RESUMO
INTRODUCTION AND OBJECTIVES: Dual antiplatelet therapy (DAPT) duration after ST-segment elevation myocardial infarction (STEMI) remains a matter of debate. METHODS: We analyzed the effect of DAPT on 5-year all-cause mortality, cardiovascular mortality, and cardiovascular readmission or mortality in a cohort of 1-year survivor STEMI patients. RESULTS: A total of 3107 patients with the diagnosis of STEMI were included: 93% of them were discharged on DAPT, a therapy that persisted in 275 high-risk patients at 5 years. Cardiovascular mortality in patients on single antiplatelet therapy vs DAPT at 5 years was 1.4% vs 3.6% (P <.01), respectively, whereas noncardiovascular mortality was 3.3% vs 5.8% (P=.049) at 5 years. Cardiovascular readmission or mortality in patients with single antiplatelet therapy vs DAPT was 11.4% vs 46.5% (P <.001). Extended DAPT was independently associated with worse 5-year all-cause mortality (HR, 2.16; 95%CI, 1.40-3.33), cardiovascular mortality (HR, 2.83; 95%CI, 1.37-5.84), and cardiovascular readmission or mortality (HR, 5.20; 95%CI, 3.96-6.82). These findings were confirmed in propensity score matching and inverse probability weighting analyses. CONCLUSIONS: Our results suggest the hypothesis that, in 1-year STEMI survivors, extending DAPT up to 5 years in high-risk patients does not improve their long-term prognosis.
Assuntos
Intervenção Coronária Percutânea , Infarto do Miocárdio com Supradesnível do Segmento ST , Humanos , Inibidores da Agregação Plaquetária/uso terapêutico , Infarto do Miocárdio com Supradesnível do Segmento ST/tratamento farmacológico , Infarto do Miocárdio com Supradesnível do Segmento ST/diagnóstico , Resultado do Tratamento , Intervenção Coronária Percutânea/métodosRESUMO
Eumelanin is the biopolymer responsible for photoprotection in living beings and holds great promise as a smart biomaterial, but its detailed structure has not been characterized experimentally. Theoretical models are urgently needed to improve our knowledge of eumelanin's function and exploit its properties, but the enormous amount of possible oligomer components has made modelling not possible until now. Here we show that the stability and lowest vertical optical absorption of 5,6-dihydroxyindole (DHI) eumelanin dimer components can be modeled with deep neural networks, using fingerprint-like molecular representations as input. In spite of the modest data set size, average errors of only 6 and 9% for stability and S1 absorption energy are obtained. Our fingerprints code the connectivity and oxidation patterns of the dimers in a straightforward, unambiguous way and can be extended to larger oligomers. This proof-of-principle work shows that machine learning can be applied to help solve the structural challenge of melanin.
RESUMO
OBJECTIVES: To identify predictors of primary angioplasty delay in patients with ST-elevation myocardial infarction (STEMI) transported from out-of-hospital sites or from hospitals without percutaneous coronary intervention (PCI) suites. MATERIAL AND METHODS: Retrospective cohort study of cases between 2008 and 2018 in a university hospital receiving patients diagnosed with STEMI who required a PCI. We performed linear and multivariate regression analyses to identify factors that predicted delay in interpreting a diagnostic electrocardiogram (ECG) until the guidewire passed the lesion (diagnosis-guidewire-crossing time). RESULTS: A total of 1039 cases were studied; 296 patients (28.4%) had delays of more than 120 minutes between STEMI diagnosis and guidewire crossing. Factors associated with PCI delay were advanced age (odds ratio [OR] = 1.02; 95% CI, 1.01-1.04]), severe heart failure on admission (OR = 2.28; 95% CI, 1.23-4.22), history of cardiac bypass surgery (OR = 10.01; 95% CI, 2.60-41.81), out-of-hospital cardiac arrest (OR = 4.34; 95% CI, 1.84-10.32), lateral ischemia (OR, 1.64; 95% CI, 1.06-2.51), first medical attention in a hospital without a PCI suite (OR = 1.52; 95% CI, 1.05-2.21), first medical attention outside regular working hours (OR = 1.46; 95% CI, 1.06-2.02), and distance in kilometers to a PCI suite (OR = 1.04; 95% CI, 1.03-1.05). CONCLUSION: Patients with STEMI who required transport to a hospital with a PCI suite experienced primary angioplasty delays. Delays were related to logistical and clinical factors as well as to infarction characteristics.
