Multi-robot cooperative navigation in real-world environments is essential in many applications, including surveillance and search-and-rescue missions. State-of-the-art methods for cooperative navigation are often tested in ideal laboratory conditions and not ready to be deployed in real-world environments, which are often cluttered with static and dynamic obstacles. In this work, we explore a graph-based framework to achieve control of real robot formations moving in a world cluttered with a variety of obstacles by introducing a new distributed algorithm for reconfiguring the formation shape. We systematically validate the reconfiguration algorithm using three real robots in scenarios of increasing complexity.


Characterizing statistical properties of solutions of inverse problems is essential for decision making. Bayesian inversion offers a tractable framework for this purpose, but current approaches are computationally unfeasible for most realistic imaging applications in the clinic. We introduce two novel deep learning based methods for solving large-scale inverse problems using Bayesian inversion: a sampling based method using a Wasserstein GAN with a novel mini-discriminator and a direct approach that trains a neural network using a novel loss function. The performance of both methods is demonstrated on image reconstruction in ultra low dose 3D helical CT. We compute the posterior mean and standard deviation of the 3D images followed by a hypothesis test to assess whether a "dark spot" in the liver of a cancer stricken patient is present. Both methods are computationally efficient and our evaluation shows very promising performance that clearly supports the claim that Bayesian inversion is usable for 3D imaging in time critical applications.


Purpose
The purpose of this paper is to understand how human resource (HR) practitioners subjectively experience emotions in their working lives and how they navigate emotionally challenging work.

Design/methodology/approach
A narrative methodology and participant-led photo-elicitation methods were used with five HR practitioners from different sectors to uncover experiences of emotion in their work.

Findings
Participants describe themselves as perceived by non-HR employees as non-emotional human beings, expected to "take" emotional expression from others, but to display little themselves. HR practitioners use emotion-focussed coping strategies, both self and team-care, to cope with the emotionally challenging work inherent in their role.

Research limitations/implications
As a pilot study of five participants, further research is needed to strengthen the findings; however, the in-depth qualitative methods used provide rich insight into their working lives.

Practical implications
HR practitioners' well-being should not be taken for granted or overlooked in organisations. Opportunities for informal networking with HR communities and training/coaching interventions could provide support on approaches to the emotional challenges faced.

Originality/value
This paper provides insights into how HR practitioners experience the challenges of their work, in contrast to mainstream research emphasising the impact of human resource management policy and practices on employees and organisations. Attention is drawn to the subjective experience of emotion, rather than the mainstream objectification, managerialisation and generalisation of emotion.


In the past years, a large number of evaluations have assessed the impact of European Union?s Cohesion Policy, the largest transnational programme for regional development promotion worldwide. In particular, great attention has been devoted to determine the effectiveness of EU funds in improving economic and labour market conditions in Objective 1 regions, the most economically disadvantaged areas receiving the large majority of the funds. However, there is no consensus yet in the literature on whether Structural Funds have successfully contributed to promote convergence by improving the economic performance of the poorest regions in the EU. In addition, a lot of attention has been devoted to determine the effect of Cohesion Policy interventions on inequality across regions and countries, but little research has been carried out to analyse the impact of the policy in reducing disparities within the targeted regions. In order to provide more conclusive answers on the effectiveness of Cohesion Policy, a new strand of evaluations has emerged in the literature. This research is based on the adoption of quasi-experimental methods to capture the causal effects of the policy. In this paper we follow this example and use a differences-in-differences strategy to estimate the impact of Cohesion Policy. By exploiting an exogenous change in eligibility for Objective 1 support occurred in the UK region of Cornwall in 2000, we are able to compare Cornwall?s unemployment trend before and after the introduction of Objective 1 support. The study is performed at three different geographical levels. First, we construct a synthetic control region that equals the treated region in all relevant characteristics except for not having received Objective 1 funding. Second, we use Local Authority Districts and compare the unemployment change of Cornwall with similar English areas. Third, we adopt propensity score matching techniques and estimate the average treatment effect at the level of electoral wards. All different methodologies provide clear evidence of a reduction in unemployment in Cornwall as compared to regions that did not benefit from Objective 1 support. In the second part of the study we identify the areas that experienced the highest unemployment change. We estimate the difference in quantile functions between Cornwall wards and control wards, constraining treatment and control to have the same unemployment distribution before the treatment. We find that the main variations occurred towards the higher end of the unemployment distribution, that is, for wards starting from a higher unemployment level. This suggests that labour market disparities have consistently reduced during the intervention period. We put forth some hypotheses to explain Cornwall?s successful management of EU funds.


We study the time-series properties of firm effects in the two-way fixed effects models popularized by Abowd, Kramarz, and Margolis (1999) (AKM) using two approaches. The first—the rolling AKM approach (R-AKM)—estimates AKM models separately for successive two-year intervals. The second—the time-varying AKM approach (TV-AKM)—is an extension of the original AKM model that allows for unrestricted interactions of year and firm indicators. We apply to both approaches the leave-one-out methodology of Kline, Saggio and Solvsten (2019) to correct for biases in the resulting variance components. Using administrative wage records from Washington State, we find, first, that firm effects for hourly wage rates and earnings are highly persistent. Specifically, the autocorrelation coefficient between firm effects in 2002 and 2014 is 0.74 for wages and 0.82 for earnings. Second, the R-AKM approach uncovers cyclicality in firm effects and worker-firm sorting. During the Great Recession the variability in firm effects increased, while the degree of worker-firm sorting decreased. Third, we document an increase in wage dispersion between 2002–2003 and 2013–2014. This increase in wage dispersion is driven by increases in the variance of worker effects and sorting, with an accompanying decrease in the variance of firm wage effects. Auxiliary analyses suggest that the misspecification of standard AKM models resulting from restricting firm effects to be fixed over time is a second-order concern.


This thesis focuses on developing a model of eco-socially conscious consumer behaviour related to choice and use of personal cars. It presents empirical evidence relating to the factors that must be considered when promoting environmentally friendly cars (noted as alternative fuel vehicles (AFVs) throughout the thesis), especially in an emerging economy such as Pakistan. The rationale and motivation behind this project is that there is an increasing rate of environmental problems such as air pollution and CO₂ in emerging economies and relatively lower competence in developing strategies aimed at improving climate change resilience. Together with changing the climate, anti-environmental anthropogenic activities make it more difficult for affected communities to prosper. To curb these environmental problems, studies reported in the academic literature have suggested taking measures to reduce the impact of human activities on the environment and regulating consumption of environmentally harmful products. In response to these emerging demands, marketers have invested heavily, regarding both product development and promotion of pro-environmental behaviours, in various domains of commercial interest. One such area is the use of personal cars, a sector that is proliferating and, given that CO₂ emissions from cars are one of the most significant sources of environmental problems (particularly global warming), there is a need to promote alternative fuel vehicles (AFVs) and eco-social behaviours in the use of personal cars. This thesis reports on two major studies to answer three underlying research questions. The first study focuses on two research questions. The first research question, RQ1, explores how automobile industry consumers (those in the personal cars segment) define eco-socially conscious behaviour (ESCCB) related to the choice and use of personal cars in Pakistan. The second research question, RQ2, attempts to identify the profiles of different customer segments based ESCCB defined in RQ1. The second study is focused on the theoretical explanation of factors that are suggested in the literature to affect ESCCB related to the choice and use of personal cars. The Theory of Planned Behaviour (TPB) and Value-Beliefs-Norms Theory (VBN) have been converged to provide a holistic explanation of ESCCB.

Based on scientific methodologies recommended for new scale development, the results reported in this thesis suggest that ESCCB related to choice and use of personal cars is a latent construct manifested in three underlying dimensions: eco-social use, eco-social purchase and eco-social conservation. A market segmentation approach using cluster and discriminant analysis suggests that three consumer segments exist in the Pakistani automobile market based on response towards eco-social behaviour and inclination towards choosing AFVs. The first segment, the conservatives, are not concerned about the environmental issues, prefer conventional cars, and are least sensitive to the eco-social use of personal cars. The second segment, the indifferents, are unsure whether they should buy AFVs and whether this will positively affect the environment. The third segment, and the largest one (51%), the enthusiasts, are highly inclined towards purchasing AFVs and eco-social use of personal cars to reduce the impact of the use of personal cars on the environment. The findings of Study 1 hold significant implications for marketing practitioners and policymakers. Some conceptual and methodological limitations are highlighted.

The results of Study 2 suggest that the Theory of Planned Behaviour (TPB), Value- Belief-Norm (VBN) Theory and the integrated model, were all found to be very strong in explaining not only ESCCB intentions but also actual behaviour, related to purchase of environmentally friendly cars and conservation of fuel. Results showed that the integrated model based on TPB and VBN was stronger in predicting ESCCB-conservation (49.7 per cent variance) than TPB (46.7 per cent variance) and VBN (26.7 per cent variance). A similar pattern of results was evident for ESCCB-purchases (integrated model: 14.8 per cent variance, TPB: 12.5 per cent variance, VBN: 10.8 per cent variance). However, the predictive power of the three models for actual eco-socially conscious consumer behaviour (ESCCB) had slightly different results. TPB was found stronger to predict actual ESCCB (33.4 per cent variance), followed by the integrated model based on TPB and VBN (31.9 per cent variance) and VBN (15.7 per cent variance).

This study contributes to both theoretical and practical aspects linked with ecosocially conscious consumer behaviour related to choice and use of personal cars. These contributions extend the theoretical literature related to eco-social behaviours and provides policy measures for marketing practitioners and public policy makers. The study findings not only provide guidelines for automobile related behaviours but can also be generalised in other areas.


