Effect of a Temperature Increase in the Non-Noxious Range on Proton-Evoked ASIC and TRPV1 Activity

Anais do II Seminário Seminário Estadual PIBID do Paraná: tecendo saberes / organizado por Dulcyene Maria Ribeiro e Catarina Costa Fernandes — Foz do Iguaçu: Unioeste; Unila, 2014Este trabalho tem como objetivo apresentar de que forma foi elaborada e aplicada uma sequência didática, produzida por alunos bolsistas do PIBID LI da UENP, Campus de Cornélio Procópio, que objetiva o aprimoramento da oralidade da língua inglesa. Trazemos como referencial teórico as Diretrizes Curriculares da Educação do Paraná (DCE), bem como autores que foram trabalhados e discutidos nos encontros semanais no programa PIBID e, que embasaram a construção das sequências didáticas, tais como Dolz e Schneuwly (2004), Cristovão (2001) e Petreche (2009

Protonation controls ASIC1a activity via coordinated movements in multiple domains.

Acid-sensing ion channels (ASICs) are neuronal Na(+)-conducting channels activated by extracellular acidification. ASICs are involved in pain sensation, expression of fear, and neurodegeneration after ischemic stroke. Functional ASICs are composed of three identical or homologous subunits, whose extracellular part has a handlike structure. Currently, it is unclear how protonation of residues in extracellular domains controls ASIC activity. Knowledge of these mechanisms would allow a rational development of drugs acting on ASICs. Protonation may induce conformational changes that control the position of the channel gate. We used voltage-clamp fluorometry with fluorophores attached to residues in different domains of ASIC1a to detect conformational changes. Comparison of the timing of fluorescence and current signals identified residues involved in movements that preceded desensitization and may therefore be associated with channel opening or early steps leading to desensitization. Other residues participated in movements intimately linked to desensitization and recovery from desensitization. Fluorescence signals of all mutants were detected at more alkaline pH than ionic currents. Their midpoint of pH dependence was close to that of steady-state desensitization, whereas the steepness of the pH fluorescence relationship was closer to that of current activation. A sequence of movements was observed upon acidification, and its backward movements during recovery from desensitization occurred in the reverse order, indicating that the individual steps are interdependent. Furthermore, the fluorescence signal of some labeled residues in the finger domain was strongly quenched by a Trp residue in the neighboring β-ball domain. Upon channel activation, their fluorescence intensity increased, indicating that the finger moved away from the β ball. This extensive analysis of activity-dependent conformational changes in ASICs sheds new light on the mechanisms by which protonation controls ASIC activity

Simultaneous OH-PLIF and schlieren imaging of flame acceleration in an obstacle-laden channel

Flame acceleration in stoichiometric H_2/O_2 at 12 and 25 kPa initial pressure in an obstacle-laden square cross-section channel was studied experimentally using planar laser-induced fluorescence imaging of hydroxyl radicals (OH-PLIF) and simultaneous high-speed schlieren imaging. Results were obtained resolving the explosion front structure as it develops immediately after ignition as a slow-flame to the eventual formation of a shock-flame complex in the fast-flame regime. The images provide a novel level of detail and allow for the determination of the effects of turbulence-flame and shock-flame interaction. In the slow-flame regime, vortex shedding off obstacle edges occurs over long time-scales, vortices are convected downstream and turbulent combustion takes place in the obstacle wakes. The fast-flame regime is marked by the presence of compression waves (and shock waves) which interact with the flame and cause macroscopic deformation of the flame and small-scale wrinkling due to Richtmyer-Meshkov instability. A quasi-steady fast-flame is characterized by the close proximity of the precursor shock and the turbulent flame. The flow-field that governs the flame shape is established impulsively by the precursor shock. Shock-flame interactions lead to flame front perturbations on both small and large scales. The OH-PLIF technique makes it possible to discern the flame front from other density interfaces that appear in the complex fast-flame structure observed in schlieren images and also eliminates the line-of-sight integration limitation

Extracellular Subunit Interactions Control Transitions between Functional States of Acid-sensing Ion Channel 1a.

