CARETS: A prototype regional environmental information system. Volume 13: Utility of CARETS products to local planners; an evaluation

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TBC1D1 Regulates Insulin- and Contraction-Induced Glucose Transport in Mouse Skeletal Muscle

OBJECTIVE: TBC1D1 is a member of the TBC1 Rab-GTPase family of proteins and is highly expressed in skeletal muscle. Insulin and contraction increase TBC1D1 phosphorylation on phospho-Akt substrate motifs (PASs), but the function of TBC1D1 in muscle is not known. Genetic linkage analyses show a TBC1D1 R125W missense variant confers risk for severe obesity in humans. The objective of this study was to determine whether TBC1D1 regulates glucose transport in skeletal muscle. RESEARCH DESIGN AND METHODS: In vivo gene injection and electroporation were used to overexpress wild-type and several mutant TBC1D1 proteins in mouse tibialis anterior muscles, and glucose transport was measured in vivo. RESULTS: Expression of the obesity-associated R125W mutant significantly decreased insulin-stimulated glucose transport in the absence of changes in TBC1D1 PAS phosphorylation. Simultaneous expression of an inactive Rab-GTPase (GAP) domain of TBC1D1 in the R125W mutant reversed this decrease in glucose transport caused by the R125W mutant. Surprisingly, expression of TBC1D1 mutated to Ala on four conserved Akt and/or AMP-activated protein kinase predicted phosphorylation sites (4P) had no effect on insulin-stimulated glucose transport. In contrast, expression of the TBC1D1 4P mutant decreased contraction-stimulated glucose transport, an effect prevented by concomitant disruption of TBC1D1 Rab-GAP activity. There was no effect of the R125W mutation on contraction-stimulated glucose transport. CONCLUSIONS: TBC1D1 regulates both insulin- and contraction-stimulated glucose transport, and this occurs via distinct mechanisms. The R125W mutation of TBC1D1 impairs skeletal muscle glucose transport, which could be a mechanism for the obesity associated with this mutation

A preliminary analysis of spatiotemporal patterns in swordfish habitat distributions.

A species distribution model (SDM) for swordfish that was in the development stage has been finalized. The model used detailed biological and oceanographic data to define the spatial distribution of Swordfish. The SDM adequately predicted Swordfish habitat (and thus fish) distributions such that it was found suitable for investigations into the spatiotemporal distribution of habitat. Results of this preliminary investigation supports the current hypothesized stock boundaries between the north and south Atlantic stocks used for management. Both the north and south Atlantic may be experiencing an expansion of habitat. This could result in decreased density of swordfish into a larger area and/or change MSY production metrics. A more detailed examination of this possibility is recommended.Versión del edito

Characterization of an Acute Muscle Contraction Model Using Cultured C2C12 Myotubes

A cultured C2C12 myotube contraction system was examined for application as a model for acute contraction-induced phenotypes of skeletal muscle. C2C12 myotubes seeded into 4-well rectangular plates were placed in a contraction system equipped with a carbon electrode at each end. The myotubes were stimulated with electric pulses of 50 V at 1 Hz for 3 ms at 997-ms intervals. Approximately 80% of the myotubes were observed to contract microscopically, and the contractions lasted for at least 3 h with electrical stimulation. Calcium ion $(Ca^{2+})$ transient evoked by the electric pulses was detected fluorescently with Fluo-8. Phosphorylation of protein kinase B/Akt (Akt), 5′ AMP-activated protein kinase (AMPK), p38 mitogen-activated protein kinase (p38), and c-Jun NH2-terminal kinase (JNK)1/2, which are intracellular signaling proteins typically activated in exercised/contracted skeletal muscle, was observed in the electrically stimulated C2C12 myotubes. The contractions induced by the electric pulses increased glucose uptake and depleted glycogen in the C2C12 myotubes. C2C12 myotubes that differentiated after exogenous gene transfection by a lipofection or an electroporation method retained their normal contractile ability by electrical stimulation. These findings show that our C2C12 cell contraction system reproduces the muscle phenotypes that arise in vivo (exercise), in situ (hindlimb muscles in an anesthetized animal), and in vitro (dissected muscle tissues in incubation buffer) by acute muscle contraction, demonstrating that the system is applicable for the analysis of intracellular events evoked by acute muscle contraction

Contraction and AICAR Stimulate IL-6 Vesicle Depletion From Skeletal Muscle Fibers In Vivo

Recent studies suggest that interleukin 6 (IL-6) is released from contracting skeletal muscles; however, the cellular origin, secretion kinetics, and signaling mechanisms regulating IL-6 secretion are unknown. To address these questions, we developed imaging methodology to study IL-6 in fixed mouse muscle fibers and in live animals in vivo. Using confocal imaging to visualize endogenous IL-6 protein in fixed muscle fibers, we found IL-6 in small vesicle structures distributed throughout the fibers under basal (resting) conditions. To determine the kinetics of IL-6 secretion, intact quadriceps muscles were transfected with enhanced green fluorescent protein (EGFP)-tagged IL-6 (IL-6-EGFP), and 5 days later anesthetized mice were imaged before and after muscle contractions in situ. Contractions decreased IL-6-EGFP–containing vesicles and protein by 62% (P < 0.05), occurring rapidly and progressively over 25 min of contraction. However, contraction-mediated IL-6-EGFP reduction was normal in muscle-specific AMP-activated protein kinase (AMPK) α2-inactive transgenic mice. In contrast, the AMPK activator AICAR decreased IL-6-EGFP vesicles, an effect that was inhibited in the transgenic mice. In conclusion, resting skeletal muscles contain IL-6–positive vesicles that are expressed throughout myofibers. Contractions stimulate the rapid reduction of IL-6 in myofibers, occurring through an AMPKα2-independent mechanism. This novel imaging methodology clearly establishes IL-6 as a contraction-stimulated myokine and can be used to characterize the secretion kinetics of other putative myokines

The acquisition of mechano-electrical transducer current adaptation in auditory hair cells requires myosin VI

Mutations in Myo6, the gene encoding the (F-actin) minus end-directed unconventional myosin, myosin VI, cause hereditary deafness in mice (Snell's waltzer) and humans. In the sensory hair cells of the cochlea, myosin VI is expressed in the cell bodies and along the stereocilia that project from the cells’ apical surface. It is required for maintaining the structural integrity of the mechanosensitive hair bundles formed by the stereocilia. In this study we investigate whether myosin VI contributes to mechano-electrical transduction. We report that Ca²+-dependent adaptation of the mechano-electrical transducer (MET) current, which serves to keep the transduction apparatus operating within its most sensitive range, is absent in outer and inner hair cells from homozygous Snell's waltzer mutant mice, which fail to express myosin VI. The operating range of the MET channels is also abnormal in the mutants, resulting in the absence of a resting MET current. We found that cadherin 23, a component of the hair bundle's transient lateral links, fails to be downregulated along the length of the stereocilia in maturing Myo6 mutant mice. MET currents of heterozygous littermates appear normal. We propose that myosin VI, by removing key molecules from developing hair bundles, is required for the development of the MET apparatus and its Ca²+-dependent adaptation