Stimulus-induced S-nitrosylation of Syntaxin 4 impacts insulin granule exocytosis

Glucose-stimulated insulin release from pancreatic islet β-cells involves increased levels of reactive oxygen and nitrogen species. Although this is normal, under pathophysiological conditions such as chronic hyperglycemia and inflammation, insulin exocytosis fails, and yet the mechanistic reason for failure is unclear. Hypothesizing that exocytotic proteins might be targets of S-nitrosylation, with their dysfunction under conditions of nitrosative stress serving as a mechanistic basis for insulin secretory dysfunction, we identified the t-SNARE protein Syntaxin 4 as a target of modification by S-nitrosylation. The cellular content of S-nitrosylated Syntaxin 4 peaked acutely, within 5 min of glucose stimulation in both human islets and MIN6 β-cells, corresponding to the time at which Syntaxin 4 activation was detectable. S-Nitrosylation was mapped to Syntaxin 4 residue Cys(141), located within the Hc domain predicted to increase accessibility for v-SNARE interaction. A C141S-Syntaxin 4 mutant resisted S-nitrosylation induced in vitro by the nitric oxide donor compound S-nitroso-L-glutathione, failed to exhibit glucose-induced activation and VAMP2 binding, and failed to potentiate insulin release akin to that of wild-type Syntaxin 4. Strikingly, S-nitrosylation of Syntaxin 4 could be induced by acute treatment with inflammatory cytokines (TNFα, IL-1β, and IFNγ), coordinate with inappropriate Syntaxin 4 activation and insulin release in the absence of the glucose stimulus, consistent with nitrosative stress and dysfunctional exocytosis, preceding the cell dysfunction and death associated with more chronic stimulation (24 h). Taken together, these data indicate a significant role for reactive nitrogen species in the insulin exocytosis mechanism in β-cells and expose a potential pathophysiological exploitation of this mechanism to underlie dysfunctional exocytosis.     

Toll-like receptor 3 signaling evokes a proinflammatory and proliferative phenotype in human vascular smooth muscle cells

Inflammation plays a key role in atherogenesis, perhaps promoted by bacterial and viral products present within the artery wall. Vascular smooth muscle cells (VSMC) can express certain bacterially responsive Toll-like receptors (TLR), which promote a proinflammatory and proliferative VSMC phenotype when activated, but it is unknown whether virally activated TLR can regulate VSMC phenotype. Here we tested the role in VSMC of TLR3, which is activated by double-stranded (dsRNA), a molecular signature of viruses. VSMC from multiple vessel types, including human coronary artery (HCoASMC) and mouse aorta (MAoSMC), expressed TLR3 constitutively, and HCoASMC were exquisitely sensitive to dsRNA-stimulated release of monocyte chemoattractant protein-1 (MCP-1) and interleukin-6. dsRNA-induced MCP-1 release was abolished by small interfering RNA-mediated TLR3 knockdown in HCoASMC and was absent in TLR3-/- MAoSMC but was unimpaired in TLR2-/- and in TLR4 signaling-deficient MAoSMC. Exposure to dsRNA also activated ERK1/2 and NF-kappaB in both human and murine SMC, but these effects were absent in SMC from TLR3-deficient mice, demonstrating a crucial role of TLR3 signaling. dsRNA also stimulated proliferation of HCoASMC, indicated by increased DNA synthesis, and induced persistent elevations in the intracellular levels of growth-promoting mediators, including interleukin-1alpha and phospho-ERK1/2. We conclude that exposure of HCoASMC to dsRNA elicits dramatic TLR3-mediated proinflammatory and proproliferative phenotypic changes, responses that could potentially be triggered by viral infection of cells within the arterial wall.     

Diffusion driven instability in an inhomogeneous domain

Diffusion driven instability in reaction-diffusion systems has been proposed as a mechanism for pattern formation in numerous embryological and ecological contexts. However, the possible effects of environmental inhomogeneities has received relatively little attention. We consider a general two species reaction-diffusion model in one space dimension, with one diffusion coefficient a step function of the spatial coordinate. We derive the dispersion relation and the solution of the linearized system. We apply our results to Turing-type models for both embryogenesis and predator-prey interactions. In the former case we derive conditions for pattern to be isolated in one part of the domain, and in the latter we introduce the concept of “environmental instability”. Our results suggest that environmental inhomogeneity could be an important regulator of biological pattern formation.     

