Multicolour correlative imaging using phosphor probes

Correlative light and electron microscopy exploits the advantages of optical methods, such as multicolour probes and their use in hydrated live biological samples, to locate functional units, which are then correlated with structural details that can be revealed by the superior resolution of electron microscopes. One difficulty is locating the area imaged by the electron beam in the much larger optical field of view. Multifunctional probes that can be imaged in both modalities and thus register the two images are required. Phosphor materials give cathodoluminescence (CL) optical emissions under electron excitation. Lanthanum phosphate containing thulium or terbium or europium emits narrow bands in the blue, green and red regions of the CL spectrum; they may be synthesised with very uniform-sized crystals in the 10- to 50-nm range. Such crystals can be imaged by CL in the electron microscope, at resolutions limited by the particle size, and with colour discrimination to identify different probes. These materials also give emissions in the optical microscope, by multiphoton excitation. They have been deposited on the surface of glioblastoma cells and imaged by CL. Gadolinium oxysulphide doped with terbium emits green photons by either ultraviolet or electron excitation. Sixty-nanometre crystals of this phosphor have been imaged in the atmospheric scanning electron microscope (JEOL ClairScope). This probe and microscope combination allow correlative imaging in hydrated samples. Phosphor probes should prove to be very useful in correlative light and electron microscopy, as fiducial markers to assist in image registration, and in high/super resolution imaging studies

Activation of mGluR5 Induces Rapid and Long-Lasting Protein Kinase D Phosphorylation in Hippocampal Neurons

Metabotropic glutamate receptors (mGluRs), including mGluR5, play a central role in regulating the strength and plasticity of synaptic connections in the brain. However, the signaling pathways that connect mGluRs to their downstream effectors are not yet fully understood. Here, we report that stimulation of mGluR5 in hippocampal cultures and slices results in phosphorylation of protein kinase D (PKD) at the autophosphorylation site Ser-916. This phosphorylation event occurs within 30 s of stimulation, persists for at least 24 h, and is dependent on activation of phospholipase C and protein kinase C. Our data suggest that activation of PKD may represent a novel signaling pathway linking mGluR5 to its downstream targets. These findings have important implications for the study of the molecular mechanisms underlying mGluR-dependent synaptic plasticity.Howard Hughes Medical InstituteFRAXA Research FoundationNational Institute of Mental Health (U.S.)Eunice Kennedy Shriver National Institute of Child Health and Human Development (U.S.

Development of high temperature, radiation hard detectors based on diamond

© 2016 Single crystal CVD diamond has many desirable properties compared to current, well developed, detector materials; exceptional radiation, chemical and physical hardness, chemical inertness, low Z (close to human tissue, good for dosimetry), wide bandgap and an intrinsic pathway to fast neutron detection through the 12C(n,α)9Be reaction. However effective exploitation of these properties requires development of a suitable metallisation scheme to give stable contacts for high temperature applications. To best utilise available processing techniques to optimise sensor response through geometry and conversion media configurations, a reliable model is required. This must assess the performance in terms of spectral response and overall efficiency as a function of detector and converter geometry. The same is also required for proper interpretation of experimental data. Sensors have been fabricated with varying metallisation schemes indented to permit high temperature operation; Present test results indicate that viable fabrication schemes for high temperature contacts have been developed and present modelling results, supported by preliminary data from partners indicate simulations provide a useful representation of response

Disability patterns over the first year after a diagnosis of epilepsy

Objective To determine the patterns and predictors of disability over the first 12 months after a diagnosis of epilepsy. Patients and methods The Sydney Epilepsy Incidence Study to Measure Illness Consequences (SEISMIC) was a prospective, multicenter, community-based study of people with newly diagnosed epilepsy in Sydney, Australia. Disability was assessed using the World Health Organization’s, Disability Assessment Schedule (WHODAS) 2.0 12-item version, at baseline (i.e. within 28 days of diagnosis) and 12 months post-diagnosis. Demographic, socioeconomic, clinical and epilepsy-related data, obtained through structured interviews, were entered into multivariable linear regression and shift analysis to determine predictors of greater disability. Results Of 259 adults (≥18 years), 190 (73%) had complete WHODAS at baseline (mean ± SD scores 4 ± 6) and follow-up (4 ± 8). After adjustment for age, sex and co-morbidity, greater overall disability at 12 months was associated with lower education (P = 0.05), economic hardship (P = 0.004), multiple antiepileptic medications (P = 0.02) and greater disability (P < 0.001) at the time of diagnosis; these variables explained 38.3% of the variance. Among the 12 WHODAS items, “being emotionally affected by health problems” was the most frequent disability problem identified at both time points (all P < 0.0001). The proportion of participants without problems in that domain improved over 12 months (from 24% to 50%, P < 0.0001), whereas the other 11 items remained relatively stable. Independent baseline predictors of a worse emotional outcome at 12 months were severe/extreme emotional distress (odds ratio [OR] 4.52, 95% confidence intervals [CI] 1.67–12.24), economic hardship (OR 2.30, 95% CI 1.24–4.25) and perceived stigma (OR 2.02, 95% CI 1.03–3.93). Conclusion Most people report problems with emotional health after a diagnosis of epilepsy but many recover over the next 12 months. Services addressing the social and psychological impact of diagnosis may be needed to improve outcome

