Acute anti-allodynic action of gabapentin in dorsal horn and primary somatosensory cortex: Correlation of behavioural and physiological data

Neuropathic pain is a debilitating consequence of neuronal injury or disease. Although first line treatments include the alpha-2-delta (a2d)-ligands, pregabalin and gabapentin (GBP), the mechanism of their anti-allodynic action is poorly understood. One specific paradox is that GBP relieves signs of neuropathic pain in animal models within 30min of an intraperitoneal (IP) injection yet its actions in vitro on spinal dorsal horn or primary afferent neurons take hours to develop. We found, using confocal Ca2þ imaging, that substantia gelatinosa neurons obtained ex vivo from rats subjected to sciatic chronic constriction injury (CCI) were more excitable than controls. We confirmed that GBP (100 mg/kg) attenuated mechanical allodynia in animals subject to CCI within 30min of IP injection.Substantia gelatinosa neurons obtained ex vivo from these animals no longer displayed CCI-induced increased excitability. Electrophysiological analysis of substantia gelatinosa neurons ex vivo suggest that rapidly developing in vivo anti-allodynic effects of GBP i) are mediated intracellularly, ii) involve actions on the neurotransmitter release machinery and iii) depend on decreased excitatory synaptic drive to excitatory neurons without major actions on inhibitory neurons or on intrinsic neuronal excitability. Experiments using in vivo Ca2þ imaging showed that 100 mg/kg GBP also suppressed the response of the S1 somatosensory cortex of CCI rats, but not that of control rats, to vibrotactile stimulation. Since the level of a2d1 protein is increased in primary afferent fibres after sciatic CCI, we suggest this dictates the rate of GBP action; rapidly developing actions can only be seen when a2d1 levels are elevated

A Plug-and-Play Platform for the Formation of Trifunctional Cysteine Bioconjugates that also Offers Control over Thiol Cleavability

Linkers that enable the site-selective synthesis of chemically modified proteins are of great interest to the field of chemical biology. Homogenous bioconjugates often show advantageous pharmacokinetic profiles and consequently increased efficacy in vivo. Cysteine residues have been exploited as a route to site-selectively modify proteins, and many successfully approved therapeutics make use of cysteine directed conjugation reagents. However, commonly used linkers, including maleimide–thiol conjugates, are not stable to the low concentrations of thiol present in blood. Furthermore, only a few cysteine-targeting reagents enable the site-selective attachment of multiple functionalities: a useful tool in the fields of theranostics and therapeutic blood half-life extension. Herein, we demonstrate the application of the pyridazinedione motif to enable site-selective attachment of three functionalities to a protein bearing a single cysteine residue. Extending upon previously documented dual modification work, here we demonstrate that by exploiting a bromide leaving group as an additional reactive point on the pyridazinedione scaffold, a thiol or aniline derivative can be added to a protein, post-conjugation. Thiol cleavability appraisal of the resultant C–S and C–N linked thio-bioconjugates demonstrated C–S functionalized linkers to be cleavable and C–N functionalized linkers to be noncleavable when incubated in an excess of glutathione. The plug-and-play trifunctional platform was exemplified by attaching clinically relevant motifs: biotin, fluorescein, a polyethylene glycol chain, and a model peptide. This platform provides a rare opportunity to combine up to three functionalities on a protein in a site-selective fashion. Furthermore, by selecting the use of a thiol or an amine for functionalization, we provide unique control over linker cleavability toward thiols, allowing this novel linker to be applied in a range of physiological environments

Use of pyridazinediones as extracellular cleavable linkers through reversible cysteine conjugation

Herein we report a retro-Michael deconjugation pathway of thiol-pyridazinedione linked protein bioconjugates to provide a novel cleavable linker technology. We demonstrate that the novel pyridazinedione linker does not suffer from off-target modification with blood thiols (e.g., glutathione, human serum albumin (HSA)), which is in sharp contrast to an analogous maleimide linker

First-food systems transformations and the ultra-processing of infant and young child diets: The determinants, dynamics and consequences of the global rise in commercial milk formula consumption