Maintenance of an adequate supply of cholesterol is important for neuronal function, whereas excess cholesterol promotes amyloid precursor protein (APP) cleavage generating toxic amyloid-β (Aβ) peptides. To gain insights into the pathways that regulate neuronal cholesterol level, we investigated the potential for reconstituted apolipoprotein E (apoE) discs, resembling nascent lipoprotein complexes in the central nervous system, to stimulate neuronal [3H]cholesterol efflux. ApoE discs potently accelerated cholesterol efflux from primary human neurons and cell lines. The process was saturable (17.5 μg of apoE/ml) and was not influenced by APOE genotype. High performance liquid chromatography analysis of cholesterol and cholesterol metabolites effluxed from neurons indicated that <25% of the released cholesterol was modified to polar products (e.g. 24-hydroxycholesterol) that diffuse from neuronal membranes. Thus, most cholesterol (∼75%) appeared to be effluxed from neurons in a native state via a transporter pathway. ATP-binding cassette transporters ABCA1, ABCA2, and ABCG1 were detected in neurons and neuroblastoma cell lines and expression of these cDNAs revealed that ABCA1 and ABCG1 stimulated cholesterol efflux to apoE discs. In addition, ABCA1 and ABCG1 expression in Chinese hamster ovary cells that stably express human APP significantly reduced Aβ generation, whereas ABCA2 did not modulate either cholesterol efflux or Aβ generation. These data indicate that ABCA1 and ABCG1 play a significant role in the regulation of neuronal cholesterol efflux to apoE discs and in suppression of APP processing to generate Aβ peptides.


The anti-allergic effect of epigallocatechin-3-O-(3-O-methyl) gallate (EGCG3"Me) and epigallocatechin-3-O-(4-O-methyl) gallate (EGCG4"Me) isolated from Japanese or Taiwanese tea (Camellia sinensis L.) leaves. These catechins strongly inhibited mast cell activation and histamine release after FcepsilonRI cross-linking through the suppression of tyrosine phosphorylation (Lyn) of cellular protein kinase, and the suppression of myosin light chain phosphorylation and high-affinity IgE receptor expression via the binding to 67kDa laminin receptor. A double blind clinical study on subjects with Japanese cedar pollinosis was carried out. At the eleventh weeks after starting to intake, the most severe cedar pollen scattering period, symptoms i.e. blowing nose, itch of eyes were significantly relieved in Benifuuki group compared with placebo group. Over one consecutive month intake of Benifuuki green containing O-methylated catechin tea was useful for reduction of some symptoms derived from Japanese cedar pollinosis, and did not affect any normal immune response in the subjects with Japanese cedar-pollinosis. From the investigation that the effects of cultivars, tea seasons of crops and manufacturing methods, green or semi-fermented teas, made from fully-matured Benifuuki in second crop season, should be consumed. It is possible to develop functional articles such as beverage or food with this Benifuuki green tea.


The indirect impact of fl ower herbivory on plant reproduction depends on the pollination environment, particularly on the presence or absence of pollinator species with the ability to discriminate damaged from undamaged fl owers. The change in pollinator assemblages, due to habitat modifi cation, may modify the impact of fl ower herbivory on plant reproductive success. In this work, we evaluate the effect of fl ower herbivory on the seed production of Viola portalesia (Gay) in two contrasting environments, a native and low-disturbed habitat and an extensively transformed habitat characterized by Pinus radiata plantations. Even though the two habitats dif fered substantially in the composition of pollinator assemblages and visitation rate, the fl ower damage performed on different petals had no impact on seed production neither within nor between habitats, indicating that change in pollinator assemblages have no indirect reproductive impact via discrimination of damaged fl owers. There was a strong habitat effect, however, for seed production, being higher in the pine plantation than in the native habitat. Seed production on a per fl ower basis increased at a higher rate with the number of fl owers in the pine plantation, which suggests a numerical response of pollinators to changes in food availability in this habitat.


Excerpt] An intensified focus on youth at the international level in recent years has brought a greater understanding of youth labour markets and led to development of a growing number of national action plans for youth employment as well as other more specific youth-related policies and programmes at the national level. But has the increased global awareness of the vulnerabilities of youth brought about any quantifiable changes in their labour market situation? Are more young people attaining their desired job? The ILO's third edition of the Global Employment Trends for Youth (October 2008) examines the most recent labour market indicators and finds that young people still suffer disproportionately from a deficit of decent work opportunities. However, progress has been seen in some regions. The report, which updates the world and regional youth labour market indicators presented in previous reports (2004 and 2006), is organized according to nine regional analyses.


OBJECTIVE To determine the long term outcome in patients treated with percutaneous transluminal septal myocardial ablation (PTSMA) for hypertrophic obstructive cardiomyopathy (HOCM). DESIGN AND SETTING Observational, single centre study. PATIENTS 25 patients (13 women, 12 men, mean (SD) age 54.7 (15.0) years) with drug treatment resistant New York Heart Association (NYHA) class 2.8 (0.6) symptoms attributed to a high left ventricular outflow gradient (LVOTG) and a coronary artery anatomy suitable for intervention. INTERVENTION PTSMA by injection of 4.1 (2.6) ml of alcohol (96%) into 1.4 (0.6) septal perforator arteries to ablate the hypertrophied interventricular septum. OUTCOME MEASURES During in-hospital follow up, enzyme rise, the frequency of atrioventricular conduction lesions requiring permanent DDD pacing, and in-hospital mortality were assessed. Long term follow up (30 (4) months, range 24–36 months) included symptoms, echocardiographic measurements of left atrial and left ventricular dimensions and function, and LVOTG. RESULTS Mean postinterventional creatine kinase rise was 780 (436) U/l. During PTSMA 13 patents developed total heart block, permanent pacing being necessary in five of them. One 86 year old patient died from ventricular fibrillation associated with intensive treatment (β mimetic and theophylline) for coexistent severe obstructive airway disease. After three months, three patients underwent re-PTSMA because of a dissatisfactory primary result, leading to LVOTG elimination in all of them. During long term follow up, LVOTG showed sustained reduction (3 (6) mm Hg at rest and 12 (19) mm Hg with provocation) associated with stable symptomatic improvement (NYHA class 1.2 (1.0)) and without significant global left ventricular dilatation. CONCLUSIONS PTSMA is an effective non-surgical technique for reduction of symptoms and LVOTG in HOCM. Prospective, long term observations of larger populations are necessary in order to determine the definitive significance of the procedure.


In urban environments great effort is directed toward alleviating traffic including the design and implementation of complex software and hardware infrastructure. We introduce the idea of an auction-based mechanism for resolving vehicle intersections using a multi-way group auction mechanism. We propose a supporting infrastructure that has promise for increasing performance and responsiveness to dynamic traffic conditions. In order to evaluate new intersections, we propose new metrics that attempt to capture a more human aspect of vehicular transportation. We demonstrate that Interchange intersections perform well in single and multi-grid configurations, are self-adapting and are responsive to a variety of traffic loads.


Studio portrait photograph of the Frederiksen family in front of a blank backdrop. An ornate table has been added on the right, on which one of the children sits. The man on the far right has been identified as Andrew C. Frederiksen. His first wife, Karen Marie Tarina Frederiksen, stands on the left with her right hand around the shoulder of Thora Marie Jorgine 'Doris" Frederiksen, her daughter. Her second daughter, a slightly older girl sitting in a small chair between her parents, has been identified as Olivia E. Frederiksen. Lastly, their son, Auderman Frederiksen, stands in the back with his left arm resting on his father's shoulder. There is a hand-written note on the back of the photograph but it cannot be read with certainty.


In the past, settlors tended to limit a trustee's discretion by setting forth a specific formula for the distribution of trust assets. Nowadays, however, settlors often prefer to vest more discretion in their trustees. This is partly due to the fact that beneficiaries tend to live longer and, therefore, trusts inevitably last longer, thereby requiring trustees to respond to changing conditions. In addition, settlors often believe that vesting increased discretion on the part of trustees will discourage beneficiaries from bringing expensive and disruptive challenges to their decisions.
Nevertheless, the trend toward increased discretion is not without its problems. First of all, there is a need to balance the wishes of the settlor against the duty of the courts to oversee the conduct of trustees and other fiduciaries. In addition, it is also necessary for courts to balance the wishes of the settlor with the right of the beneficiaries to receive fair and impartial treatment. Finally, it is necessary to determine when, if ever, creditors should be able to reach a beneficiary's interest in a discretionary trust. The article begins with a description of the various linguistic formulas that settlors have typically used to describe the scope of a trustee's discretion. It concludes that no language, however broad, can completely shield a trustee from judicial scrutiny. It then examines some of standards courts invoke when they purport to review the exercise of discretion by trustees. These standards be classified as subjective, objective or a combination of both.
The article also examines the ability of trust beneficiaries to challenge a trustee's exercise of discretion. In addition, the article discusses the rights of creditors and concludes that discretionary and support trusts are treated like spendthrift trusts. This means that providers of necessary goods and services, as well as spouse, ex-spouses and minor children, can often reach a beneficiary's interest in a trust. Finally, the article suggests some improvements in certain problem areas and advises drafters to be more specific about what a trustee can and cannot do when exercising discretion.


