Circadian variation in gastric vagal afferent mechanosensitivity

Food intake is coordinated to cellular metabolism by clock gene expression with a master clock in the suprachiasmatic nucleus synchronized by light exposure. Gastric vagal afferents play a role in regulating food intake, but it is unknown whether they exhibit circadian variation in their mechanosensitivity. We aimed to determine whether gastric vagal afferents express clock genes and whether their response to mechanical stimuli oscillates throughout the light/dark cycle. Nodose ganglia were collected from 8-week-old female C57BL/6 mice every 3 h starting at lights off (1800 h) to quantify Bmal1, Per1, Per2, and Nr1d1 mRNA by qRT-PCR. Additionally in vitro single-fiber recordings of gastric vagal mechanoreceptors were taken at all time points. Per1, Per2, Bmal1, and Nr1d1 mRNA is expressed in the nodose ganglia and levels oscillated over a 24 h period. In mice fed ad libitum, gastric content was 3 times higher at 0000 h and 0300 h than 1200 h. The response of tension receptors to 3 g stretch was reduced by up to 70% at 2100 h, 0000 h, and 0300 h compared with 1200 h. Gastric mucosal receptor response to stroking with a 50 mg von Frey hair was 3 times greater at 1200 h and 1500 h than the response at 0000 h. Similar findings were obtained in mice fasted for 6 h or maintained in darkness for 3 d before study. Therefore, these changes do not result from food intake or the light/dark cycle. Thus, gastric vagal mechanoreceptors display circadian rhythm, which may act to control food intake differentially at different times of the day.Stephen J. Kentish, Claudine L. Frisby, David J. Kennaway, Gary A. Wittert, and Amanda J. Pag

Synthesis and self-assembly of polyimide/poly(dimethylsiloxane) brush triblock copolymers

A series of novel brush triblock copolymers containing 'glassy' fluorinated polyimide, poly((4,4'-hexafluoroisopropylidene diphthalic anhydride)-co-(2,3,5,6-tetramethyl-1,4-phenylenediamine)) (poly(6FDA-co-TMPD)), and 'rubbery' polydimethylsiloxane monomethacrylate (PDMS-MA) were synthesized and characterized. Well-defined difunctional poly(6FDA-co-TMPD) with a,w-amino end-groups was initially prepared via step-growth polymerization using precise control of the diamine (TMPD) to dianhydride (6FDA) ratio. Subsequent functionalization with 2-bromoisobutyryl bromide afforded a telechelic macroinitiator suitable for atom transfer radical polymerization (ATRP). The macroinitiator and its diamino poly(6FDA-co-TMPD) precursor were characterized via gel permeation chromatography (GPC), 1H nuclear magnetic resonance (NMR) spectroscopic analysis and matrix assisted laser desorption ionization time-of-flight (MALDI ToF) mass spectroscopy. ATRP of PDMS-MA using the macroinitiator in different molar ratios afforded a series of brush triblock copolymers with high monomer conversions (88-94%) and varying PDMS weight fractions. Self-assembly of the triblock brush copolymers in dimethylformamide (DMF) afforded nanoparticles with hydrodynamic diameters (dH) ranging from 87 to 109 nm, as determined by dynamic light scattering (DLS) analysis. Cross-linking of the nanoparticles was achieved via hydrogen abstraction through the thermal degradation of benzoyl peroxide. Transmission electron microscopy (TEM) and scanning transmission electron microscopy (STEM) revealed that the self-assemblies and their cross-linked derivatives had spherical morphologies

Functional Analysis of a Unique Troponin C Mutation, GLY159ASP, that Causes Familial Dilated Cardiomyopathy, Studied in Explanted Heart Muscle

Background-Familial dilated cardiomyopathy can be caused by mutations in the proteins of the muscle thin filament. In vitro, these mutations decrease Ca2+ sensitivity and cross-bridge turnover rate, but the mutations have not been investigated in human tissue. We studied the Ca2+-regulatory properties of myocytes and troponin extracted from the explanted heart of a patient with inherited dilated cardiomyopathy due to the cTnC G159D mutation.Methods and Results-Mass spectroscopy showed that the mutant cTnC was expressed approximately equimolar with wild-type cTnC. Contraction was compared in skinned ventricular myocytes from the cTnC G159D patient and nonfailing donor heart. Maximal Ca2+-activated force was similar in cTnC G159D and donor myocytes, but the Ca2+ sensitivity of cTnC G159D myocytes was higher (EC50 G159D/donor=0.60). Thin filaments reconstituted with skeletal muscle actin and human cardiac tropomyosin and troponin were studied by in vitro motility assay. Thin filaments containing the mutation had a higher Ca2+ sensitivity (EC(50)G159D/donor=0.55 +/- 0.13), whereas the maximally activated sliding speed was unaltered. In addition, the cTnC G159D mutation blunted the change in Ca2+ sensitivity when TnI was dephosphorylated. With wild-type troponin, Ca2+ sensitivity was increased (EC50 P/unP=4.7 +/- 1.9) but not with cTnC G159D troponin (EC50 P/unP=1.2 +/- 0.1).Conclusions-We propose that uncoupling of the relationship between phosphorylation and Ca2+ sensitivity could be the cause of the dilated cardiomyopathy phenotype. The differences between these data and previous in vitro results show that native phosphorylation of troponin I and troponin T and other posttranslational modifications of sarcomeric proteins strongly influence the functional effects of a mutation. (Circ Heart Fail. 2009;2:456-464.

