso terapêutico , Proteínas do Tecido Nervoso/antagonistas & inibidores , Neurônios/efeitos dos fármacos , Neurônios/imunologia , Neurônios/metabolismo , Neurônios/patologia , Sistema Nervoso Simpático/efeitos dos fármacos , Sistema Nervoso Simpático/imunologia , Sistema Nervoso Simpático/patologia , Regulação para Cima/efeitos dos fármacosRESUMO
By using pseudorabies virus expressing green fluorescence protein, we found that efferent bone marrow-neural connections trace to sympathetic centers of the central nervous system in normal mice. However, this was markedly reduced in type 1 diabetes, suggesting a significant loss of bone marrow innervation. This loss of innervation was associated with a change in hematopoiesis toward generation of more monocytes and an altered diurnal release of monocytes in rodents and patients with type 1 diabetes. In the hypothalamus and granular insular cortex of mice with type 1 diabetes, bone marrow-derived microglia/macrophages were activated and found at a greater density than in controls. Infiltration of CD45(+)/CCR2(+)/GR-1(+)/Iba-1(+) bone marrow-derived monocytes into the hypothalamus could be mitigated by treatment with minocycline, an anti-inflammatory agent capable of crossing the blood-brain barrier. Our studies suggest that targeting central inflammation may facilitate management of microvascular complications.
Assuntos
Medula Óssea/inervação , Medula Óssea/patologia , Sistema Nervoso Central/patologia , Diabetes Mellitus Tipo 1/complicações , Diabetes Mellitus Tipo 1/patologia , Inflamação/patologia , Animais , Medula Óssea/efeitos dos fármacos , Sistema Nervoso Central/efeitos dos fármacos , Sistema Nervoso Central/metabolismo , Citocinas/metabolismo , Ensaio de Imunoadsorção Enzimática , Proteínas de Fluorescência Verde/metabolismo , Hematopoese/efeitos dos fármacos , Herpesvirus Suídeo 1/efeitos dos fármacos , Herpesvirus Suídeo 1/fisiologia , Humanos , Inflamação/complicações , Inflamação/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microglia/efeitos dos fármacos , Microglia/metabolismo , Microglia/patologia , Minociclina/farmacologia , Modelos Biológicos , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , Monócitos/patologia , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Neurônios/patologia , Neurotransmissores/metabolismo , Proteínas Proto-Oncogênicas c-fos/metabolismo , Ratos , Ratos Wistar , Sistema Nervoso Simpático/efeitos dos fármacos , Sistema Nervoso Simpático/metabolismo , Sistema Nervoso Simpático/patologiaRESUMO
Our objective is to develop a new therapy for the treatment of stroke. Currently, the only effective therapy for acute ischemic stroke is the thrombolytic agent recombinant tissue plasminogen activator. α1-Antitrypsin (AAT), a serine proteinase inhibitor with potent anti-inflammatory, anti-apoptotic, antimicrobial, and cytoprotective activities, could be beneficial in stroke. The goal of this study is to test whether AAT can improve ischemic stroke outcome in an established rat model. Middle cerebral artery occlusion was induced in male rats via intracranial (i.c.) microinjection of endothelin-1. Five to 10 minutes after stroke induction, rats received either i.c. or intravenous delivery of human AAT. Cylinder and vibrissae tests were used to evaluate sensorimotor function before and 72 hours after middle cerebral artery occlusion. Infarct volumes were examined via either 2,3,5-triphenyltetrazolium chloride assay or magnetic resonance imaging 72 hours after middle cerebral artery occlusion. Despite equivalent initial strokes, at 72 hours, the infarct volumes of the human AAT treatment groups (local and systemic injection) were statistically significantly reduced by 83% and 63% (P < .0001 and P < .05, respectively) compared with control rats. Human AAT significantly limited sensory motor system deficits. Human AAT could be a potential novel therapeutic drug for the protection against neurodegeneration after ischemic stroke, but more studies are needed to investigate the protective mechanisms and efficacy in other animal models.
