Laner 2016 Abstract Mito Xmas Meeting Innsbruck

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OXPHOS and ET capacity in permeabilized fibers of canine superathletes.

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Laner V, Boushel RC, Hamilton KL, Miller BF, Williamson KK, Davis MS, Gnaiger E (2016)

Event: Mito Xmas Meeting 2016 Innsbruck AT

Comparative mitochondrial physiology strongly relies on quantitative data sets for comparison of OXPHOS capacities and respiratory control patterns between species and tissues. Combination and interpretation of a wide variety of studies requires standardization of respiratory protocols, implementation of quality control criteria, and consistency of normalization. Previously we described a reference method for the application of a cytochrome c threshold as exclusion criterion in mitochondrial OXPHOS analyses [1]. Alaskan sled dogs (N=6) were studied 72 to 120 h after finishing a competitive 1,000 mile race within less than nine days. Permeabilized fibers (0.81-1.28 mg ± 0.12 SD wet weight per assay) were prepared from needle biopsies and immediately studied by high-resolution respirometry [2] using 12 chambers in parallel (Oroboros Oxygraph-2k). Compared to human skeletal muscle fibers, the canine samples were more delicate to handle, highly sticky and appeared to be fragile, disintegrating to various degrees during substrate-uncoupler-inhibitor titration (SUIT) protocols in mt-respiration medium MiR06Cr. Two substrate-uncoupler-inhibitor titration protocols were applied (Fig. 1). SUIT1 emphasized pathway control with fatty acid oxidation (F) versus carbohydrate oxidation capacity, whereas the focus of SUIT2 was on coupling control with N-linked substrates. Both protocols were designed to provide a common reference state of NS-linked ET capacity, in comparison to separate N- and S-linked pathway control states (N versus S). NS-linked ET capacity was 262±41 pmol∙s-1∙mg-1 Ww independent of the presence or absence of 0.2 mM octanoyl carnitine (F). This is the highest value so far reported for mammalian skeletal muscle. Top human endurance athletes have a NS-linked ET capacity approaching 200 pmol∙s-1∙mg-1 Ww [3], compared to 153±19 pmol∙s-1∙mg-1 Ww in competitive racing horses [4].


O2k-Network Lab: AT Innsbruck MitoFit, AT Innsbruck Oroboros, CA Vancouver Boushel RC, US CO Fort Collins Miller BF, US OK Stillwater Davis MS, AT Innsbruck Gnaiger E, US CO Fort Collins Hamilton K


Labels: MiParea: Respiration, Comparative MiP;environmental MiP, Exercise physiology;nutrition;life style 


Organism: Dog  Tissue;cell: Skeletal muscle  Preparation: Permeabilized tissue 


Coupling state: LEAK, OXPHOS, ET  Pathway: F, N, S, NS, ROX  HRR: Oxygraph-2k  Event: Poster 


Affiliations

Laner V(1), Boushel RC(2), Hamilton KL(3), Miller BF(3), Williamson KK(4), Davis MS(5), Gnaiger E(1,6)
  1. Oroboros Instruments Corp, Innsbruck, Austria
  2. School Kinesiology, Univ British Columbia, Vancouver, BC, Canada
  3. Dept Health Exercise Sc, Colorado State Univ, Fort Collins, CO, USA
  4. Waypoint Veterinary Education, Edmond, OK, USA
  5. Dept Physiological Sc, Oklahoma State Univ, Stillwater, OK, USA
  6. Dept Visceral, Transplant Thoracic Surgery, D. Swarovski Research Lab, Medical Univ Innsbruck, Austria


References

  1. Laner V, Boushel RC, Hamilton KL, Miller BF, Williamson KK, Davis MS, Gnaiger E (2014) Cytochrome c flux control factor as a quality criterion in respiratory OXPHOS analysis in canine permeabilized fibers. Mitochondr Physiol Network 19.13:63-4.
  2. Pesta D, Gnaiger E (2012) High-resolution respirometry. OXPHOS protocols for human cells and permeabilized fibers from small biopsies of human muscle. Methods Mol Biol 810:25-58.
  3. Gnaiger E (2009) Capacity of oxidative phosphorylation in human skeletal muscle. New perspectives of mitochondrial physiology. Int J Biochem Cell Biol 41:1837-45.
  4. Votion DM, Gnaiger E, Lemieux H, Mouithys-Mickalad A, Serteyn D (2012) Physical fitness and mitochondrial respiratory capacity in horse skeletal muscle. PLoS One 7:e34890.