Nanoscale-targeted patch-clamp recordings of functional presynaptic ion channels

Important modulatory roles have been attributed to presynaptic NMDA receptors (NMDARs) located on cerebellar interneuron terminals. Evidence supporting a presynaptic location includes an increase in the frequency of mini events following the application of NMDA and gold particle-labelled NMDA receptor antibody localisation. However, more recent work, using calcium indicators, casts doubt on the idea of presynaptic NMDARs because basket cell varicosities did not show the expected calcium rise following either the local iontophoresis of L-aspartate or the two-photon uncaging of glutamate. (In theory such calcium imaging is sensitive enough to detect the calcium rise from even a single activated receptor.) It has therefore been suggested that the effects of NMDA are mediated via the activation of somatodendritic channels, which subsequently cause a subthreshold depolarization of the axon. Here we report results from a vibrodissociated preparation of cerebellar Purkinje cells, in which the interneuron cell bodies are no longer connected but many of their terminal varicosities remain attached and functional. This preparation can retain both inhibitory and excitatory inputs. We find that the application of NMDA increases the frequency of both types of synaptic event. The characteristics of these events suggest they can originate from interneuron, parallel fiber and even climbing fiber terminals. Interestingly, retrograde signalling seems to activate only the inhibitory terminals. Finally, antibody staining of these cells shows NMDAR-like immunoreactivity co-localised with synaptic markers. Since the Purkinje cells show no evidence of postsynaptic NMDAR-mediated currents, we conclude that functional NMDA receptors are located on presynaptic terminals

Moderate hypothermia within 6 h of birth plus inhaled xenon versus moderate hypothermia alone after birth asphyxia (TOBY-Xe): a proof-of-concept, open-label, randomised controlled trial

Background Moderate cooling after birth asphyxia is associated with substantial reductions in death and disability, but additional therapies might provide further benefit. We assessed whether the addition of xenon gas, a promising novel therapy, after the initiation of hypothermia for birth asphyxia would result in further improvement. Methods Total Body hypothermia plus Xenon (TOBY-Xe) was a proof-of-concept, randomised, open-label, parallel-group trial done at four intensive-care neonatal units in the UK. Eligible infants were 36–43 weeks of gestational age, had signs of moderate to severe encephalopathy and moderately or severely abnormal background activity for at least 30 min or seizures as shown by amplitude-integrated EEG (aEEG), and had one of the following: Apgar score of 5 or less 10 min after birth, continued need for resuscitation 10 min after birth, or acidosis within 1 h of birth. Participants were allocated in a 1:1 ratio by use of a secure web-based computer-generated randomisation sequence within 12 h of birth to cooling to a rectal temperature of 33·5°C for 72 h (standard treatment) or to cooling in combination with 30% inhaled xenon for 24 h started immediately after randomisation. The primary outcomes were reduction in lactate to N-acetyl aspartate ratio in the thalamus and in preserved fractional anisotropy in the posterior limb of the internal capsule, measured with magnetic resonance spectroscopy and MRI, respectively, within 15 days of birth. The investigator assessing these outcomes was masked to allocation. Analysis was by intention to treat. This trial is registered with ClinicalTrials.gov, number NCT00934700, and with ISRCTN, as ISRCTN08886155. Findings The study was done from Jan 31, 2012, to Sept 30, 2014. We enrolled 92 infants, 46 of whom were randomly assigned to cooling only and 46 to xenon plus cooling. 37 infants in the cooling only group and 41 in the cooling plus xenon group underwent magnetic resonance assessments and were included in the analysis of the primary outcomes. We noted no significant differences in lactate to N-acetyl aspartate ratio in the thalamus (geometric mean ratio 1·09, 95% CI 0·90 to 1·32) or fractional anisotropy (mean difference −0·01, 95% CI −0·03 to 0·02) in the posterior limb of the internal capsule between the two groups. Nine infants died in the cooling group and 11 in the xenon group. Two adverse events were reported in the xenon group: subcutaneous fat necrosis and transient desaturation during the MRI. No serious adverse events were recorded. Interpretation Administration of xenon within the delayed timeframe used in this trial is feasible and apparently safe, but is unlikely to enhance the neuroprotective effect of cooling after birth asphyxia