Assuntos
Encéfalo/efeitos dos fármacos , Infarto da Artéria Cerebral Média/tratamento farmacológico , Fármacos Neuroprotetores/farmacologia , alfa 1-Antitripsina/farmacologia , Animais , Encéfalo/irrigação sanguínea , Encéfalo/patologia , Encéfalo/fisiopatologia , Citoproteção , Modelos Animais de Doenças , Endotelina-1 , Humanos , Infarto da Artéria Cerebral Média/induzido quimicamente , Infarto da Artéria Cerebral Média/diagnóstico , Infarto da Artéria Cerebral Média/fisiopatologia , Injeções Intravenosas , Imageamento por Ressonância Magnética , Masculino , Microinjeções , Atividade Motora/efeitos dos fármacos , Fármacos Neuroprotetores/administração & dosagem , Ratos Sprague-Dawley , Limiar Sensorial/efeitos dos fármacos , Fatores de Tempo , alfa 1-Antitripsina/administração & dosagemRESUMO
We established a novel brain slice assay to test the ability of acetylcholinesterase (AChE) reactivators to prevent ACh-induced M1 muscarinic acetylcholine receptor (mAChR) dependent hyperexcitability observed after exposure to the organophosphate (OP)-based AChE inhibitor and sarin surrogate 4-nitrophenyl isopropyl methylphosphonate (NIMP). Whole-cell patch clamp recordings were used to evaluate the response of pyramidal neurons in the rat basolateral amygdala (BLA) to brief (1 min) bath application of ACh (100 µM), either in control conditions, or after exposure to NIMP ± an AChE reactivator. Bath application of ACh produced atropine- and pirenzepine-sensitive inward currents in voltage clamped BLA pyramidal neurons, and increased the frequency of spontaneous EPSCs, suggesting robust activation of M1 mAChRs. Responses to ACh were increased ~3-5 fold in slices that had been preincubated in NIMP, and these effects were reversed in a concentration dependent manner by exposure to a commercially available AChE reactivator. The current work outlines a simple assay that can be used to evaluate the efficacy of both known and novel AChE reactivators in an area of the limbic system that likely contributes to seizures after acute exposure to OP-based AChE inhibitors.
RESUMO
A family of 20 tris-azaaromatic quaternary ammonium (AQA) compounds were tested for their inhibition of alpha7 nicotinic acetylcholine receptors (nAChRs) expressed in Xenopus laevis oocytes. The potency of inhibitory activity was related to the hydrophobic character of the tris head groups. Two tris-AQA compounds were studied in detail: the highly effective inhibitor 1,3,5-tri-[5-(1-quinolinum)-pent-1-yn-1-yl]-benzene tribromide (tPyQB) and the less potent antagonist 1,3,5,-tri-{5-[1-(2-picolinium)]-pent-1-yn-1-yl}benzene tribromide (tPy2PiB). In addition, we evaluated 1,2,4,5-tetra-{5-[1-(3-benzyl)pyridinium]pent-1-yl}benzene tetrabromide (tkP3BzPB), a tetrakis-AQA with very hydrophobic headgroups. We compared the activity of the AQA compounds to the frequently used alpha7-antagonist methyllycaconitine (MLA). Both tPyQB and tkP3BzPB were selective antagonists of alpha7. However, although inhibition by tPyQB was reversible within 5 min, the recovery time constant for tkP3BzPB inhibition was 26.6 +/- 0.8 min, so that the equilibrium inhibition in the prolonged presence of nanomolar concentrations of tkP3BzPB was nearly 100%. The potency, selectivity, and slow reversibility of tkP3BzPB were comparable with or greater than that of MLA. The inhibitory actions of tPyQB, tPy2PiB, and tkP3BzPB were evaluated on the acetylcholine (ACh)-evoked responses of native nAChRs in rat brain slices. The alpha7-mediated responses of hippocampal interneurons were effectively reduced by 1 microM tPyQB and tkP3BzPB but not tPy2PiB. In rat medial septum, tkP3BzPB produced a greater inhibition of ACh-evoked responses of cells with fast inward currents (type I) than of cells with predominantly slow kinetics (type II), suggesting that tkP3BzPB can block alpha7 yet preserve the responsiveness of non-alpha7 receptors. These agents might be helpful in elucidating complex receptor responses in brain regions with mixed populations of nAChRs.