Grassland birds have recently undergone substantial population declines throughout much of their historic ranges in North America. The majority of remaining grassland bird habitat is rangeland managed for livestock production, so grazing management has implications for grassland bird conservation efforts. Populations of mesocarnivores have recently seen range expansions and increased abundance as a result of anthropogenic extirpation of apex predators in grassland ecosystems. Mesocarnivores are often major predators of grassland birds and their nests, so considering the effects of management actions on mesocarnivore occupancy is important within grassland bird conservation efforts. I evaluated the relative effects of three livestock grazing systems on grassland bird abundance and mesocarnivore occupancy in a northern mixed-grass prairie ecosystem of eastern Montana, USA. During 2016–17, I conducted replicated point-count surveys at 150 locations on a 3,000-ha ranch managed with rest-rotation cattle grazing, and 155 locations on adjacent reference properties (4,300-ha) employing season-long or 2-pasture summer-rotational grazing systems to identify grazing management influences on grassland bird abundances. During 2016–17, I deployed remote cameras at 45 locations within rest-rotation grazing systems, and at 45 locations on reference properties to identity grazing management influences on mesocarnivore occupancy. Overall, there was no noticeable benefit of rest-rotation grazing on abundance or species diversity of grassland birds relative to season-long and summer-rotation grazing systems. Speciesspecific responses to livestock grazing system were found for each of three obligate grassland birds. Support for interactions between grazing system and local rangeland production potential were found, limiting the ability to recommend general livestock management practices for the benefit of grassland bird populations. Additionally, abundance of grasshopper sparrow (Ammodramus savannarum; β = -0.10 ± 0.03) and western meadowlark (Sturnella neglecta; β = -0.09 ± 0.03) showed a negative relationship with increasing stocking rate. Occupancy of mesocarnivores was highest in rest-rotation grazing systems, followed by season-long and summer-rotation systems, respectively, and showed a positive relationship with increasing stocking rate (β = 1.64 ± 1.10). Regional guidelines for livestock grazing management may be inappropriate in terms of grassland bird conservation efforts in the northern mixed-grass prairie.


Fires occur extensively across Zambia every year, a problem recognized as a major threat to biodiversity. Yet, basic tools for mapping at a spatial and temporal scale that provide useful information for understanding and managing this problem are not available. The objectives of this research were: to develop a method to map the spatio-temporal seasonal fire occurrence using satellite imagery, to develop a technique for estimating missing data in the satellite imagery considering the possibility of change in land cover over time, and to demonstrate applicability of these new tools by analyzing the fine-scale seasonal patterns of landscape fires in eastern Zambia. A new approach for mapping burned areas uses multitemporal image analysis with a fuzzy clustering algorithm to automatically select spectral-temporal signatures that are then used to classify the images to produce the desired spatio-temporal burned area information. Testing with Landsat data (30m resolution) in eastern Zambia showed accuracies in predicting burned areas above 92%. The approach is simple to implement, data driven, and can be automated, which can facilitate quicker production of burned area information. A profile-based approach for filling missing data uses multitemporal imagery and exploits the similarity in land cover temporal profiles and spatial relationships to reliably estimate missing data even in areas with significant changes. Testing with simulated missing data from an 8-image spectral index sequence showed highly correlated (R of 0.78-0.92) and precise estimates (deviations 4-7%) compared to actual values. The profile-based approach overcomes the common requirement of gap-filling methods that there is gradual or no change in land cover, and provides accurate gap-filling under conditions of both gradual and abrupt changes. The spatio-temporal progression of landscape burning was evaluated for the 2009 and 2012 fire seasons (June-November) using Landsat data. Results show widespread burning (~ 60%) with most fires occurring late (August-October) in the season. Fire occurrence and burn patch sizes decreased with increasing settlement density and landscape fragmentation reflecting human influences and fuel availability. Small fires (< 5ha) are predominant and were significantly under-detected (>50%) by a global dataset (MODIS Burned Area Product (500m resolution)), underscoring the critical need of higher geometric resolution imagery such as Landsat imagery for mapping such fine-scale fire activity.


In this study, a preliminary screening of cellular extracts of S. solfataricus allowed the identification of an α-N-acetylglucosaminidase activity. However, the characterization of the corresponding recombinant enzyme revealed a novel GlcNAc de-N-acetylase, which, in cooperation with the α-glucosidase, catalyzed the hydrolysis of O-α-GlcNAc glycosides. In addition, we show that the product of a gene flanking the one encoding the de-N-acetylase is a putative glycosyltransferase, suggesting the involvement of the two enzymes in the metabolism of GlcNAc. The discovery and functional analysis of novel enzymatic activities involved in the modification of this essential sugar represent a powerful strategy to shed light on the physiology and metabolism of Archaea. ABSTRACT Sulfolobus solfataricus is an aerobic crenarchaeal hyperthermophile with optimum growth at temperatures greater than 80°C and pH 2 to 4. Within the crenarchaeal group of Sulfolobales, N-acetylglucosamine (GlcNAc) has been shown to be a component of exopolysaccharides, forming their biofilms, and of the N-glycan decorating some proteins. The metabolism of GlcNAc is still poorly understood in Archaea, and one approach to gaining additional information is through the identification and functional characterization of carbohydrate active enzymes (CAZymes) involved in the modification of GlcNAc. The screening of S. solfataricus extracts allowed the detection of a novel α-N-acetylglucosaminidase (α-GlcNAcase) activity, which has never been identified in Archaea. Mass spectrometry analysis of the purified activity showed a protein encoded by the sso2901 gene. Interestingly, the purified recombinant enzyme, which was characterized in detail, revealed a novel de-N-acetylase activity specific for GlcNAc and derivatives. Thus, assays to identify an α-GlcNAcase found a GlcNAc de-N-acetylase instead. The α-GlcNAcase activity observed in S. solfataricus extracts did occur when SSO2901 was used in combination with an α-glucosidase. Furthermore, the inspection of the genomic context and the preliminary characterization of a putative glycosyltransferase immediately upstream of sso2901 (sso2900) suggest the involvement of these enzymes in the GlcNAc metabolism in S. solfataricus. IMPORTANCE In this study, a preliminary screening of cellular extracts of S. solfataricus allowed the identification of an α-N-acetylglucosaminidase activity. However, the characterization of the corresponding recombinant enzyme revealed a novel GlcNAc de-N-acetylase, which, in cooperation with the α-glucosidase, catalyzed the hydrolysis of O-α-GlcNAc glycosides. In addition, we show that the product of a gene flanking the one encoding the de-N-acetylase is a putative glycosyltransferase, suggesting the involvement of the two enzymes in the metabolism of GlcNAc. The discovery and functional analysis of novel enzymatic activities involved in the modification of this essential sugar represent a powerful strategy to shed light on the physiology and metabolism of Archaea.


Like other international financial bodies, the International Organization of Securities Commissions (IOSCO) has responded to the financial crisis of 2008. IOSCO thus revised its Objectives and Principles and added eight new Principles, including two that specifically focused on systemic risk. IOSCO's ongoing efforts to support these new Principles are parallel to efforts by other financial regulators to deal with systemic risk. Yet, IOSCO's efforts focus on somewhat different issues in the capital markets than the issues of interest to bank regulators. This article will outline IOSCO's Objectives and Principles and explain how they were revised in response to the financial crisis of 2008. The article also will discuss certain key initiatives where a lack of harmonization would be detrimental to effective regulation, in the regulation of hedge funds, credit rating agencies, short selling, and technological innovations, including direct electronic access, dark pools, and high frequency trading. These topics have been selected because they are not within the traditional purview of bank regulators and they are securities regulatory concerns related to systemic risk in the capital markets. An important issue is whether IOSCO can successfully raise standards in these controversial areas in the face of political pressure from market players and competition between capital market centers. This article concludes that IOSCO harmonization efforts tend to be at a level of generality that may be an insufficient prod to regulatory reform. When national interests are at stake, securities regulators follow those interests rather than IOSCO directives. Since IOSCO has no enforcement mechanisms aside from peer pressure, and its members are so numerous and varied, it is unrealistic to expect rigorous and detailed harmonization of new standards of conduct or regulation. Nevertheless, IOSCO can play a useful role in highlighting critical emerging areas where securities regulation is in need of reform and it has done so with regard to a number of systemic risk issues in the trading markets.


Multiple sources abound in language, at all levels of linguistic analysis (phonology, syntax, semantics, etc.), and in a range of historical pursuits, including etymology and variationist investigations. From a methodological standpoint, moreover, recognizing multiple sources is often good historical linguistic practice (contrary to inclinations towards neat and elegant solutions that satisfy Occam's Razor). That is, if we can identify multiple pressures on some part of a language system, it cannot always readily be excluded that some or even all might have played a role in shaping a particular development; if all of the factors represent reasonable pressures that speakers could have been aware of and influenced by, excluding any could simply be arbitrary. In this paper, accordingly, I survey the breadth of multiple sources in a variety of areas of language change, and advance one particular consequence that multiple sources can lead to, namely the hypothesis that recognizing multiple sources can be a basis for positing proto-language variation that is realized in variation within single languages and across related languages.


Wastewater treatment plant (WWTP) effluents are known contributors of chemical mixtures into the environment. Of particular concern are endocrine‐disrupting compounds, such as estrogens, which can affect the hypothalamic‐pituitary‐gonadal axis function in exposed organisms. The present study examined reproductive effects in fathead minnows exposed for 21 d to a historically estrogenic WWTP effluent. Fathead minnow breeding pairs were held in control water or 1 of 3 effluent concentrations (5%, 20%, and 100%) in a novel onsite, flow‐through system providing real‐time exposure. The authors examined molecular and biochemical endpoints representing key events along adverse outcome pathways linking estrogen receptor activation and other molecular initiating events to reproductive impairment. In addition, the authors used chemical analysis of the effluent to construct a chemical‐gene interaction network to aid in targeted gene expression analyses and identifying potentially impacted biological pathways. Cumulative fecundity was significantly reduced in fish exposed to 100% effluent but increased in those exposed to 20% effluent, the approximate dilution factor in the receiving waters. Plasma vitellogenin concentrations in males increased in a dose‐dependent manner with effluent concentration; however, male fertility was not impacted. Although in vitro analyses, analytical chemistry, and biomarker responses confirmed the effluent was estrogenic, estrogen receptor agonists were unlikely the primary driver of impaired reproduction. The results provide insights into the significance of pathway‐based effects with regard to predicting adverse reproductive outcomes. Environ Toxicol Chem 2016;35:702–716. Published 2015 by Wiley Periodicals Inc. on behalf of SETAC. This article is a US Government work, and as such, is in the public domain in the United States of America.


