Consumption and activity decline in Northern Pike (Esox lucius) during and after silver nanoparticle addition to a lake.

Silver nanoparticles (AgNPs) are antimicrobial additives in many consumer products with high potential for release into aquatic ecosystems. Though AgNPs have been shown to have negative impacts on fish in laboratory experiments, these effects are rarely observed at ecologically relevant concentrations or in situ in field settings. To evaluate ecosystem-level effects of this contaminant, AgNPs were added to a lake at the IISD Experimental Lakes Area (IISD-ELA) during 2014 and 2015. Mean total silver (TAg) concentrations in the water column were 4 µg L-1 during additions. The growth of Northern Pike (Esox lucius) declined, and their primary prey, Yellow Perch (Perca flavescens) became less abundant after AgNP exposure. Here, we used a combined contaminant-bioenergetics modeling approach to show that individual activity and both individual and population-level consumption of Northern Pike declined significantly in the lake dosed with AgNPs, which, combined with other evidence, suggests that observed declines in body size were likely a result of indirect effects (i.e., reduced prey availability). Further, we found the contaminant-bioenergetics approach was sensitive to modelled elimination rates of mercury, overestimating consumption and activity by 43% and 55%, respectively, when using the mercury elimination rate commonly used in these models versus field-derived estimates for this species. This study contributes to the growing evidence of potentially long-term negative impacts on fish from chronic exposure to environmentally relevant concentrations of AgNPs in a natural setting.

Neuronal cell types, projections, and spatial organization of the central amygdala.

The central amygdala (CEA) has been richly studied for interpreting function and behavior according to specific cell types and circuits. Such work has typically defined molecular cell types by classical inhibitory marker genes; consequently, whether marker-gene-defined cell types exhaustively cover the CEA and co-vary with connectivity remains unresolved. Here, we combined single-cell RNA sequencing, multiplexed fluorescent in situ hybridization, immunohistochemistry, and long-range projection mapping to derive a "bottom-up" understanding of CEA cell types. In doing so, we identify two major cell types, encompassing one-third of all CEA neurons, that have gone unresolved in previous studies. In spatially mapping these novel types, we identify a non-canonical CEA subdomain associated with Nr2f2 expression and uncover an Isl1-expressing medial cell type that accounts for many long-range CEA projections. Our results reveal new CEA organizational principles across cell types and spatial scales and provide a framework for future work examining cell-type-specific behavior and function.

Spatially patterned excitatory neuron subtypes and projections of the claustrum.

The claustrum is a functionally and structurally complex brain region, whose very spatial extent remains debated. Histochemical-based approaches typically treat the claustrum as a relatively narrow anatomical region that primarily projects to the neocortex, whereas circuit-based approaches can suggest a broader claustrum region containing projections to the neocortex and other regions. Here, in the mouse, we took a bottom-up and cell-type-specific approach to complement and possibly unite these seemingly disparate conclusions. Using single-cell RNA-sequencing, we found that the claustrum comprises two excitatory neuron subtypes that are differentiable from the surrounding cortex. Multicolor retrograde tracing in conjunction with 12-channel multiplexed in situ hybridization revealed a core-shell spatial arrangement of these subtypes, as well as differential downstream targets. Thus, the claustrum comprises excitatory neuron subtypes with distinct molecular and projection properties, whose spatial patterns reflect the narrower and broader claustral extents debated in previous research. This subtype-specific heterogeneity likely shapes the functional complexity of the claustrum.

Elucidating memory in the brain via single-cell transcriptomics.

Elucidating the neural mechanisms of memory in the brain is a central goal of neuroscience. Here, we discuss modern-day transcriptomics methodologies, and how they are well-poised to revolutionize our insight into memory mechanisms at unprecedented resolution and throughput. Focusing on the hippocampus and amygdala, two regions extensively examined in memory research, we show how single-cell transcriptomics technologies have been leveraged to understand the naïve state of these brain regions. Building upon this foundation, we show that these technologies can be applied to single-trial learning paradigms to comprehensively identify molecules and cells that participate in the encoding and retrieval of memory. Transcriptomics also provides an opportunity to understand the cell-type organization of the human hippocampus and amygdala, and due to conservation of these brain regions between humans and rodents, to infer behavioral and causal contributions in the human brain by leveraging rodent cell-type homologies and interventions. Ultimately, such transcriptomic technologies are poised to usher in a qualitatively novel understanding of memory in the brain.

