Regulation of hnRNP A2/B1 and hnRNP K by synaptic activity and BDNF in the hippocampus

Tese de doutoramento em Biociências, apresentada à Faculdade de Ciências e Tecnologia da Universidade de CoimbraSynaptic plasticity describes the process by which connections between neurons, or synapses, change in strength. By definition, it is a functional term referring to an increase or decrease in synaptic efficacy, which is accompanied by structural changes at synapses.Long-term potentiation (LTP) is the most studied form of synaptic plasticity and it has been widely recognized that synaptic changes that underpin certain forms of learning and memory may be similar to those involved in the expression of LTP. Some of the structural, biochemical and functional modifications at the synapse associated with synaptic plasticity require activity-dependent transport and translation of dendritic-localized mRNAs, with concomitant alterations in the synaptic proteome. It is becoming evident that dendritic protein synthesis has a crucial role in several forms of synaptic plasticity, including brain-derived neurotrophic factor (BDNF)-mediated LTP. Dendritic transcripts required for local protein synthesis are transported in a repressed state along the microtubule cytoskeleton as components of large messenger ribonucleoprotein complexes (mRNPs). In order to be transported, dendritic mRNAs must contain a cis-acting element in their sequence that is recognized by specific RNA binding proteins that, together with other factors, form a functional mRNP granule that engage with motor proteins for the transport. When they reach their destination, usually in, or in the vicinity of activated synapses, the translational-block is relieved and the mRNAs are translated upon neuronal activation. Among the molecular components of neuronal mRNPs, there are several members of the heterogeneous nuclear ribonucleoprotein (hnRNP) family of proteins. hnRNP A2/B1 is one of the best described trans-acting factor involved in the transport of dendritic-localized mRNAs in neurons, but whether this is a constitutive or a regulated process is unknown. hnRNP K is another member of the hnRNP family of proteins present in neuronal mRNPs, but the function of hnRNP K in neurons, and whether this protein plays a role in dendritic mRNA metabolism, remains to be determined. Here we show that both hnRNP A2/B1 and hnRNP K accumulate in dendrites and at the synapse following neuronal activation in hippocampal neurons. Importantly, the activity-dependent delivery of hnRNP A2/B1 into synaptic sites requires BDNF. In addition, we observed that this neurotrophin also upregulates the dendritic and synaptic levels of hnRNP K and hnRNP A2/B1 in primary cultures of hippocampal neurons. Previous studies from our laboratory identified several transcripts associated with hnRNP K in cultured hippocampal neurons, including mRNAs coding for proteins with roles in synaptic plasticity such as GluA1, GluN1, BDNF and CaMKIIẞ. In addition, we demonstrated that the neurotrophin BDNF induces the release of these transcripts from the hnRNPK-containing mRNPs in hippocampal neurons. Herein we show that BDNF also promotes the dissociation of hnRNPK-bound mRNAs locally at the synapse. These results suggest a key role for hnRNP K in the local translation of several proteins that contribute to the late phase of LTP. Accordingly, we demonstrate that hnRNP K-associated mRNAs are differentially regulated during high-frequency stimulation (HFS)-induced LTP in the perforant path-dentate gyrus synapse in live anesthetized rats. Importantly, we show that this form of synaptic potentiation requires TrkB (tropomyosin-related kinase B) signaling. Although our results indicate that BDNF induces the release of the transcripts bound to mRNPs containing hnRNP K, at this time point it is not possible to rule out the contribution of other RNA-binding proteins that were found to co-immunoprecipitate with hnRNP K in a proteomic screen. Altogether, our data support the evidence available pointing to a prominent role of hnRNP A2/B1 in the delivery of transcripts into dendritic domains and suggest that this process is regulated by synaptic activity and by the neurotrophin BDNF. Furthermore, we show that BDNF is required for the activity-induced synaptic delivery of hnRNP A2/B1. We also show that hnRNP K is another hnRNP that is likely to have a major role in the regulation of local mRNA metabolism in dendrites. Ourresults show that hnRNP K and hnRNP K-associated mRNAs are regulated by synaptic activity and BDNF, both in vitro and in vivo,and suggest hnRNP K as an important regulator of neuronal function, in particularin BDNF-induced plasticity events.A plasticidade sináptica descreve um processo