The inappropriate marketing and aggressive promotion of breastmilk substitutes (BMS) undermines breastfeeding and harms child and maternal health in all country contexts. Although a global milk formula ‘sales boom’ is reportedly underway, few studies have investigated its dynamics and determinants. This study takes two steps. First, it describes trends and patterns in global formula sales volumes (apparent consumption), by country income and region. Data are reported for 77 countries, for the years 2005–19, and for the standard (0–6 months), follow‐up (7‐12 m), toddler (13‐36 m), and special (0‐6 m) categories. Second, it draws from the literature to understand how transformations underway in first‐food systems – those that provision foods for children aged 0–36 months – explain the global transition to higher formula diets. Total world formula sales grew by 115% between 2005 and 2019, from 3.5 to 7.4 kg/child, led by highly‐populated middle‐income countries. Growth was rapid in South East and East Asia, especially in China, which now accounts for one third of world sales. This transition is linked with factors that generate demand for BMS, including rising incomes, urbanisation, the changing nature of woman's work, social norms, media influences and medicalisation. It also reflects the globalization of the baby food industry and its supply chains, including the increasing intensity and sophistication of its marketing practices. Policy and regulatory frameworks designed to protect, promote and support breastfeeding are partially or completely inadequate in the majority of countries, hence supporting industry expansion over child nutrition. The results raise serious concern for global child and maternal health

Fructose transport-deficient Staphylococcus aureus reveals important role of epithelial glucose transporters in limiting sugar-driven bacterial growth in airway surface liquid.

Hyperglycaemia as a result of diabetes mellitus or acute illness is associated with increased susceptibility to respiratory infection with Staphylococcus aureus. Hyperglycaemia increases the concentration of glucose in airway surface liquid (ASL) and promotes the growth of S. aureus in vitro and in vivo. Whether elevation of other sugars in the blood, such as fructose, also results in increased concentrations in ASL is unknown and whether sugars in ASL are directly utilised by S. aureus for growth has not been investigated. We obtained mutant S. aureus JE2 strains with transposon disrupted sugar transport genes. NE768(fruA) exhibited restricted growth in 10 mM fructose. In H441 airway epithelial-bacterial co-culture, elevation of basolateral sugar concentration (5-20 mM) increased the apical growth of JE2. However, sugar-induced growth of NE768(fruA) was significantly less when basolateral fructose rather than glucose was elevated. This is the first experimental evidence to show that S. aureus directly utilises sugars present in the ASL for growth. Interestingly, JE2 growth was promoted less by glucose than fructose. Net transepithelial flux of D-glucose was lower than D-fructose. However, uptake of D-glucose was higher than D-fructose across both apical and basolateral membranes consistent with the presence of GLUT1/10 in the airway epithelium. Therefore, we propose that the preferential uptake of glucose (compared to fructose) limits its accumulation in ASL. Pre-treatment with metformin increased transepithelial resistance and reduced the sugar-dependent growth of S. aureus. Thus, epithelial paracellular permeability and glucose transport mechanisms are vital to maintain low glucose concentration in ASL and limit bacterial nutrient sources as a defence against infection

A systematic review of evidence for fitness-to-drive among people with the mental health conditions of schizophrenia, stress/anxiety disorder, depression, personality disorder and obsessive compulsive disorder.

Background: Limited evidence exists regarding fitness-to-drive for people with the mental health conditions of schizophrenia, stress/anxiety disorder, depression, personality disorder and obsessive compulsive disorder (herein simply referred to as 'mental health conditions'). The aim of this paper was to systematically search and classify all published studies regarding driving for this population, and then critically appraise papers addressing assessment of fitness-to-drive where the focus was not on the impact of medication on driving. Methods: A systematic search of three databases (CINAHL, PSYCHINFO, EMBASE) was completed from inception to May 2016 to identify all articles on driving and mental health conditions. Papers meeting the eligibility criteria of including data relating to assessment of fitness-to-drive were critically appraised using the American Academy of Neurology and Centre for Evidence-Based Medicine protocols. Results: A total of 58 articles met the inclusion criteria of driving among people with mental health conditions studied, and of these, 16 contained data and an explicit focus on assessment of fitness-to-drive. Assessment of fitness-to-drive was reported in three ways: 1) factors impacting on the ability to drive safely among people with mental health conditions, 2) capability and perception of health professionals assessing fitness-to-drive of people with mental health conditions, and 3) crash rates. The level of evidence of the published studies was low due to the absence of controls, and the inability to pool data from different diagnostic groups. Evidence supporting fitness-todrive is conflicting. Conclusions: There is a relatively small literature in the area of driving with mental health conditions, and the overall quality of studies examining fitness-to-drive is low. Large-scale longitudinal studies with age-matched controls are urgently needed in order to determine the effects of different conditions on fitness-to-drive

Plasticity of the human visual system after retinal gene therapy in patients with Leber's congenital amaurosis.