Tritium in fish from remote lakes in northwestern Ontario, Canada.

Tritium is most commonly generated as a by-product of nuclear reactors. As such, environmental concentrations are typically only reported near regions of interest, and background concentrations in areas unaffected by anthropogenic disturbance are not well characterized. To provide information on background levels of tritium in the natural environment, tissue-free water tritium (TFWT) and organically-bound tritium (OBT) were measured in the flesh of 106 fish collected within three lakes located at the IISD-Experimental Lakes Area (ELA) in Ontario, Canada in 2014. For the three ELA lakes studied, water tritium (HTO) activity concentration was determined to be below reliably detectable levels (0.6 Bq/L). Fish TFWT was found to be below 0.7 Bq/L, similar to the surrounding water tritium activity concentration. Fish OBT activity concentrations, at below 5 Bq/L, were also very low. Fish size was significantly related to OBT activity in Lake Whitefish and White Sucker from Lake 302, but not in other lakes. Though we observed significant differences in potential tritium exposure to humans among lakes, the levels of tritium reported here are below the Canadian natural background radiation of 1.8 mSv/y. These results provide information on background levels of tritium in freshwater fishes in Canada.

Point-of-care fluorescence imaging predicts the presence of pathogenic bacteria in wounds: a clinical study.

OBJECTIVE: Bacteria in chronic wounds are invisible to the naked eye and can lead to delayed wound healing. Point-of-care bacterial fluorescence imaging illuminates a wound with 405nm light, triggering bacteria to produce red fluorescence and enabling real-time bacterial localisation. Prospective, single-blind clinical trials (clinicaltrials.gov #NCT02682069, #NCT03091361) were conducted to determine the positive predictive value (PPV) of this red fluorescence for detecting bacteria in chronic wounds. METHOD: Lower limb chronic wounds were imaged for bacterial fluorescence using the MolecuLight i:X imaging device. Regions positive for red fluorescence were discretely sampled using either biopsy or curettage to correlate red fluorescence signals to bacterial presence and analysed via gold standard quantitative polymerase chain reaction (qPCR) or via semi-quantitative culture analysis respectively. RESULTS: A total of 60 lower limb chronic wounds were imaged. Quantitative PCR analysis of wound tissue biopsies obtained from regions of red fluorescence yielded a PPV of 100%. Total bacterial load in these areas was ≥104 CFU/g. Semi-quantitative culture analysis of curettage scrapings from regions of red fluorescence yielded a PPV of 100%, with predominately moderate or heavy bacterial growth. There were nine distinct bacterial species detected, all common pathogens in chronic wounds. Staphylococcus aureus was the most prevalent species. CONCLUSION: Bacterial fluorescence image-guided curettage or biopsy sampling positively predicts bacterial presence in wounds at potentially harmful levels, entirely eliminating the risk of false negative sampling. Fluorescence imaging of wounds offers clinicians real-time information on a wound's bacterial burden, insight which can influence treatment decisions at the point-of care.

Respirometry increases cortisol levels in rainbow trout Oncorhynchus mykiss: implications for measurements of metabolic rate.

This study aimed to assess the extent to which chasing, handling and confining Oncorhynchus mykiss to a small respirometer chamber during respirometric experiments is stressful and affects metabolic measurements. The study observed increased cortisol levels in animals tested using a chase protocol and subsequent intermittent-flow respirometry, suggesting that this procedural treatment may stress animals.

Spatial memory is impaired by peripubertal GnRH agonist treatment and testosterone replacement in sheep.