no qual a força da interacção entre neurónios, ou sinapses, é alterada. Por definição, é um termo funcional que se refere ao aumento ou decréscimo na eficácia sináptica, sendo acompanhado por alterações estruturais nas sinapses.A potenciação sináptica de longa duração (LTP) é a forma mais estudada de plasticidade sináptica e tem sido amplamente reconhecido que as alterações sinápticas responsáveis por certas formas de aprendizagem e memória podem ser semelhantes aquelas em que a expressão da LTP se baseia. Algumas das modificações estruturais, bioquímicas e funcionais nas sinapses associadas à plasticidade sináptica dependem do transporte e da tradução de RNAs mensageiros (mRNAs) localizados nas dendrites, em resposta à actividade sináptica, com a concomitante alteração local do proteoma. É cada vez mais evidente que a síntese de proteínas nas dendrites desempenha um papel crucial em diversas formas de plasticidade sináptica, incluindo na potenciação de longa duração mediada pelo BDNF (factor neurotrófico derivado do cérebro). Os transcritos dendríticos são transportados ao longo dos microtúbulos, numa forma em que a tradução está bloqueada, como componentes de complexos ribonucleoproteicos mensageiros (mRNPs). Para serem transportados, os mRNAs necessitam de conter na sua sequência um elemento cis-acting que é reconhecido por proteínas que ligam RNA e que, juntamente com outros factores, formam um complexo funcional que se liga a proteínas motoras para o transporte. Quando chegam ao seu destino, normalmente em sinapses ou na imediação de sinapses activadas, o bloqueio da tradução é libertado e os mRNAs são traduzidos após activação neuronal. Entre os constituintes moleculares que compõem os mRNPs em neurónios estão vários membros da família de proteínas hnRNP (ribonucleoproteinas nucleares heterogéneas).A proteína hnRNP A2/B1 é um dos mais bem descritos factores trans-acting envolvidos no transporte de mRNAs ao longo das dendrites em neurónios, contudo não se sabe se este é um processo constitutivo ou regulado. A hnRNP K também está presente em mRNPs em neurónios mas a sua função, e se desempenha algum papel no metabolismo de mRNAs nas dendrites, ainda está por determinar. Neste trabalho mostramos que tanto a hnRNP A2/B1 como a hnRNP K se acumulam nas dendrites e nas sinapses após actividade sináptica em neurónios do hipocampo em cultura. Verificou-se também que o endereçamento de hnRNP A2/B1 para a sinapse em resposta à actividade neuronal depende de BDNF. De acordo com estes resultados, a estimulação de neurónios do hipocampo com BDNF também aumentou os níveis de hnRNP K e hnRNP A2/B1 nas dendrites e nas sinapses. Estudos anteriores realizados no nosso laboratório identificaram diversos transcritos associados com a hnRNP K em neurónios do hipocampo, incluindo mRNAs que codificam proteínas importantes para a plasticidade sináptica como GluA1, GluN1, BDNF e CaMKIIẞ. De seguida verificou-se também que a neurotrofina BDNF induz a libertação desses transcritos de mRNPs que contêm hnRNP K em neurónios do hipocampo. Neste trabalho mostrámos que o BDNF também promove a dissociação dos mRNAs associados à hnRNP K localmente na sinapse. Estes resultados sugerem um papel fundamental da hnRNP K na tradução local de várias proteínas que contribuem para a fase tardia da LTP. De acordo com estas evidências experimentais, demonstrámos que os mRNAs associados à hnRNP K são regulados diferencialmente durante a potenciação de longa duração (LTP) induzida pela estimulação de alta frequência nas sinapses da via perforante-giro dentado em ratos vivos anestesiados. Demonstrámos também que esta forma de potenciação sináptica requer a activação dos receptores TrkB. Apesar de os resultados obtidos indicarem que o BDNF induz a libertação dos transcritos associados a mRNPs contendo a hnRNP K, não é possível excluir a contribuição de outras ribonucleoproteínas que co-imunoprecipitaram com a hnRNP K num estudo de proteómica efectuado. No seu conjunto, os resultados obtidos apoiam o modelo que propõe uma função para a hnRNP A2/B1 na entrega de transcritos em regiões específicas das dendrites e sugerem que este é um processo regulado pela actividade sináptica e pelo BDNF. Demonstrámos também que a acumulação de hnRNP A2/B1 na sinapse após actividade sináptica depende de BDNF. A hnRNP K é outra hnRNP que parece desempenhar um papel crucial na regulação do metabolismo do mRNA localmente nas dendrites. Os nossos resultados demonstram que a interacção entre a hnRNP K e os mRNAs a ela associados é regulada pela actividade sináptica e pelo BDNF, tanto in vitro como in vivo, o que sugere a hnRNP K como um importante regulador da função neuronal, em particular em fenómenos de plasticidade sináptica induzidos pelo BDNF