Assuntos
Compostos de Amônio Quaternário/farmacologia , Receptores Nicotínicos/efeitos dos fármacos , Acetilcolina/metabolismo , Acetilcolina/farmacologia , Animais , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Oócitos , Ratos , Receptores Nicotínicos/biossíntese , Xenopus laevis , Receptor Nicotínico de Acetilcolina alfa7RESUMO
We examined the contribution of N-methyl-D-aspartate receptor (NMDAR) subunits in the redox-mediated decline in NMDAR function during aging. GluN2A and GluN2B selective antagonists decreased peak NMDAR currents to a similar extent in young and aged animals, indicating that a shift in diheteromeric GluN2 subunits does not underlie the age-related decrease in the NMDAR synaptic function. Application of dithiothreitol (DTT) in aged animals, increased peak NMDAR synaptic currents, prolonged the decay time, and increased the sensitivity of the synaptic response to the GluN2B antagonist, ifenprodil, indicating that DTT increased the contribution of GluN2B subunits to the synaptic response. The DTT-mediated increase in NMDAR function was inhibited by partial blockade of NMDARs, and this inhibition was rescued by increasing Ca2+ concentration in the recording medium. The results indicate that DTT-mediated potentiation requires Ca2+ influx through NMDAR activity. Finally, redox regulation of NMDAR function depends on the activity of Ca2+/calmodulin-dependent protein kinase II (CaMKII). The results indicate that activity-dependent NMDAR synaptic plasticity is suppressed by redox-mediated inhibition of CaMKII activation during aging. The redox regulation of NMDARs represents a suppression of a metaplasticity mechanism, which can disrupt synaptic plasticity and cognition associated with neurological or psychiatric diseases, and aging.
Assuntos
Envelhecimento , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Plasticidade Neuronal , Células Piramidais/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Transmissão Sináptica , Animais , Região CA1 Hipocampal/citologia , Ditiotreitol/farmacologia , Masculino , Modelos Animais , Oxidantes/farmacologia , Oxirredução , Piperidinas/farmacologia , Subunidades Proteicas/antagonistas & inibidores , Subunidades Proteicas/metabolismo , Ratos , Ratos Endogâmicos F344 , Receptores de N-Metil-D-Aspartato/antagonistas & inibidoresRESUMO
One approach to treatment of negative cognitive effects associated with Alzheimer's disease and schizophrenia may involve activation of neuronal alpha7 nicotinic acetylcholine receptors (nAChRs). We used the alpha7-selective partial agonist 3-(4-hydroxy, 2-methoxy-benzylidene)anabaseine (4OH-GTS-21), the alpha7 modulator 5-hydroxyindole (5-HI), and recombinant adeno-associated virus (rAAV)-mediated alpha7 gene transfer in order to test the hypothesis whether combining these strategies would significantly increase indirect measures of alpha7 nAChR function, including measures of spontaneous synaptic events in CA1 pyramidal cells. 5-HI (1 mM), and 5-HI (1 mM)+4OH-GTS-21 (5 microM) increased the frequency of APV- and NBQX-sensitive currents, while 5-HI+4OH-GTS-21 increased the frequency and amplitude of bicuculline-sensitive currents. Effects on EPSCs were blocked with tetrodotoxin (TTX) (1 microM), but not by methyllycaconitine (MLA) (50 nM). Neither TTX nor MLA reduced the potentiation of IPSC frequencies. However, TTX blocked, and in some cases MLA reduced, the potentiation of IPSC amplitudes. These data suggest that effects of 5-HI+4OH-GTS-21 on EPSC frequency were associated with action potential-dependent transmitter release produced by 5HI, and that potentiation of IPSC amplitudes resulted at least in part, from activation of alpha7 nAChRs. Finally, rAAV-mediated alpha7 gene transfer did not alter the magnitude of effects produced by 5-HI or 5-HI+4OH-GTS-21. Thus, although we previously showed that direct measures of alpha7 nAChR function were enhanced by alpha7 gene transfer, indirect measures of alpha7 nAChRs function were not significantly enhanced by combining alpha7 gene transfer with either agonist activation or positive allosteric modulation of alpha7 nAChRs.