Chronic gonadotropin-releasing hormone agonist (GnRHa) is used therapeutically to block activity within the reproductive axis through down-regulation of GnRH receptors within the pituitary gland. GnRH receptors are also expressed in non-reproductive tissues, including areas of the brain such as the hippocampus and amygdala. The impact of long-term GnRHa-treatment on hippocampus-dependent cognitive functions, such as spatial orientation, learning and memory, is not well studied, particularly when treatment encompasses a critical window of development such as puberty. The current study used an ovine model to assess spatial maze performance and memory of rams that were untreated (Controls), had both GnRH and testosterone signaling blocked (GnRHa-treated), or specifically had GnRH signaling blocked (GnRHa-treated with testosterone replacement) during the peripubertal period (8, 27 and 41 weeks of age). The results demonstrate that emotional reactivity during spatial tasks was compromised by the blockade of gonadal steroid signaling, as seen by the restorative effects of testosterone replacement, while traverse times remained unchanged during assessment of spatial orientation and learning. The blockade of GnRH signaling alone was associated with impaired retention of long-term spatial memory and this effect was not restored with the replacement of testosterone signaling. These results indicate that GnRH signaling is involved in the retention and recollection of spatial information, potentially via alterations to spatial reference memory, and that therapeutic medical treatments using chronic GnRHa may have effects on this aspect of cognitive function.

The necessity-concerns framework: a multidimensional theory benefits from multidimensional analysis.

BACKGROUND: Patients' medication-related concerns and necessity-beliefs predict adherence. Evaluation of the potentially complex interplay of these two dimensions has been limited because of methods that reduce them to a single dimension (difference scores). PURPOSE: We use polynomial regression to assess the multidimensional effect of stroke-event survivors' medication-related concerns and necessity beliefs on their adherence to stroke-prevention medication. METHODS: Survivors (n = 600) rated their concerns, necessity beliefs, and adherence to medication. Confirmatory and exploratory polynomial regression determined the best-fitting multidimensional model. RESULTS: As posited by the necessity-concerns framework (NCF), the greatest and lowest adherence was reported by those necessity weak concerns and strong concerns/weak Necessity-Beliefs, respectively. However, as could not be assessed using a difference-score model, patients with ambivalent beliefs were less adherent than those exhibiting indifference. CONCLUSIONS: Polynomial regression allows for assessment of the multidimensional nature of the NCF. Clinicians/Researchers should be aware that concerns and necessity dimensions are not polar opposites.

Poor tolerability of thalidomide in end-stage oesophageal cancer.

Oesophageal cancer cachexia is a significant clinical problem, resulting in excessive morbidity and mortality. In a pilot study, 10 patients with cachexia due to advanced cancer of the oesophagus gained weight, including lean tissue, after 14-day treatment with thalidomide. Here, we present randomised placebo controlled trial data over a 6-week period to test the hypothesis that thalidomide is superior to placebo in terms of weight gain in patients with cachexia caused by oesophageal cancer. Thalidomide, 200 mg daily, or an identical placebo was given to patients with advanced oesophageal cancer. Total body weight and lean body mass were assessed in addition to drug tolerability and performance indices. Thirty-four patients were recruited. Of these, six given thalidomide and 16 given placebo completed the protocol; all withdrawals were due to adverse drug reactions or complications of disease. Thalidomide showed no benefit over placebo in participants who completed the protocol. These data suggest that thalidomide is poorly tolerated in patients with advanced cancer of the oesophagus and may not ameliorate the progression of cachexia. In the absence of hard supportive evidence, off-licence treatment with thalidomide should be used with great caution as an adjunct to nutritional support in patients with advanced cancer.

Resistance exercise enhances mTOR and MAPK signalling in human muscle over that seen at rest after bolus protein ingestion.

AIM: Feeding protein after resistance exercise enhances the magnitude and duration of myofibrillar protein synthesis (MPS) over that induced by feeding alone. We hypothesized that the underlying mechanism for this would be a greater and prolonged phosphorylation of signalling involved in protein translation. METHODS: Seven healthy young males performed unilateral resistance exercise followed immediately by the ingestion of 25 g of whey protein to maximally stimulate MPS in a rested and exercised leg. RESULTS: Phosphorylation of p70 ribosomal protein S6 kinase (p70S6K) was elevated (P<0.05) above fasted at 1 h at rest whereas it was elevated at 1, 3 and 5 h after exercise with protein ingestion and displayed a similar post-exercise time course to that shown by MPS. Extracellular regulated kinase1/2 (ERK1/2) and p90 ribosomal S6 kinase (p90RSK) phosphorylation were unaltered after protein ingestion at rest but were elevated (P < 0.05) above fasted early in recovery (1 h) and were greater for the exercised-fed leg than feeding alone (main effect; P < 0.01). Eukaryotic elongation factor 2 (eEF2) phosphorylation was also less (main effect; P<0.05) in the exercised-fed leg than in the rested leg suggesting greater activity after exercise. Eukaryotic initiation 4E binding protein-1 (4EBP-1) phosphorylation was increased (P<0.05) above fasted to the same extent in both conditions. CONCLUSION: Our data suggest that resistance exercise followed by protein feeding stimulates MPS over that induced by feeding alone in part by enhancing the phosphorylation of select proteins within the mammalian target of rapamycin (p70S6K, eEF2) and by activating proteins within the mitogen-activated protein kinase (ERK1/2, p90RSK) signalling.