Neuronal Activity Induces Synaptic Delivery of hnRNP A2/B1 by a BDNF-Dependent Mechanism in Cultured Hippocampal Neurons

Dendritic protein synthesis plays a critical role in several forms of synaptic plasticity, including BDNF (brain-derived neurotrophic factor)-mediated long-term synaptic potentiation (LTP). Dendritic transcripts are typically transported in a repressed state as components of large ribonucleoprotein complexes, and then translated upon stimulation at, or in the vicinity, of activated synapses. Heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNP A2/B1) is a trans-acting factor involved in dendritic mRNA trafficking, but how the distribution of the protein in dendrites is regulated has not been characterized. Here we found that a fraction of hnRNP A2/B1 is present at the synapse under resting conditions in cultured hippocampal neurons. Accordingly, this ribonucleoprotein was detected in free mRNP, monosomal, and polyribosomal fractions obtained from synaptoneurosomes. Neuronal activity and BDNF treatment increased hnRNP A2/B1 protein levels in the cell body and dendritic compartments, and induced the delivery of this protein to synaptic sites. The activitydependent accumulation of hnRNP A2/B1 at the synapse required, at least in part, the activation of TrkB receptors, presumably by BDNF. This neurotrophin also upregulated the hnRNP A2/B1 mRNA in the soma but was without effect on the abundance of neuritic hnRNP A2/B1 transcripts. These results show that the distribution of hnRNP A2/B1 is regulated by BDNF and by neuronal activity, an effect that may have a role in BDNF-induced synaptic plasticity events

Noite de terror na cidade luz: os atos terroristas em Paris e a avaliação da imagem de destino por turistas brasileiros

Contemporary terrorism may be considered one of the ills of the 21st century and its actions have reached an increasing number of non-combatant targets. The extremist groups are looking for places with large circulation of people as sights to carry out their attacks. The attacks aim to reach a greater number of kafirs, term to define the unbelievers or infidels of the Islamic doctrine, or to obtain a greater attention of the media. Thus, the present study aimed to understand how Brazilian tourists, who were in Paris, perceive the interference of the night of the attacks in November 2015 in the tourist image of the city. The research was qualitative with the triangulation of data: documentary research, open questionnaires and testimonials in videos, with the use of content analysis. The results show a medium impact on tourism activity, but little importance in the image of the city of Paris in the months following the attacks, with the flow of visitors reestablished in the short term. The primary relevance of this article was to understand the impact on tourist demand after terrorist attacks from the perception of tourists under the image of the destination.El terrorismo contemporáneo puede ser considerado uno de los males del siglo XXI y sus acciones han alcanzado un número cada vez mayor de blancos no combatientes. Los grupos extremistas están buscando lugares con gran circulación de personas como puntos turísticos para realizar sus ataques. Los atentados anhelan alcanzar un mayor número de kafirs, término para definir a los incrédulos o infieles de la doctrina islámica, o obtener una mayor atención de los medios. Así, el presente estudio tuvo como objetivo entender cómo los turistas brasileños, que se encontraban en París, perciben la interferencia de la noche de los atentados en noviembre de 2015 en la imagen turística de la ciudad. La investigación fue cualitativa con la triangulación de los datos: investigación documental, cuestionarios abiertos y testimonios en videos, con el uso del análisis de contenido. Los resultados muestran un impacto mediano en la actividad turística, pero poco importante en la imagen de la ciudad de París en los meses posteriores a los atentados, siendo el flujo de visitantes restablecido a corto plazo. La relevancia primordial de este artículo fue comprender el impacto en la demanda turística post-atentados terroristas a partir de la percepción de los turistas bajo la imagen del destino.O terrorismo contemporâneo pode ser considerado um dos males do século XXI e suas ações têm atingido um número cada vez maior de alvos não combatentes. Os grupos extremistas estão à procura de lugares com grande circulação de pessoas como pontos turísticos para realizar os seus ataques. Os atentados almejam alcançar um maior número de kafirs, termo para definir os incrédulos ou infiéis da doutrina islâmica, ou obter uma maior atenção da mídia. Assim, o presente estudo objetivou entender como turistas brasileiros, que estavam em Paris, percebem a interferência da noite dos atentados em 13 e 14 de novembro de 2015 na imagem turística da cidade. A investigação foi qualitativa exploratória com a triangulação dos dados: pesquisa documental, questionários abertos e depoimentos em vídeos, com o uso da análise de conteúdo. Os resultados mostram um impacto mediano na atividade turística, porém pouco importante na imagem da cidade de Paris nos meses subsequentes aos atentados, sendo o fluxo de visitantes reestabelecido em curto prazo. A relevância primordial deste artigo foi compreender o impacto na demanda turística pós-atentados terroristas a partir da percepção dos turistas sob a imagem do destino

