Direct In Vivo Manipulation and Imaging of Calcium Transients in Neutrophils Identify a Critical Role for Leading-Edge Calcium Flux.

Calcium signaling has long been associated with key events of immunity, including chemotaxis, phagocytosis, and activation. However, imaging and manipulation of calcium flux in motile immune cells in live animals remain challenging. Using light-sheet microscopy for in vivo calcium imaging in zebrafish, we observe characteristic patterns of calcium flux triggered by distinct events, including phagocytosis of pathogenic bacteria and migration of neutrophils toward inflammatory stimuli. In contrast to findings from ex vivo studies, we observe enriched calcium influx at the leading edge of migrating neutrophils. To directly manipulate calcium dynamics in vivo, we have developed transgenic lines with cell-specific expression of the mammalian TRPV1 channel, enabling ligand-gated, reversible, and spatiotemporal control of calcium influx. We find that controlled calcium influx can function to help define the neutrophil's leading edge. Cell-specific TRPV1 expression may have broad utility for precise control of calcium dynamics in other immune cell types and organisms.

Experience-induced preference for oviposition repellents derived from a non-host plant by a specialist herbivore

Foraging adults of phytophagous insects are attracted by host-plant volatiles and supposedly repelled by volatiles from non-host plants. In behavioural control of pest insects, chemicals derived from non-host plants applied to crops are expected to repel searching adults and thereby reduce egg laying. How experience by searching adults of non-host volatiles affects their subsequent searching and oviposition behaviour has been rarely tested. In laboratory experiments, we examined the effect of experience of a non-host-plant extract on the oviposition behaviour of the diamondback moth (DBM), Plutella xylostella, a specialist herbivore of cruciferous plants. Naive ovipositing DBM females were repelled by an extract of dried leaves of Chrysanthemum morifolium, a non-host plant of DBM, but experienced females were not repelled. Instead they were attracted by host plants treated with the non-host-plant extract and laid a higher proportion of eggs on treated than on untreated host plants. Such behavioural changes induced by experience could lead to host-plant range expansion in phytophagous insects and play an important role in determining outcome for pest management of some behavioural manipulation methods.

Mycetoma caused by Nocardia brasiliensis in an immunocompetent patient

Dendritic cells play a central role in the regulation of immunological reactivity. The existence of functionally specialized populations of dendritic cells in the skin is a consequence of qualitatively different attacks on our organism. slanDCs are human inflammatory dendritic cells that are characterized by the specific expression of the carbohydrate 6-sulfo LacNAc (slan). After phenotypic maturation, slanDCs are capable of producing very high amounts of proinflammatory mediators such as IL-12, TNF-, IL-1 and IL-23. Recent data describe a potential role of slanDCs in a number of different diseases like psoriasis, lupus erythematosus, and tumors, thus opening up new areas of research on their respective pathogenesis. Furthermore, a slanDC-specific targeting system has been developed as a basis for direct therapeutic manipulation. Future challenges of slanDC research include deepening our understanding of the significance of slanDCs in the regulation of adaptive and innate immune responses, as well as translating this knowledge into therapeutic options.

Influence of manure scrapers on dairy cows in cubicle housing systems

Manure scrapers are widely used in dairy cow loose-housing systems. In order to evaluate the effects of the scrapers on the cows, we assessed their impact on the animals' cardiac activity, feeding behaviour, and the behavioural reactions of cows confronted with different types of scrapers. In part I of the study, we measured cardiac activity (mean R–R interval and RMSSD, a parameter of heart-rate variability) whilst observing the behaviour of 29 focal cows on three farms during situations with and without active manure scrapers. Lower RMSSD values were observed during scraping events while cows were either lying, standing or walking in the alleyway, standing completely in the lying cubicle, or standing half in the lying cubicle (P=0.03), but only tended to differ while directly confronted with the scraper (P=0.06). This indicates that dairy cows experienced at least some mild stress during manure-scraping events. In part II, the feeding behaviour of 12 cows on each of two farms was recorded by means of a jaw-movement sensor and compared between situations with the manure-scraping event following forage provision either within or outside the main daily feeding period (i.e. within 1 or after 2 h from forage provisioning, respectively). The duration of night-time feeding (P=0.049) and the number of feeding bouts (P=0.036) were higher when a manure-scraping event took place within the main daily feeding period, indicating that the cows' feeding behaviour had been disturbed. In part III, we observed the cows' behaviour on 15 farms during eight manure scraping events per farm, where each of five farms had one of three different scraper types. We assessed the cows' immediate reactions when confronted with the scraper. In addition, we recorded the number of animals present in the alleyways before and after the manure-scraping events. The more cows that were present in the alleyways before the scraping event, the lower the proportion of cows showing direct behavioural reactions both with (P=0.017) and without (P=0.028) scraper contact, and the higher the number of cows that left the alleyways (P<0.001). Scraper type did not influence the proportion of cows showing behavioural reactions. In conclusion, our results show that dairy cows perceive the manure-scraping event negatively in some situations, that feeding behaviour may be disturbed when scrapers are active during the main feeding period, and that cows avoid the scraper during crowded situations.