Acid-sensing ion channels (ASICs) are neuronal, voltage-independent Na(+) channels that are transiently activated by extracellular acidification. They are involved in pain sensation, the expression of fear, and in neurodegeneration after ischemic stroke. Our study investigates the role of extracellular subunit interactions in ASIC1a function. We identified two regions involved in critical intersubunit interactions. First, formation of an engineered disulfide bond between the palm and thumb domains leads to partial channel closure. Second, linking Glu-235 of a finger loop to either one of two different residues of the knuckle of a neighboring subunit opens the channel at physiological pH or disrupts its activity. This suggests that one finger-knuckle disulfide bond (E235C/K393C) sets the channel in an open state, whereas the other (E235C/Y389C) switches the channel to a non-conducting state. Voltage-clamp fluorometry experiments indicate that both the finger loop and the knuckle move away from the β-ball residue Trp-233 during acidification and subsequent desensitization. Together, these observations reveal that ASIC1a opening is accompanied by a distance increase between adjacent thumb and palm domains as well as a movement of Glu-235 relative to the knuckle helix. Our study identifies subunit interactions in the extracellular loop and shows that dynamic changes of these interactions are critical for normal ASIC function

Assessment of lung density in pediatric patients using three-dimensional ultrashort echo-time and four-dimensional zero echo-time sequences.

PURPOSE Lung magnetic resonance imaging (MRI) using conventional sequences is limited due to strong signal loss by susceptibility effects of aerated lung. Our aim is to assess lung signal intensity in children on ultrashort echo-time (UTE) and zero echo-time (ZTE) sequences. We hypothesize that lung signal intensity can be correlated to lung physical density. MATERIALS AND METHODS Lung MRI was performed in 17 children with morphologically normal lungs (median age: 4.7 years, range 15 days to 17 years). Both lungs were manually segmented in UTE and ZTE images and the average signal intensities were extracted. Lung-to-background signal ratios (LBR) were compared for both sequences and between both patient groups using non-parametric tests and correlation analysis. Anatomical region-of-interest (ROI) analysis was performed for the normal cohort for assessment of the anteroposterior lung gradient. RESULTS There was no significant difference between LBR of normal lungs using UTE and ZTE (p < 0.05). Both sequences revealed a LBR age-dependency with a high negative correlation for UTE (Rs =  - 0.77; range 2.98-1.41) and ZTE (Rs =  - 0.82; range 2.66-1.38)). Signal-to-noise (SNR) and contrast-to-noise ratios (CNR) were age-dependent for both sequences. SNR was higher for children up to 2 years old with 3D UTE Cones while for the rest it was higher with 4D ZTE. CNR was similar for both sequences. Posterior lung areas exhibited higher signal intensity compared to anterior ones (UTE 9.4% and ZTE 12% higher), both with high correlation coefficients (R2UTE = 0.94, R2ZTE = 0.97). CONCLUSION The ZTE sequence can measure signal intensity similarly to UTE in pediatric patients. Both sequences reveal an age- and gravity-dependency of LBR

Conformational dynamics and role of the acidic pocket in ASIC pH-dependent gating.

Acid-sensing ion channels (ASICs) are proton-activated Na &lt;sup&gt;+&lt;/sup&gt; channels expressed in the nervous system, where they are involved in learning, fear behaviors, neurodegeneration, and pain sensation. In this work, we study the role in pH sensing of two regions of the ectodomain enriched in acidic residues: the acidic pocket, which faces the outside of the protein and is the binding site of several animal toxins, and the palm, a central channel domain. Using voltage clamp fluorometry, we find that the acidic pocket undergoes conformational changes during both activation and desensitization. Concurrently, we find that, although proton sensing in the acidic pocket is not required for channel function, it does contribute to both activation and desensitization. Furthermore, protonation-mimicking mutations of acidic residues in the palm induce a dramatic acceleration of desensitization followed by the appearance of a sustained current. In summary, this work describes the roles of potential pH sensors in two extracellular domains, and it proposes a model of acidification-induced conformational changes occurring in the acidic pocket of ASIC1a

Simultaneous OH-PLIF and schlieren imaging of flame acceleration in an obstacle-laden channel

Flame acceleration in stoichiometric H_2/O_2 at 12 and 25 kPa initial pressure in an obstacle-laden square cross-section channel was studied experimentally using planar laser-induced fluorescence imaging of hydroxyl radicals (OH-PLIF) and simultaneous high-speed schlieren imaging. Results were obtained resolving the explosion front structure as it develops immediately after ignition as a slow-flame to the eventual formation of a shock-flame complex in the fast-flame regime. The images provide a novel level of detail and allow for the determination of the effects of turbulence-flame and shock-flame interaction. In the slow-flame regime, vortex shedding off obstacle edges occurs over long time-scales, vortices are convected downstream and turbulent combustion takes place in the obstacle wakes. The fast-flame regime is marked by the presence of compression waves (and shock waves) which interact with the flame and cause macroscopic deformation of the flame and small-scale wrinkling due to Richtmyer-Meshkov instability. A quasi-steady fast-flame is characterized by the close proximity of the precursor shock and the turbulent flame. The flow-field that governs the flame shape is established impulsively by the precursor shock. Shock-flame interactions lead to flame front perturbations on both small and large scales. The OH-PLIF technique makes it possible to discern the flame front from other density interfaces that appear in the complex fast-flame structure observed in schlieren images and also eliminates the line-of-sight integration limitation