Optimal Behavioral Hierarchy

Human behavior has long been recognized to display hierarchical structure: actions fit together into subtasks, which cohere into extended goal-directed activities. Arranging actions hierarchically has well established benefits, allowing behaviors to be represented efficiently by the brain, and allowing solutions to new tasks to be discovered easily. However, these payoffs depend on the particular way in which actions are organized into a hierarchy, the specific way in which tasks are carved up into subtasks. We provide a mathematical account for what makes some hierarchies better than others, an account that allows an optimal hierarchy to be identified for any set of tasks. We then present results from four behavioral experiments, suggesting that human learners spontaneously discover optimal action hierarchies.     

Trigeminal Ganglion Neurons of Mice Show Intracellular Chloride Accumulation and Chloride-Dependent Amplification of Capsaicin-Induced Responses

Intracellular Cl⁻ concentrations ([Cl⁻]_i) of sensory neurons regulate signal transmission and signal amplification. In dorsal root ganglion (DRG) and olfactory sensory neurons (OSNs), Cl⁻ is accumulated by the Na⁺-K⁺-2Cl⁻ cotransporter 1 (NKCC1), resulting in a [Cl⁻]_i above electrochemical equilibrium and a depolarizing Cl⁻ efflux upon Cl⁻ channel opening. Here, we investigate the [Cl⁻]_i and function of Cl⁻ in primary sensory neurons of trigeminal ganglia (TG) of wild type (WT) and NKCC1^−/− mice using pharmacological and imaging approaches, patch-clamping, as well as behavioral testing. The [Cl⁻]_i of WT TG neurons indicated active NKCC1-dependent Cl⁻ accumulation. Gamma-aminobutyric acid (GABA)_A receptor activation induced a reduction of [Cl⁻]_i as well as Ca²⁺ transients in a corresponding fraction of TG neurons. Ca²⁺ transients were sensitive to inhibition of NKCC1 and voltage-gated Ca²⁺ channels (VGCCs). Ca²⁺ responses induced by capsaicin, a prototypical stimulus of transient receptor potential vanilloid subfamily member-1 (TRPV1) were diminished in NKCC1^−/− TG neurons, but elevated under conditions of a lowered [Cl⁻]_o suggesting a Cl⁻-dependent amplification of capsaicin-induced responses. Using next generation sequencing (NGS), we found expression of different Ca²⁺-activated Cl⁻ channels (CaCCs) in TGs of mice. Pharmacological inhibition of CaCCs reduced the amplitude of capsaicin-induced responses of TG neurons in Ca²⁺ imaging and electrophysiological recordings. In a behavioral paradigm, NKCC1^−/− mice showed less avoidance of the aversive stimulus capsaicin. In summary, our results strongly argue for a Ca²⁺-activated Cl⁻-dependent signal amplification mechanism in TG neurons that requires intracellular Cl⁻ accumulation by NKCC1 and the activation of CaCCs.     

Vitamin E status and metabolism in adult and aged aryl hydrocarbon receptor null mice