AIM
Keyhole endoscopy is a promising therapeutic option for spontaneous intracerebral hemorrhage (ICH). We sought to compare the clinical outcomes between keyhole endoscopy surgery and craniotomy for basal ganglia ICH.

MATERIAL AND METHODS
The authors performed a retrospective analysis of the clinical and radiographic data obtained in 28 keyhole endoscopic procedures and 30 craniotomy procedures. Hematoma evacuation rate, infection rate, rebleeding and mean operation time were recorded as primary end points. Outcome Scale (GOS) values were recorded at the 3-month postoperative follow-up. The operation time from symptom onset is also studied between < 8 hours group and 8-24 hours group.

RESULTS
The evacuation rate was significantly higher in the endoscopy group compared with the craniotomy group (P < 0.05), and infectious rate was lower in the endoscopy group compared with the craniotomy group( P < 0.05). Mortality rates between the 2 groups did not show statistically significant differences. The patients operated within 8h had better outcome (GOS 4 and 5) than that operated between 8-24h (p < 0.05).

CONCLUSION
The data indicate that in patients with ICH, keyhole endoscopic surgery is safe and feasible, while operation within 8h can promote recovery of patients. These preliminary results warrant further study in a large, prospective, randomized trial in the near future.


Assessing Computer-Mediated Communication Discourse of a Traumatic Brain Injury Survivor Cheryl L. Fickey Prichard Speech-language pathologists and audiologists have begun using computer technology for information storage and retrieval and as one means of providing clinical services to communication-disordered patients. The purpose of this study is to develop a protocol to assess the written communication of brain-injured clients whose communication therapy milieu includes interactions with the world outside the treatment room through computer-mediated communication (CMC). A panel of experts reviewed a series of discourse analysis procedures and rated their perceptions of validity, reliability, and ease-of use for the procedures as means of evaluating CMC discourse. The Computer-Mediated Communication Evaluation Protocol (CMC-EP) was developed based on the results of the analysis of the panel ratings and a post-rating consensus inquiry. A series of e-mail messages and on-line text chats generated over a period of three years were evaluated using the CMC-EP. The results of the CMC-EP were examined to determine if patterns of change were revealed in the CMC discourse of a traumatic brain injury. The CMC-EP consists of four procedures: T-unit analysis, cohesion analysis, Correct Information Unit analysis, and three scales of the Rating of Communication Behaviors. Using the CMC-EP enables speech-language pathologists to complete surface / sentential analysis, cohesion analysis, informational analysis, and conversation analysis of CMC discourse.


Assume that several competing methods are available to estimate a parameter in a given statistical model. The aim of estimator averaging is to provide a new estimator, built as a linear combination of the initial estimators, that achieves better properties, under the quadratic loss, than each individual initial estimator. This contribution provides an accessible and clear overview of the method, and investigates its performances on standard spatial point process models. It is demonstrated that the average estimator clearly improves on standard procedures for the considered models. For each example, the code to implement the method with the R software (which only consists of few lines) is provided.


Aims To develop human organotypic retinal cultures (HORCs) to study retinal ganglion cell (RGC) death in response to ischaemic and excitotoxic insults, both known to cause loss of RGCs and proposed as mechanisms involved in glaucomatous retinal neurodegeneration. Methods Human donor eyes were obtained within 24 h post mortem. The retina was isolated and explants cultured using two techniques. THY-1 mRNA (assessed by real-time quantitative PCR) and neuronal nuclei (NeuN) (assessed by immunohistochemistry) were used as markers of RGCs. Apoptosis was assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL). Results The distribution of THY-1 mRNA and NeuN-labelling within the human retina was consistent with the expected distribution of RGCs. Gross morphology and retinal architecture remained stable over a 96 h culture period. THY-1 mRNA and NeuN-labelled RGC layer cells decreased over the culture period, and there was an increase in TUNEL-labelling with time, but HORCs cultured in serum-free DMEM/HamF12 medium were useful for up to 48 h in culture. N-methyl-d-aspartate (10 μM) caused a reduction in THY-1 mRNA by 24 h and decreased the numbers of NeuN-labelled RGC layer neurons by 48 h, suggesting that the loss of THY-1 mRNA was a marker of RGC stress prior to death. Simulated ischaemia (60 min oxygen/glucose deprivation) caused a reduction at 24 h in both THY-1 mRNA and the numbers of NeuN-labelled neurons of HORCs. Conclusion HORCs provide a useful model to investigate RGC insult by neurodegenerative mechanisms that may lead to glaucoma in human eyes.


The freezing of individual droplets of very pure supercooled water, ranging in diameter from 5 to 120 μm, has been photographed and measured as they fall through a vertical temperature gradient established in cold, purified helium, air and other gases. From measurements made on a few hundreds of droplets of the same size, it is possible to deduce the median (or mean) freezing temperature of the group and the statistical spread about the mean temperature. When the 'observed' freezing temperatures are corrected for the thermal inertia of the falling droplets and adjusted to a standard rate of cooling, they are in good agreement with earlier work in which the droplets were cooled in a liquid supporting medium. The observed variation of freezing temperature with droplet diameter may be satisfactorily interpreted in terms of the theory of homogeneous nucleation of supercooled water if the specific surface free energy of the crystal/liquid interface is taken to be 20 erg cm-2 between —35 and —40°C. The experimental results are also consistent with, and offer additional support for, the recent Némethy-Scheraga statistical thermodynamical treatment of the 'flickering-cluster' model of liquid water.


This document provides a short literature review on the complementarity (and antagonisms) between liability rules, safety regulation and insurance and their effect on safety management. It draws on a range of disciplines, with a focus on economic analysis of law and regulation theory. Some of the issues discussed are rather complex; this document attempts to provide simple explanations together with references to the professional literature for the interested reader. Some issues are the subject of ongoing debate between scholars; in such situations, we have attempted to present the various points of view. The document provides background information concerning the topics discussed during the NeTWork'2012 workshop, and draws on some of the contributions of workshop participants and the rich discussion which took place during the three days. The first chapter presents issues related to regulation, starting with the classical economic justifications for state intervention (presence of externalities, information failures and moral hazard). A number of obstacles to the effectiveness of safety regulation are presented. Finally, some alternatives or complements to regulation, including self-regulation, are briefly discussed. Chapter 2 presents an overview of liability law, starting with some introductory definitions. Factors which weaken the effectiveness of liability as an incentive to invest in prevention are discussed, as are negative effects of liability regimes on safety management. A number of case studies illustrating the liability of regulators are briefly presented. Chapter 3 discusses the impact of insurance and reinsurance on firms' and individuals' safety management. The last chapter briefly analyzes firms' and individuals' sources of motivation to take care.


Men Speak Out: Views on Gender, Sex, and Power (2nd ed.) highlights new essays on pornography, pop culture, queer identity, Muslim masculinity, and the war on women. With personal candor and political insight, this collection of diverse authors explores sex work, digital activism, incarceration, domestic violence, surviving incest, and standing firmly as male allies facing the backlash against women's reproductive rights. Featuring eleven new essays and six revised thematic sections, this second edition of a favorite anthology continues to encourage robust discussion and vibrant debate about masculinity and the possibilities for progressive change. The contemporary, compelling essays in Men Speak Out appeal to students, scholars, activists, and everyday readers.


During ion implantation, positive charge accumulates due to excess positive beam ions incident on the wafer as well as through the emission of secondary electrons from the wafer surface. This positive charge build-up is counteracted by the flow of electrons onto the wafer surface. These electrons are drawn from the beam plasma, from supplemental sources of electrons ("flood guns"), and from current flows along the wafer surface and through the bulk [1]. In a typical situation, these charge flows are not exactly balanced, especially in local regions (such as on isolated polysilicon structures), leading to voltage build-up and degradation or destruction of gate oxides and other dielectric films. A successful charge neutralization technology strives to balance the total charged flux into the region near the wafer surface and minimize net charge flow to the wafer. The potential differences between the ion beam plasma and the structures on the wafer surface influence the charge flow to the surface and must be minimized. Since build-up of excess electrons on the wafer surface may also lead to damage in sensitive dielectric films ("overflooding"), the energy of the electrons that are used to balance the positive charge flow must be lower than those potentials that can lead to oxide wearout and breakdown problems. In ion implanters, the beam generates too few electrons by collisional ionization of background gases to balance the positive ion beam current. To neutralize the beam, an auxiliary source of electrons--an electron "flood gun"--must provide additional negative charge.