OBJETIVO: Identificar variables predictoras del retraso hasta la angioplastia primaria, en los pacientes con infarto agudo de miocardio con elevación del ST (IAMEST) trasladados desde el medio extrahospitalario o desde hospitales sin hemodinámica. METODO: Estudio de cohortes, retrospectivo, realizado entre 2008 y 2018 en un hospital universitario receptor de pacientes con diagnóstico de IAMEST y que requirieron angioplastia primaria. Se realizó un análisis multivariable de regresión logística y lineal para identificar variables predictoras de demora de tiempo de electrocardiograma (ECG) diagnóstico hasta el paso de guía. RESULTADOS: Se incluyeron 1.039 pacientes en el estudio. Doscientos noventa y seis pacientes (28,4%) presentaban tiempos ECG diagnóstico-paso de guía > 120 minutos. Las variables asociadas a tiempos prolongados de angioplastia primaria fueron la edad avanzada [odds ratio (OR) = 1,02; IC 95%: 1,01-1,04] la insuficiencia cardiaca grave al ingreso (OR = 2,28; IC 95%: 1,23-4,22), la cirugía cardiaca previa de bypass (OR = 10,01; IC 95%: 2,60-41,81), la muerte súbita extrahospitalaria recuperada (OR = 4,34; IC 95%: 1,84-10,32), la localización lateral del infarto (OR = 1,64; IC 95%: 1,06-2,51), el primer contacto con hospital sin disponibilidad de hemodinámica (OR = 1,52; IC 95%: 1,05- 2,21), la atención fuera de horas (OR = 1,46; IC 95%: 1,06-2,02) y finalmente la distancia en kilómetros al centro con hemodinámica (OR = 1,04; IC 95%: 1,03-1,05). CONCLUSIONES: En los pacientes con IAMEST que requirieron traslado a un centro con hemodinámica, la demora en la realización de la angioplastia primaria se relacionó con factores clínicos, con características del infarto y logísticas.
Assuntos
Intervenção Coronária Percutânea , Infarto do Miocárdio com Supradesnível do Segmento ST , Angioplastia , Eletrocardiografia , Hospitais , Humanos , Estudos Retrospectivos , Infarto do Miocárdio com Supradesnível do Segmento ST/diagnóstico , Infarto do Miocárdio com Supradesnível do Segmento ST/cirurgiaRESUMO
Right-sided infective endocarditis (RSIE), classically associated with intravenous drug use or intracardiac devices, is considered a good-prognosis infective endocarditis (IE) form. However, predisposing factors and prognosis for "NODID" RSIE (NOt associated with cardiac Devices or Intravenous Drug use) remain unclear. The aim of this study was to evaluate predisposing factors and prognosis of NODID RSIE compared to other RSIE forms. A retrospective cohort study (January 2008-January 2019) was conducted in a reference center on 300 patients diagnosed with IE. Endocarditis-related events were defined as related to IE in mortality or open-heart surgery during follow-up. A review and meta-analysis of associated literature (January 2008-January 2019) were also performed. Fifty-seven patients presented RSIE (19%), 22 of which were NODID RSIE (39%). Use of intravascular catheters (23% vs 3%; p = 0.027) and congenital heart diseases (18% vs 0%; p = 0.019) were associated with NODID RSIE. This group had a higher in-hospital mortality (23% vs 3%; p = 0.027) and endocarditis-related event rates (41% vs 6%; p = 0.001) than non-NODID RSIE. Furthermore, NODID RSIE was independently associated with in-hospital endocarditis-related events (OR = 19.29; 95%CI:2.23-167.16; p = 0.007). Our meta-analysis evaluated four studies and identified 96 cases (30%) of NODID RSIE from 320 total RSIE cases. NODID RSIE patients demonstrated higher in-hospital mortality (RR = 2.81; 95%CI:1.61-4.90; p < 0.001; I2 = 0.0%) and necessity of open-heart surgery (RR = 13.89; 95%CI:4.14-46.60; p < 0.001; I2 = 0.0%) than non-NODID RSIE cases. Our study suggests that NODID RSIE has the highest endocarditis-related event rate and in-hospital mortality among RSIE cases and therefore should not be considered a good-prognosis IE.
