Inexpensive methods for live imaging of central pattern generator activity in the Drosophila larval locomotor system

This work was supported by EPSRC Doctoral Training grant EP/L505079/1) and a European Research Council Grant to MCG (640012) It was also supported by a British Council GREAT Scholarship (to VS) as well as grants from the St Andrews Learning and Teaching Initiative and the McCall-McBain Foundation to SRP. This work was also supported by the open source DIY communities as described throughout the paper.Central pattern generators (CPGs) are neural networks that produce rhythmic motor activity in the absence of sensory input. CPGs produce ‘fictive’ behaviours in vitro which parallel activity seen in intact animals. CPG networks have been identified in a wide variety of model organisms and have been shown to be critical for generating rhythmic behaviours such as swimming, walking, chewing and breathing. Work with CPG preparations has led to fundamental advances in neuroscience; however, most CPG preparations involve intensive dissections and require sophisticated electrophysiology equipment, making export to teaching laboratories problematic. Here we present an integrated approach for bringing the study of locomotor CPGs in Drosophila larvae into teaching laboratories. First, we present freely available genetic constructs that enable educators to express genetically encoded calcium indicators in cells of interest in the larval central nervous system. Next, we describe how to isolate the larval central nervous system and prepare it for live imaging. We then show how to modify standard compound microscopes to enable fluorescent imaging using 3D printed materials and inexpensive optical components. Finally, we show how to use the free image analysis programme ImageJ and freely available features in the signal analysis programme DataView to analyse rhythmic CPG activity in the larval CNS. Comparison of results to those obtained on research equipment shows that signal-to-noise levels are comparable and core features of larval CPG activity can be observed. Overall, this work shows the viability of exporting live imaging experiments to low cost environments and paves the way for new teaching laboratory exercises revolving around optical imaging of CPG activity.Publisher PDFPeer reviewe

Interpreting the extended emission around three nearby debris disc host stars

Cool debris discs are a relic of the planetesimal formation process around their host star, analogous to the solar system's Edgeworth-Kuiper belt. As such, they can be used as a proxy to probe the origin and formation of planetary systems like our own. The Herschel Open Time Key Programmes "DUst around NEarby Stars" (DUNES) and "Disc Emission via a Bias-free Reconnaissance in the Infrared/Submillimetre" (DEBRIS) observed many nearby, sun-like stars at far-infrared wavelengths seeking to detect and characterize the emission from their circumstellar dust. Excess emission attributable to the presence of dust was identified from around $\sim$ 20% of stars. Herschel's high angular resolution ($\sim$ 7" FWHM at 100 $\mu$m) provided the capacity for resolving debris belts around nearby stars with radial extents comparable to the solar system (50 to 100 au). As part of the DUNES and DEBRIS surveys, we obtained observations of three debris disc stars, HIP 22263 (HD 30495), HIP 62207 (HD 110897), and HIP 72848 (HD 131511), at far-infrared wavelengths with the Herschel PACS instrument. Combining these new images and photometry with ancilliary data from the literature, we undertook simultaneous multi-wavelength modelling of the discs' radial profiles and spectral energy distributions using three different methodologies: single annulus, modified black body, and a radiative transfer code. We present the first far-infrared spatially resolved images of these discs and new single-component debris disc models. We characterize the capacity of the models to reproduce the disc parameters based on marginally resolved emission through analysis of two sets of simulated systems (based on the HIP 22263 and HIP 62207 data) with the noise levels typical of the Herschel images. We find that the input parameter values are recovered well at noise levels attained in the observations presented here.Comment: 13 pages, 5 figures, 5 tables, accepted for publication in A&