Assuntos
Anabasina/análogos & derivados , Dependovirus/fisiologia , Hipocampo/efeitos dos fármacos , Indóis/farmacologia , Receptores Nicotínicos/genética , Sinapses/efeitos dos fármacos , Anabasina/farmacologia , Animais , Animais Recém-Nascidos , Interações Medicamentosas , Antagonistas de Aminoácidos Excitatórios/farmacologia , Técnicas de Transferência de Genes , Hipocampo/fisiologia , Técnicas In Vitro , Técnicas de Patch-Clamp , Quinoxalinas/farmacologia , Ratos , Ratos Sprague-Dawley , Bloqueadores dos Canais de Sódio/farmacologia , Sinapses/fisiologia , Transmissão Sináptica/efeitos dos fármacos , Transmissão Sináptica/fisiologia , Transmissão Sináptica/efeitos da radiação , Tetrodotoxina/farmacologia , Valina/análogos & derivados , Valina/farmacologia , Receptor Nicotínico de Acetilcolina alfa7RESUMO
S 24795 evoked methyllycaconitine-sensitive inward currents in voltage-clamped hippocampal interneurons with maximum amplitude about 14% that of ACh-evoked responses. Experiments with rat alpha7 receptors expressed in Xenopus oocytes confirmed that S 24795 is a partial agonist of alpha7 nAChR with an EC(50) of 34+/-11 microM and I(max) of approximately 10% relative to ACh. When 60 microM ACh was co-applied to alpha7-expressing oocytes along with increasing concentrations of S 24795, there was a progressive decrease in response compared to the responses to 60 microM ACh alone (IC(50) 45+/-9 microM). The positive allosteric modulator 5-hydroxyindole potentiated ACh- and S 24795-evoked responses of alpha7 receptors in both oocytes and hippocampal interneurons. In hippocampal slice experiments, depending on the ACh concentrations in the application pipette and the ratio of ACh to S 24795, co-application of S 24795 with ACh variously increased, decreased, or had no effect on responses, compared to ACh alone. In order to estimate the effective dilution factor for the pressure application experiments, we tested alpha7 receptors in oocytes with ACh alone and in co-application with S 24795 at the same ratios as in the slice experiments, but at varying dilution factors. The pattern of interaction seen in the slice experiments was most closely matched under the conditions of a 3:100 dilution, suggesting that the pipette solution was diluted approximately 30-fold at the site of action. This dilution factor was consistent with the potency of ACh and S 24795 in the oocyte expression system (EC(50)s approximately 30 microM).
Assuntos
Agonistas Colinérgicos/farmacologia , Hipocampo/citologia , Interneurônios/efeitos dos fármacos , Receptores Nicotínicos/fisiologia , Acetilcolina/farmacologia , Aconitina/análogos & derivados , Aconitina/farmacologia , Animais , Animais Recém-Nascidos , Antagonistas Colinérgicos/farmacologia , Relação Dose-Resposta a Droga , Interações Medicamentosas , Feminino , Técnicas In Vitro , Indóis/farmacologia , Concentração Inibidora 50 , Masculino , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Potenciais da Membrana/efeitos da radiação , Oócitos , Técnicas de Patch-Clamp , Compostos de Piridínio/farmacologia , Ratos , Ratos Sprague-Dawley , Xenopus , Receptor Nicotínico de Acetilcolina alfa7RESUMO
The widespread nature of diabetes affects all organ systems of an individual including the bone marrow. Long-term damage to the cellular and extracellular components of the bone marrow leads to a rapid decline in the bone marrow-hematopoietic stem/progenitor cells (HS/PCs) compartment. This review will highlight the importance of bone marrow microenvironment in maintaining bone marrow HS/PC populations and the contribution of these key populations in microvascular repair during the natural history of diabetes. The autonomic nervous system can initiate and propagate bone marrow dysfunction in diabetes. Systemic pharmacological strategies designed to protect the bone marrow-HS/PC population from diabetes induced-oxidative stress and advanced glycation end product accumulation represent a new approach to target diabetic retinopathy progression. Protecting HS/PCs ensures their participation in vascular repair and reduces the risk of vasogdegeneration occurring in the retina.