Assuntos
Procedimentos Cirúrgicos Cardíacos , Endocardite Bacteriana/mortalidade , Endocardite Bacteriana/cirurgia , Mortalidade Hospitalar , Adulto , Intervalo Livre de Doença , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Abuso de Substâncias por Via Intravenosa/mortalidade , Abuso de Substâncias por Via Intravenosa/cirurgia , Taxa de SobrevidaRESUMO
INTRODUCTION AND OBJECTIVES: In recent years, public access defibrillation programs have exponentially increased the availability of automatic external defibrillators (AED) in public spaces but there are no data on their performance in our setting. We conducted a descriptive analysis of the performance of AED since the launch of a public defibrillation program in our region. METHODS: A retrospective analysis was conducted of electrocardiographic tracings and the performance of AED in a public defibrillation program from June 2011 to June 2015 in the province of Girona, Spain. RESULTS: There were 231 AED activations. Full information was available on 188 activations, of which 82% corresponded to mobile devices and 18% to permanent devices. Asystole was the most prevalent rhythm (42%), while ventricular fibrillation accounted for 23%. The specificity of the device in identifying a shockable rhythm was 100%, but there were 8 false negatives (sensitivity 83%). There were 47 shockable rhythms, with a spontaneous circulation recovery rate of 49% (23 cases). There were no accidents related to the use of the device. CONCLUSIONS: Nearly half of the recorded rhythms were asystole. The AED analyzed showed excellent safety and specificity, with moderate sensitivity. Half the patients with a shockable rhythm were successfully treated by the AED.
Assuntos
Desfibriladores , Cardioversão Elétrica/instrumentação , Serviços Médicos de Emergência , Avaliação de Programas e Projetos de Saúde , Fibrilação Ventricular/terapia , Cardioversão Elétrica/estatística & dados numéricos , Eletrocardiografia , Humanos , Incidência , Estudos Retrospectivos , Espanha/epidemiologia , Fibrilação Ventricular/epidemiologia , Fibrilação Ventricular/fisiopatologiaRESUMO
OBJECTIVES: Long distance from a hospital with a catheterization laboratory is associated with a poorer prognosis in patients who undergo primary angioplasty for ST-elevation myocardial infarction (STEMI). An invasive pharmacologic strategy could offer an alternative treatment for these patients. We aimed to establish whether prognosis was better with primary angioplasty or fibrinolysis for reperfusion in cases of STEMI occurring far from a catheterization laboratory. MATERIAL AND METHODS: Prospective registry study of patients with STEMI admitted to our cardiology critical care unit. Patients were included over a 5-year period if they received reperfusion therapy and had required transport of more than 50 km to reach a hospital with a catheterization laboratory. We recorded characteristics of the STEMI event, treatment times, and short- and long-term mortality. The data was used for survival analysis. RESULTS: We registered 584 patients; 194 were treated with primary angioplasty and 390 with fibrinolysis. The mean time between first physician contact and balloon insertion was 160 minutes. The mean time between first physician contact and needle insertion for fibrinolysis was 30 minutes. The 2-year mortality rate was higher in patients treated with angioplasty (12.2%) than with those who underwent fibrinolysis (7.0%) ) (P=.04). Survival analysis showed that risk for death was higher in the primary angioplasty group (hazard ratio, 1.97 (95% CI, 0.64-0.95; P=.001). CONCLUSION: When STEMI occurs more than 50 km from a catheterization laboratory, reperfusion by means of balloon angioplasty delays care considerably and is associated with a higher mortality rate than reperfusion by fibrinolysis.