RELATO DE EXPERIÊNCIA NA COMUNIDADE INDÍGENA DE CATU, CANGUARETAMA/RN: os diferentes grupos humanos que formaram, convivem e conflitam no Brasil

EXPERIENCE REPORT IN THE INDIGENOUS COMMUNITY OF CATU, CANGUARETAMA/RN: the different human groups who formed, live and conflited in BrazilINFORME DE EXPERIENCIA EN COMUNIDAD INDÍGENA DE CATU, CANGUARETAMA/RN: los diferentes grupos humanos que se formaron, viven y se encuentran en conflicto en BrasilO referido trabalho foi direcionado aos estudantes dos 7º Anos A, B e C da Escola Municipal Professora Terezinha Paulino de Lima, localizada no bairro de Nossa Senhora da Apresentação, em Natal/RN, no ano de 2016. Com a premissa de ensinar História e Geografia a partir das discussões sobre os problemas da sociedade atual, procurou-se estimular o debate sobre as diversidades étnico-raciais em defesa de uma educação para a cidadania. Abordou-se sobre as características contemporâneas das comunidades indígenas potiguares e os conflitos atuais relacionados à questão da demarcação de terras, aos estereótipos e aos preconceitos decorrentes de um senso comum sobre a falsa inexistência de índios no estado do Rio Grande do Norte. Procedimentalmente, foram feitas atividades interdisciplinares, dentro da sala de aula formal e fora dela, através de uma sequência didática que, inicialmente, contou com levantamento bibliográfico acerca da temática diversidade étnico-racial em autores clássicos e nos livros didáticos, abordando-se, inclusive, manifestações culturais de grupos sociais afrodescendentes e outros - como meio de ampliação da percepção sobre as diferenças. Em seguida, houve aulas de campo na comunidade indígena de Catu, município de Canguaretama, onde os alunos obtiveram, através de relatos orais, entre outras vivências, informações sobre a história e os dilemas socioambientais dessa comunidade e ao Estuário Potengi, possibilitando análises sobre os impactos da ocupação no território potiguar. Por fim, novamente em sala de aula formal, os alunos sintetizaram suas conclusões e as compartilharam com toda a comunidade escolar, sendo avaliados como tendo alcançado os objetivos propostos para este trabalho adequadamente.Palavras-chave: Povos Índígenas; Povos Formadores do Brasil; Diversidades Étnico-raciais; Comunidade Indígena Catu.ABSTRACTThis report of experience was directed to students of the 7th Years A, B and C of the Municipal School Professor Terezinha Paulino de Lima, located in the neighborhood of Nossa Senhora da Apresentação, in Natal/RN, in the year 2016. With the premise of teaching History and Geography from the discussions about the problems of the current society, we tried to stimulate the debate on ethnic-racial diversities in defense of an education for citizenship. We spoke about the contemporary characteristics of the indigenous communities of Potiguares and the current conflicts related to the land demarcation issue, the stereotypes and the prejudices arising from a common sense about the false inexistence of Indians in the state of Rio Grande do Norte. Interdisciplinary activities were carried out, both within and outside the formal classroom, through a didactic sequence that initially had a bibliographical survey about ethnic-racial diversity in classic authors and in textbooks, including, cultural manifestations of afrodescendent social groups and others - as a means of increasing the perception about the differences. Then, there were field lessons to the indigenous community of Catu, municipality of Canguaretama, where the students obtained, through oral reports, among other experiences, information on the history and socio-environmental dilemmas of this community and the Potengi Estuary, making possible impacts of occupation in the territory of the state. Finally, again in the formal classroom, the students synthesized their conclusions and shared them with the whole school community, being evaluated as having achieved the objectives proposed for this work properly.Keywords: Indigenous Peoples; Formative Peoples of Brazil; Ethnic-racial Diversity; Indigenous Community of Catu.RESUMENEl trabajo fue dirigido a los estudiantes de los 7º A, B y C de la Escuela Municipal Profesora Terezinha Paulino de Lima, ubicada en el barrio de Nuestra Señora de la Presentación, en Natal/RN, en el año 2016. Con la premisa de enseñar Historia y Geografía a partir de las discusiones sobre los problemas de la sociedad actual, se intentó estimular el debate sobre las diversidades étnico-raciales en defensa de una educación para la ciudadanía. Se habló sobre las características contemporáneas de las comunidades indígenas potiguares y los conflictos actuales relacionados con la cuestión de la demarcación de tierras, los estereotipos y los prejuicios derivados de un sentido común sobre la falsa inexistencia de indios en el estado de Rio Grande do Norte. En el aula formal y fuera de ella, se realizaron actividades interdisciplinares, a través de una secuencia didáctica que, inicialmente, contó con levantamiento bibliográfico acerca de la temática diversidad étnico-racial en autores clásicos y en los libros didácticos, abordándose, inclusive, manifestaciones culturales de grupos sociales afrodescendientes y otros - como medio de ampliación de la percepción sobre las diferencias. A continuación, hubo clases de campo a la comunidad indígena de Catu, municipio de Canguaretama, donde los alumnos obtuvieron, a través de relatos orales, entre otras vivencias, informaciones sobre la historia y los dilemas socioambientales de esa comunidad y el Estuario Potengi, posibilitando análisis sobre los. impactos de la ocupación en el territorio potiguar. Por último, nuevamente en el aula formal, los alumnos sintetizaron sus conclusiones y las compartieron con toda la comunidad escolar, siendo evaluados como habiendo alcanzado los objetivos propuestos para este trabajo adecuadamente.Palabras clave: Pueblos Indígenas; Pueblos Originales de Brasil; La Diversidad Étnica y Racial; Comunidad Indígena Catu