I. INTRODUCTION II. BACKGROUND: HOW AND WHY THE "NEW" CONSTITUTION OF JAPAN WAS ADOPTED A. An Overview B. The Adoption Process III. THE REINTERPRETATION OF ARTICLE 9 AND THE REMILITARIZATION OF JAPAN IV. BOTH SIDES OF THE ARGUMENT FOR REMILITARIZATION A. Arguments in Favor of Remilitarization B. Arguments Opposing Remilitarization V. THE FUTURE OF ARTICLE 9 A. Debate and the Possibility of Constitutional Amendment B. A Historic Shift in Politics: The Future of Article 9 Under the Democratic Party of Japan VI. U.S.-JAPAN RELATIONS: NOW AND IN THE FUTURE VII. CONCLUSION I. INTRODUCTION In response to the recent threats emanating from China and North Korea, as well as the ongoing war in Afghanistan, the United States is urging Japan to rebuild its long-dismantled military forces. (1) In 2009, following a historic political shift within Japan's government, the controversial topic of amending Article 9 of Japan's Constitution emerged once again. (2) The irony of U.S. support of Japanese "remilitarization" is that the United States designed and wrote the framework for Japan's post-WWII pacifist Constitution (sometimes called the "Peace Constitution"), which expressly prohibited Japan from ever again maintaining an offensive military. (3) Since WWII however, the United States has dramatically changed its stance on the Japanese military because of Japan's economic strength and the strong U.S.--Japan bonds that have developed. (4) The ideological shift toward remilitarization has been met with mixed opinions both in Japan and abroad. (5) This comment will examine Article 9's history, the possibility of amendment under the Democratic Party of Japan, changing interpretations of Japan's constitutional potential for war, both sides of the remilitarization debate, and the implications on U.S.--Japan military and economic relations, both now and in the future. I propose that Article 9 be amended and remilitarization allowed in Japan. Japan is well aware of its aggressive military history, and although the Japanese people tend to support Japan's post-WWII pacifist stance, (6) the time has come for Japan to take greater responsibility for its own defense. The United States has not formally "occupied" Japan for quite some time, but it continues to bolster Japan's military resources by providing potential offensive capabilities should the need arise. (7) I further propose that remilitarization has already become a reality and that legal and political semantics are distracting attention from this truth. I will also examine the question of whether it is politically and economically feasible for the United States to continue to provide the level of military support it has been providing since WWII. In my opinion, the answer to this question is a resounding no, and a limited allowance for Japanese military buildup is the only realistic option at this point. II. BACKGROUND: HOW AND WHY THE "NEW" CONSTITUTION OF JAPAN WAS ADOPTED A. An Overview Japan's Peace Constitution came into effect on May 3, 1947 and is uniquely pacifist in nature. (8) Article 9 renounces war, but has been interpreted to allow Japan Jieitai ([TEXT NOT REPRODUCIBLE IN ASCII]), which translates to "Self-Defense Forces" (SDF), a broad designation that includes the Air SDF, Maritime SDF, and Ground SDF. (9) The Peace Constitution is also sometimes referred to as the "MacArthur Constitution," because it was heavily influenced by General Douglas MacArthur, Supreme Commander for the Allied Powers. (10) After U.S. occupation of Japan began, MacArthur was instrumental to the architecture of Article 9, which was designed to ensure that Japan would never go to war again: Japan was to remain disarmed, and Japan's postwar military policies were to be subject to strict U.N. oversight. (11) Japan's new Constitution reflected significant changes in the views of the post-WWII international community, and was revolutionary at its inception. …


In this study, human and rat cancer cells were used to investigate the expression of p53 and p21/WAF1/CIP1 and their association with apoptosis after exposure to nitric oxide (NO). It was found that NO induced nuclear accumulation of p53 protein in a dose‐ and time‐dependent manner. The level of p53 protein was elevated by about fivefold compared with that of mock‐treated cells 48 h after exposure to 300 ppm NO. The induction of p53 by NO was found by pulse‐chase analysis to be mainly regulated by post‐translational modification. The correlation between p53 status and apoptosis induced by NO in human cancer cells was also investigated in this study. We found that apoptosis was easily induced in cells containing wild‐type p53 (COLO 205 and Hep G2) after exposure to NO. The p21/WAF1/CIP1 protein was induced by NO in cells containing wild‐type p53 (Hep G2) but not in cells without p53 (Hep 3B) or with mutated p53 (HT‐29). Our results indicate that wild‐type p53 and p21/WAF1/CIP1 expression was elevated in human cancer cells by exposure to NO and suggest that this may eventually promote apoptosis. © 1996 Wiley‐Liss, Inc.


An inability to reconcile society's need for protection from juvenile crime with the use of nonpunitive measures has troubled the juvenile justice system since its inception. Society long ago adopted a paternalistic attitude toward juvenile crime, treating such behavior not as a question of law enforcement, but as a social and psychological problem requiring therapeutic interventions and state assumption of parental rights and duties.1 The juvenile court was conceived as a kind of social welfare agency rather than as an instrument for the enforcement of the criminal laws. With the mantle of benevolence bestowed and in the name of "individualized treatment," the juvenile courts were given broad jurisdiction over both criminal and noncriminal misbehavior and were vested with virtually unlimited discretion to impose limitations and sanctions on a child's conduct 2 They were to operate a "no fault" process, geared to providing treatment and rehabilitation to children whose overt misbehavior manifested underlying problems. 3 Juvenile court procedures were to be informal and nonadversarial, and the court was to make dispositions based on the best interests of the child. 4


This research explores the role and value of nurse practitioners to UK general practice from the perspective of nurses working in these advanced roles. Nursing has had a presence in general practice for decades, but it is only over the last twenty years that it has extended into a traditional medical domain of care and treatment. Research has understandably focused on the ability of nurses to substitute for doctors and there has been relatively little investigation of what nursing at an advanced level contributes.The study is located within a qualitative interpretive paradigm utilising a Social Constructionist (SC) approach which recognises that knowledge is not based solely on objective observations of the world, but is generated between individuals in the course of their everyday life. The theoretical perspective grounded in this epistemological paradigm is symbolic interactionism (SI). This emphasises the construction of the social world and meaning through the use of symbols, particularly language.Thematic Analysis (TA) is utilised deliberately as a research strategy guiding sampling, data generation, collection and analysis. A purposive sample of ten nurse practitioners was selected. Semi-structured interviews were conducted, digitally recorded, transcribed and the data analysed using Braun and Clarke's model.Four broad themes were identified from the narratives; the enactment and development of the nurse practitioner role, its value to the organisation and function of general practice, the impact of nurse consultation upon the patient experience and finally, how the role has integrated into the primary health care team.The findings demonstrate that rather than one generic nurse practitioner role in general practice there are multiple constructs, driven at macro level by political necessity, negotiated at micro level by the needs of individual general practices and framed within a professional vacuum of non-regulation. This has not been fully explained before. The research provides a clear and original understanding of what nurse practitioners can contribute to general practice through the diversification of their roles, not as substitute but as part of a diverse, fluid team working collaboratively to address the needs of the general practice population.


Abstract Third party apps that work on top of personal cloud services, such as Google Drive and Drop-box, require access to the user's data in order to provide some functionality. Through detailed analysis of a hundred popular Google Drive apps from Google's Chrome store, we discover that the existing permission model is quite often misused: around two-thirds of analyzed apps are over-privileged, i.e., they access more data than is needed for them to function. In this work, we analyze three different permission models that aim to discourage users from installing over-privileged apps. In experiments with 210 real users, we discover that the most successful permission model is our novel ensemble method that we call Far-reaching Insights. Far-reaching Insights inform the users about the data-driven insights that apps can make about them (e.g., their topics of interest, collaboration and activity patterns etc.) Thus, they seek to bridge the gap between what third parties can actually know about users and users' perception of their privacy leakage. The efficacy of Far-reaching Insights in bridging this gap is demonstrated by our results, as Far-reaching Insights prove to be, on average, twice as effective as the current model in discouraging users from installing over-privileged apps. In an effort to promote general privacy awareness, we deployed PrivySeal, a publicly available privacy-focused app store that uses Far-reaching Insights. Based on the knowledge extracted from data of the store's users (over 115 gigabytes of Google Drive data from 1440 users with 662 installed apps), we also delineate the ecosystem for 3rd party cloud apps from the standpoint of developers and cloud providers. Finally, we present several general recommendations that can guide other future works in the area of privacy for the cloud. To the best of our knowledge, ours is the first work that tackles the privacy risk posed by 3rd party apps on cloud platforms in such depth.


Immunopotentiators in Modern Vaccines, Second Edition, provides in-depth insights and overviews of the most successful adjuvants, those that have been included in licensed products, also covering the most promising technologies that have emerged in recent years. In contrast to existing books on the subject, the chapters here provide summaries of key data on the mechanisms of action of the individual vaccine adjuvants. In addition, the book covers key aspects of how the technologies might be further developed and what might be their limitations, while also giving an overview of what made the most advanced adjuvant technologies successful. Provides contributions from leading international authorities in the field. Features immunopotentiators classified by function, with well-illustrated, informative figures presenting the interaction between the immunopotentiators and the host immune system. Lists advantages and potential hurdles for achieving a practical application for each specific immunopotentiator. Offers US FDA perspectives which highlight how future adjuvants will be approved for new generation vaccines.


Policy rules that are consistent with inflation targeting are examined in a small macroeconomic model of the US economy. We compare the properties and outcomes of explicit instrument rules' as well as targeting rules.' The latter, which imply implicit instrument rules, may be closer to actual operating procedures of inflation-targeting central banks. We find that inflation forecasts are central for good policy rules under inflation targeting. Some simple instrument and targeting rules do remarkably well relative to the optimal rule; others, including some that are often used as representing inflation targeting, do less well.


Enterprise integration is a broad activity that involves solving a range of issues relating to business process definition, common data standards, architectural compatibility, technical interoperability, and organizational alignment. Enterprise Architecture and Integration: Methods, Implementation, and Technologies provides a detailed analysis of the important strategies for integrating IT systems into fields such as e-business and customer-relationship management. This Premier Reference Source supplies readers with a comprehensive survey of existing enterprise architecture and integration approaches, and presents case studies that illustrate best practices. It takes a holistic view of enterprise integration, describing innovative methods, tools, and architectures with which organizations can systematically achieve enterprise integration.


