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Mitochondrial Redox Changes during Rest-Work Transition in Dog Gracilis Muscle

  • J. Olgin
  • R. J. Connect
  • B. Chance
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 200)

Abstract

The state of reduction of mitochondrial pyridine nucleotides (PN) and flavoproteins (FP) can in principle be estimated in tissue from their fluorescence.1 Although this technique has teen applied quantitatively to isolated mitochondria, there have been major problems in obtaining quantitative estimates of the state of mitochondrial reduction in vivo. Two basic categories of difficulties occur in the application to intact tissue: (1) heterogeneity of the signal due to nonspecific tissue reflectance and/or fluorescence, limited spatial resolution, variable mitochondrial content and interference by extraneous chromophores such as hemoglobin and myoglobin; and (2) lack of reproducible, absolute calibration of the recording apparatus for maximally oxidized and reduced tissue signals. We have developed techniques to overcome some of the above limitations. By using frozen tissue samples and making measurements at liquid N2 temperatures with laser enhanced activation light, greater fluorescence yields are obtained and the spatial resolution is improved.2 The use of the ratio of two indicators, flavoprotein and pyridine nucleotides, helps minimize interference by variable mitochondrial concentrations and the effect of extraneous chromophores.2 Finally, samples corresponding to full oxidation and full reduction can be made available for calibration. This report describes the application of these techniques and other methodological improvements to quantitate the state of mitochondrial redox during the transition from rest to heavy work in dog gracilis muscle.

Keywords

Light Guide Metabolic Ratio Pyridine Nucleotide Gracilis Muscle Toad Muscle 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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References