Desarrollo de tecnologías de telemanipulación con alto grado de escalado orientadas a la interacción hombre-robot en entornos nucleares

Esta tesis se centra en desarrollo de tecnologías para la interacción hombre-robot en entornos nucleares de fusión. La problemática principal del sector de fusión nuclear radica en las condiciones ambientales tan extremas que hay en el interior del reactor, y la necesidad de que los equipos cumplan requisitos muy restrictivos para poder aguantar esos niveles de radiación, magnetismo, ultravacío, temperatura... Como no es viable la ejecución de tareas directamente por parte de humanos, habrá que utilizar dispositivos de manipulación remota para llevar a cabo los procesos de operación y mantenimiento. En las instalaciones de ITER es obligatorio tener un entorno controlado de extrema seguridad, que necesita de estándares validados. La definición y uso de protocolos es indispensable para regir su buen funcionamiento. Si nos centramos en la telemanipulación con algo grado de escalado, surge la necesidad de definir protocolos para sistemas abiertos que permitan la interacción entre equipos y dispositivos de diversa índole. En este contexto se plantea la definición del Protocolo de Teleoperación que permita la interconexión entre dispositivos maestros y esclavos de distinta tipología, pudiéndose comunicar bilateralmente entre sí y utilizar distintos algoritmos de control según la tarea a desempeñar. Este protocolo y su interconectividad se han puesto a prueba en la Plataforma Abierta de Teleoperación (P.A.T.) que se ha desarrollado e integrado en la ETSII UPM como una herramienta que permita probar, validar y realizar experimentos de telerrobótica. Actualmente, este Protocolo de Teleoperación se ha propuesto a través de AENOR al grupo ISO de Telerobotics como una solución válida al problema existente y se encuentra bajo revisión. Con el diseño de dicho protocolo se ha conseguido enlazar maestro y esclavo, sin embargo con los niveles de radiación tan altos que hay en ITER la electrónica del controlador no puede entrar dentro del tokamak. Por ello se propone que a través de una mínima electrónica convenientemente protegida se puedan multiplexar las señales de control que van a través del cableado umbilical desde el controlador hasta la base del robot. En este ejercicio teórico se demuestra la utilidad y viabilidad de utilizar este tipo de solución para reducir el volumen y peso del cableado umbilical en cifras aproximadas de un 90%, para ello hay que desarrollar una electrónica específica y con certificación RadHard para soportar los enormes niveles de radiación de ITER. Para este manipulador de tipo genérico y con ayuda de la Plataforma Abierta de Teleoperación, se ha desarrollado un algoritmo que mediante un sensor de fuerza/par y una IMU colocados en la muñeca del robot, y convenientemente protegidos ante la radiación, permiten calcular las fuerzas e inercias que produce la carga, esto es necesario para poder transmitirle al operador unas fuerzas escaladas, y que pueda sentir la carga que manipula, y no otras fuerzas que puedan influir en el esclavo remoto, como ocurre con otras técnicas de estimación de fuerzas. Como el blindaje de los sensores no debe ser grande ni pesado, habrá que destinar este tipo de tecnología a las tareas de mantenimiento de las paradas programadas de ITER, que es cuando los niveles de radiación están en sus valores mínimos. Por otro lado para que el operador sienta lo más fielmente posible la fuerza de carga se ha desarrollado una electrónica que mediante el control en corriente de los motores permita realizar un control en fuerza a partir de la caracterización de los motores del maestro. Además para aumentar la percepción del operador se han realizado unos experimentos que demuestran que al aplicar estímulos multimodales (visuales, auditivos y hápticos) aumenta su inmersión y el rendimiento en la consecución de la tarea puesto que influyen directamente en su capacidad de respuesta. Finalmente, y en referencia a la realimentación visual del operador, en ITER se trabaja con cámaras situadas en localizaciones estratégicas, si bien el humano cuando manipula objetos hace uso de su visión binocular cambiando constantemente el punto de vista adecuándose a las necesidades visuales de cada momento durante el desarrollo de la tarea. Por ello, se ha realizado una reconstrucción tridimensional del espacio de la tarea a partir de una cámara-sensor RGB-D, lo cual nos permite obtener un punto de vista binocular virtual móvil a partir de una cámara situada en un punto fijo que se puede proyectar en un dispositivo de visualización 3D para que el operador pueda variar el punto de vista estereoscópico según sus