In this masterly new work, acclaimed historian Giles MacDonogh explores the moment when Hitler gambled everything. Until 1938, Hitler could be dismissed as a ruthless but efficient dictator, a problem to Germany alone; after 1938 he was clearly a threat to the entire world. In that year The Third Reich came of age and the Fuhrer showed his hand - bringing Germany into line with Nazi ideology and revealing long-held plans to take back those parts of Europe lost to 'Greater Germany' after the First World War. The sequence of events began in January with the purging of the army, and escalated with the merger with Austria - the Anschluss, and the first persecutions of Viennese Jewry. In the following months Hitler moulded the nation to his will. Elections brought him a 99 per cent approval rating. MacDonogh gives a full account of the nationalist opposition that failed to topple Hitler in September 1938. By the end of the year the brutal reality of the Nazi regime was revealed by Joseph Goebbels in Kristallnacht, a nationwide assault on Germany's native Jewish population. MacDonogh's access to many new sources gives insights into what life was like under the eye of the regime, revealing the role of the Anglican Church after the Anschluss, saving those Jews who were willing to convert, and also the Kendrick Affair - the still-secret details of the Austrian double agent who brought down the whole MI6 operation in Austria and Germany, just as the Chamberlain government began negotiations with Hitler at Munich. A remarkable and revealing account of Hitler's opening moves to war.


The Baltic Sea Region is rapidly becoming one of the world's more competitive regions. The region is capitalising on its strengths and making the most of its diversity to stimulate innovation, build a strong pool of skilled labour and foster entrepreneurship. A deep spirit of co-operation and integration has led the Baltic Sea countries to set up ambitious governance frameworks to pursue economic development objectives jointly from Oslo to St. Petersburg. Yet major challenges at the local level lie ahead for the Eastern shores of the Baltic, where economic transition still needs to be accompanied by more innovative strategic planning, new forms of governance and dynamic civic entrepreneurship. Policies will need to be made more adaptable and capacities will need to be strengthened if prosperity and living standards are to increase on the Baltic Rim. Fortunately, the Baltic Sea Region includes some of the world's most innovative countries. From Denmark to Finland, the Region possesses a breadth of experience in facilitating policy co-ordination, adjusting policy to local conditions and involving business and civil society in shaping policy measures. There is a great deal that other countries can learn from this experience in setting up partnerships and other forms of governance. The learning process has already started, with the Baltic Rim becoming a unique laboratory for economic and employment development. This book analyses the new developments in the Baltic States and Northwest Russia and provides suggestions on how to speed up this progress. It is essential reading for all stakeholders in the Baltic Sea Region and for those elsewhere wishing to apply emerging lessons to their region of the world. (Less)


In recent years, massive numbers of distributed energy resources are being installed in distribution feeders at both the utility and customer levels. These integrations are creating bi-directional power flow complexities in all the existing functionalities of distribution management system (DMS) and outage management system (OMS). Therefore, a significant interest is growing among utilities to combine all the DMS and OMS functionalities with distributed energy resources management system. This combined structure is known as advanced distribution management system (ADMS). In this paper, major ADMS functionalities, such as, conservation voltage reduction, fault location, isolation, and service restoration, and demand response are described with existing and aspiring implementation and assessment. Future challenges are also described from the utility perspective.


TITLE: The characterization and analysis of in-vitro and elevated temperature repassivation of Ti-6Al-4V via AFM techniques AUTHOR: ABSTRACT The Characterization and Analysis of In-vitro and Elevated Temperature Repassivation of Research in the corrosion of orthopaedic implants is a growing research field where implants have been known to show adverse effects in patients who have encountered the unfortunate dissolution of their implants due to corrosion. Once corrosion begins within the body, many adverse biological reactions can occur such as late onset infections resulting in severe health complications. The focus of this research is specifically related to the problem of late onset infections caused by localized corrosion of orthopaedic implants. In medical implants today the most common form of corrosion protection is the implant materials' ability to impede corrosion through the formation of an oxide layer. This ability to passivate and quickly repassivate a uniform and stable oxide layer dictates how well an orthopaedic implant will survive in-vivo. To better understand the repassivation of orthopaedic implant materials, research was conducted at the nanoscale via atomic force microscopy (AFM) on anodized Ti-6Al-4V. Using an Asylum Research MFP-3D TM AFM and AFM lithography techniques, nano scratch test methods were created simulating in-vitro surface repassivation conditions. These nano-scratches were created and characterized in Hank's balanced saline solution (HBSS) with the AFM in contact mode at 1 and 3 Hz scan rates. HBSS was used as it best simulates the pH, ionic compounds, and constituents that are commonly found in blood. It was discovered that the AFM was successful in creating in-vitro repassivation conditions. However, the ability of the AFM to successfully observe repassivation was limited by the speed of the AFM scanner. Using the same AFM scratch methods, experiments were performed in air and in-vitro and characterized with AFM conductance measurements at 20, 37, & 45 °C. The conductance measurements were taken using an AFM conductance module and allowed for observations of v decreasing current measurements over time. The current data was then used to calculate current density, resistivity, conductance, and electron mobility and compared to similar experiments This study highlights the ability of the AFM to create and characterize repassivation and shows promise in developing further capability to use the AFM for characterization of repassivation on the nanoscale. vi ACKNOWLEDGEMENTS To my family, thank you for the unconditional love and support throughout my life and education. I am who I am today because of …


Protein-metal complexes may be transferred from solution into the gas phase by electrospray ionization (ESI), such that they can be directly analyzed by mass spectrometry (MS). In principle, therefore, ESI-MS represents a simple and elegant approach for gaining insights into the binding stoichiometry and affinity of these assemblies. Unfortunately, the formation of nonspecific metal adducts during ESI can be a severe problem, often leading to binding levels that are dramatically higher than those in bulk solution. Focusing on several calcium binding proteins as test systems, this work explores the suitability of different salts to serve as metal source. Despite their widespread use in previous ESI-MS studies, calcium chloride and acetate induce extensive nonspecific adduction. In contrast, much lower levels of artifactual metal binding are observed in the presence of calcium tartrate. In the case of high and intermediate affinity proteins, the resulting ESI-MS data are in excellent agreement with the calcium binding behavior in bulk solution. The situation is more challenging when studying proteins with very low affinities, but in the presence of tartrate qualitative information on protein-metal interactions can still be obtained. The beneficial effects of tartrate also extend to zinc binding experiments. This work does not directly explore the mechanism by which tartrate suppresses nonspecific metalation. However, it seems likely that weak chelators such as tartrate sequester metal ions within rapidly shrinking droplets during the final stages of ESI, thereby reducing nonspecific metal adduction to protein carboxylates. The use of tartrate and possibly other weak chelators will greatly enhance the reliability of future ESI-MS studies on the interactions of proteins with divalent metal ions.


A surface plasmon-polariton (SPP) polarizer with a shorter polarizer length can be realized by strong coupling of a metal film to a core, that is, by matching of the phase constant of the SPP wave in the metal film to that of the guided mode in the core and by selection of a suitably thin buffer-layer thickness. To calculate the polarizer's extinction ratio accurately, we have considered three TM supermodes of a SPP polarizer with a symmetric seven-layered slab structure rather than only a fundamental supermode that matches an input field well for large buffer thicknesses. When the buffer layer becomes thinner, the characteristics of the SPP polarizer cannot be simply estimated when only the propagation constant of the fundamental supermode is known, since its field distribution becomes quite different from that of the input field. It is shown that the field distribution beats in the SPP polarizer because of the interference effect of these supermodes. Finally, the total extinction ratio and insertion loss, including coupling losses at input and output ends, are estimated.


We present the first systematic investigation of the polar cap boundary in Saturn's high‐latitude magnetosphere through a multi‐instrument assessment of various Cassini in situ data sets gathered between 2006 and 2009. We identify 48 polar cap crossings where the polar cap boundary can be clearly observed in the step in upper cutoff of auroral hiss emissions from the plasma wave data, a sudden increase in electron density, an anisotropy of energetic electrons along the magnetic field, and an increase in incidence of higher‐energy electrons from the low‐energy electron spectrometer measurements as we move equatorward from the pole. We determine the average level of coincidence of the polar cap boundary identified in the various in situ data sets to be 0.34° ± 0.05° colatitude. The average location of the boundary in the southern (northern) hemisphere is found to be at 15.6° (13.3°) colatitude. In both hemispheres we identify a consistent equatorward offset between the poleward edge of the auroral upward directed field‐aligned current region of ~1.5–1.8° colatitude to the corresponding polar cap boundary. We identify atypical observations in the boundary region, including observations of approximately hourly periodicities in the auroral hiss emissions close to the pole. We suggest that the position of the southern polar cap boundary is somewhat ordered by the southern planetary period oscillation phase but that it cannot account for the boundary's full latitudinal variability. We find no clear evidence of any ordering of the northern polar cap boundary location with the northern planetary period magnetic field oscillation phase.


This paper reports on a refractometer, which is based on measuring the throughput of a bent channel waveguide. A new type of channel waveguide, designed in SiON technology makes the throughput strongly dependent on the refractive index of a measurand fluid. This new waveguide can be designed for any measurand refractive index range between 1.00<n<2.00. In addition, monomode input and multimode output waveguides matching the sensor waveguide were designed which significantly simplify the fabrication process by eliminating the occurrence of critical alignment steps during the processing. Preliminary measurements on the device show a high sensing resolution of /spl Delta/n=1/spl times/10/sup -5/ in an aqueous solution with a refractive index of n=1.37. The sensor characteristics, make this simple sensor very interesting for controlling a chemical concentration during chemical processes, e.g., in the food and beverages industry.


The almost 7 decades lasting futile attempts to understand the possible physical content of the third Wigner representation class (the infinite spin class) came to a partial solution with the 2006 discovery of existence of string-localized spacetime covariantizations . This has led to a still ongoing vast generalization of renormalizability to fields with arbitrary high spin and a better understanding of the origin of partial invisibility as observed in the confinement of gluons and quarks. The present note explains the total (non-gravitational) invisibility of fields associated to the third Wigner representation class. The last section presents a critical look at the possibility that third class Wigner matter may play a role in dark matter formation.