Assuntos
Medula Óssea/fisiologia , Retinopatia Diabética/fisiopatologia , Células-Tronco Hematopoéticas/fisiologia , Vasos Retinianos/fisiologia , Nicho de Células-Tronco/fisiologia , Humanos , RejuvenescimentoRESUMO
We recently found indicators of hypothalamic inflammation and neurodegeneration linked to the loss of neuroprotective factors including insulin-like growth factor (IGF-1) and IGF binding protein-2 (IGFBP-3) in mice made diabetic using streptozotocin (STZ). In the current work, a genetic model of type-1 diabetes (Ins2(Akita) mouse) was used to evaluate changes in neuronal activity and concomitant changes in the proinflammatory mediator high-mobility group box-1 (HMBG1). We found basal hypothalamic neuronal activity as indicated by manganese-enhanced magnetic resonance imaging (MEMRI) was significantly decreased in 8 months old, but not 2 months old Ins2(Akita) diabetic mice compared to controls. In tissue from the same animals we evaluated the expression of HMBG1 using immunohistochemistry and confocal microscopy. We found decreased HMBG1 nuclear localization in the paraventricular nucleus of the hypothalamus (PVN) in 8 months old, but not 2 months old diabetic animals indicating nuclear release of the protein consistent with an inflammatory state. Adjacent thalamic regions showed little change in HMBG1 nuclear localization and neuronal activity as a result of diabetes. This work extends our previous findings demonstrating changes consistent with hypothalamic neuroinflammation in STZ treated animals, and shows active inflammatory processes are correlated with changes in basal hypothalamic neuronal activity in Ins2(Akita) mice.
Assuntos
Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 1/fisiopatologia , Proteína HMGB1/metabolismo , Hipotálamo/metabolismo , Hipotálamo/fisiopatologia , Neurônios/fisiologia , Transporte Ativo do Núcleo Celular , Animais , Núcleo Celular/metabolismo , Citoplasma/metabolismo , Diabetes Mellitus Tipo 1/genética , Inflamação/metabolismo , Inflamação/fisiopatologia , Masculino , Camundongos , Núcleo Hipotalâmico Paraventricular/metabolismoRESUMO
In this study, the role of CX3CR1 in the progression of diabetic retinopathy (DR) was investigated. The retinas of wild-type (WT), CX3CR1 null (CX3CR1gfp/gfp, KO), and heterozygous (CX3CR1+/gfp, Het) mice were compared in the presence and absence of streptozotocin (STZ)-induced diabetes. CX3CR1 deficiency in STZ-KO increased vascular pathology at 4 months of diabetes, as a significant increase in acellular capillaries was observed only in the STZ-KO group. CX3CR1 deficiency and diabetes had similar effects on retinal neurodegeneration measured by an increase in DNA fragmentation. Retinal vascular pathology in STZ-KO mice was associated with increased numbers of monocyte-derived macrophages in the retina. Furthermore, compared to STZ-WT, STZ-KO mice exhibited increased numbers of inflammatory monocytes in the bone marrow and impaired homing of monocytes to the spleen. The induction of retinal IL-10 expression by diabetes was significantly less in KO mice, and when bone marrow-derived macrophages from KO mice were maintained in high glucose, they expressed significantly less IL-10 and more TNF-α in response to LPS stimulation. These findings support that CX3CR1 deficiency accelerates the development of vascular pathology in DR through increased recruitment of proinflammatory myeloid cells that demonstrate reduced expression of anti-inflammatory IL-10. KEY MESSAGES: ⢠CX3CR1 deletion in STZ-diabetic mice accelerated the onset of diabetic retinopathy (DR). ⢠The early onset of DR was associated with increased retinal cell apoptosis. ⢠The early onset of DR was associated with increased recruitment of bone marrow-derived macrophages to the retina. ⢠Bone marrow-derived macrophages from CX3CR1 KO diabetic mice expressed more TNF-α and less IL-10. ⢠The role of IL-10 in protection from progression of DR is highlighted.