preferencias. La correcta integración de estas tecnologías para la interacción hombre-robot en la P.A.T. ha permitido validar mediante pruebas y experimentos para verificar su utilidad en la aplicación práctica de la telemanipulación con alto grado de escalado en entornos nucleares de fusión. Abstract This thesis focuses on developing technologies for human-robot interaction in nuclear fusion environments. The main problem of nuclear fusion sector resides in such extreme environmental conditions existing in the hot-cell, leading to very restrictive requirements for equipment in order to deal with these high levels of radiation, magnetism, ultravacuum, temperature... Since it is not feasible to carry out tasks directly by humans, we must use remote handling devices for accomplishing operation and maintenance processes. In ITER facilities it is mandatory to have a controlled environment of extreme safety and security with validated standards. The definition and use of protocols is essential to govern its operation. Focusing on Remote Handling with some degree of escalation, protocols must be defined for open systems to allow interaction among different kind of equipment and several multifunctional devices. In this context, a Teleoperation Protocol definition enables interconnection between master and slave devices from different typologies, being able to communicate bilaterally one each other and using different control algorithms depending on the task to perform. This protocol and its interconnectivity have been tested in the Teleoperation Open Platform (T.O.P.) that has been developed and integrated in the ETSII UPM as a tool to test, validate and conduct experiments in Telerobotics. Currently, this protocol has been proposed for Teleoperation through AENOR to the ISO Telerobotics group as a valid solution to the existing problem, and it is under review. Master and slave connection has been achieved with this protocol design, however with such high radiation levels in ITER, the controller electronics cannot enter inside the tokamak. Therefore it is proposed a multiplexed electronic board, that through suitable and RadHard protection processes, to transmit control signals through an umbilical cable from the controller to the robot base. In this theoretical exercise the utility and feasibility of using this type of solution reduce the volume and weight of the umbilical wiring approximate 90% less, although it is necessary to develop specific electronic hardware and validate in RadHard qualifications in order to handle huge levels of ITER radiation. Using generic manipulators does not allow to implement regular sensors for force feedback in ITER conditions. In this line of research, an algorithm to calculate the forces and inertia produced by the load has been developed using a force/torque sensor and IMU, both conveniently protected against radiation and placed on the robot wrist. Scaled forces should be transmitted to the operator, feeling load forces but not other undesirable forces in slave system as those resulting from other force estimation techniques. Since shielding of the sensors should not be large and heavy, it will be necessary to allocate this type of technology for programmed maintenance periods of ITER, when radiation levels are at their lowest levels. Moreover, the operator perception needs to feel load forces as accurate as possible, so some current control electronics were developed to perform a force control of master joint motors going through a correct motor characterization. In addition to increase the perception of the operator, some experiments were conducted to demonstrate applying multimodal stimuli (visual, auditory and haptic) increases immersion and performance in achieving the task since it is directly correlated with response time. Finally, referring to the visual feedback to the operator in ITER, it is usual to work with 2D cameras in strategic locations, while humans use binocular vision in direct object manipulation, constantly changing the point of view adapting it to the visual needs for performing manipulation during task procedures. In this line a three-dimensional reconstruction of non-structured scenarios has been developed using RGB-D sensor instead of cameras in the remote environment. Thus a mobile virtual binocular point of view could be generated from a camera at a fixed point, projecting stereoscopic images in 3D display device according to operator preferences. The successful integration of these technologies for human-robot interaction in the T.O.P., and validating them through tests and experiments, verify its usefulness in practical application of high scaling remote handling at nuclear fusion environments.