OBJETIVO: En los pacientes con infarto agudo de miocardio con elevación del ST (IAMEST) sometidos a angioplastia primaria (AP), la distancia al centro de hemodinámica alarga los tiempos y empeora el pronóstico. En estos pacientes, la estrategia fármaco-invasiva podría ser una alternativa. Nuestro objetivo fue establecer si el pronóstico es diferente en función del tipo de tratamiento de reperfusión en los pacientes alejados de un laboratorio de hemodinámica. METODO: Registro prospectivo durante 5 años de los pacientes con IAMEST ingresados en la unidad de cuidados críticos cardiológicos. Se seleccionaron pacientes que recibieron terapias de reperfusión y estaban a más de 50 Km del centro con hemodinámica. Se recogieron las características del infarto, tiempos de tratamiento y la mortalidad a corto y largo plazo. Finalmente se realizó un modelo de supervivencia. RESULTADOS: Se registraron 584 pacientes; 194 recibieron AP y 390 fibrinolisis (FL). En los pacientes sometidos a AP, la mediana del tiempo desde el primer contacto médico (PCM)-balón fue 160 minutos. Los tratados con FL presentaron un PCM-aguja de 30 minutos. La mortalidad en el seguimiento a dos años fue superior en los pacientes tratados con AP (12,2%) frente a los que recibieron FL (7,0%) (p = 0,04). Existió un aumento del riesgo de mortalidad en el grupo tratado con AP con hazard ratio (HR) de 1,97 (IC 95%: 1,04-3,70; p = 0,035). CONCLUSIONES: En los pacientes que sufren un IAMEST a más de 50 Km de un centro con hemodinámica, la reperfusión mediante AP tiene importantes retrasos y se asocia con mayor mortalidad respecto a la FL.
Assuntos
Angioplastia com Balão , Infarto do Miocárdio com Supradesnível do Segmento ST/terapia , Terapia Trombolítica , Adulto , Idoso , Idoso de 80 Anos ou mais , Cateterismo Cardíaco , Feminino , Fibrinolíticos/uso terapêutico , Acessibilidade aos Serviços de Saúde , Humanos , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Prognóstico , Modelos de Riscos Proporcionais , Estudos Prospectivos , Sistema de Registros , Infarto do Miocárdio com Supradesnível do Segmento ST/tratamento farmacológico , Tempo para o Tratamento , ViagemRESUMO
The case study analyzes the use of social media as a component of disaster response during and after the Louisiana Floods of August 2016. The study analyzes the survey responses of thirty social media users on a series of questions regarding social networks they regularly used during the flooding events, the extent to which users contacted government agencies via those networks, other uses of social media connected with the disaster, and whether social media served as a primary means of communication during cell carrier service interruptions. The results of this study show that there was a correlation between service disruption and increased use of social media as a means of communication. Additionally, the survey showed that social media networks have been utilized for a wide range of purposes during disasters, including locating family and loved ones, requesting help, disseminating information, and psychosocial interaction. Finally, a majority of respondents did not use social media to contact government agencies, and a number of respondents rated federal government engagement through social media as either dissatisfactory or were neutral on the question.
Assuntos
Desastres/estatística & dados numéricos , Inundações , Socorro em Desastres/organização & administração , Trabalho de Resgate/métodos , Mídias Sociais , Comunicação , Órgãos Governamentais , Humanos , Disseminação de Informação/métodos , Louisiana , Melhoria de Qualidade , Mídias Sociais/organização & administração , Mídias Sociais/normas , Mídias Sociais/estatística & dados numéricos , Rede Social , Inquéritos e QuestionáriosRESUMO
Freeze-fracture electron microscopy has been a major technique in ultrastructural research for over 40 years. However, the lack of effective means to study the molecular composition of membranes produced a significant decline in its use. Recently, there has been a major revival in freeze-fracture electron microscopy thanks to the development of effective ways to reveal integral membrane proteins by immunogold labeling. One of these methods is known as detergent-solubilized Freeze-fracture Replica Immunolabeling (FRIL). The combination of the FRIL technique with optogenetics allows a correlated analysis of the structural and functional properties of central synapses. Using this approach it is possible to identify and characterize both pre- and postsynaptic neurons by their respective expression of a tagged channelrhodopsin and specific molecular markers. The distinctive appearance of the postsynaptic membrane specialization of glutamatergic synapses further allows, upon labeling of ionotropic glutamate receptors, to quantify and analyze the intrasynaptic distribution of these receptors. Here, we give a step-by-step description of the procedures required to prepare paired replicas and how to immunolabel them. We will also discuss the caveats and limitations of the FRIL technique, in particular those associated with potential sampling biases. The high reproducibility and versatility of the FRIL technique, when combined with optogenetics, offers a very powerful approach for the characterization of different aspects of synaptic transmission at identified neuronal microcircuits in the brain. Here, we provide an example how this approach was used to gain insights into structure-function relationships of excitatory synapses at neurons of the intercalated cell masses of the mouse amygdala. In particular, we have investigated the expression of ionotropic glutamate receptors at identified inputs originated from the thalamic posterior intralaminar and medial geniculate nuclei. These synapses were shown to relay sensory information relevant for fear learning and to undergo plastic changes upon fear conditioning.