Development of high temperature, radiation hard detectors based on diamond

Single crystal CVD diamond has many desirable properties compared to current, well developed, detector materials; exceptional radiation, chemical and physical hardness, chemical inertness, low Z (close to human tissue, good for dosimetry), wide bandgap and an intrinsic pathway to fast neutron detection through the 12C(n,α)9Be reaction. However effective exploitation of these properties requires development of a suitable metallisation scheme to give stable contacts for high temperature applications. To best utilise available processing techniques to optimise sensor response through geometry and conversion media configurations, a reliable model is required. This must assess the performance in terms of spectral response and overall efficiency as a function of detector and converter geometry. The same is also required for proper interpretation of experimental data. Sensors have been fabricated with varying metallisation schemes indented to permit high temperature operation; Present test results indicate that viable fabrication schemes for high temperature contacts have been developed and present modelling results, supported by preliminary data from partners indicate simulations provide a useful representation of response.The EPSRC (Engineering and Physical Sciences Research Council) (Grant no: EP/L504671/1) and InnovateUK

Activation of Akt by the Bacterial Inositol Phosphatase, SopB, is Wortmannin Insensitive

Salmonella enterica uses effector proteins translocated by a Type III Secretion System to invade epithelial cells. One of the invasion-associated effectors, SopB, is an inositol phosphatase that mediates sustained activation of the pro-survival kinase Akt in infected cells. Canonical activation of Akt involves membrane translocation and phosphorylation and is dependent on phosphatidyl inositide 3 kinase (PI3K). Here we have investigated these two distinct processes in Salmonella infected HeLa cells. Firstly, we found that SopB-dependent membrane translocation and phosphorylation of Akt are insensitive to the PI3K inhibitor wortmannin. Similarly, depletion of the PI3K regulatory subunits p85α and p85ß by RNAi had no inhibitory effect on SopB-dependent Akt phosphorylation. Nevertheless, SopB-dependent phosphorylation does depend on the Akt kinases, PDK1 and rictor-mTOR. Membrane translocation assays revealed a dependence on SopB for Akt recruitment to Salmonella ruffles and suggest that this is mediated by phosphoinositide (3,4) P2 rather than phosphoinositide (3,4,5) P3. Altogether these data demonstrate that Salmonella activates Akt via a wortmannin insensitive mechanism that is likely a class I PI3K-independent process that incorporates some essential elements of the canonical pathway

"I won't be staying here for long": a qualitative study on the retention of migrant nurses in Ireland

<p>Abstract</p> <p>Background</p> <p>Although international nurse recruitment campaigns have succeeded in attracting large numbers of migrant nurses to countries such as Ireland, where domestic supply has not kept pace with demand, the long-term success of such initiatives from a workforce planning perspective will depend on the extent to which these nurses can be retained in destination countries.</p> <p>Methods</p> <p>This paper draws on qualitative, in-depth interviews undertaken with 21 migrant nurses in Ireland, focusing specifically on their future migration intentions.</p> <p>Results</p> <p>Our findings indicate that more than half of the respondents are considering migration onwards, for the most part because the destination country has failed to provide them with sufficient stability, particularly in terms of citizenship and family reunification. In considering onward migration, factors outside the health system were of most concern to those interviewed.</p> <p>Conclusion</p> <p>This demonstrates the need for destination countries to take a broader and more long-term approach to international nurse recruitment, rather than regarding it as an inexpensive way to fill gaps within the health care system.</p

Ionotropic Glutamate Receptor AMPA 1 Is Associated with Ovulation Rate

Ionotropic glutamate receptors mediate most excitatory neurotransmission in the central nervous system by opening ion channels upon the binding of glutamate. Despite the essential roles of glutamate in the control of reproduction and anterior pituitary hormone secretion, there is a limited understanding of how glutamate receptors control ovulation. Here we reveal the function of the ionotropic glutamate receptor AMPA-1 (GRIA1) in ovulation. Based on a genome-wide association study in Bos taurus, we found that ovulation rate is influenced by a variation in the N-terminal leucine/isoleucine/valine-binding protein (LIVBP) domain of GRIA1, in which serine is replaced by asparagine. GRIA1Asn has a weaker affinity to glutamate than GRIA1Ser, both in Xenopus oocytes and in the membrane fraction of bovine brain. This single amino acid substitution leads to the decreased release of gonadotropin-releasing hormone (GnRH) in immortalized hypothalamic GT1-7 cells. Cows with GRIA1Asn have a slower luteinizing hormone (LH) surge than cows with GRIA1Ser. In addition, cows with GRIA1Asn possess fewer immature ovarian follicles before superovulation and have a lower response to hormone treatment than cows with GRIA1Ser. Our work identified that GRIA1 is a critical mediator of ovulation and that GRIA1 might be a useful target for reproductive therapy