Fluids in cosmology

We review the role of fluids in cosmology by first introducing them in General Relativity and then by applying them to a FRW Universe's model. We describe how relativistic and non-relativistic components evolve in the background dynamics. We also introduce scalar fields to show that they are able to yield an inflationary dynamics at very early times (inflation) and late times (quintessence). Then, we proceed to study the thermodynamical properties of the fluids and, lastly, its perturbed kinematics. We make emphasis in the constrictions of parameters by recent cosmological probes.Comment: 34 pages, 4 figures, version accepted as invited review to the book "Computational and Experimental Fluid Mechanics with Applications to Physics, Engineering and the Environment". Version 2: typos corrected and references expande

Effects of cash transfers on Children's health and social protection in Sub-Saharan Africa: differences in outcomes based on orphan status and household assets

BACKGROUND: Unconditional and conditional cash transfer programmes (UCT and CCT) show potential to improve the well-being of orphans and other children made vulnerable by HIV/AIDS (OVC). We address the gap in current understanding about the extent to which household-based cash transfers differentially impact individual children’s outcomes, according to risk or protective factors such as orphan status and household assets. METHODS: Data were obtained from a cluster-randomised controlled trial in eastern Zimbabwe, with random assignment to three study arms – UCT, CCT or control. The sample included 5,331 children ages 6-17 from 1,697 households. Generalized linear mixed models were specified to predict OVC health vulnerability (child chronic illness and disability) and social protection (birth registration and 90% school attendance). Models included child-level risk factors (age, orphan status); household risk factors (adults with chronic illnesses and disabilities, greater household size); and household protective factors (including asset-holding). Interactions were systematically tested. RESULTS: Orphan status was associated with decreased likelihood for birth registration, and paternal orphans and children for whom both parents’ survival status was unknown were less likely to attend school. In the UCT arm, paternal orphans fared better in likelihood of birth registration compared with non-paternal orphans. Effects of study arms on outcomes were not moderated by any other risk or protective factors. High household asset-holding was associated with decreased likelihood of child’s chronic illness and increased birth registration and school attendance, but household assets did not moderate the effects of cash transfers on risk or protective factors. CONCLUSION: Orphaned children are at higher risk for poor social protection outcomes even when cared for in family-based settings. UCT and CCT each produced direct effects on children’s social protection which are not moderated by other child- and household-level risk factors, but orphans are less likely to attend school or obtain birth registration. The effects of UCT and CCT are not moderated by asset-holding, but greater household assets predict greater social protection outcomes. Intervention efforts need to focus on ameliorating the additional risk burden carried by orphaned children. These efforts might include caregiver education, and additional incentives based on efforts made specifically for orphaned children

Combinatorial Smad2/3 Activities Downstream of Nodal Signaling Maintain Embryonic/Extra-Embryonic Cell Identities during Lineage Priming

Epiblast cells in the early post-implantation stage mammalian embryo undergo a transition described as lineage priming before cell fate allocation, but signaling pathways acting upstream remain ill defined. Genetic studies demonstrate that Smad2/3 double-mutant mouse embryos die shortly after implantation. To learn more about the molecular disturbances underlying this abrupt failure, here we characterized Smad2/3-deficient embryonic stem cells (ESCs). We found that Smad2/3 double-knockout ESCs induced to form epiblast-like cells (EpiLCs) display changes in naive and primed pluripotency marker gene expression, associated with the disruption of Oct4-bound distal regulatory elements. In the absence of Smad2/3, we observed enhanced Bmp target gene expression and de-repression of extra-embryonic gene expression. Cell fate allocation into all three embryonic germ layers is disrupted. Collectively, these experiments demonstrate that combinatorial Smad2/3 functional activities are required to maintain distinct embryonic and/or extra-embryonic cell identity during lineage priming in the epiblast before gastrulation. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved

Optical detection of single non-absorbing molecules using the surface plasmon of a gold nanorod

Current optical detection schemes for single molecules require light absorption, either to produce fluorescence or direct absorption signals. This severely limits the range of molecules that can be detected, because most molecules are purely refractive. Metal nanoparticles or dielectric resonators detect non-absorbing molecules by a resonance shift in response to a local perturbation of the refractive index, but neither has reached single-protein sensitivity. The most sensitive plasmon sensors to date detect single molecules only when the plasmon shift is amplified by a highly polarizable label or by a localized precipitation reaction on the particle's surface. Without amplification, the sensitivity only allows for the statistical detection of single molecules. Here we demonstrate plasmonic detection of single molecules in realtime, without the need for labeling or amplification. We monitor the plasmon resonance of a single gold nanorod with a sensitive photothermal assay and achieve a ~ 700-fold increase in sensitivity compared to state-of-the-art plasmon sensors. We find that the sensitivity of the sensor is intrinsically limited due to spectral diffusion of the SPR. We believe this is the first optical technique that detects single molecules purely by their refractive index, without any need for photon absorption by the molecule. The small size, bio-compatibility and straightforward surface chemistry of gold nanorods may open the way to the selective and local detection of purely refractive proteins in live cells

The genetic basis of multiple sclerosis: a model for MS susceptibility

<p>Abstact</p> <p>Background</p> <p>MS-pathogenesis is known to involve both multiple environmental events, and several independent genetic risk-factors.</p> <p>Methods</p> <p>A model of susceptibility is developed and a mathematical analysis undertaken to elucidate the nature of genetic susceptibility to MS and to understand the constraints that are placed on the genetic basis of MS, both by the known epidemiological facts of this disease and by the known frequency of the HLA DRB1*1501 allele in the general populations of northern Europe and North America.</p> <p>Results</p> <p>For the large majority of cases (possibly all), MS develops, in part, because an individual is genetically susceptible. Nevertheless, 2.2% or less of the general population is genetically susceptible. Moreover, from the model, the number of susceptibility-loci that need to be in a "susceptible allelic state" to produce MS-susceptibility is small (11-18), whereas the total number of such susceptibility-loci is large (50-200), and their "frequency of susceptibility" is low (i.e., ≤ 0.12). The optimal solution to the model equations (which occurs when 80% of the loci are recessive) predicts the epidemiological data quite closely.</p> <p>Conclusions</p> <p>The model suggests that combinations of only a small number of genetic loci in a "susceptible allelic state" produce MS-susceptibility. Nevertheless, genome-wide associations studies with hundreds of thousands of SNPs, are plagued by both false-positive and false-negative identifications and, consequently, emphasis has been rightly placed on the replicability of findings. Nevertheless, because genome-wide screens don't distinguish between true susceptibility-loci and disease-modifying-loci, and because only true susceptibility-loci are constrained by the model, unraveling the two will not be possible using this approach.</p> <p>The model also suggests that HLA DRB1 may not be as uniquely important for MS-susceptibility as currently believed. Thus, this allele is only one among a hundred or more loci involved in MS susceptibility. Even though the "frequency of susceptibility" at the HLA DRB1 locus is four-fold that of other loci, the penetrance of those susceptible genotypes that include this allele is no different from those that don't. Also, almost 50% of genetically-susceptible individuals, lack this allele. Moreover, of those who have it, only a small fraction (≤ 5.2%) are even susceptible to getting MS.</p

Incident venous thromboembolic events in the Prospective Study of Pravastatin in the Elderly at Risk (PROSPER)