Could better jobs improve mental health? A prospective study of change in work conditions and mental health in mid-aged adults.

OBJECTIVES: To investigate the extent improvement or deterioration in employee job security, control or workload is associated with a change in mental health. DESIGN: Self-report panel data (2000, 2004) on mental health (symptoms of depression and generalised anxiety) and job demands, control and insecurity. Changes in exposures and outcomes were calculated by subtracting wave 1 from wave 2 scores. Changes in mental health were regressed onto changes in work conditions, adjusting for confounders. Sensitivity analyses assessed reverse causation, floor and ceiling effects. SETTING: Two adjoining cities in south-east Australia. PARTICIPANTS: 1975 employees aged 40-48 years, 50% (n=995) male. RESULTS: Improvements and deterioration in each work condition were associated with corresponding improvements or deterioration in mental health. The association between changes in job insecurity and symptoms of depression was B=0.386 (95% CI 0.245 to 0.527) and with anxiety symptoms was B=0.434 (95% CI 0.267 to 0.601). Similarly, changes in job control were associated with changes in depressive (B=-0.548; 95% CI -0.791 to -0.304) and anxiety symptoms (B=-0.608; 95% CI -0.896 to -0.319) as were changes in job demands (B depression=0.386; 95% CI 0.245 to 0.527; B anxiety=0.434; 95% CI 0.267 to 0.601). Excluding people with severe symptoms at baseline did not alter the findings; however, path analyses indicated that depression may precede a worsening of work conditions. CONCLUSION: Among mid-aged employees, deteriorating work conditions may amplify population health burdens, especially anxiety. Furthermore, better quality jobs, combining an array of positive conditions, could alleviate major population health burdens.

Prey life-history and bioenergetic responses across a predation gradient.

To evaluate the importance of non-consumptive effects of predators on prey life histories under natural conditions, an index of predator abundance was developed for naturally occurring populations of a common prey fish, the yellow perch Perca flavescens, and compared to life-history variables and rates of prey energy acquisition and allocation as estimated from mass balance models. The predation index was positively related to maximum size and size at maturity in both male and female P. flavescens, but not with life span or reproductive investment. The predation index was positively related to size-adjusted specific growth rates and growth efficiencies but negatively related to model estimates of size-adjusted specific consumption and activity rates in both vulnerable (small) and invulnerable (large) size classes of P. flavescens. These observations suggest a trade-off between growth and activity rates, mediated by reduced activity in response to increasing predator densities. Lower growth rates and growth efficiencies in populations with fewer predators, despite increased consumption suggests either 1) a reduction in prey resources at lower predator densities or 2) an intrinsic cost of rapid prey growth that makes it unfavourable unless offset by a perceived threat of predation. This study provides evidence of trade-offs between growth and activity rates induced by predation risk in natural prey fish populations and illustrates how behavioural modification induced through predation can shape the life histories of prey fish species.

Comparative structural and hemodynamic analysis of vascular trees.

The availability of detailed three-dimensional images of vascular trees from mammalian organs provides a wealth of essential data for understanding the processes and mechanisms of vascular patterning. Using this detailed geometric data requires the ability to compare individual representations of vascular trees in statistically meaningful ways. This article provides some comparisons of geometry and also of simulated hemodynamics, enabling the identification of similarities and differences among 10 individual specimens (5 placenta specimens and 5 lung specimens). Similar comparisons made with a series of models (starting with the simplest and increasing in complexity) enable the identification of essential features that are needed to account for the patterns and function of vascular arborization.

Facts, noise and wishful thinking: muscle protein turnover in aging and human disuse atrophy.