ASIC1a affects hypothalamic signaling and regulates the daily rhythm of body temperature in mice

The body temperature of mice is higher at night than during the day. We show here that global deletion of acid-sensing ion channel 1a (ASIC1a) results in lower body temperature during a part of the night. ASICs are pH sensors that modulate neuronal activity. The deletion of ASIC1a decreased the voluntary activity at night of mice that had access to a running wheel but did not affect their spontaneous activity. Daily rhythms of thyrotropin-releasing hormone mRNA in the hypothalamus and of thyroid-stimulating hormone β mRNA in the pituitary, and of prolactin mRNA in the hypothalamus and pituitary were suppressed in ASIC1a-/- mice. The serum thyroid hormone levels were however not significantly changed by ASIC1a deletion. Our findings indicate that ASIC1a regulates activity and signaling in the hypothalamus and pituitary. This likely leads to the observed changes in body temperature by affecting the metabolism or energy expenditure

Optimizing the reliability and resource efficiency of MapReduce-based systems

Debido al gran incremento de datos digitales que ha tenido lugar en los últimos años, ha surgido un nuevo paradigma de computación paralela para el procesamiento eficiente de grandes volúmenes de datos. Muchos de los sistemas basados en este paradigma, también llamados sistemas de computación intensiva de datos, siguen el modelo de programación de Google MapReduce. La principal ventaja de los sistemas MapReduce es que se basan en la idea de enviar la computación donde residen los datos, tratando de proporcionar escalabilidad y eficiencia. En escenarios libres de fallo, estos sistemas generalmente logran buenos resultados. Sin embargo, la mayoría de escenarios donde se utilizan, se caracterizan por la existencia de fallos. Por tanto, estas plataformas suelen incorporar características de tolerancia a fallos y fiabilidad. Por otro lado, es reconocido que las mejoras en confiabilidad vienen asociadas a costes adicionales en recursos. Esto es razonable y los proveedores que ofrecen este tipo de infraestructuras son conscientes de ello. No obstante, no todos los enfoques proporcionan la misma solución de compromiso entre las capacidades de tolerancia a fallo (o de manera general, las capacidades de fiabilidad) y su coste. Esta tesis ha tratado la problemática de la coexistencia entre fiabilidad y eficiencia de los recursos en los sistemas basados en el paradigma MapReduce, a través de metodologías que introducen el mínimo coste, garantizando un nivel adecuado de fiabilidad. Para lograr esto, se ha propuesto: (i) la formalización de una abstracción de detección de fallos; (ii) una solución alternativa a los puntos únicos de fallo de estas plataformas, y, finalmente, (iii) un nuevo sistema de asignación de recursos basado en retroalimentación a nivel de contenedores. Estas contribuciones genéricas han sido evaluadas tomando como referencia la arquitectura Hadoop YARN, que, hoy en día, es la plataforma de referencia en la comunidad de los sistemas de computación intensiva de datos. En la tesis se demuestra cómo todas las contribuciones de la misma superan a Hadoop YARN tanto en fiabilidad como en eficiencia de los recursos utilizados. ABSTRACT Due to the increase of huge data volumes, a new parallel computing paradigm to process big data in an efficient way has arisen. Many of these systems, called dataintensive computing systems, follow the Google MapReduce programming model. The main advantage of these systems is based on the idea of sending the computation where the data resides, trying to provide scalability and efficiency. In failure-free scenarios, these frameworks usually achieve good results. However, these ones are not realistic scenarios. Consequently, these frameworks exhibit some fault tolerance and dependability techniques as built-in features. On the other hand, dependability improvements are known to imply additional resource costs. This is reasonable and providers offering these infrastructures are aware of this. Nevertheless, not all the approaches provide the same tradeoff between fault tolerant capabilities (or more generally, reliability capabilities) and cost. In this thesis, we have addressed the coexistence between reliability and resource efficiency in MapReduce-based systems, looking for methodologies that introduce the minimal cost and guarantee an appropriate level of reliability. In order to achieve this, we have proposed: (i) a formalization of a failure detector abstraction; (ii) an alternative solution to single points of failure of these frameworks, and finally (iii) a novel feedback-based resource allocation system at the container level. Finally, our generic contributions have been instantiated for the Hadoop YARN architecture, which is the state-of-the-art framework in the data-intensive computing systems community nowadays. The thesis demonstrates how all our approaches outperform Hadoop YARN in terms of reliability and resource efficiency