Assuntos
Receptor 1 de Quimiocina CX3C/deficiência , Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 1/patologia , Retinopatia Diabética/metabolismo , Retinopatia Diabética/patologia , Animais , Apoptose , Peso Corporal , Células da Medula Óssea/metabolismo , Receptor 1 de Quimiocina CX3C/metabolismo , Modelos Animais de Doenças , Deleção de Genes , Hemoglobinas Glicadas/metabolismo , Homeostase , Hipotálamo/metabolismo , Inflamação/patologia , Mediadores da Inflamação/metabolismo , Interleucina-10/metabolismo , Macrófagos/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Monócitos/metabolismo , Células Mieloides/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Retina/metabolismo , Retina/patologia , EstreptozocinaRESUMO
The medial septum-diagonal band (MS/DB) contains primarily cholinergic and GABAergic neurons that project to the hippocampus, and are important for learning and memory. Whole-cell patch clamp methods with brain slices from p11--p20 rats were used to measure MS/DB cell responses to focal somatic application of 1mM acetylcholine (ACh) and a series of current pulses was applied in order to assess firing frequencies and the presence of hyperpolarization-activated currents (Ih). We identified three types of cells: (1) cells with fast inward currents blocked by methyllycaconitine (MLA) with slow firing rates (3--12 Hz), accommodating action potentials, and no Ih (n=20); (2) cells with currents that had both fast (MLA-sensitive) and slow components that were blocked with mecamylamine (MEC) that showed fast firing (up to 60 Hz) and slow firing (up to 3 Hz), with accommodating and non-accommodating action potentials (n=46), 33% of which had Ih; and (3) cells not responsive to ACh with moderate firing rates (10--42 Hz), some with accommodating action potentials and some without (n=19), of which 92% had Ih. These results are among the first to demonstrate functional nicotinic receptors in the MS/DB. The data suggest that these receptors include alpha 7 and non-alpha 7 subtypes and that the expression of each is correlated with firing frequency and the presence of Ih. Responses to ACh were not affected by tetrodotoxin (TTX) and CdC l(2) but were blocked by MLA or MLA and MEC, suggesting that these currents involve direct activation of nicotinic receptors.
Assuntos
Potenciais de Ação/fisiologia , Feixe Diagonal de Broca/fisiologia , Neurônios/fisiologia , Receptores Nicotínicos/metabolismo , Núcleos Septais/fisiologia , Animais , Masculino , Ratos , Ratos Sprague-Dawley , Estatística como AssuntoRESUMO
PURPOSE: Age-related macular degeneration (AMD) is a leading cause of visual impairment worldwide. Genetics and diet contribute to the relative risk for developing AMD, but their interactions are poorly understood. Genetic variations in Complement Factor H (CFH), and dietary glycemic index (GI) are major risk factors for AMD. We explored the effects of GI on development of early AMD-like features and changes to central nervous system (CNS) inflammation in Cfh-null mice. METHODS: Aged 11-week-old wild type (WT) C57Bl/6J or Cfh-null mice were group pair-fed high or low GI diets for 33 weeks. At 10 months of age, mice were evaluated for early AMD-like features in the neural retina and RPE by light and electron microscopy. Brains were analyzed for Iba1 macrophage/microglia immunostaining, an indicator of inflammation. RESULTS: The 10-month-old WT mice showed no retinal abnormalities on either diet. The Cfh-null mice, however, showed distinct early AMD-like features in the RPE when fed a low GI diet, including vacuolation, disruption of basal infoldings, and increased basal laminar deposits. The Cfh-null mice also showed thinning of the RPE, hypopigmentation, and increased numbers of Iba1-expressing macrophages in the brain, irrespective of diet. CONCLUSIONS: The presence of early AMD-like features by 10 months of age in Cfh-null mice fed a low GI diet is surprising, given the apparent protection from the development of such features in aged WT mice or humans consuming lower GI diets. Our findings highlight the need to consider gene-diet interactions when developing animal models and therapeutic approaches to treat AMD.
Assuntos
Fator H do Complemento/genética , Carboidratos da Dieta/farmacologia , Índice Glicêmico/genética , Degeneração Macular/genética , Animais , Glicemia/metabolismo , Fator H do Complemento/metabolismo , Modelos Animais de Doenças , Genótipo , Imuno-Histoquímica , Degeneração Macular/dietoterapia , Degeneração Macular/patologia , Camundongos , Camundongos Endogâmicos C57BL , Microscopia Eletrônica , Epitélio Pigmentado da Retina/ultraestruturaRESUMO
In this study, we assessed whether Per2 clock gene-mutant mice exhibit a vascular phenotype similar to diabetes. Per2 (B6.129-Per2(tm1Drw)/J) or wild-type control mice 4 and 12 months of age were used. To evaluate diabetes-like phenotype in Per2 mutant mice, retina was quantified for mRNA expression, and degree of diabetic retinopathy was evaluated. Bone marrow neuropathy was studied by staining femurs for tyrosine hydroxylase (TH) and neurofilament 200 (NF-200). The rate of proliferation and quantification of bone marrow progenitor cells (BMPCs) was performed, and a threefold decrease in proliferation and 50% reduction in nitric oxide levels were observed in Per2 mutant mice. TH-positive nerve processes and NF-200 staining were reduced in Per2 mutant mice. Both retinal protein and mRNA expression of endothelial nitric oxide synthase were decreased by twofold. Other endothelial function genes (VEGFR2, VEGFR1) were downregulated (1.5-2-fold) in Per2 mutant retinas, whereas there was an upregulation of profibrotic pathway mediated by transforming growth factor-ß1. Our studies suggest that Per2 mutant mice recapitulate key aspects of diabetes without the metabolic abnormalities, including retinal vascular damage, neuronal loss in the bone marrow, and diminished BMPC function.