Surprisingly little is known about the mechanisms of muscle atrophy with aging and disuse in human beings, in contrast to rodents, from which much has been extrapolated to explain the human condition. However, this extrapolation is likely unwarranted because the time course, extent of wasting, muscle fiber involvement and alterations of muscle protein turnover are all quite different in rodent and human muscle. Furthermore, there is little evidence that static indices of protein turnover represent dynamic changes and may be misleading. With disuse there are reductions in the rate of muscle protein synthesis (MPS) large enough to explain the atrophic loss of muscle protein without a concomitant increase in proteolysis. In aging, there is no evidence that there are marked alterations in basal muscle protein turnover in healthy individuals but instead the ability to maintain muscle after feeding is compromised. This anabolic resistance is evident with physical inactivity, which exacerbates the inability to maintain muscle mass with aging. The main conclusion of this review is that in uncomplicated, non-inflammatory disuse atrophy, the facilitative change causing loss of muscle mass is a depression of MPS, exacerbated by anabolic resistance during feeding, with possible adaptive depressions, rather than increases, of muscle proteolysis.

Alterations of protein turnover underlying disuse atrophy in human skeletal muscle.

Unloading-induced atrophy is a relatively uncomplicated form of muscle loss, dependent almost solely on the loss of mechanical input, whereas in disease states associated with inflammation (cancer cachexia, AIDS, burns, sepsis, and uremia), there is a procatabolic hormonal and cytokine environment. It is therefore predictable that muscle loss mainly due to disuse alone would be governed by mechanisms somewhat differently from those in inflammatory states. We suggest that in vivo measurements made in human subjects using arterial-venous balance, tracer dilution, and tracer incorporation are dynamic and thus robust by comparison with static measurements of mRNA abundance and protein expression and/or phosphorylation in human muscle. In addition, measurements made with cultured cells or in animal models, all of which have often been used to infer alterations of protein turnover, appear to be different from results obtained in immobilized human muscle in vivo. In vivo measurements of human muscle protein turnover in disuse show that the primary variable that changes facilitating the loss of muscle mass is protein synthesis, which is reduced in both the postabsorptive and postprandial states; muscle proteolysis itself appears not to be elevated. The depressed postprandial protein synthetic response (a phenomenon we term "anabolic resistance") may even be accompanied by a diminished suppression of proteolysis. We therefore propose that most of the loss of muscle mass during disuse atrophy can be accounted for by a depression in the rate of protein synthesis. Thus the normal diurnal fasted-to-fed cycle of protein balance is disrupted and, by default, proteolysis becomes dominant but is not enhanced.

Cyclic stretch reduces myofibrillar protein synthesis despite increases in FAK and anabolic signalling in L6 cells.

Muscle protein synthesis is increased after exercise, but evidence is now accruing that during muscular activity it is suppressed. In life, muscles are subjected to shortening forces due to contraction, but may also be subject to stretching forces during lengthening. It would be biologically inefficient if contraction and stretch have different effects on muscle protein turnover, but little is known about the metabolic effects of stretch. To investigate this, we assessed myofibrillar and sarcoplasmic protein synthesis (MPS, SPS, respectively) by incorporation of [1-13C]proline (using gas chromatography-mass spectrometry) and anabolic signalling (by phospho-immunoblotting and kinase assays) in cultured L6 skeletal muscle cells during 30 min of cyclic stretch and over 30 min intervals for up to 120 min afterwards. SPS was unaffected, whereas MPS was suppressed by 40 +/- 0.03% during stretch, before returning to basal rates by 90-20 min afterwards. Paradoxically, stretch stimulated anabolic signalling with peak values after 2-30 min: e.g. focal adhesion kinase (FAK Tyr576/577; +28 +/- 6%), protein kinase B activity (Akt; +113 +/- 31%), p70S6K1 (ribosomal S6 kinase Thr389; 25 +/- 5%), 4E binding protein 1 (4EBP1 Thr37/46; 14 +/- 3%), eukaryotic elongation factor 2 (eEF2 Thr56; -47 +/- 4%), extracellular regulated protein kinase 1/2 (ERK1/2 Tyr202/204; +65% +/- 9%), eukaryotic initiation factor 2alpha (eIF2alpha Ser51; -20 +/- 5%, P < 0.05) and eukaryotic initiation factor 4E (eIF4E Ser209; +33 +/- 10%, P < 0.05). After stretch, except for Akt activity, stimulatory phosphorylations were sustained: e.g. FAK (+26 +/- 11%) for > or =30 min, eEF2 for > or =60 min (peak -45 +/- 4%), 4EBP1 for > or =90 min (+33 +/- 5%), and p70S6K1 remained elevated throughout (peak +64 +/- 7%). Adenosine monophosphate-activated protein kinase (AMPK) phosphorylation was unchanged throughout. We report for the first time that acute cyclic stretch specifically suppresses MPS, despite increases in activity/phosphorylation of elements thought to increase anabolism.