Measuring impacts of an holistic farm business management training program

A case study approach within an action research framework incorporating qualitative and quantitative domains was adopted to explore the impact on Queensland farmers of a farm business management extension programme. Three new indices were developed to quantify changes perceived by participants. The first measure, the Bennett Change Index, provided statistically significant evidence that attitudinal and behavioural changes were more frequent in participants with less formal education, but also more frequent in participants who had high urbanisation and self-directed learning index scores. The other 2 new indices, Management Constructs Change and Management Objectives Change, provided evidence of statistically significant changes in participant beliefs about, and attitudes towards, farm business management. Although highly correlated with each other, these changes were unrelated statistically to any of 6 other commonly used biographical or psychometric indices employed; including level of formal education. It is concluded that these new measures, with context-relevant modifications, have potential as aids to programme impact evaluation in a range of agricultural and wider applications. They may provide insights into personal psychological issues that complement direct behavioural measures of change.

Gamma-hydroxybutyrate does not maintain self-administration but induces conditioned place preference when injected in the ventral tegmental area

Gamma-hydroxybutyric acid (GHB) is an endogenous brain substance that has diverse neuropharmacological actions, including rewarding properties in different animal species and in humans. As other drugs of abuse, GHB affects the firing of ventral tegmental neurons (VTA) in anaesthetized animals and hyperpolarizes dopaminergic neurons in VTA slices. However, no direct behavioural data on the effects of GHB applied in the VTA or in the target regions of its dopaminergic neurons, e.g. the nucleus accumbens (NAc), are available. Here, we investigated the effects of various doses of intravenous GHB in maintaining self-administration (from 0.001 to 10 mg/kg per infusion), and its ability to induce conditioned place preference (CPP) in rats when given orally (175-350 mg/kg) or injected directly either in the VTA or NAc (from 10 to 300 microg/0.5 microl per side). Our results indicate that while only 0.01 mg/kg per infusion GHB maintained self-administration, although not on every test day, 350 mg/kg GHB given orally induced CPP. CPP was also observed when GHB was injected in the VTA (30-100 microg/0.5 microl per side) but not in the NAc. Together with recent in-vitro findings, these results suggest that the rewarding properties of GHB mainly occur via disinhibition of VTA dopaminergic neurons.

Deliberate and spontaneous sensations of disembodiment:capacity or flaw?

Introduction: Hallucinations that involve shifts in the subjectively experienced location of the self, have been termed “out-of-body experiences” (OBEs). Early psychiatric accounts cast OBEs as a specific instance of depersonalisation and derealisation disorder (DPD-DR). However, during feelings of alienation and lack of body realism in DPD-DR the self is experienced within the physical body. Deliberate forms of “disembodiment” enable humans to imagine another’s visuo-spatial perspective taking (VPT), thus, if a strong relationship between deliberate and spontaneous forms of disembodiment could be revealed, then uncontrolled OBEs could be “the other side of the coin” of a uniquely human capacity. Methods: We present a narrative review of behavioural and neuroimaging work emphasising methodological and theoretical aspects of OBE and VPT research and a potential relationship. Results: Results regarding a direct behavioural relationship between VPT and OBE are mixed and we discuss reasons by pointing out the importance of using realistic tasks and recruiting genuine OBEers instead of general DPD-DR patients. Furthermore, we review neuroimaging evidence showing overlapping neural substrates between VPT and OBE, providing a strong argument for a relationship between the two processes. Conclusions: We conclude that OBE should be regarded as a necessary implication of VPT ability in humans, or even as a necessary and potentially sufficient condition for the evolution of VPT.

Validation of a new method for human nerve decellularization: toward a new tool in peripheral nerve reconstructive surgery.