Assuntos
Tonsila do Cerebelo/citologia , Técnica de Fratura por Congelamento/métodos , Optogenética/métodos , Receptores de Glutamato , Animais , Camundongos , Reprodutibilidade dos Testes , SinapsesRESUMO
Optogenetic approaches are now widely used to study the function of neural populations and circuits by combining targeted expression of light-activated proteins and subsequent manipulation of neural activity by light. Channelrhodopsins (ChRs) are light-gated cation-channels and when fused to a fluorescent protein their expression allows for visualization and concurrent activation of specific cell types and their axonal projections in defined areas of the brain. Via stereotactic injection of viral vectors, ChR fusion proteins can be constitutively or conditionally expressed in specific cells of a defined brain region, and their axonal projections can subsequently be studied anatomically and functionally via ex vivo optogenetic activation in brain slices. This is of particular importance when aiming to understand synaptic properties of connections that could not be addressed with conventional electrical stimulation approaches, or in identifying novel afferent and efferent connectivity that was previously poorly understood. Here, a few examples illustrate how this technique can be applied to investigate these questions to elucidating fear-related circuits in the amygdala. The amygdala is a key region for acquisition and expression of fear, and storage of fear and emotional memories. Many lines of evidence suggest that the medial prefrontal cortex (mPFC) participates in different aspects of fear acquisition and extinction, but its precise connectivity with the amygdala is just starting to be understood. First, it is shown how ex vivo optogenetic activation can be used to study aspects of synaptic communication between mPFC afferents and target cells in the basolateral amygdala (BLA). Furthermore, it is illustrated how this ex vivo optogenetic approach can be applied to assess novel connectivity patterns using a group of GABAergic neurons in the amygdala, the paracapsular intercalated cell cluster (mpITC), as an example.
Assuntos
Tonsila do Cerebelo/fisiologia , Extinção Psicológica/fisiologia , Medo/fisiologia , Córtex Pré-Frontal/fisiologia , Animais , Axônios/fisiologia , Channelrhodopsins , Dependovirus/genética , Estimulação Elétrica , Proteínas de Fluorescência Verde/genética , Camundongos , Camundongos Endogâmicos C57BL , Optogenética/métodos , Técnicas de Patch-Clamp , Técnicas EstereotáxicasRESUMO
KEY POINTS: Throughout life, fear learning is indispensable for survival and neural plasticity in the lateral amygdala underlies this learning and storage of fear memories. During development, properties of fear learning continue to change into adulthood, but currently little is known about changes in amygdala circuits that enable these behavioural transitions. In recordings from neurons in lateral amygdala brain slices from infant up to adult mice, we show that spontaneous and evoked excitatory and inhibitory synaptic transmissions mature into adolescence. At this time, increased inhibitory activity and signalling has the ability to restrict the function of excitation by presynaptic modulation, and may thus enable precise stimulus associations to limit fear generalization from adolescence onward. Our results provide a basis for addressing plasticity mechanisms that underlie altered fear behaviour in young animals. ABSTRACT: Convergent evidence suggests that plasticity in the lateral amygdala (LA) participates in acquisition and storage of fear memory. Sensory inputs from thalamic and cortical areas activate principal neurons and local GABAergic interneurons, which provide feed-forward inhibition that tightly controls LA activity and plasticity via pre- and postsynaptic GABAA and GABAB receptors. GABAergic control is also critical during fear expression, generalization and extinction in adult animals. During rodent development, properties of fear and extinction learning continue to change into early adulthood. Currently, few studies have assessed physiological changes in amygdala circuits that may enable these behavioural transitions. To obtain first insights, we investigated changes in spontaneous and sensory input-evoked inhibition onto LA principal neurons and then focused on GABAB receptor-mediated modulation of excitatory sensory inputs in infant, juvenile, adolescent and young adult mice. We found that spontaneous and sensory-evoked inhibition increased during development. Physiological changes were accompanied by changes in dendritic morphology. While GABAB heteroreceptors were functionally expressed on sensory afferents already early in development, they could only be physiologically recruited by sensory-evoked GABA release to mediate heterosynaptic inhibition from adolescence onward. Furthermore, we found GABAB -mediated tonic inhibition of sensory inputs by ambient GABA that also emerged in adolescence. The observed increase in GABAergic drive may be a substrate for providing modulatory GABA. Our data suggest that GABAB -mediated tonic and evoked presynaptic inhibition can suppress sensory input-driven excitation in the LA to enable precise stimulus associations and limit generalization of conditioned fear from adolescence onward.