Disassociation between the effects of amino acids and insulin on signaling, ubiquitin ligases, and protein turnover in human muscle.

We determined the effects of intravenous infusion of amino acids (AA) at serum insulin of 5, 30, 72, and 167 mU/l on anabolic signaling, expression of ubiquitin-proteasome components, and protein turnover in muscles of healthy young men. Tripling AA availability at 5 mU/l insulin doubled incorporation of [1-(13)C]leucine [i.e., muscle protein synthesis (MPS), P < 0.01] without affecting the rate of leg protein breakdown (LPB; appearance of d(5)-phenylalanine). While keeping AA availability constant, increasing insulin to 30 mU/l halved LPB (P < 0.05) without further inhibition at higher doses, whereas rates of MPS were identical to that at 5 mU/l insulin. The phosphorylation of PKB Ser(473) and p70(S6k) Thr(389) increased concomitantly with insulin, but whereas raising insulin to 30 mU/l increased the phosphorylation of mTOR Ser(2448), 4E-BP1 Thr(37/46), or GSK3beta Ser(9) and decreased that of eEF2 Thr(56), higher insulin doses to 72 and 167 mU/l did not augment these latter responses. MAFbx and proteasome C2 subunit proteins declined as insulin increased, with MuRF-1 expression largely unchanged. Thus increasing AA and insulin availability causes changes in anabolic signaling and amounts of enzymes of the ubiquitin-proteasome pathway, which cannot be easily reconciled with observed effects on MPS or LPB.

Rheumatoid cachexia: a clinical perspective.

Rheumatoid cachexia is under-recognized in clinical practice. The loss of lean body tissue, which characterizes cachexia, is often compensated for by gain in body fat-so called 'cachectic obesity'-so that 85% or more RA patients have a normal BMI. Severe cachexia with loss of weight leads to increased morbidity and premature mortality but loss of muscle bulk with a normal BMI also associates with poor clinical outcomes. Increasing BMI, even into the obese range, is associated with less joint damage and reduced mortality. Measurement of body composition using DXA and other techniques is feasible but the results must be interpreted with care. Newer techniques such as whole-body MRI will help define with more confidence the mass and distribution of fat and muscle and help elucidate the relationships between body composition and outcomes. Cachexia shows little response to diet alone but progressive resistance training and anti-TNF therapies show promise in tackling this potentially disabling extra-articular feature of RA.

Exercise- and nutrient-controlled mechanisms involved in maintenance of the musculoskeletal mass.

The mechanisms of maintenance of the protein mass of muscle and associated connective tissue and bone are becoming more accessible as a result of the use of a combination of well-established techniques for measurement of protein turnover and measurement of protein expression and phosphorylation state of signalling molecules involved in anabolic and catabolic responses. Amino acids, hormones and physical activity appear to be the major short-term physiological regulators of muscle mass, mainly through their actions on protein synthesis and breakdown, on a time scale of minutes to hours, with duration of changes in gene expression up to weeks. Amino acids are the main components in the diet regulating protein turnover, having marked effects in stimulating muscle protein synthesis and with almost no effect on muscle protein breakdown. Branched-chain amino acids, and in particular leucine, simulate protein synthesis via signalling pathways involving mTOR (mammalian target of rapamycin) in a dose-response manner. Insulin has little effect on protein synthesis in human muscle, but it has a marked inhibitory effect on protein breakdown. The amino acid simulation of anabolism is not dependent on the presence of insulin, IGF-1 (insulin-like growth factor-1) or growth hormone. Exercise not only stimulates protein synthesis in muscle, but also in tendon; and disuse atrophy is accompanied by marked decreases of both muscle and tendon collagen protein synthesis. Bone collagen synthesis appears to be nutritionally regulated by the availability of amino acids, but not lipid or glucose.