Phosphomimetic Mutation of Ser-187 of SNAP-25 Increases both Syntaxin Binding and Highly Ca2+-sensitive Exocytosis

The phosphorylation targets that mediate the enhancement of exocytosis by PKC are unknown. PKC phosporylates the SNARE protein SNAP-25 at Ser-187. We expressed mutants of SNAP-25 using the Semliki Forest Virus system in bovine adrenal chromaffin cells and then directly measured the Ca2+ dependence of exocytosis using photorelease of caged Ca2+ together with patch-clamp capacitance measurements. A flash of UV light used to elevate [Ca2+]i to several μM and release the highly Ca2+-sensitive pool (HCSP) of vesicles was followed by a train of depolarizing pulses to elicit exocytosis from the less Ca2+-sensitive readily releasable pool (RRP) of vesicles. Carbon fiber amperometry confirmed that the amount and kinetics of catecholamine release from individual granules were similar for the two phases of exocytosis. Mimicking PKC phosphorylation with expression of the S187E SNAP-25 mutant resulted in an approximately threefold increase in the HCSP, whereas the response to depolarization increased only 1.5-fold. The phosphomimetic S187D mutation resulted in an ∼1.5-fold increase in the HCSP but a 30% smaller response to depolarization. In vitro binding assays with recombinant SNARE proteins were performed to examine shifts in protein–protein binding that may promote the highly Ca2+-sensitive state. The S187E mutant exhibited increased binding to syntaxin but decreased Ca2+-independent binding to synaptotagmin I. Mimicking phosphorylation of the putative PKA phosphorylation site of SNAP-25 with the T138E mutation decreased binding to both syntaxin and synaptotagmin I in vitro. Expressing the T138E/ S187E double mutant in chromaffin cells demonstrated that enhancing the size of the HCSP correlates with an increase in SNAP-25 binding to syntaxin in vitro, but not with Ca2+-independent binding of SNAP-25 to synaptotagmin I. Our results support the hypothesis that exocytosis triggered by lower Ca2+ concentrations (from the HCSP) occurs by different molecular mechanisms than exocytosis triggered by higher Ca2+ levels

Distracted , Stressed, and Confused: The Effects of Distraction and Stress on Memory Retention

Previous research has tested the effects of test anxiety and memory impairment due to stress, but little research has investigated the interplay of these factors with environmental distraction. Our study aims to fill in this gap by examining the association between distraction, stress and information recall in an effort to identify optimal testing experiences within the academic environment. Professors and administrators can improve academic success based upon the knowledge of how stressful stimuli affects test performance. In this experiment, we seek to establish a connection between stress, distraction, and memory recall within academic testing. By employing distracting stimuli in a controlled environment, we hope to uncover further insight into whether students perform better or worse in stressful environments. Understanding how added stressors influence student performance can help professors and administrators structure effective learning environments. Likewise, our findings might also add to our knowledge of the influence of marginal distractibility on the recall of class content. We speculate that there is an optimal range of distractibility and stress within the academic setting. Understanding how added stressors and distractions influence student performance can help college administrators structure effective learning environments. During the experimental phase, our team examined the effects of stress on material recollection by using a graded versus an ungraded exam. We expected that the combination of stress and distracted conditions will result in a significantly diminished student memory performance, compared to the marginal declines caused by stress or distraction alone. Additionally, we explored the role of environmental distraction on memory performance by measuring how students responded to the sudden announcement of a graded test. We hypothesized that if a sudden stressor is presented, one will respond negatively, unable to recall the desired information. Thus, we expect the student to perform less successfully as one would without the stressor. We suspected that environmental noise and distractions increased, academic anxiety will increase as well, causing memory deficits for the affected students. Findings are under review, but are expected to support the stated hypotheses

Громадянське суспільство як категорія етнополітології

У статті «громадянське суспільство» розглядається як категорія етнополітології, акцентується увага на визначальному впливі громадянського суспільства на формування сучасної нації.В статье «гражданское общество» рассматривается как категория этнополитологии, акцентируется внимание на определяющем влиянии гражданского общества на формирование современной нации.In the article «civil society» is considered as a category ethnopolitology, emphasized the decisive influence of civil society on the formation of modern nation

Hydrothermal cooling of the ocean crust: Insights from ODP Hole 1256D

publisher: Elsevier articletitle: Hydrothermal cooling of the ocean crust: Insights from ODP Hole 1256D journaltitle: Earth and Planetary Science Letters articlelink: http://dx.doi.org/10.1016/j.epsl.2017.01.010 content_type: article copyright: © 2017 The Author(s). Published by Elsevier B.V

Characterisation of insulin analogues therapeutically available to patients

The structure and function of clinical dosage insulin and its analogues were assessed. This included ‘native insulins’ (human recombinant, bovine, porcine), ‘fast-acting analogues’ (aspart, glulisine, lispro) and ‘slow-acting analogues’ (glargine, detemir, degludec). Analytical ultracentrifugation, both sedimentation velocity and equilibrium experiments, were employed to yield distributions of both molar mass and sedimentation coefficient of all nine insulins. Size exclusion chromatography, coupled to multi-angle light scattering, was also used to explore the function of these analogues. On ultracentrifugation analysis, the insulins under investigation were found to be in numerous conformational states, however the majority of insulins were present in a primarily hexameric conformation. This was true for all native insulins and two fast-acting analogues. However, glargine was present as a dimer, detemir was a multi-hexameric system, degludec was a dodecamer (di-hexamer) and glulisine was present as a dimer-hexamer-dihexamer system. However, size-exclusion chromatography showed that the two hexameric fast-acting analogues (aspart and lispro) dissociated into monomers and dimers due to the lack of zinc in the mobile phase. This comprehensive study is the first time all nine insulins have been characterised in this way, the first time that insulin detemir have been studied using analytical ultracentrifugation and the first time that insulins aspart and glulisine have been studied using sedimentation equilibrium. The structure and function of these clinically administered insulins is of critical importance and this research adds novel data to an otherwise complex functional physiological protein