Far-infrared and sub-millimetre imaging of HD 76582's circumstellar disc

Debris discs, the tenuous rocky and icy remnants of planet formation, are believed to be evidence for planetary systems around other stars. The JCMT/SCUBA-2 debris disc legacy survey ‘SCUBA-2 Observations of Nearby Stars’ (SONS) observed 100 nearby stars, amongst them HD 76582, for evidence of such material. Here, we present imaging observations by JCMT/SCUBA-2 and Herschel/PACS at sub-millimetre and far-infrared wavelengths, respectively. We simultaneously model the ensemble of photometric and imaging data, spanning optical to sub-millimetre wavelengths, in a self-consistent manner. At far-infrared wavelengths, we find extended emission from the circumstellar disc providing a strong constraint on the dust spatial location in the outer system, although the angular resolution is too poor to constrain the interior of the system. In the sub-millimetre, photometry at 450 and 850 µm reveals a steep fall-off that we interpret as a disc dominated by moderately sized dust grains (amin = 36 µm), perhaps indicative of a non-steady-state collisional cascade within the disc. A disc architecture of three distinct annuli, comprising an unresolved component at 20 au and outer components at 80 and 270 au, along with a very steep particle size distribution (γ = 5), is proposed to match the observations

Rapid nano-gram scale screening method of micro-arrays to evaluate drug-polymer blends using high-throughput printing technology

A miniaturized, high-throughput assay was optimized to screen polymer-drug solid dispersions using a 2-D Ink-jet printer. By simply printing nanoliter amounts of polymer and drug solutions onto an inert surface, drug:polymer micro-dots of tunable composition were produced in an easily-addressable micro-array format. The amount of material printed for each dried spot ranged from 25 ng to 650 ng. These arrays were used to assess the stability of drug:polymer dispersions with respect to recrystallization, using polarized light microscopy. One array with a panel of 6 drugs formulated at different ratios with Poly (vinylpyrrolidone-vinyl acetate) copolymer (PVPVA) was developed to estimate a possible bulk (gram-scale) approximation threshold from the final printed nano amount of formulation. Another array was printed at a fixed final amount of material to establish a literature comparison of one drug formulated with different commercial polymers for validation. This new approach may offer significant efficiency in pharmaceutical formulation screening, with each experiment in the nano-micro-array format requiring from 3 up to 6 orders of magnitude lower amounts of sample than conventional screening methods

Repositioning of special schools within a specialist, personalised educational marketplace - the need for a representative principle

This paper considers how notions of inclusive education as defined in the United Nations Educational, Scientific and Cultural Organization (UNESCO) Salamanca Agreement (1994) have become dissipated, and can be developed and reframed to encourage their progress. It analyses the discourse within a range of academic, legal and media texts, exploring how this dissipation has taken place within the UK. Using data from 78 specialist school websites it contextualises this change in the use of the terms and ideas of inclusion with the rise of two other constructs, the 'specialist school' and 'personalisation'. It identifies the need for a precisely defined representative principle to theorise the type of school which inclusion aims to achieve, which cannot be subsumed by segregated providers. It suggests that this principle should not focus on the individual, but draw upon a liberal/democratic view of social justice, underlining inclusive education's role in removing social barriers that prevent equity, access and participation for all

Plant communities affect arbuscular mycorrhizal fungal diversity and community composition in grassland microcosms

The diversity of arbuscular mycorrhizal (AM) fungi was investigated in an unfertilized limestone grassland soil supporting different synthesized vascular plant assemblages that had developed for 3 yr. The experimental treatments comprised: bare soil; monocultures of the nonmycotrophic sedge Carex flacca; monocultures of the mycotrophic grass Festuca ovina; and a species-rich mixture of four forbs, four grasses and four sedges. The diversity of AM fungi was analysed in roots of Plantago lanceolata bioassay seedlings using terminal-restriction fragment length polymorphism (T-RFLP). The extent of AM colonization, shoot biomass and nitrogen and phosphorus concentrations were also measured. The AM diversity was affected significantly by the floristic composition of the microcosms and shoot phosphorus concentration was positively correlated with AM diversity. The diversity of AM fungi in P. lanceolata decreased in the order: bare soil > C. flacca > 12 species > F. ovina. The unexpectedly high diversity in the bare soil and sedge monoculture likely reflects differences in the modes of colonization and sources of inoculum in these treatments compared with the assemblages containing established AM-compatible plants

Novel Approaches to Detect Serum Biomarkers for Clinical Response to Interferon-β Treatment in Multiple Sclerosis