Role of the Proteasome in Excitotoxicity-Induced Cleavage of Glutamic Acid Decarboxylase in Cultured Hippocampal Neurons

Glutamic acid decarboxylase is responsible for synthesizing GABA, the major inhibitory neurotransmitter, and exists in two isoforms—GAD65 and GAD67. The enzyme is cleaved under excitotoxic conditions, but the mechanisms involved and the functional consequences are not fully elucidated. We found that excitotoxic stimulation of cultured hippocampal neurons with glutamate leads to a time-dependent cleavage of GAD65 and GAD67 in the N-terminal region of the proteins, and decrease the corresponding mRNAs. The cleavage of GAD67 was sensitive to the proteasome inhibitors MG132, YU102 and lactacystin, and was also abrogated by the E1 ubiquitin ligase inhibitor UBEI-41. In contrast, MG132 and UBEI-41 were the only inhibitors tested that showed an effect on GAD65 cleavage. Excitotoxic stimulation with glutamate also increased the amount of GAD captured in experiments where ubiquitinated proteins and their binding partners were isolated. However, no evidences were found for direct GADs ubiquitination in cultured hippocampal neurons, and recombinant GAD65 was not cleaved by purified 20S or 26S proteasome preparations. Since calpains, a group of calcium activated proteases, play a key role in GAD65/67 cleavage under excitotoxic conditions the results suggest that GADs are cleaved after ubiquitination and degradation of an unknown binding partner by the proteasome. The characteristic punctate distribution of GAD65 along neurites of differentiated cultured hippocampal neurons was significantly reduced after excitotoxic injury, and the total GAD activity measured in extracts from the cerebellum or cerebral cortex at 24h postmortem (when there is a partial cleavage of GADs) was also decreased. The results show a role of the UPS in the cleavage of GAD65/67 and point out the deregulation of GADs under excitotoxic conditions, which is likely to affect GABAergic neurotransmission. This is the first time that the UPS has been implicated in the events triggered during excitotoxicity and the first molecular target of the UPS affected in this cell death process

The RNA-binding protein hnRNP K mediates the effect of BDNF on dendritic mRNA metabolism and regulates synaptic NMDA receptors in hippocampal neurons

