Skip to main content
SpringerLink
Log in

Effects of hydrogen peroxide on mitochondrial enzyme function studied in situ in rat heart myocytes

  • Original Contributions
  • Published:
Basic Research in Cardiology Aims and scope Submit manuscript

Summary

Our previous work indicated that energy transduction, as measured by myocyte respiration, was inhibited by hydrogen peroxide, but the mitochondrial membrane potential was relatively unaffected. Therefore, we determined in the present study the critical steps in mitochondrial energy transduction by measuring the sensitivity to hydrogen peroxide of NADH-CoQ reductase, ATP synthase, and adenine nucleotide translocasein situ in myocytes. Adult rat heart cells were isolated using collagenase and incubated in the presence of 0.1–10 mM hydrogen peroxide for 30 min. Activities of NADH-CoQ reductase and oligomycin-sensitive ATP synthase were assayed enzymatically with sonicated myocytes, and adenine nucleotide translocase activities were determined by atractylosideinhibitable [14C]ADP uptake of myocytes, permeabilized by saponin. The NADH-CoQ reductase and ATP synthase activities were inhibited to 77% and 67% of control, respectively, following an exposure to 10 mM hydrogen peroxide for 30 min. The adenine nucleotide translocase activities were inhibited in a concentration- and time-dependent manner and by 10 mM hydrogen peroxide to 44% of control. The dose-response relationship indicated that the translocase was the most susceptible to hydrogen peroxide among the three enzymes studied. Combined treatment of myocytes with 3-amino-1,2,4-triazole, 1,3-bis(2-chloroethyl)-1-nitrosourea and diethyl maleate (to inactivate catalase, to inhibit glutathione reductase activity, and to deplete glutathione, respectively) enhanced the sensitivity of translocase to hydrogen peroxide, supporting the view that the cellular defense mechanism is a significant factor in determining the toxicity of hydrogen peroxide. The results indicate that hydrogen peroxide can cause dysfunction in mitochondrial energy transduction, principally as the result of inhibition of adenine nucleotide translocase.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Balaban RS (1990) Regulation of oxidative phosphorylation in the mammalian cell. Am J Physiol 258:C377-C389

    Google Scholar 

  2. Chance B, Sies H, Boveris A (1979) Hydroperoxide metabolism in mammalian organs. Physiol Rev 59:527–6055

    Google Scholar 

  3. Chevion M (1988) A site-specific mechanism for free radical induced biological damage: The essential role of redox-active transition metals. Free Radicals Biol Med 5:27–37

    Google Scholar 

  4. Cotgreave IA, Moldeus P, Orrenius S (1988) Host biochemical defense mechanisms against prooxidants. Ann Rev Pharmacol Toxicol 28:189–212

    Google Scholar 

  5. Das AM, Harris DA (1990) Control of mitochondrial ATP synthesis in heart cells: inactive to active transitions caused by beating or positive inotropic agents. Cardiovasc Res 24:411–417

    Google Scholar 

  6. Davies KJA, Lin SW (1988) Oxidatively denatured proteins are degraded by an ATP-independent proteolytic pathway in Escherichia coli. Free Radicals Biol Med 5:225–236

    Google Scholar 

  7. Deneke SM, Fanburg BL (1989) Regulation of cellular glutathione. Am J Physiol 257:L163-L173

    Google Scholar 

  8. Duan J, Karmazyn M (1988) Relationship between oxidative phosphorylation and adenine nucleotide translocase activity of two populations of cardiac mitochondria and mechanical recovery of ischemic hearts following reperfusion. Can J Physiol Pharmacol 67:704–709

    Google Scholar 

  9. Duee ED, Vignais PV (1969) Kinetics and specificity of the adenine nucleotide translocation in the rat liver mitochondria. J Biol Chem 244:3920–3931

    Google Scholar 

  10. Hassinen IE (1986) Mitochondrial respiratory control in the myocardium. Biochem Biophys Acta 853:135–151

    Google Scholar 

  11. Hatefi Y, Rieske JS (1967) Preparation and properties on DPNH-coenzymes Q reductase (complex I of respiratory chain). Methods Enzym 10:235–239

    Google Scholar 

  12. Humphrey SM, Buckman JE, Holliss DG (1990) Subcellular distribution of energy metabolites in the pre-ischaemic and post-ischaemic perfused working rat heart. Eur J Biochem 191:755–759

    Google Scholar 

  13. Hyslop PA, Hinshaw DB, Schraufstatter IU, Sklar LA, Spragg RG, Cochrane CG (1986) Intracellular calcium homeostasis during hydrogen peroxide injury to cultured P388D1 cells. J Cell Physiol 129:356–366

    Google Scholar 

  14. Imlay JA, Linn S (1988) DNA damage and oxygen radical toxicity. Science 240:1302–1309

    Google Scholar 

  15. Jenkinson SG, Jordan JM, Lawrence RA (1988) BCNU-induced protection from hyperbaric hyperoxia: role of glutathione metabolism. J Appl Physiol 65:2531–2535

    Google Scholar 

  16. Kaminishi T, Matsuoka T, Yanagishita T, Kako KJ (1989) Increase vs decrease of calcium uptake by isolated heart cells induced by H2O2 vs HOCl. Am J Physiol 256:C598-C607

    Google Scholar 

  17. Karmazyn M, Mailer K, Currie W (1990) Acquisition and decay of heat-shock-enhanced postischemic ventricular recovery. Am J Physiol 28:H424-H431