Interferon beta (IFNβ) is the most common immunomodulatory treatment for relapsing-remitting multiple sclerosis (RRMS). However, some patients fail to respond to treatment. In this study, we identified putative clinical response markers in the serum and plasma of people with multiple sclerosis (MS) treated with IFNβ. In a discovery-driven approach, we use 2D-difference gel electrophoresis (DIGE) to identify putative clinical response markers and apply power calculations to identify the sample size required to further validate those markers. In the process we have optimized a DIGE protocol for plasma to obtain cost effective and high resolution gels for effective spot comparison. APOA1, A2M, and FIBB were identified as putative clinical response markers. Power calculations showed that the current DIGE experiment requires a minimum of 10 samples from each group to be confident of 1.5 fold difference at the p<0.05 significance level. In a complementary targeted approach, Cytometric Beadarray (CBA) analysis showed no significant difference in the serum concentration of IL-6, IL-8, MIG, Eotaxin, IP-10, MCP-1, and MIP-1α, between clinical responders and non-responders, despite the association of these proteins with IFNβ treatment in MS

Real time Raman imaging to understand dissolution performance of amorphous solid dispersions

We have employed for the first time Raman spectroscopic imaging along with multi-variate curve resolution (MCR) analysis to investigate in real time and in-situ the dissolution mechanisms that underpin amorphous solid dispersions, with data being collected directly from the dosage form itself. We have also employed a novel rotating disk dissolution rate (RDDR) methodology to track, through the use of high-performance liquid chromatography (HPLC), the dissolution trends of both drug and polymer simultaneously in multi-component systems. Two formulations of poorly water-soluble felodipine in a polymeric matrix of copovidone VA64 which have different drug loadings of 5% and 50% w/w were used as models with the aim of studying the effects of increasing the amount of active ingredient on the dissolution performance. It was found that felodipine and copovidone in the 5% dispersion dissolve with the same dissolution rate and that no Raman spectral changes accompanied the dissolution, indicating that the two components dissolve as single entity, whose behaviour is dominated by water-soluble copovidone. For the 50% drug-loaded dispersion, partial RDDR values of both felodipine and copovidone were found to be extremely low. MCR Raman maps along with classical Raman/X-ray powder diffraction (XRPD) characterisation revealed that after an initial loss of copovidone from the extrudate the drug re-crystallises, pointing to a release dynamics dependent on the low water solubility and high hydrophobicity of felodipine. Raman imaging revealed different rates of transition from amorphous to crystalline felodipine at different locations within the dosage form

Characterising online museum users: a study of the National Museums Liverpool museum website

Museums are increasing access to their collections and providing richer user experiences via web-based interfaces. However, they are seeing high numbers of users looking at only one or two pages within 10 s and then leaving. To reduce this rate, a better understanding of the type of user who visits a museum website is required. Existing models for museum website users tend to focus on groups that are readily accessible for study or provide little detail in their definitions of the groups. This paper presents the results of a large-scale user survey for the National Museums Liverpool museum website in which data on a wide range of user characteristics were collected regarding their current visit to provide a better understanding of their motivations, tasks, engagement and domain knowledge. Results show that the frequently understudied general public and non-professional users make up the majority (approximately 77%) of the respondents

The Halo Occupation Distribution of Black Holes: Dependence on Mass

We investigate the halo occupation distribution (HOD) of black holes within a hydrodynamic cosmological simulation that directly follows black hole growth. Similar to the HOD of galaxies/subhalos, we find that the black hole occupation number can be described by the form N_BH proportional to 1+ (M_Host)^alpha where alpha evolves mildly with redshift indicating that a given mass halo (M_Host) at low redshift tends to host fewer BHs than at high redshift (as expected as a result of galaxy and BH mergers). We further divide the occupation number into contributions from black holes residing in central and satellite galaxies within a halo. The distribution of M_BH within halos tends to consist of a single massive BH (distributed about a peak mass strongly correlated with M_Host), and a collection of relatively low-mass secondary BHs, with weaker correlation with M_Host. We also examine the spatial distribution of BHs within their host halos, and find they typically follow a power-law radial distribution (i.e. much more centrally concentrated than the subhalo distribution). Finally, we characterize the host mass for which BH growth is feedback dominated (e.g. star formation quenched). We show that halos with M_Host > 3 * 10^12 M_sun have primary BHs that are feedback dominated by z~3 with lower mass halos becoming increasingly more affected at lower redshift.Comment: 10 pages, 7 figures, submitted to MNRA