Paradoxical family practices: LGBTQ+ young people, mental health and wellbeing

This article will explore how LGBTQ+ young people sustain, and in some cases survive, family relationships. We develop the concept of ‘paradoxical family practices’ and use this to demonstrate the ways in which LGBTQ+ young people manage family life through everyday emotion work. This highlights: (1) how families ordinarily navigate heteronormativity and ‘issues’ of gender/sexuality; (2) the efficacy of ‘paradoxical family practices’ as a conceptual tool; (3) the value of emotion-centred multiple qualitative methods to explore the lives of LGBTQ+ young people and mental health. Findings derive from a small-scale UK study funded by the Wellcome Trust (UNS39780) and were generated through a two-stage methodology comprising digital/paper emotion maps and qualitative interviews with LGBTQ+ young people aged 16–25 (n = 12) followed by diary methods and follow-up interviews (n = 9). Interviews were also completed with ‘family members’ (n = 7)

Characterization of the in situ magnetic architecture of oceanic crust (Hess Deep) using near-source vector magnetic data

Marine magnetic anomalies are a powerful tool for detecting geomagnetic polarity reversals, lithological boundaries, topographic contrasts, and alteration fronts in the oceanic lithosphere. Our aim here is to detect lithological contacts in fast-spreading lower crust and shallow mantle by characterizing magnetic anomalies and investigating their origins. We conducted a high-resolution, near-bottom, vector magnetic survey of crust exposed in the Hess Deep “tectonic window” using the remotely operated vehicle (ROV) Isis during RRS James Cook cruise JC21 in 2008. Hess Deep is located at the western tip of the propagating rift of the Cocos-Nazca plate boundary near the East Pacific Rise (EPR) (2°15′N, 101°30′W). ROV Isis collected high-resolution bathymetry and near-bottom magnetic data as well as seafloor samples to determine the in situ lithostratigraphy and internal structure of a section of EPR lower crust and mantle exposed on the steep (~20°dipping) south facing slope just north of the Hess Deep nadir. Ten magnetic profiles were collected up the slope using a three-axis fluxgate magnetometer mounted on ROV Isis. We develop and extend the vertical magnetic profile (VMP) approach of Tivey (1996) by incorporating, for the first time, a three-dimensional vector analysis, leading to what we here termed as “vector vertical magnetic profiling” approach. We calculate the source magnetization distribution, the deviation from two dimensionality, and the strike of magnetic boundaries using both the total field Fourier-transform inversion approach and a modified differential vector magnetic analysis. Overall, coherent, long-wavelength total field anomalies are present with a strong magnetization contrast between the upper and lower parts of the slope. The total field anomalies indicate a coherently magnetized source at depth. The upper part of the slope is weakly magnetized and magnetic structure follows the underlying slope morphology, including a “bench” and lobe-shaped steps, imaged by microbathymetry. The lower part of the slope is strongly magnetized, with a gradual reduction in amplitude from east to west across the slope. Surface morphology and recent drilling results indicate that the slope has been affected by mass wasting, but the observation of internally coherent magnetization distributions within the upper and lower slopes suggest that the disturbance is surficial. We attribute the spatial differences in magnetization distribution to the combination of changes in in situ lithology and depth to the source. These survey lines document the first magnetic profiles that capture the gabbro-ultramafic and possibly dike-gabbro boundaries in fast-spreading lower crust

Resurrecting biodiversity: advanced assisted reproductive technologies and biobanking

Biodiversity is defined as the presence of a variety of living organisms on the Earth that is essential for human survival. However, anthropogenic activities are causing the sixth mass extinction, threatening even our own species. For many animals, dwindling numbers are becoming fragmented populations with low genetic diversity, threatening long-term species viability. With extinction rates 1000–10,000 times greater than natural, ex situ and in situ conservation programmes need additional support to save species. The indefinite storage of cryopreserved (−196°C) viable cells and tissues (cryobanking), followed by assisted or advanced assisted reproductive technology (ART: utilisation of oocytes and spermatozoa to generate offspring; aART: utilisation of somatic cell genetic material to generate offspring), may be the only hope for species’ long-term survival. As such, cryobanking should be considered a necessity for all future conservation strategies. Following cryopreservation, ART/aART can be used to reinstate lost genetics back into a population, resurrecting biodiversity. However, for this to be successful, species-specific protocol optimisation and increased knowledge of basic biology for many taxa are required. Current ART/aART is primarily focused on mammalian taxa; however, this needs to be extended to all, including to some of the most endangered species: amphibians. Gamete, reproductive tissue and somatic cell cryobanking can fill the gap between losing genetic diversity today and future technological developments. This review explores species prioritisation for cryobanking and the successes and challenges of cryopreservation and multiple ARTs/aARTs. We here discuss the value of cryobanking before more species are lost and the potential of advanced reproductive technologies not only to halt but also to reverse biodiversity loss

Fertility, Living Arrangements, Care and Mobility

There are four main interconnecting themes around which the contributions in this book are based. This introductory chapter aims to establish the broad context for the chapters that follow by discussing each of the themes. It does so by setting these themes within the overarching demographic challenge of the twenty-first century – demographic ageing. Each chapter is introduced in the context of the specific theme to which it primarily relates and there is a summary of the data sets used by the contributors to illustrate the wide range of cross-sectional and longitudinal data analysed