Assuntos
Tonsila do Cerebelo/fisiologia , GABAérgicos/farmacologia , Tonsila do Cerebelo/efeitos dos fármacos , Animais , Baclofeno/farmacologia , Benzilaminas/farmacologia , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Potenciais Pós-Sinápticos Inibidores/efeitos dos fármacos , Masculino , Camundongos Endogâmicos C57BL , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Ácidos Nipecóticos/farmacologia , Ácidos Fosfínicos/farmacologia , Picrotoxina/farmacologia , Propanolaminas/farmacologiaRESUMO
Increasing evidence suggests that parallel plastic processes in the amygdala involve inhibitory elements to control fear and extinction memory. GABAergic medial paracapsular intercalated cells (mpITCs) are thought to relay activity from basolateral nucleus (BLA) and prefrontal cortex to inhibit central amygdala output during suppression of fear. Recently, projection diversity and differential behavioral activation of mpITCs in distinct fear states suggest additional functions. Here, we show that mpITCs receive convergent sensory thalamic and cortical inputs that undergo fear learning-related changes and are dynamically modulated via presynaptic GABAB receptors recruited by GABA released from the mpITC network. Among mpITCs, we identify cells that inhibit but are also mutually activated by BLA principal neurons. Thus, mpITCs take part in fear learning-modulated feedforward and feedback inhibitory circuits to simultaneously control amygdala input and output nuclei. Our findings place mpITCs in a unique position to gate acquired amygdala-dependent behaviors via their direct sensory inputs.
Assuntos
Tonsila do Cerebelo/fisiologia , Medo/fisiologia , Aprendizagem/fisiologia , Receptores de GABA/metabolismo , Vias Aferentes/fisiologia , Animais , Complexo Nuclear Basolateral da Amígdala/fisiologia , Córtex Cerebral/fisiologia , Condicionamento Psicológico/fisiologia , Extinção Psicológica/fisiologia , Técnicas In Vitro , Masculino , Camundongos , Camundongos Transgênicos , Técnicas de Patch-Clamp , Núcleos Talâmicos/fisiologiaRESUMO
Prepulse inhibition (PPI) of startle occurs when intensity stimuli precede stronger startle-inducing stimuli by 10-1000 ms. PPI deficits are found in individuals with schizophrenia and other psychiatric disorders, and they correlate with other cognitive impairments. Animal research and clinical studies have demonstrated that both PPI and cognitive function can be enhanced by nicotine. PPI has been shown to be mediated, at least in part, by mesopontine cholinergic neurons that project to pontine startle neurons and activate muscarinic and potentially nicotine receptors (nAChRs). The subtypes and anatomical location of nAChRs involved in mediating and modulating PPI remain unresolved. We tested the hypothesis that nAChRs that are expressed by pontine startle neurons contribute to PPI. We also explored whether or not these pontine receptors are responsible for the nicotine enhancement of PPI. While systemic administration of nAChR antagonists had limited effects on PPI, PnC microinfusions of the non-α7nAChR preferring antagonist TMPH, but not of the α7nAChR antagonist MLA, into the PnC significantly reduced PPI. Electrophysiological recordings from startle-mediating PnC neurons confirmed that nicotine affects excitability of PnC neurons, which could be antagonized by TMPH, but not by MLA, indicating the expression of non-α7nAChR. In contrast, systemic nicotine enhancement of PPI was only reversed by systemic MLA and not by TMPH or local microinfusions of MLA into the PnC. In summary, our data indicate that non-α7nAChRs in the PnC contribute to PPI at stimulus intervals of 100 ms or less, whereas activation of α7nAChRs in other brain areas is responsible for the systemic nicotine enhancement of PPI. This is important knowledge for the correct interpretation of behavioral, preclinical, and clinical data as well as for developing drugs for the amelioration of PPI deficits and the enhancement of cognitive function.