Assuntos
Medula Óssea/inervação , Diabetes Mellitus/fisiopatologia , Retinopatia Diabética/etiologia , Proteínas Circadianas Period/fisiologia , Animais , Medula Óssea/patologia , Ritmo Circadiano , Células-Tronco Hematopoéticas/fisiologia , Camundongos , Mutação , Óxido Nítrico/análise , Óxido Nítrico Sintase Tipo III/análise , Proteínas Circadianas Period/genética , Permeabilidade , Fenótipo , Retina/enzimologia , Fator de Crescimento Transformador beta1/fisiologia , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/fisiologiaRESUMO
Oxidative stress in the brain is implicated in increased sympathetic drive, inflammatory status, and vascular dysfunctions, associated with development and establishment of hypertension. However, little is known about the mechanism of this impaired brain-vascular communication. Here, we tested the hypothesis that increased oxidative stress in the brain cardioregulatory areas, such as the paraventricular nucleus of the hypothalamus, is driven by mitochondrial reactive oxygen species and leads to increased inflammatory cells (ICs) and decreased/dysfunctional endothelial progenitor cells (EPCs), thereby compromising vasculature repair and accelerating hypertension. Chronic angiotensin II infusion resulted in elevated blood pressure and sympathetic vasomotor drive, decreased spontaneous baroreflex gain, and increased microglia activation in the paraventricular nucleus. This was associated with 46% decrease in bone marrow (BM)-derived EPCs and 250% increase in BM ICs, resulting in 5-fold decrease of EPC/IC ratio in the BM. Treatment with mitochondrial-targeted antioxidant, a scavenger of mitochondrial O(2)(-·), intracerebroventricularly but not subcutaneously attenuated angiotensin II-induced hypertension, decreased activation of microglia in the paraventricular nucleus, and normalized EPCs/ICs. This functional communication between the brain and BM was confirmed by retrograde neuronal labeling from the BM with green fluorescent protein-tagged pseudorabies virus. Administration of green fluorescent protein-tagged pseudorabies virus into the BM resulted in predominant labeling of paraventricular nucleus neurons within 3 days, with some fluorescence in the nucleus tractus solitarius, the rostral ventrolateral medulla, and subfornical organ. Taken together, these data demonstrate that inhibition of mitochondrial reactive oxygen species attenuates angiotensin II-induced hypertension and corrects the imbalance in EPCs/ICs in the BM. They suggest that an imbalance in vascular reparative and ICs may perpetuate vascular pathophysiology in this model of hypertension.