Brain-derived neurotrophic factor (BDNF) is an important mediator of long-term synaptic potentiation (LTP) in the hippocampus. The local effects of BDNF depend on the activation of translation activity, which requires the delivery of transcripts to the synapse. In this work, we found that neuronal activity regulates the dendritic localization of the RNA-binding protein heterogeneous nuclear ribonucleoprotein K (hnRNP K) in cultured rat hippocampal neurons by stimulating BDNF-Trk signaling. Microarray experiments identified a large number of transcripts that are coimmunoprecipitated with hnRNP K, and about 60% of these transcripts are dissociated from the protein upon stimulation of rat hippocampal neurons with BDNF. In vivo studies also showed a role for TrkB signaling in the dissociation of transcripts from hnRNP K upon high-frequency stimulation (HFS) of medial perforant path-granule cell synapses of male rat dentate gyrus (DG). Furthermore, treatment of rat hippocampal synaptoneurosomes with BDNF decreased the coimmunoprecipitation of hnRNP K with mRNAs coding for glutamate receptor subunits, Ca2+- and calmodulin-dependent protein kinase IIβ (CaMKIIβ) and BDNF. Downregulation of hnRNP K impaired the BDNF-induced enhancement of NMDA receptor (NMDAR)-mediated mEPSC, and similar results were obtained upon inhibition of protein synthesis with cycloheximide. The results demonstrate that BDNF regulates specific populations of hnRNP-associated mRNAs in neuronal dendrites and suggests an important role of hnRNP K in BDNF-dependent forms of synaptic plasticity.publishe

BDNF-Live-Exon-Visualization (BLEV) Allows Differential Detection of BDNF Transcripts in vitro and in vivo

Bdnf exon-IV and exon-VI transcripts are driven by neuronal activity and are involved in pathologies related to sleep, fear or memory disorders. However, how their differential transcription translates activity changes into long-lasting network changes is elusive. Aiming to trace specifically the network controlled by exon-IV and -VI derived BDNF during activity-dependent plasticity changes, we generated a transgenic reporter mouse for BDNF-live-exon-visualization (BLEV), in which expression of Bdnf exon-IV and -VI can be visualized by co-expression of CFP and YFP. CFP and YFP expression was differentially activated and targeted in cell lines, primary cultures and BLEV reporter mice without interfering with BDNF protein synthesis. CFP and YFP expression, moreover, overlapped with BDNF protein expression in defined hippocampal neuronal, glial and vascular locations in vivo. So far, activity-dependent BDNF cannot be explicitly monitored independent of basal BDNF levels. The BLEV reporter mouse therefore provides a new model, which can be used to test whether stimulus-induced activity-dependent changes in BDNF expression are instrumental for long-lasting plasticity modifications

Experimental Study of Infill Walls with Joint Reinforcement Subjected to In-Plane Lateral Load

The results of an experimental study of four infilled frames with brick masonry walls subject to reversal cyclic lateral load are presented. The variables studied were the height to length aspect ratio of the wall and the use of joint reinforcement. The investigation was motivated by the fact that the Mexican code establishes the same specifications about the use of joint reinforcement for infill walls as for confined walls, because there is not enough experimental evidence on joint reinforced infill walls. To investigate the possible interaction of the study variables in the seismic performance of the walls, two pairs of specimens, scaled 1:2, with different aspect ratios (H/L = 0.75, 0.41) were tested. The specimens in each pair were identical except that one of them included steel bars into the bed-joints as reinforcement leading to amount  phfyh=0.6 MPa. The infill walls with H/L = 0.41 were included from a previous study. The behavior of the specimens was defined in terms of lateral strength, ductility, displacement capacity, deformation of the joint reinforcement and crack pattern. The results indicate that joint reinforcement increases the strength of the system; however, the increase was more pronounced in longer walls. Ductility was reduced with horizontal reinforcement and this behavior was more important for longer walls. As occurred in confined walls, the joint reinforcement generates a more distributed cracking and reduces the width of the cracks. The experiments are described and this and other results are discussed in detail

BDNF upregulates hnRNP A2/B1 mRNA in the cell body compartment of hippocampal neurons.

<p>Cultured hippocampal neurons were stimulated or not with 100 ng/ml BDNF, for 30 min or 2 h. The cell body mRNA was mechanically separated from the transcripts of neurites and 500–1000 ng of RNA from each compartment was used in the reverse transcription reaction. The analysis of hnRNP A2/B1 mRNA levels was performed by qRT-PCR using <i>Ppia</i> as internal control gene. The results are the average ± SEM of five (cell body compartment) or seven (neurite compartment) independent transcription reactions, performed in distinct preparations. Statistical analysis of logtransformed expression data was performed by one-way ANOVA, followed by Dunnet's test. * P<0.05.</p