Much of our knowledge of the mechanisms underlying plasticity in the visual cortex in response to visual impairment, vision restoration, and environmental interactions comes from animal studies. We evaluated human brain plasticity in a group of patients with Leber's congenital amaurosis (LCA), who regained vision through gene therapy. Using non-invasive multimodal neuroimaging methods, we demonstrated that reversing blindness with gene therapy promoted long-term structural plasticity in the visual pathways emanating from the treated retina of LCA patients. The data revealed improvements and normalization along the visual fibers corresponding to the site of retinal injection of the gene therapy vector carrying the therapeutic gene in the treated eye compared to the visual pathway for the untreated eye of LCA patients. After gene therapy, the primary visual pathways (for example, geniculostriate fibers) in the treated retina were similar to those of sighted control subjects, whereas the primary visual pathways of the untreated retina continued to deteriorate. Our results suggest that visual experience, enhanced by gene therapy, may be responsible for the reorganization and maturation of synaptic connectivity in the visual pathways of the treated eye in LCA patients. The interactions between the eye and the brain enabled improved and sustained long-term visual function in patients with LCA after gene therapy

Early and Middle Holocene Hunter-Gatherer Occupations in Western Amazonia: The Hidden Shell Middens

We report on previously unknown early archaeological sites in the Bolivian lowlands, demonstrating for the first time early and middle Holocene human presence in western Amazonia. Multidisciplinary research in forest islands situated in seasonally-inundated savannahs has revealed stratified shell middens produced by human foragers as early as 10,000 years ago, making them the oldest archaeological sites in the region. The absence of stone resources and partial burial by recent alluvial sediments has meant that these kinds of deposits have, until now, remained unidentified. We conducted core sampling, archaeological excavations and an interdisciplinary study of the stratigraphy and recovered materials from three shell midden mounds. Based on multiple lines of evidence, including radiocarbon dating, sedimentary proxies (elements, steroids and black carbon), micromorphology and faunal analysis, we demonstrate the anthropogenic origin and antiquity of these sites. In a tropical and geomorphologically active landscape often considered challenging both for early human occupation and for the preservation of hunter-gatherer sites, the newly discovered shell middens provide evidence for early to middle Holocene occupation and illustrate the potential for identifying and interpreting early open-air archaeological sites in western Amazonia. The existence of early hunter-gatherer sites in the Bolivian lowlands sheds new light on the region's past and offers a new context within which the late Holocene "Earthmovers" of the Llanos de Moxos could have emerged. © 2013 Lombardo et al

A random cell motility gradient downstream of FGF controls elongation of amniote embryos

Vertebrate embryos are characterized by an elongated antero-posterior (AP) body axis, which forms by progressive cell deposition from a posterior growth zone in the embryo. Here, we used tissue ablation in the chicken embryo to demonstrate that the caudal presomitic mesoderm (PSM) has a key role in axis elongation. Using time-lapse microscopy, we analysed the movements of fluorescently labelled cells in the PSM during embryo elongation, which revealed a clear posterior-to-anterior gradient of cell motility and directionality in the PSM. We tracked the movement of the PSM extracellular matrix in parallel with the labelled cells and subtracted the extracellular matrix movement from the global motion of cells. After subtraction, cell motility remained graded but lacked directionality, indicating that the posterior cell movements associated with axis elongation in the PSM are not intrinsic but reflect tissue deformation. The gradient of cell motion along the PSM parallels the fibroblast growth factor (FGF)/mitogen-activated protein kinase (MAPK) gradient1, which has been implicated in the control of cell motility in this tissue2. Both FGF signalling gain- and loss-of-function experiments lead to disruption of the motility gradient and a slowing down of axis elongation. Furthermore, embryos treated with cell movement inhibitors (blebbistatin or RhoK inhibitor), but not cell cycle inhibitors, show a slower axis elongation rate. We propose that the gradient of random cell motility downstream of FGF signalling in the PSM controls posterior elongation in the amniote embryo. Our data indicate that tissue elongation is an emergent property that arises from the collective regulation of graded, random cell motion rather than by the regulation of directionality of individual cellular movements