&lt;p&gt;Background: Venous thromboembolic events (VTE), including deep venous thrombosis and pulmonary embolism, are common in older age. It has been suggested that statins might reduce the risk of VTE however positive results from studies of middle aged subjects may not be generalisable to elderly people. We aimed to determine the effect of pravastatin on incident VTE in older people; we also studied the impact of clinical and plasma risk variables.&lt;/p&gt; &lt;p&gt;Methods: This study was an analysis of incident VTE using data from the Prospective Study of Pravastatin in the Elderly at Risk (PROSPER), a randomized, double-blind, placebo-controlled trial of pravastatin in men and women aged 70-82. Mean follow-up was 3.2 years. Risk for VTE was examined in non-warfarin treated pravastatin (n = 2834) and placebo (n = 2865) patients using a Cox's proportional hazard model, and the impact of other risk factors assessed in a multivariate forward stepwise regression analysis. Baseline clinical characteristics, blood biochemistry and hematology variables, plasma levels of lipids and lipoproteins, and plasma markers of inflammation and adiposity were compared. Plasma markers of thrombosis and hemostasis were assessed in a nested case (n = 48) control (n = 93) study where the cohort was those participants, not on warfarin, for whom data were available.&lt;/p&gt; &lt;p&gt;Results: There were 28 definite cases (1.0%) of incident VTE in the pravastatin group recipients and 20 cases (0.70%) in placebo recipients. Pravastatin did not reduce VTE in PROSPER compared to placebo [unadjusted hazard ratio (95% confidence interval) 1.42 (0.80, 2.52) p = 0.23]. Higher body mass index (BMI) [1.09 (1.02, 1.15) p = 0.0075], country [Scotland vs Netherlands 4.26 (1.00, 18.21) p = 0.050 and Ireland vs Netherlands 6.16 (1.46, 26.00) p = 0.013], lower systolic blood pressure [1.35 (1.03, 1.75) p = 0.027] and lower baseline Mini Mental State Examination (MMSE) score [1.19 (1.01, 1.41) p = 0.034] were associated with an increased risk of VTE, however only BMI, country and systolic blood pressure remained significant on multivariate analysis. In a nested case control study of definite VTE, plasma Factor VIII levels were associated with VTE [1.52 (1.01, 2.28), p = 0.044]. However no other measure of thrombosis and haemostasis was associated with increased risk of VTE.&lt;/p&gt; &lt;p&gt;Conclusions: Pravastatin does not prevent VTE in elderly people at risk of vascular disease. Blood markers of haemostasis and inflammation are not strongly predictive of VTE in older age however BMI, country and lower systolic blood pressure are independently associated with VTE risk.&lt;/p&gt

Inhibition of Y1 receptor signaling improves islet transplant outcome

Failure to secrete sufficient quantities of insulin is a pathological feature of type-1 and type-2 diabetes, and also reduces the success of islet cell transplantation. Here we demonstrate that Y1 receptor signaling inhibits insulin release in β-cells, and show that this can be pharmacologically exploited to boost insulin secretion. Transplanting islets with Y1 receptor deficiency accelerates the normalization of hyperglycemia in chemically induced diabetic recipient mice, which can also be achieved by short-term pharmacological blockade of Y1 receptors in transplanted mouse and human islets. Furthermore, treatment of non-obese diabetic mice with a Y1 receptor antagonist delays the onset of diabetes. Mechanistically, Y1 receptor signaling inhibits the production of cAMP in islets, which via CREB mediated pathways results in the down-regulation of several key enzymes in glycolysis and ATP production. Thus, manipulating Y1 receptor signaling in β-cells offers a unique therapeutic opportunity for correcting insulin deficiency as it occurs in the pathological state of type-1 diabetes as well as during islet transplantation.Islet transplantation is considered one of the potential treatments for T1DM but limited islet survival and their impaired function pose limitations to this approach. Here Loh et al. show that the Y1 receptor is expressed in β- cells and inhibition of its signalling, both genetic and pharmacological, improves mouse and human islet function.info:eu-repo/semantics/publishe