Defects of the peripheral nervous system are extremely frequent in trauma and surgeries and have high socioeconomic costs. In case of peripheral nerve injury, the first approach is primary neurorrhaphy, which is direct nerve repair with epineural microsutures of the two stumps. However, this is not feasible in case of stump retraction or in case of tissue loss (gap > 2 cm), where the main surgical options are autologous grafts, allogenic grafts, or nerve conduits. While the gold standard is the autograft, it has disadvantages related to its harvesting, with an inevitable donor site morbidity and functional deficit. Fresh nerve allografts have therefore become a viable alternative option, but they require immunosuppression, which is often contraindicated. Acellular Nerve Allografts (ANA) represent a valid alternative, they do not need immunosuppression and appear to be safe and effective based on recent studies. The purpose of this study is to propose and develop an innovative method of nerve decellularization (Rizzoli method), conforming to cleanroom requirements in order to perform the direct tissue manipulation step and the nerve decellularization process within five hours, so as to accelerate the detachment of myelin and cellular debris, without detrimental effects on nerve architecture. In this study, the safety and the efficacy of the new method are evaluated in vitro and in vivo by histological, immunohistochemical, and histomorphometric studies in rabbits and humans. The new method is rapid, safe, and cheaper if compared with available commercial ANAs. The present study shows that the method, previously optimized in vitro and in vivo on animal model presented by our group, can be applied on human nerve samples. This work represents the first step in providing a novel, safe, and inexpensive tool for use by European tissue banks to democratize the use of nerve tissue transplantation for nerve injury reconstruction.

Endosperm balance number manipulation for direct in vivo germplasm introgression to potato from a sexually isolated relative (Solanum commersonii Dun.)

Diploid (2n = 2x = 24) Solanum species with endosperm balance number (EBN) = 1 are sexually isolated from diploid 2EBN species and both tetraploid (2n = 4x = 48, 4EBN) and haploid (2n = 2x = 24, 2EBN) S. tuberosum Group Tuberosum. To sexually overcome these crossing barriers in the diploid species S. commersonii (1EBN), the manipulation of the EBN was accomplished by scaling up and down ploidy levels. Triploid F1 hybrids between an in vitro-doubled clone of S. commersonii (2n = 4x = 48, 2EBN) and diploid 2EBN clones were successfully used in 3x × 4x crosses with S. tuberosum Group Tuberosum, resulting in pentaploid/near pentaploid BC1 progenies. This provided evidence of 2n (3x) egg formation in the triploid female parents. Two selected BC1 pentaploid hybrids were successfully backcrossed both as male and as female parents with S. tuberosum Group Tuberosum. The somatic chromosome number varied greatly among the resulting BC2 progenies, which included hyperaneuploids, but also a number (4.8%) of 48-chromosome plants. The introgression of S. commersonii genomes was confirmed by the presence of S. commersonii-specific randomly amplified polymorphic DNA markers in the BC2 population analyzed. The results clearly demonstrate the feasibility of germplasm introgression from sexually isolated diploid 1EBN species into the 4x (4EBN) gene pool of the cultivated potato using sexual hybridization. Based on the amount and type of genetic variation generated, cumbersomeness, general applicability, costs, and other factors, it would be interesting to compare the approach reported here with other in vitro or in vivo, direct or indirect, approaches previously reported.

Fast And Direct Na And K Determination In Table, Marine, And Low-sodium Salts By X-ray Fluorescence And Chemometrics.

X-ray fluorescence (XRF) is a fast, low-cost, nondestructive, and truly multielement analytical technique. The objectives of this study are to quantify the amount of Na(+) and K(+) in samples of table salt (refined, marine, and light) and to compare three different methodologies of quantification using XRF. A fundamental parameter method revealed difficulties in quantifying accurately lighter elements (Z < 22). A univariate methodology based on peak area calibration is an attractive alternative, even though additional steps of data manipulation might consume some time. Quantifications were performed with good correlations for both Na (r = 0.974) and K (r = 0.992). A partial least-squares (PLS) regression method with five latent variables was very fast. Na(+) quantifications provided calibration errors lower than 16% and a correlation of 0.995. Of great concern was the observation of high Na(+) levels in low-sodium salts. The presented application may be performed in a fast and multielement fashion, in accordance with Green Chemistry specifications.