  1. 1.
    Chance, B., J.R. Williamson, D. Jamieson, and B. Schoener. Properties and kinetics of reduced pyridine nucleotide fluorescence of the isolated and in vivo rat heart. Biochem. Z. 341: 357 (1965).Google Scholar
  2. 2.
    Chance, B., B. Schoener, R. Oshino, F. Itschak, and Y. Nakase. Oxidation-reduction ratio studies of mitochondria in freeze-trapped samples. J. Biol. Chem. 254: 4764 (1979).Google Scholar
  3. 3.
    Gayeski, T.E.J., R.J. Connett, and C.R. Honig. Oxygen transport in rest-work transition illustrates new functions for myoglobin. Am. J. Physiol. 248: H914 (1985).Google Scholar
  4. 4.
    Connett, R.J., T.E.J. Gayeski, and C.R. Honig. Lactate accumulation in fully aerobic, working dog gracilis muscle. Am. J. Physiol. 246: H120 (1984).Google Scholar
  5. 5.
    Quistorff, B. and B. Chance. Two and three dimensional analysis of brain oxygen delivery. In Oxygen and Physiological Function, F.F. Jobsis, ed. Professional Information Library, p. 100 (1977).Google Scholar
  6. 6.
    Chance, B., U. Legalais, J. Sorge, and N. Graham. A versatfle time-sharing multichannel spectrophotometer, reflectometer, and fluorometer. Analytical Biochem. 66: 498 (1975).CrossRefGoogle Scholar
  7. 7.
    Chance, B. and B. Schoener. Fluorometric studies of flavin component of the respiratory chain. In Flavins and Fiavoproteins, E.C. Slater, ed. Elsevier, Amsterdam, p. 510 (1966).Google Scholar
  8. 8.
    Hassinen, I. and B. Chance. Oxidation-reduct,rn properties of the mitochondrial flavoprotein chain. Biochem. Biophys. Res. Comm. 31(6): 895 (1968).Google Scholar
  9. 9.
    Chance, B., L. Mela, and D. Wong. Fiavoproteins of the respiratory chain. In Flavins and Flavoproteins K. Yasi, ed. University Park Press, Baltimore, p. 107 (1968).Google Scholar
  10. 10.
    Clark, A. Jr., and P.A.A. Clark. Capture of spatially homogeneous chemical reactions in tissue by freezing. Biophvs. J. 42: 25 (1983).CrossRefGoogle Scholar
  11. 11.
    Haselgrove, J.C. C.H. Barlow, and B. Chance. The 3-D distribution of metabolic states in the gerbil brain during the course of spreading depression. In Cerebral Metabolism and Neuronal Function, J.U. Passonneau, R.A. Hawkins, W.D. Lust, and F.A. Welsh,Williams & Wilkins, p. 72 (1980).Google Scholar
  12. 12.
    Haselgrove, J.C., C. Barlow, B. Chance, and E. Joyce. Three dimensional display of the ischemic region of a rat heart. In Frontiers of Biological’Energetics, Vol. II, P.L. Dutton, J.S. Leigh, and A. Scarpa, p. 1515 (1978).Google Scholar
  13. 13.
    Quistorff, B., B. Chance, and T. Takeda. Two and three dimensional redox heterogeneity of rat liver. In Frontiers of Biological Energetics, Vol. II, P.L. Dutton, J.S. Leigh, and A. Scarpa,p. 1487 (1978).Google Scholar
  14. 14.
    Chance, B. and B. Quistorff. Study of tissue oxygen gradients by single and multiple indicators. In Oxygen Transport to Tissue III, I.A. Silver, M. Ericinska, and H. Bicher,Plenum Press, p. 331 (1978).Google Scholar
  15. 15.
    Shapiro, I.M., E.E. Golub, S. Kakuta, J.C. Haselgrove, J. Haveri, B. Chance, and P. Frasca. Initiation of endochondrial calcification is related to changes in redox state of hypertrophie chrondrocytes. Science 217: 950 (1982).PubMedCrossRefGoogle Scholar
  16. 16.
    Chance, B., C.H. Barlow, J.C. Haselgrove, Y. Nakase, B. Quistorff, A. Matchinsky, and A. Mayevsky. Microheterogeneities of redox states of perfused intact organs. In Microheterogeneities and Metabolic Compartmentation, P.A. Srere and R.W. Esterbrook Academic Press, p. 131 (1978).Google Scholar
  17. 17.
    Quistorff, B. and B. Chance. Redox scanning in the study of metabolic zonation of the liver. In Regulation of HepaticMetabolism: Intra and Intercellular Compartmentation,R. Thurman, F. Kauffman, and K. Jungermann, Plenum Press, in press.Google Scholar
  18. 18.
    Gayeski, T.E.J. A cryogenic microspectrophotometric method for measuring myoglobin saturation in subcellular volumes; Application to resting dog gracilis muscle. Ph.D. Dissertation, University of Rochester, Rochester, NY (1982).Google Scholar
  19. 19.
    Schultz, R., T. Thurman, J.R. Williamson, B. Chance, and Th. Bucher. Flavin and pyridine nucleotide oxydation-reduction changes in perfused rat liver. Anoxia and subcellular localization of fluorescent flavoproteins. J. Biol. Chem. 244 (09): 2317 (1969).Google Scholar
  20. 20.
    Jobsis, F.F. and J.C. Duffield. Oxidative and glycolytic recovery metabolism in muscle. J. Gen. Physiol. 50: 1009 (1967).Google Scholar
  21. 21.
    Chapman, J.B. Fluorometric studies of oxidative metabolism in isolated papillary muscle of the rabbit. J. Gen. Physiol. 59: 135 (1972).Google Scholar
  22. 22.
    Williamson, J.R. Glycolytic control mechanisms. J. Biol. Chem. 240: 2308 (1965).Google Scholar
  23. 23.
    O’Connor, M.J., F. Welsh, L. Komarnicky, T. Davis, J. Stevens, D. Lewis, and C. Herman. Origin of labile NADH tissue fluorescence. In Oxygen and Physiological Function, F.F. Jobsis, ed. Professional Information Library, p. 909 (1977).Google Scholar
  24. 24.
    Jong, Y.A. and E.J. Davis. Reconstruction of steady-state in cell-free systems. Arch. Biochem. Biophys. 222: 179 (1983).Google Scholar
  25. 25.
    Connett, R.J., T.E.J. Gayeski, and C.R. Honig. Energy sources in fully aerobic rest-work transitions: A new role for glycolysis. Am. J. Physiol. 248: H922, 1985.Google Scholar
  26. 26.
    Connett, R.J., T.E.J. Gayeski, and C.R. Honig. An upper bound on the minimum P02 for 02 consumption in red muscle. Adv. Exp.Med. Biol., in press.Google Scholar
  27. 27.
    Chance, B. Reaction of oxygen with the respiratory chain in cells and tissues. J. Gen. Physiol. 49: 163 (1965).PubMedCrossRefGoogle Scholar
  28. 28.
    Chance, B., G. Mauriello, and Hubert, X.M. Muscle as a Tissue, C. Radai and S.M. Horvath, eds. McGraw Hill, pp. 128–145 (1962).Google Scholar
  29. 29.
    Chance, B. The response of mitochondria to muscular contraciton. Annals N.Y. Acad. Sci. 81: 4077 (1949).Google Scholar

Copyright information

© Plenum Press, New York 1986

Authors and Affiliations

  • J. Olgin
    • 2
  • R. J. Connect
    • 1
  • B. Chance
    • 2
  1. 1.School of Medicine and DentistryThe University of RochesterRochesterUSA
  2. 2.Department of Biochemistry and BiophysicsUniversity of PennsylvaniaPhiladelphiaUSA

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