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Suspended animation of C. Elegans embryos using carbon monoxide

Suspended animation of C. Elegans embryos using carbon monoxide. By Cyndi Edwards. Hypoxia. Normal air: ~21 kPa O 2 (101 kPa = 1 atm) Mild hypoxia: 0.25-1.0 kPa O 2 Intermediate hypoxia: 0.001-0.25 kPa O 2 During mild hypoxia: HIF-1 (Hypoxia Inducible Factor-1)

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Suspended animation of C. Elegans embryos using carbon monoxide

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  1. Suspended animation of C. Elegans embryosusing carbon monoxide By Cyndi Edwards

  2. Hypoxia • Normal air: ~21 kPa O2 (101 kPa = 1 atm) • Mild hypoxia: 0.25-1.0 kPa O2 • Intermediate hypoxia: 0.001-0.25 kPa O2 • During mild hypoxia: • HIF-1 (Hypoxia Inducible Factor-1) • Anaerobic energy production genes up-regulated • glycolytic enzymes, glucose transporters • Antioxidant genes upregulated • catalase, superoxide dismutase

  3. Anoxia • <0.001 kPa O2 • HIF-1 response alone is not sufficient to protect the cells from free radicals • Oxidative phosphorylation ceases • ATP demand must also decrease • Cells of some organisms enter into a state called “suspended animation”

  4. Suspended Animation • No observable movement is visible • Developmental progression ceases • Turtle hepatocytes in this state are capable of reducing their ATP demand by 94% • limit protein synthesis, ion channel activity, and anabolic pathways • Zebrafish embryos and C. elegans embryos stop moving and developing

  5. C. Elegans Embryo Viability

  6. Carbon Monoxide: A Competitive Inhibitor of Oxygen Binding • CO binds and inhibits cytochrome c oxidase (complex IV), causing free radical production • Thus CO effectively lowers the effective amount of oxygen electron carriers CO

  7. Can carbon monoxide induce suspended animation in damaging low oxygen conditions?

  8. Experimental Setup Embryos put in 30 mL leak-proof glass syringes with gas inlets Embryos exposed to various levels of hypoxia or to anoxia by changing gas inflow Two-cell embryos were harvested from adults of the Bristol N2 strain Embryos were then removed and immediately photographed Embryos were moved to culture plates to assay their viability Embryos were exposed to the same oxygen level for 24 hours

  9. Intermediate Hypoxic Exposure Resulted in Abnormal Development

  10. Carbon monoxide was effective in inducing suspended animation

  11. Take Home Message • C. elegans embryos are capable of surviving in anoxia and mild hypoxia, but intermediate hypoxic conditions are lethal • CO induced suspended animation in embryos experiencing intermediate hypoxia • Embryos that had entered CO-induced suspended animation were highly viable

  12. References Article • Nystul TG, Roth MB. Proc Natl Acad Sci USA. 2004 Jun 15;101(24):9133-6. Epub 2004 Jun 7. Carbon monoxide-induced suspended animation protects against hypoxic damage in Caenorhabditis elegans. Images • http://biology.mcgill.ca/labs/roy/Researchinterestspage.html • http://en.wikipedia.org/wiki/Carbon_monoxide • http://www.castlefordschools.com/kent/07-08%20lessons/Lessons/Chemistry%20Lessons/chp%201/Chemistry_files/image045.jpg

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