Assuntos
Pressão Sanguínea/fisiologia , Medula Óssea/fisiopatologia , Encéfalo/fisiopatologia , Hipertensão/fisiopatologia , Angiotensina II , Animais , Animais Recém-Nascidos , Sistema Nervoso Autônomo/fisiopatologia , Pressão Sanguínea/efeitos dos fármacos , Medula Óssea/metabolismo , Medula Óssea/patologia , Encéfalo/metabolismo , Encéfalo/patologia , Células Cultivadas , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Citometria de Fluxo , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Células-Tronco Hematopoéticas/metabolismo , Células-Tronco Hematopoéticas/patologia , Hipertensão/induzido quimicamente , Hipertensão/prevenção & controle , Infusões Intraventriculares , Linfócitos/metabolismo , Linfócitos/patologia , Macrófagos/metabolismo , Macrófagos/patologia , Masculino , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Compostos Organofosforados/administração & dosagem , Compostos Organofosforados/farmacologia , Núcleo Hipotalâmico Paraventricular/metabolismo , Núcleo Hipotalâmico Paraventricular/patologia , Núcleo Hipotalâmico Paraventricular/fisiopatologia , Piperidinas/administração & dosagem , Piperidinas/farmacologia , Ratos , Ratos Sprague-DawleyRESUMO
Endothelial progenitor cells (EPCs), critical for mediating vascular repair, are dysfunctional in a hyperglycemic and/or hypercholesterolemic environment. Their dysfunction contributes to the progression of diabetic macro- and microvascular complications. Activation of "cholesterol-sensing" nuclear receptors, the liver X receptors (LXRα/LXRß), protects against atherosclerosis by transcriptional regulation of genes important in promoting cholesterol efflux and inhibiting inflammation. We hypothesized that LXR activation with a synthetic ligand would correct diabetes-induced EPC dysfunction and improve diabetic retinopathy. Studies were performed in streptozotocin (STZ)-injected DBA/2J mice fed a high-fat Western diet (DBA/STZ/WD) and treated with the LXR agonist GW3965 and in LXRα(-/-), LXRß(-/-), and LXRα/ß(-/-) mice. Retinas were evaluated for number of acellular capillaries and glial fibrillary acidic protein (GFAP) immunoreactivity. Bone marrow EPCs were analyzed for migratory function and gene expression. Compared with vehicle-treated DBA/STZ/WD mice, GW3965 treated mice showed fewer acellular capillaries and reduced GFAP expression. These mice also exhibited enhanced EPC migration and restoration of inflammatory and oxidative stress genes toward nondiabetic levels. LXRα(-/-), LXRß(-/-), and LXRα/ß(-/-) mice developed acellular capillaries and EPC dysfunction similar to the DBA/STZ/WD mice. These studies support a key role for LXR in retinal and bone marrow progenitor dysfunction associated with type 1 diabetes. LXR agonists may represent promising pharmacologic targets for correcting retinopathy and EPC dysfunction.
Assuntos
Diabetes Mellitus Experimental/metabolismo , Retinopatia Diabética/metabolismo , Receptores Nucleares Órfãos/metabolismo , Animais , Benzoatos/farmacologia , Benzoatos/uso terapêutico , Benzilaminas/farmacologia , Benzilaminas/uso terapêutico , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/genética , Retinopatia Diabética/tratamento farmacológico , Retinopatia Diabética/genética , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Receptores X do Fígado , Masculino , Camundongos , Camundongos Knockout , Receptores Nucleares Órfãos/agonistas , Receptores Nucleares Órfãos/genética , Células-Tronco/metabolismo , Células-Tronco/patologiaRESUMO
BACKGROUND: To date, recombinant tissue plasminogen activator (rtPA) is the only approved drug for ischemic stroke. It is intravenously administered functioning as a thrombolytic agent and is used to obtain reperfusion of the affected area of the brain. Excitotoxicity, inflammation and apoptosis are all involved in delayed neuronal death following stroke and offer multiple opportunities to intervene with neuroprotective agents. Gelsolin (GSN) is an actin- and calcium-binding protein mediating the disassembly of actin filaments and activity of calcium channels. It also functions as a regulator of apoptosis and inflammatory responses. This study tests the hypothesis that increasing the concentration of the form of GSN known as plasma GSN (pGSN) near an infarct will provide neuroprotection following ischemic stroke. METHODS: We induced middle cerebral artery occlusion (MCAO) in male rats via intracranial injection of endothelin-1 (ET-1), a potent vasoconstrictor, and then treated with local delivery of pGSN. Whole brain laser Doppler perfusion imaging was performed through the skull to assess MCAO effectiveness. Cylinder and vibrissae tests evaluated sensorimotor function before and 72 h after MCAO. Infarct volumes were examined 72 h after MCAO via 2, 3, 5-triphenyltetrazolium chloride (TTC) assay. RESULTS: Estimates of relative cerebral perfusion were significantly decreased in all groups receiving MCAO with no differences detected between treatments. Despite equivalent initial strokes, the infarct volume of the pGSN treatment group was significantly reduced compared with the untreated MCAO rats at 72 h. ET-1 induced significant deficits in both cylinder and vibrissae tests while pGSN significantly limited these deficits. CONCLUSION: Gelsolin could be a promising drug for protection against neurodegeneration following ischemic stroke.