The Influence of monovalent cations on myofibrillar function

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Revisiting Frank–Starling: regulatory light chain phosphorylation alters the rate of force redevelopment (ktr) in a length-dependent fashion

Force and power in cardiac muscle have a known dependence on phosphorylation of the myosin-associated regulatory light chain (RLC). We explore the effect of RLC phosphorylation on the ability of cardiac preparations to redevelop force (ktr ) in maximally activating [Ca2+ ]. Activation was achieved by rapidly increasing the temperature (temperature-jump of 0.5-20ºC) of permeabilized trabeculae over a physiological range of sarcomere lengths (1.85-1.94 μm). The trabeculae were subjected to shortening ramps over a range of velocities and the extent of RLC phosphorylation was varied. The latter was achieved using an RLC-exchange technique, which avoids changes in the phosphorylation level of other proteins. The results show that increasing RLC phosphorylation by 50% accelerates ktr by ∼50%, irrespective of the sarcomere length, whereas decreasing phosphorylation by 30% slows ktr by ∼50%, relative to the ktr obtained for in vivo phosphorylation. Clearly, phosphorylation affects the magnitude of ktr following step shortening or ramp shortening. Using a two-state model, we explore the effect of RLC phosphorylation on the kinetics of force development, which proposes that phosphorylation affects the kinetics of both attachment and detachment of cross-bridges. In summary, RLC phosphorylation affects the rate and extent of force redevelopment. These findings were obtained in maximally activated muscle at saturating [Ca2+ ] and are not explained by changes in the Ca2+ -sensitivity of acto-myosin interactions. The length-dependence of the rate of force redevelopment, together with the modulation by the state of RLC phosphorylation, suggests that these effects play a role in the Frank-Starling law of the heart.Published versio

Caspase-2 deficiency enhances whole-body carbohydrate utilisation and prevents high-fat diet-induced obesity

Published online 26 October 2017Caspase-2 has been shown to be involved in metabolic homeostasis. Here, we show that caspase-2 deficiency alters basal energy metabolism by shifting the balance in fuel choice from fatty acid to carbohydrate usage. At 4 weeks of age, whole-body carbohydrate utilisation was increased in Casp2-/- mice and was maintained into adulthood. By 17 weeks of age, Casp2-/- mice had reduced white adipose mass, smaller white adipocytes decreased fasting blood glucose and plasma triglycerides but maintained normal insulin levels. When placed on a 12-week high-fat diet (HFD), Casp2-/- mice resisted the development of obesity, fatty liver, hyperinsulinemia and insulin resistance. In addition, HFD-fed Casp2-/- mice had reduced white adipocyte hypertrophy, apoptosis and expansion of both subcutaneous and visceral adipose depots. Increased expression of UCP1 and the maintenance of adiponectin levels in white adipose tissue of HFD-fed Casp2-/- mice indicated increased browning and adipocyte hyperplasia. We found that while the preference for whole-body carbohydrate utilisation was maintained, HFD-fed Casp2-/- mice were not impaired in their ability to switch to utilising fats as a fuel source. Our findings suggest that caspase-2 impacts basal energy metabolism by regulating adipocyte biology and fat expansion, most likely via a non-apoptotic function. Furthermore, we show that caspase-2 deficiency shifts the balance in fuel choice towards increased carbohydrate utilisation and propose that this is due to mild energy stress. As a consequence, Casp2-/- mice show an adaptive remodelling of adipose tissue that protects from HFD-induced obesity and improves glucose homeostasis while paradoxically increasing their susceptibility to oxidative stress induced damage and premature ageing.Claire H Wilson, Andrej Nikolic, Stephen J Kentish, Marianne Keller, George Hatzinikolas, Loretta Dorstyn, Amanda J Page and Sharad Kuma

A New Governance Approach to Designing an Effective Arrangement for the Sustainable Management of Renewable Marine Resources in the Eastern Caribbean States

The study\u27s main purpose is to propose a governance framework that meets the priority of sustainable development for the regulation of offshore renewable resources in the OECS region. The study develops an analytical framework for evaluating the recently adopted Round 3 model of governance for the regulation of offshore wind and other marine activities in the United Kingdom. The focus is on the licensing procedures applicable to offshore wind development. Thereafter, the study examines the appropriateness of the application of the Round 3 model to the regulation of marine renewables in the OECS, and makes recommendations in that regard. Additionally, through the study of marine renewable resource development, this thesis looks at general conditions for effective ocean governance. In this regard, the thesis argues that strict hierarchical governance of the marine environment is not a desirable approach to effective ocean governance