The paper investigates the effect of co-operative membership on people's capability to participate in household decision-making and on domestic gender relations. Our hypothesis is that the democratisation process activated in genuine co-operatives, authentic member-owned forms of business, may then be transferred to the household. We tested this in the "Coppalj" co-operative in Brazil, where we collected primary data. Both the techniques employed, regression and propensity score matching, support our hypotheses in a number of life domains. Though results vary slightly according to the domain and the outcome indicator, they show that members of the co-operative have a statistically significant higher capability to participate in decision-making and to share their decisions with partners than non-members (the control group). We then triangulated these quantitative outcomes with qualitative ones: the latter confirm an improvement in gender relations between co-operators and their partners, highlighting the fundamental role of Coppalj in fostering gender equality.


We report on the synthesis of the newly discovered cubic phase of tin sulfide π-SnS and compare its properties to the well-known phase of tin sulfide, α-SnS. Shape control was achieved by the variation of synthesis parameters, resulting in cubic, rhombic dodecahedral and tetrahedral shapes of the π-SnS nanoparticles. X-ray diffraction provided authentication of the proposed model and refined determination of the lattice parameter a = 11.595 A. Raman spectroscopy showed a substantial shift towards higher energies and peak splitting for π-SnS. Optical absorption spectroscopy indicated an indirect band gap of 1.53 eV, in good agreement with density functional theory (DFT) calculations indicating a band gap greater than that of α-SnS. DFT total energy calculations show that the π-SnS phase is energetically similar to α-SnS, and is significantly more stable than the hypothetical ideal rocksalt structure of SnS.


The Arab-German cooperation in film began after World War I in 1919/1920 when the first Egyptians came to learn the then brand new art in Germany, and has been continuing with different Arab partners ever since. Yet there is neither a public nor a professional awareness of this history. When Arab and German film professionals meet at international co-production platforms today, they practically get together as strangers. Despite its richness, the common history does not serve as a point of reference. It is not written. This paper, therefore, attempts to shed light on this forgotten period of cooperation. It looks at how and why such a collaboration was initiated. Moreover, it describes its different formats and also why the Egyptian-German encounter eventually came to an end.


This thesis explores numerical methods for solving nonlinear partial differential equations (PDEs) that arise in option pricing problems. The goal is to develop or identify robust and efficient techniques that converge to the financially relevant solution for both one and two factor problems. To illustrate the underlying concepts, two nonlinear models are examined in detail: uncertain volatility and passport options.
For any nonlinear model, implicit timestepping techniques lead to a set of discrete nonlinear equations which must be solved at each timestep. Several iterative methods for solving these equations are tested. In the cases of uncertain volatility and passport options, it is shown that the frozen coefficient method outperforms two different Newton-type methods. Further, it is proven that the frozen coefficient method is guaranteed to converge for a wide class of one factor problems.
A major issue when solving nonlinear PDEs is the possibility of multiple solutions. In a financial context, convergence to the viscosity solution is desired. Conditions under which the one factor uncertain volatility equations are guaranteed to converge to the viscosity solution are derived. Unfortunately, the techniques used do not apply to passport options, primarily because a positive coefficient discretization is shown to not always be achievable.
For both uncertain volatility and passport options, much work has already been done for one factor problems. In this thesis, extensions are made for two factor problems. The importance of treating derivative estimates consistently between the discretization and an optimization procedure is discussed.
For option pricing problems in general, non-smooth data can cause convergence difficulties for classical timestepping techniques. In particular, quadratic convergence may not be achieved. Techniques for restoring quadratic convergence for linear problems are examined. Via numerical examples, these techniques are also shown to improve the stability of the nonlinear uncertain volatility and passport option problems.
Finally, two applications are briefly explored. The first application involves static hedging to reduce the bid-ask spread implied by uncertain volatility pricing. While static hedging has been carried out previously for one factor models, examples for two factor models are provided. The second application uses passport option theory to examine trader compensation strategies. By changing the payoff, it is shown how the expected distribution of trading account balances can be modified to reflect trader or bank preferences.


Objective: Parental rejection in childhood is associated with the increased risk of aggression in adulthood and is thought to contribute to the development of inaccurate beliefs regarding own or others' behavior (i.e., cognitive distortion) as well. Different forms of aggression are thought to be linked to different types of cognitive distortions. This, however, is unclear in adults. Additionally, it is unknown if parental rejection predicts the presence of aggression and cognitive distortions in adults displaying severe aggression. Method: One hundred twenty-three adult forensic psychiatric outpatients with aggression regulation problems were recruited. Least absolute shrinkage and selection operator (Lasso) regression analysis and path analyses were conducted to investigate distinct patterns of cognitive distortions, as measured by use of the How I Think Questionnaire, and the role of perceived parental rejection, as measured by use of the Parental Rejection and Acceptance Questionnaire. Results: Cognitive distortions related to opposition-defiance (e.g., disrespecting rules) and to physical aggression were most strongly associated with the disposition to act aggressively. Furthermore, a direct association was found between parental rejection and this current disposition. This association was partially mediated by cognitive distortions related to opposition-defiance. Conclusion: The current study supports the notion that parental rejection has profound consequences on adult behavior. Acknowledgment of the impact of cognitive distortions on current aggression might be of importance for treatment. A stronger focus on altering distinct cognitive distortions may be more successful in reducing aggression.


As a composite material, the performance of concrete materials can be expected to depend on the properties of the interfaces between its two major components, aggregate and cement paste. The microstructure at the interfacial transition zone (ITZ) is assumed to be different from the bulk material. In general, properties of conventional concrete have been found favoured by optimum packing density of the aggregate. Particle size is a common denominator in such studies. Size segregation in the ITZ among the binder particles in the fresh state, observed in simulation studies by concurrent algorithm-based SPACE system, additionally governs density as well as physical bonding capacity inside these shell-like zones around aggregate particles. These characteristics have been demonstrated qualitatively pertaining also after maturation of the concrete. Such properties of the ITZs have direct impact on composite properties. Despite experimental approaches revealed effects of aggregate grain shape on different features of material structure (among which density), and as a consequence on mechanical properties, it is still an underrated factor in laboratory studies, probably due to the general feeling that a suitable methodology for shape characterization is not available. A scientific argument hindering progress is the interconnected nature of size and shape. Presently, a practical problem preventing shape effects to be emphasized is the limitation of most computer simulation systems in concrete technology to spherical particles. New developments at Delft University of Technology will make it possible in the near future to generate jammed states, or other high-density fresh particle mixtures of non-spherical particles, which thereupon can be subjected to hydration algorithms. This paper will sketch the outlines of a methodological approach for shape assessment of loose (non-embedded) aggregate grains, and demonstrate its use for two types of aggregate, allowing evaluation of usefulness of the various methods.


Context awareness refers to systems that can both sense and react based on their environment. The complexity of these systems makes necessary to apply software engineering techniques in their development, such as Model-Driven Soft- ware development (MDD). One of the main difficulties that developers of context- aware systems must tackle is how to manage the needed context information. In this paper, we present MLContext, a textual Domain Specific Language (DSL) which is specially tailored for modeling context information and automatically generating software artefacts from context models. It has been designed to provide a high-level abstraction, to be an easy to learn, and to promote reuse of context models. We have built a toolkit including an editor and a parser to convert MLContext textual specifications into models. As a proof of concept, we have automatically gener- ated ontologies and Java code for the OCP middleware. MLContext models can be reused in applications with the same context because they do not include details re- lated to the platforms or the implementation. These context models can be specified by non-developers users because MLContext provides high-level abstractions of the domain.


On July 5, 2016, Amibera District Health office was alerted of excess of malaria cases in two villages. Within July 5 to 17, 2016 malaria outbreak and associations factors were investigated. A malaria case is when a person with fever or fever with headache is confirmed either microscopically to have Plasmodium parasites or by Rapid Diagnostic Test (RDT) to detect the malaria antigens. In the last five years malaria data were reviewed from 2011 to 2015 at the district level. Cases were identified by using line list and house to house active case search was done on a daily basis. Unmatched case-control study was conducted in 1:1 ratio conveniently selected cases (117) and community controls (117). The control was the neighbor of those who have not developed symptoms of malaria in the last three months. Data were collected through a structured interview administered questionnaire, entered into Epi-info version7  and analyzed using SPSS version 20. A total of 415 confirmed malaria cases with no death was identified. The predominant species was Plasmodium Falciparum accounting for 95.5%. The median age was 27 years (ranges 2- 63 years). The highest age specific attack rate was  above 15 years (63.7/1000 populations). Poor insecticide treated bed net utilization, absence of indoor residual spray, sleeping outside of their homes at nights and presence of stagnant water were factors associated with the disease contraction. In this study the current malaria outbreak was higher in magnitude than that of three years ago.

 Key words: Amibera, malaria, outbreak, Plasmodium Falciparum.


From the colourful abstraction of the Rietveld chair to the dry wit of the 'milkbottle lamp' by design cooperative Droog, modern design in the Netherlands has always been a hotbed of experimentation. Dutch designers have consistently pushed to the limits anything from posters to postage-stamps, home furnishings to street signage, ceramics to city airports. Indeed, in the last decade or so, Dutch design has become a worldwide phenomenon, almost a brand in itself, with regular publications in magazines and books promoting the remarkable output of this small country. This book takes an in-depth look not just at designs made in the Netherlands, but behind the works created throughout the twentieth century and beyond. Author Mienke Simon Thomas, a curator at one of Holland's leading museums, provides a compelling thematic account, guiding the reader through the beginnings of crafts education, the debates of design as art, the moral and social ideals of modernism, the new profession of industrial designer, state-sponsored initiatives, and conceptual design objects and 'anti-design'. As she argues, Dutch design seems to have been inspired by the wish to be functional, simple and affordable, but she also reveals how it has simultaneously embraced luxury, decoration and even exclusivity. A much-needed introduction to Dutch designs and their creators as well as the clients who commissioned them and the state initiatives that supported them this book will be essential reading for designers, historians and the general public with an interest in design.