RESUMO
Many lines of evidence suggest that a reciprocally interconnected network comprising the amygdala, ventral hippocampus (vHC), and medial prefrontal cortex (mPFC) participates in different aspects of the acquisition and extinction of conditioned fear responses and fear behavior. This could at least in part be mediated by direct connections from mPFC or vHC to amygdala to control amygdala activity and output. However, currently the interactions between mPFC and vHC afferents and their specific targets in the amygdala are still poorly understood. Here, we use an ex-vivo optogenetic approach to dissect synaptic properties of inputs from mPFC and vHC to defined neuronal populations in the basal amygdala (BA), the area that we identify as a major target of these projections. We find that BA principal neurons (PNs) and local BA interneurons (INs) receive monosynaptic excitatory inputs from mPFC and vHC. In addition, both these inputs also recruit GABAergic feedforward inhibition in a substantial fraction of PNs, in some neurons this also comprises a slow GABAB-component. Amongst the innervated PNs we identify neurons that project back to subregions of the mPFC, indicating a loop between neurons in mPFC and BA, and a pathway from vHC to mPFC via BA. Interestingly, mPFC inputs also recruit feedforward inhibition in a fraction of INs, suggesting that these inputs can activate dis-inhibitory circuits in the BA. A general feature of both mPFC and vHC inputs to local INs is that excitatory inputs display faster rise and decay kinetics than in PNs, which would enable temporally precise signaling. However, mPFC and vHC inputs to both PNs and INs differ in their presynaptic release properties, in that vHC inputs are more depressing. In summary, our data describe novel wiring, and features of synaptic connections from mPFC and vHC to amygdala that could help to interpret functions of these interconnected brain areas at the network level.
RESUMO
Inositol phosphates are key signaling molecules affecting a large variety of cellular processes. Inositol-polyphosphate multikinase (IPMK) is a central component of the inositol phosphate biosynthetic routes, playing essential roles during development. IPMK phosphorylates inositol 1,4,5-trisphosphate to inositol tetrakisphosphate and subsequently to inositol pentakisphosphate and has also been described to function as a lipid kinase. Recently, a catalytically inactive mammalian IPMK was reported to be involved in nutrient signaling by way of mammalian target of rapamycin and AMP-activated protein kinase. In yeast, the IPMK homologue, Arg82, is the sole inositol-trisphosphate kinase. Arg82 has been extensively studied as part of the transcriptional complex regulating nitrogen sensing, in particular arginine metabolism. Whether this role requires Arg82 catalytic activity has long been a matter of contention. In this study, we developed a novel method for the real time study of promoter strength in vivo and used it to demonstrate that catalytically inactive Arg82 fully restored the arginine-dependent transcriptional response. We also showed that expression in yeast of catalytically active, but structurally very different, mammalian or plant IPMK homologue failed to restore arginine regulation. Our work indicates that inositol phosphates do not regulate arginine-dependent gene expression.