    Google Scholar 

  18. Konno N, Kako KJ (1991) Effects of hydrogen peroxide and hypochlorite on membrane potential of mitochondria in situ in rat heart cells. Can J Physiol Pharmacol 69:1705–1712

    Google Scholar 

  19. Konno N, Kako KJ (1992) Role of cellular defense against hydrogen peroxide-induced inhibition of myocyte respiration. Basic Res Cardiol 87:239–249

    Google Scholar 

  20. LaNoue KF, Watts JA, Koch CD (1981) Adenine nucleotide transport during cardiac ischemia. Am J Physiol 241:H663–671

    Google Scholar 

  21. Malis CD, Bonventre JV (1986) Mechanism of calcium potentiation of oxygen free radical injury to renal mitochondria. J Biol Chem 261:14201–14208

    Google Scholar 

  22. Matsuoka T, Kato M, Kako KJ (1990) Effect of oxidants on Na,K,ATPase and its reversal. Basic Res Cardiol 85:330–341

    Google Scholar 

  23. McCune SA, Harris RA (1979) Mechanism responsible for 5-(tetradecyloxy)-2-furoic acid inhibition of hepatic lipogenesis. J Biol Chem 254:10095–10101

    Google Scholar 

  24. Pascoe GA, Reed DJ (1989) Cell calcium, vitamin E, and the thiol redox system in cytotoxicity. Free Radicals Biol Med 6:209–224

    Google Scholar 

  25. Reimer KA, Jennings RB (1986) Myocardial ischemia, hypoxia and infarction. In: Fozard HA, Jennings RB, Haber E, Katz AM, Morgan HE (eds) The Heart and Cardiovascular System, v 2. Raven Press, New York, pp 1133–1201

    Google Scholar 

  26. Rouslin W (1983) Mitochondrial complexes I, II, III, IV, and V in myocardial ischemia and autolysis. Am J Physiol 244:H743-H748

    Google Scholar 

  27. Shlafer M, Myers CL, Adkins S (1987) Mitochondrial hydrogen peroxide generation and activities of glutathione peroxidase and superoxide dismutase following global ischemia. J Mol Cell Cardiol 19:1195–1206

    Google Scholar 

  28. Tager JM, Wanders RJA, Groen AK, Kunz W, Bohnensack R, Kuster U, Letko G, Bohme G, Duszynski J, Wojtczak L (1983) Control of mitochondrial respiration. FEBS Letters 151:1–9

    Google Scholar 

  29. Thomas CE, Aust SD (1989) Role of metals in oxygen radical reactions and oxidative stress. In: Miquel J (ed) CRC Handbook of Free Radicals and Antioxidants in Biomedicine, v 1. CRC press Inc, Boca Raton, pp 37–48

    Google Scholar 

  30. Timerman AP, Altschuld RA, Hohl CM, Brierly GP, Merola AJ (1990) Cellular glutathione and response of adult rat heart myocytes to oxidant stress. J Moll Cell Cardiol 22:565–575

    Google Scholar 

  31. Wiswedel I, Trumper L, Schild L, Augustin W (1988) Injury of mitochondrial respiration and membrane potenial during iron/ascorbate-induced peroxidation. Biochim Biophys Acta 934:80–86

    Google Scholar 

  32. Woods JS, Calas CA, Aicher LD (1990) Stimulation of porphyrinogen oxidation by mercuric ion. II. Promotion of oxidation from the interaction of mercuric ions, glutathione, and mitochondriagenerated H2O2. Mol Pharmacol 38:261–266

    Google Scholar 

  33. Yanagishita T, Matsuoka T, Kako KJ (1989) Pharmacological intervention in oxidant-induced calcium pump dysfunction of dog heart. Biochem Internt 18:1111–1119

    Google Scholar 

  34. Zhang Y, Marcillat O, Giulivi C, Ernster L, Davies KJA (1990) The oxidative inactivation of mitochondrial electron transport chain components and ATPase. J Biol Chem 265:16330–16336

    Google Scholar 

  35. Zuurendonk PF, Tager JM (1974) Rapid separation of particulate components and soluble cytoplasm of isolated rat-liver cells. Biochem Biophys Acta 333:393–399

    Google Scholar 

  36. Zweier JL, Kuppusamy P, Williams R, Rayburn BK, Smith D, Weisfeldt ML, Flaherty JT (1989) Measurement and characterization of postischemic free radical generation in the isolated perfused heart. J Biol Chem 264:18890–18895

    Google Scholar 

Download references

Author information

Author notes

  1. K. J. Kako

    Present address: Department of Physiology, Faculty of Medicine, University of Ottawa, 451 Smyth Road, K1H 8M5, Ottawa, Ontario, Canada

  2. T. Tatsumi (post-doctoral fellow of the Heart and Stroke Foundation of Ontario)

    Present address: II. Department of Medicine, Kyoto Prefectural University of Medicine, Kawaramachi-Hirokoji, 602, Kyoto, Japan

Authors and Affiliations

  1. Department of Physiology, Faculty of Medicine, University of Ottawa, 451 Smyth Road, K1H 8M5, Ottawa, Ontario, Canada

    T. Tatsumi (post-doctoral fellow of the Heart and Stroke Foundation of Ontario)

Authors
  1. T. Tatsumi
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. K. J. Kako
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Tatsumi, T., Kako, K.J. Effects of hydrogen peroxide on mitochondrial enzyme function studied in situ in rat heart myocytes. Basic Res Cardiol 88, 199–211 (1993). https://doi.org/10.1007/BF00794993

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00794993

Key words

Search

Navigation