The role of illness, resources, appraisal, and coping strategies in adjustment to HIV/AIDS: The direct and buffering effects

This study examined the utility of a stress and coping model of adjustment to HIV/AIDS. A total of 114 HN-infected gay or bisexual men were interviewed and they completed self-administered scales. Predictors included illness variables (disease stage and number of symptoms), coping resources (optimism and social support), appraisal (threat, challenge, and controllability), and coping strategies (problem-and emotion-focused). Adjustment outcomes were depression, global distress, social adjustment, and subjective health status. Results from hierarchical regression analyses indicated that better adjustment was related to an asymptomatic illness stage, fewer HN-related symptoms, greater social support, challenge and controllability appraisals, problem-focused coping, and lower threat appraisals and reliance on emotion focused coping. There was limited support for the stress-buffering effects of optimism. Findings support the utility of a stress and coping model of adjustment to HIV/AIDS.

Robosonic: Randomness-Based Manipulation of Sounds Assisted by Robots

In this text, we intend to explore the possibilities of sound manipulation in a context of augmented reality (AR) through the use of robots. We use the random behaviour of robots in a limited space for the real-time modulation of two sound characteristics: amplitude and frequency. We add the possibility of interaction with these robots, providing the user the opportunity to manipulate the physical interface by placing markers in the action space, which alter the behaviour of the robots and, consequently, the audible result produced. We intend to demonstrate through the agents, programming of random processes and direct manipulation of this application, that it is possible to generate empathy in interaction and obtain specific audible results, which would be difficult to otherwise reproduce due to the infinite loops that the interaction promotes.

Parent-child agreement and prevalence estimates of diagnoses in childhood: direct interview versus family history method.

Diagnostic information on children is typically elicited from both children and their parents. The aims of the present paper were to: (1) compare prevalence estimates according to maternal reports, paternal reports and direct interviews of children [major depressive disorder (MDD), anxiety and attention-deficit and disruptive behavioural disorders]; (2) assess mother-child, father-child and inter-parental agreement for these disorders; (3) determine the association between several child, parent and familial characteristics and the degree of diagnostic agreement or the likelihood of parental reporting; (4) determine the predictive validity of diagnostic information provided by parents and children. Analyses were based on 235 mother-offspring, 189 father-offspring and 128 mother-father pairs. Diagnostic assessment included the Kiddie-schedule for Affective Disorders and Schizophrenia (K-SADS) (offspring) and the Diagnostic Interview for Genetic Studies (DIGS) (parents and offspring at follow-up) interviews. Parental reports were collected using the Family History - Research Diagnostic Criteria (FH-RDC). Analyses revealed: (1) prevalence estimates for internalizing disorders were generally lower according to parental information than according to the K-SADS; (2) mother-child and father-child agreement was poor and within similar ranges; (3) parents with a history of MDD or attention deficit hyperactivity disorder (ADHD) reported these disorders in their children more frequently; (4) in a sub-sample followed-up into adulthood, diagnoses of MDD, separation anxiety and conduct disorder at baseline concurred with the corresponding lifetime diagnosis at age 19 according to the child rather than according to the parents. In conclusion, our findings support large discrepancies of diagnostic information provided by parents and children with generally lower reporting of internalizing disorders by parents, and differential reporting of depression and ADHD by parental disease status. Follow-up data also supports the validity of information provided by adolescent offspring.

Cleaning wrasse species vary with respect to dependency on the mutualism and behavioural adaptations in interactions

Interspecific mutualisms are an essential feature of life on earth, yet we know little about their evolution and stability. In many mutualisms several species are available as partners, raising questions about the similarity in function and behavioural repertoire depending on the partner species. Furthermore, variation between species in the quantity and quality of interactions resulting in variation in payoffs may allow us to infer the potential evolutionary origin of a multispecies mutualism complex. We addressed these issues in the marine cleaning mutualism, in which so-called 'cleaners' remove ectoparasites from so-called 'client' reef fish. We measured several parameters concerning the quantity and quality of cleaning interactions in six sympatric cleaner wrasse species. We found significant variation between cleaner species with respect to client diversity, the number of interactions with predatory clients, the duration of interactions, the frequency of client jolts as a correlate of 'cheating' by cleaners, and behaviours used for manipulation of client decisions. Exploratory correlations between cleaner species' dependency and our variables of interest suggest that cleaning originated as a conflict-free by-product mutualism and evolved towards more sophisticated behaviours, including strategic behaviours for interactions with predators, cheating and manipulation specifically adapted to the client type.