The aryl hydrocarbon receptor (AhR) is involved in regulation of mechanisms for detoxification of xenobiotics, as well as vitamin A metabolism. Vitamin E is a fat-soluble nutrient whose metabolism is initialized via the cytochrome P450 system. Thus, AhR absence could alter hepatic regulation of α-tocopherol metabolism. To test this hypothesis, we assessed vitamin E status in adult (2-5 m) and old (21-22 m), wild-type and AhR-null mice. Plasma α-tocopherol concentrations in AhR-null mice (2.3±1.2 μmol/L, n=19) were lower than those of wild-type mice (3.2±1.2, n=17, P=.0131); those in old mice (3.2±1.2, n=20) were higher than those of adults (2.2±1.0, n=16, P=.0075). Hepatic α-tocopherol concentrations were not different between genotypes, but were nearly double in old (32±8 nmol/g, n=20) as compared with adult mice (17±2, n=16, P<.0001). Hepatic Cyp3a concentrations in AhR-null mice were greater than those in wild-type mice (P=.0011). Genotype (P=.0047), sex (P<.0001) and age (P<.0001) were significant modifiers of liver α-tocopherol metabolite (α-CEHC) concentrations. In general, Cyp3a concentrations correlated with hepatic α-tocopherol (r=0.3957, P<.05) and α-CEHC (r=0.4260, P<.05) concentrations. Since there were no significant genotype differences in the hepatic α- or γ-tocopherol concentrations, AhR-null mice did not have dramatically altered vitamin E metabolism. Since they did have higher hepatic α-CEHC concentrations, these data suggest metabolism was up-regulated in the AhR-null mice in order to maintain the hepatic tocopherol concentrations similar to those of wild-type mice.     

Revisiting dental fluctuating asymmetry in neandertals and modern humans

Previous studies have suggested that Neandertals experienced greater physiological stress and/or were less capable of mitigating stress than most prehistoric modern human populations. The current study compares estimates of dental fluctuating asymmetry (DFA) for prehistoric Inupiat from Point Hope Alaska, the Late Archaic, and Protohistoric periods from Ohio and West Virginia, and a modern sample from Ohio to Neandertals from Europe and Southwest Asia. DFA results from developmental perturbation during crown formation and is thus an indicator of developmental stress, which previous studies have found to be higher in Neandertals than in several modern human populations. Here, we use recent methodological improvements in the analysis of fluctuating asymmetry suggested by Palmer and Strobeck (Annu Rev Ecol Syst 17 ( 1986 ) 391–421, Developmental instability: causes and consequences ( 2003a ) v.1–v.36, Developmental instability: causes and consequences ( 2003b ) 279–319) and compare the fit of Neandertal DFA Index values with those of modern humans. DFA estimates for each of the modern population samples exceeded measurement error, with the Inupiat exhibiting the highest levels of DFA for most tooth positions. All significant Neandertal z‐scores were positive, exceeding the estimates for each of the modern prehistoric groups. Neandertals exhibited the fewest significant differences from the Inupiat (9.2% of values are significant at P < 0.05), while for the other modern prehistoric groups more than 10% of the Neandertal z‐scores are significant at P < 0.05, more than 90% of these significant scores at P < 0.01. These results suggest that the Inupiat experienced greater developmental stress than the other prehistoric population samples, and that Neandertals were under greater developmental stress than all other prehistoric modern human samples. Am J Phys Anthropol 149:193–204, 2012. © 2012 Wiley Periodicals, Inc.     

CaV2.1 α1 Subunit Expression Regulates Presynaptic CaV2.1 Abundance and Synaptic Strength at a Central Synapse

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The Prevalence of Non-Alcoholic Fatty Liver Disease in Children and Adolescents: A Systematic Review and Meta-Analysis

MethodsWe conducted a systematic review and meta-analysis of all studies reporting a prevalence of NAFLD based on any diagnostic method in participants 1–19 years old, regardless of whether assessing NAFLD prevalence was the main aim of the study.ResultsThe pooled mean prevalence of NAFLD in children from general population studies was 7.6% (95%CI: 5.5% to 10.3%) and 34.2% (95% CI: 27.8% to 41.2%) in studies based on child obesity clinics. In both populations there was marked heterogeneity between studies (I² = 98%). There was evidence that prevalence was generally higher in males compared with females and increased incrementally with greater BMI. There was evidence for differences between regions in clinical population studies, with estimated prevalence being highest in Asia. There was no evidence that prevalence changed over time. Prevalence estimates in studies of children/adolescents attending obesity clinics and in obese children/adolescents from the general population were substantially lower when elevated alanine aminotransferase (ALT) was used to assess NAFLD compared with biopsies, ultrasound scan (USS) or magnetic resonance imaging (MRI).ConclusionsOur review suggests the prevalence of NAFLD in young people is high, particularly in those who are obese and in males.