Assuntos
Arginina/farmacologia , Regulação Fúngica da Expressão Gênica/efeitos dos fármacos , Fosfatos de Inositol/biossíntese , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Proteínas de Saccharomyces cerevisiae/genética , Aldeído Oxirredutases/genética , Aldeído Oxirredutases/metabolismo , Arginina/metabolismo , Biocatálise , Técnicas de Inativação de Genes , Teste de Complementação Genética , Humanos , Inositol 1,4,5-Trifosfato/metabolismo , Complexos Multienzimáticos/genética , Complexos Multienzimáticos/metabolismo , Mutação , Ornitina/metabolismo , Ornitina/farmacologia , Ornitina Carbamoiltransferase/genética , Ornitina Carbamoiltransferase/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Fosfotransferases (Aceptor do Grupo Carboxila)/genética , Fosfotransferases (Aceptor do Grupo Carboxila)/metabolismo , Regiões Promotoras Genéticas/genética , Compostos de Amônio Quaternário/metabolismo , Compostos de Amônio Quaternário/farmacologia , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crescimento & desenvolvimento , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Fatores de Tempo , Transcrição Gênica/genéticaRESUMO
Myo-inositol is present in nature either unmodified or in more complex phosphorylated derivates. Of the latest, the two most abundant in eukaryotic cells are inositol pentakisphosphate (IP(5;)) and inositol hexakisphosphate (phytic acid or IP(6;)). IP(5;) and IP(6;) are the precursors of inositol pyrophosphate molecules that contain one or more pyrophosphate bonds(1). Phosphorylation of IP(6;) generates diphoshoinositolpentakisphosphate (IP(7;) or PP-IP(5;)) and bisdiphoshoinositoltetrakisphosphate (IP(8;) or (PP)(2;)-IP(4;)). Inositol pyrophosphates have been isolated from all eukaryotic organisms so far studied. In addition, the two distinct classes of enzymes responsible for inositol pyrophosphate synthesis are highly conserved throughout evolution(2-4). The IP(6;) kinases (IP(6;)Ks) posses an enormous catalytic flexibility, converting IP(5;) and IP(6;) to PP-IP(4;) and IP(7;) respectively and subsequently, by using these products as substrates, promote the generation of more complex molecules(5,6). Recently, a second class of pyrophosphate generating enzymes was identified in the form of the yeast protein VIP(1;) (also referred as PP-IP(5;)K), which is able to convert IP(6;) to IP(7;) and IP(8;)(7,8). Inositol pyrophosphates regulate many disparate cellular processes such as insulin secretion(9), telomere length(10,11), chemotaxis(12), vesicular trafficking(13), phosphate homeostasis(14) and HIV-1 gag release(15). Two mechanisms of actions have been proposed for this class of molecules. They can affect cellular function by allosterically interacting with specific proteins like AKT(16). Alternatively, the pyrophosphate group can donate a phosphate to pre-phosphorylated proteins(17). The enormous potential of this research field is hampered by the absence of a commercial source of inositol pyrophosphates, which is preventing many scientists from studying these molecules and this new post-translational modification. The methods currently available to isolate inositol pyrophosphates require sophisticated chromatographic apparatus(18,19). These procedures use acidic conditions that might lead to inositol pyrophosphate degradation(20) and thus to poor recovery. Furthermore, the cumbersome post-column desalting procedures restrict their use to specialized laboratories. In this study we describe an undemanding method for the generation, isolation and purification of the products of the IP(6;)-kinase and PP-IP(5;)-kinases reactions. This method was possible by the ability of polyacrylamide gel electrophoresis (PAGE) to resolve highly phosphorylated inositol polyphosphates(20). Following IP(6;)K1 and PP-IP(5;)K enzymatic reactions using IP(6;) as the substrate, PAGE was used to separate the generated inositol pyrophosphates that were subsequently eluted in water.
Assuntos
Fosfatos de Inositol/química , Eletroforese em Gel de Poliacrilamida/métodos , Escherichia coli/enzimologia , Inositol/química , Inositol/metabolismo , Fosfatos de Inositol/síntese química , Fosfatos de Inositol/isolamento & purificação , Fosfatos de Inositol/metabolismo , Fosforilação , Fosfotransferases (Aceptor do Grupo Fosfato)/química , Fosfotransferases (Aceptor do Grupo Fosfato)/metabolismoRESUMO
Inorganic polyphosphate (poly-P) consists of just a chain of phosphate groups linked by high energy bonds. It is found in every organism and is implicated in a wide variety of cellular processes (e.g. phosphate storage, blood coagulation, and pathogenicity). Its metabolism has been studied mainly in bacteria while remaining largely uncharacterized in eukaryotes. It has recently been suggested that poly-P metabolism is connected to that of highly phosphorylated inositol species (inositol pyrophosphates). Inositol pyrophosphates are molecules in which phosphate groups outnumber carbon atoms. Like poly-P they contain high energy bonds and play important roles in cell signaling. Here, we show that budding yeast mutants unable to produce inositol pyrophosphates have undetectable levels of poly-P. Our results suggest a prominent metabolic parallel between these two highly phosphorylated molecules. More importantly, we demonstrate that DDP1, encoding diadenosine and diphosphoinositol phosphohydrolase, possesses a robust poly-P endopolyphosphohydrolase activity. In addition, we prove that this is an evolutionarily conserved feature because mammalian Nudix hydrolase family members, the three Ddp1 homologues in human cells (DIPP1, DIPP2, and DIPP3), are also capable of degrading poly-P.