In the 1990s, the immigrant population in the United States dispersed to non-traditional settlement locations (what have become known as "new immigrant destinations"). This paper examines whether the allure of new destinations persisted in the 2000s with a particular focus on the internal migration of the foreign born during the recent deep recessionary period and its aftermath. Three specific questions motivate the analysis. First, are immigrants, much like the US-born population, becoming less migratory within the country over time? Second, is immigrant dispersal from traditional gateways via internal migration continuing despite considerable economic contraction in many new destination metropolitan areas? Third, is immigration from aboard a substitute for what appears to be declining immigrant internal migration to new destinations? The findings reveal a close correlation between the declining internal migration propensity of the US-born and immigrants in the last two decades. We also observe parallels between the geographies of migration of native- and foreign-born populations with both groups moving to similar metropolitan areas in the 1990s. This redistributive association, however, weakened in the subsequent decade as new destination metropolitan areas lost their appeal for both groups, especially immigrants. There is no evidence to suggest that immigration from abroad is substituting for the decline in immigrant redistribution through internal migration to new destinations. Across destination types the relationship between immigration from abroad and the internal migration of the foreign born remained the same before, during, and after the Great Recession.


STUDY OBJECTIVES
The main energy reserve of the brain is glycogen, which is almost exclusively localized in astrocytes. We previously reported that cerebral expression of certain genes related to glycogen metabolism changed following instrumental sleep deprivation in mice. Here, we extended our investigations to another set of genes related to glycogen and glucose metabolism. We also compared the effect of instrumentally and pharmacologically induced prolonged wakefulness, followed (or not) by 3 hours of sleep recovery, on the expression of genes related to brain energy metabolism.

DESIGN
Sleep deprivation for 6-7 hours.

SETTING
Animal sleep research laboratory.

PARTICIPANTS
Adults OF1 mice.

INTERVENTIONS
Wakefulness was maintained by "gentle sleep deprivation" method (GSD) or by administration of the wakefulness-promoting drug modafinil (MOD) (200 mg/kg i.p.).

MEASUREMENTS AND RESULTS
Levels of mRNAs encoding proteins related to energy metabolism were measured by quantitative real-time PCR in the cerebral cortex. The mRNAs encoding protein targeting to glycogen (PTG) and the glial glucose transporter were significantly increased following both procedures used to prolong wakefulness. Glycogenin mRNA levels were increased only after GSD, while neuronal glucose transporter mRNA only after MOD. These effects were reversed after sleep recovery. A significant enhancement of glycogen synthase activity without any changes in glycogen levels was observed in both conditions.

CONCLUSIONS
These results indicate the existence of a metabolic adaptation of astrocytes aimed at maintaining brain energy homeostasis during the sleep-wake cycle.


ABSTRACT Ecosystem management requires an understanding of how landscapes vary in space and time, how this variation can be affected by management decisions or stochastic events, and the potential consequences for species. Landscape trajectory analysis, coupled with a basic knowledge of species habitat selection, offers a straightforward approach to ecological risk analysis and can be used to project the effects of management decisions on species of concern. The fisher (Martes pennanti) occurs primarily in late-successional forests which, in the Sierra Nevada mountains, are susceptible to high-intensity wildfire. Understanding the effects of fuels treatments and fire on the distribution of fisher habitat is a critical conservation concern. We assumed that the more a treated landscape resembled occupied female fisher home ranges, the more likely it was to be occupied by a female and therefore the lower the risk to the population. Thus, we characterized important vegetation attributes within the home ranges of 16 female fishers and used the distribution of these attributes as a baseline against which the effects of forest management options could be compared. We used principal components analysis to identify the major axes defining occupied female fisher home ranges and these, in addition to select univariate metrics, became our reference for evaluating the effects of landscape change. We demonstrated the approach at two management units on the Sierra National Forest by simulating the effects of both no action and forest thinning, with and without an unplanned fire, on vegetation characteristics over a 45-yr period. Under the no action scenario, landscapes remained similar to reference conditions for approximately 30-yr until forest succession resulted in a loss of landscape heterogeneity. Comparatively, fuel treatment resulted in the reduction of certain forest elements below those found in female fisher home ranges yet little overall change in habitat suitability. Adding a wildfire to both scenarios resulted in divergence from reference conditions, though in the no action scenario the divergence was 4 × greater and the landscape did not recover within the 45-yr timeframe. These examples demonstrate that combining the results of forest growth and disturbance modeling with habitat selection data may be used to quantify the potential effects of vegetation management activities on wildlife habitat.


Freshwater availability is the major constraint to agriculture in arid and semi-arid regions. The groundwater and energy conservation of applying reclaimed water for irrigation was analysed, using Southern California as the spatial domain for model testing. An extensive compilation of the most recent publicly available datasets was used to calculate the energy intensity for each water supply source, the associated carbon footprint reduction and the monetary savings associated with using reclaimed water over groundwater. Our results indicate that for 1998–2010 in California the fractional water use for agriculture is 0.81 and for urban use is 0.19. During this same period, an average of 4.2 × 10 10 m 3 of water were used for crop irrigation, of which 1%, 46.8% and 52.2% came from reclaimed water, groundwater, and surface water, respectively. Each of these three main water sources is associated with a range of energy intensity (in kWh m −3 ), depending on the process and environmental characteristics of the end-use location. Our analysis of multiple process and environmental configurations produced a detailed energy intensity database, with the associated carbon footprint. These databases are used to quantify the energy and carbon footprint difference between applying the current groundwater source and reclaimed water for irrigation.


From Rabelais's celebration of wine to Proust's madeleine and Virginia Woolf's boeuf en daube in To the Lighthouse, food has figured prominently in world literature. But perhaps nowhere has it played such a vital role as in the Italian novel. In a book flowing with descriptions of recipes, ingredients, fragrances, country gardens, kitchens, dinner etiquette, and even hunger, Gian- Paolo Biasin examines food images in the modern Italian novel so as to unravel their function and meaning. As a sign for cultural values and social and economic relationships, food becomes a key to appreciating the textual richness of works such as Lampedusa's The Leopard, Manzoni's The Betrothed, Primo Levi's Survival in Auschwitz, and Calvino's Under the Jaguar Sun.The importance of the culinary sign in fiction, argues Biasin, is that it embodies the oral relationship between food and language while creating a sense of materiality. Food contributes powerfully to the reality of a text by making a fictional setting seem credible and coherent: a Lombard peasant eats polenta in The Betrothed, whereas a Sicilian prince offers a monumental macaroni timbale at a dinner in The Leopard. Similarly, Biasin shows how food is used by writers to connote the psychological traits of a character, to construct a story by making the protagonists meet during a meal, and even to call attention to the fictionality of the story with a metanarrative description. Drawing from anthropology, psychoanalysis, sociology, science, and philosophy, the author gives special attention to the metaphoric and symbolic meanings of food. Throughout he blends material culture with observations on thematics and narrativity to enlighten the readerwho enjoys the pleasures of the text as much as those of the palate.


Muscarinic acetylcholine receptors (M1-M5) belong to the class A family of transmembrane G protein coupled receptors (GPCRs) and mediate various signalling processes. M1, M3 and M5 predominantly couple to Gq and promote intracellular calcium ion release from the endoplasmic reticulum. M2 and M4 preferentially couple Gi inhibiting adenylyl cyclase activity to thus decrease cAMP production and acting to regulate various ion channels. There is growing evidence that many GPCRs can exist as dimers or higher-order oligomers (Milligan, 2013) and muscarinic receptors are no exception (Alvarez-Curto et al., 2010). Herein, combinations of homomers and heteromers of co-expressed human M2 (hM2WT) and a RASSL (Receptor Activated Solely by Synthetic Ligand) form of the human M3 receptor (hM3RASSL) (Alvarez-Curto et al., 2011) were demonstrated to occur using N-terminal SNAP and CLIP tags in combination with homogeneous time resolved FRET (htrFRET).
Stable Flp-In™ T-REx™ 293 cell lines able to inducibly express each of these receptor forms upon addition of doxycycline, and a cell line able to express both the hM3RASSL constitutively and hM2WT in a doxycycline inducible manner were generated. In these cells both hM3RASSL and hM2WT were detected after treatment with different concentrations of doxycycline via Western blots using tag-specific antibodies. Radioligand binding using [3H]-QNB indicated that similar amounts of hM2WT and hM3RASSL were expressed following induction with 5 ng.ml-1 doxycycline in the cells co-expressing the two receptors. Expression of the receptors was observed at the surface of live cells following labelling of the expressed receptors with SNAP and CLIP-specific cell impermeant substrates. Following induction with doxycycline each of hM2WT and hM3RASSL homomers and hM2WT-hM3RASSL heteromers were identified. Detection of oligomers was achieved following co-labelling with htrFRET-compatible substrates.
Occupancy of hM2WT-hM3RASSL heteromers with the hM2WT agonist carbachol resulted in a marked, time and concentration-dependent decrease in detected heteromers and a concomitant, concentration-dependent increase in hM2WT homomers. The dynamics of interchange between heteromers and homomers was investigated by using a multiplex labelling approach and htrFRET. This method involved labelling with one energy donor and two energy acceptors capable of emitting at distinct wavelengths. Results confirmed the hM2WT-hM3RASSL heteromer to hM2WT homomer transition upon selective carbachol-mediated activation of hM2WT. A small increase in the hM3RASSL homomer was detected upon activation of the hM3RASSL with the selective agonist clozapine-N-oxide, but this was only observed in the absence of heteromers.
Despite the presence of hM2WT-hM3RASSL heteromers the functional pharmacology of hM2WT and hM3RASSL receptor specific agonists was largely unaltered.