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Biochemical lesions of respiratory enzymes and configurational changes of mitochondria in vivo

I. The effect of fluoroacetate: A study by phase-contrast microscopy and time-lapse cinemicrography

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Summary

Chick embryo heart fragments in primary hanging-drop culture were treated with sodium fluoroacetate to induce inhibition of aconitate hydratase, a mitochondrial enzyme of the tricarboxylic acid cycle. The mitochondria were analyzed in the living myoblasts by phase-contrast time-lapse cinemicrography. The results were recorded in a 16 mm film. After 20–30 minutes contact of the cells with the inhibitor some mitochondria became thickened and swollen. The swelling was polymorphous, asynchronous and reversible; the same mitochondrion could swell and shrink many times. Some mitochondria seemed not to respond to fluoroacetate and remained rod-like. Mitochondria appeared the only cell components to be morphologically affected by fluoroacetate and the changes were specifically caused by the inhibitor. The type of mitochondrial swelling differed from the large-amplitude respiration-dependent swelling of the isolated mitochondria in vitro and from the configurational changes of isolated mitochondria associated with the respiratory states. The evidence pointed to a specific connection between the biochemical lesion caused by fluoroacetate and the configurational changes of the mitochondria. The mitochondrial swelling was to a large extent reversed by washing the cultures with Tyrode physiological saline solution and the reversal was further accentuated by incubation of the cultures in fresh nutrient medium.

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References

  • Allsopp, C.B., Fell, H.B.: The effects of fluoroacetate on beating embryo chick heart in tissue culture. Exp. Cell Res. 1, 590–594 (1950).

    Google Scholar 

  • Avers, C.J., Lin, F.H., Pfeffer, C.R.: Histochemical studies of mitochondrial variation during aerobic growth of respiration-normal baker's yeast. J. Histochem. Cytochem. 13, 344–349 (1965).

    Google Scholar 

  • Barasa, A.: Sulle modalità di movimento e sulle presunte relazioni tra mitocondri, lipocondri e goccie di grasso nelle cellule viventi in coltura. Monit. zool. ital. 68, Suppl. 254–263 (1960).

    Google Scholar 

  • Barratt, J.O.W.: Changes in chondriosomes occuring in pathological conditions. Quart. J. micr. Sci. 58, 553–566 (1913).

    Google Scholar 

  • Bovis, R., Kasten, F.H., Okigaki, T.: Electron microscopic study of the toxic effect of sodium fluoroacetate on rat myocardial cultures. Exp. Cell Res. 43, 611–621 (1966).

    Google Scholar 

  • Brooks, S.E.H., Audretsch, J.J.: Studies on hypoglycin toxicity in rats. I. Changes in hepatic ultrastructure. Amer. J. Path. 59, 161–179 (1970).

    Google Scholar 

  • Buffa, P., Ferrari, N., Pasquali-Ronchetti, I.: Fenomeni di reversibilità degli effetti del fluoroacetato nelle cellule coltivate in vitro. Atti Soc. ital. Patol. Vol. X, 10° Congr. Naz., Aprile 1967a, Parte 2a, 435–447.

  • Buffa, P., Godina, G., Barasa, A.: Mitochondrial changes induced by fluoroacetate in living cells. 5th Internat. Congr. Biochem., Moskow, 1961. Abstracts of Communications, 9.4.1509.

  • Buffa, P., Guarriera-Bobyleva, V., Muscatello, U., Pasquali-Ronchetti, I.: Conformational changes of mitochondria associated with uncoupling of oxidative phosphorylation in vivo and in vitro. Nature (Lond.) 226, 272–274 (1970).

    Google Scholar 

  • Buffa, P., Guarriera-Bobyleva, V., Pasquali-Ronchetti, I.: Biochemical effects of fluoroacetate poisoning in rat liver. In: Carbon-fluorine compounds. Chemistry, biochemistry and biological activities (K. Elliott and J. Birch eds.), p. 303–330. Amsterdam: Elsevier. Excerpta Medica. North Holland. 1972.

    Google Scholar 

  • Buffa, P., Muscatello, U., Godina, G., Barasa, A.: Mitochondrial modifications associated with the biochemical lesion caused by fluoroacetate. 3rd Europ. Regional Conf. Electr. Micr., Prague, 1964, p. 125–126.

  • Buffa, P., Pasquali-Ronchetti, I.: Ultrastructural modulations of mitochondria correlated to biochemical activity in vivo. 7th Internat. Congr. Electr. Micr., Grenoble, 1970. Vol. III, 147–148.

    Google Scholar 

  • Buffa, P., Pasquali-Ronchetti, I., Coppi, G.: Effects of uncoupling of oxidative phosphorylation on the ultrastructure of mitochondria in vivo. Atti VI Congr. ital. Micros. Elettr., Siena, 1967b, 7–11.

  • Buffa, P., Peters, R.A.: The in vivo formation of citrate induced by fluoroacetate and its significance. J. Physiol. (Lond.) 110, 488–500 (1949).

    Google Scholar 

  • Carey, N.H., Greville, G.D.: Mitochondria from embryonic tissues of the chick. 1. Preparation, characterization and some enzymic properties. Biochem. J. 71, 159–166 (1959a).

    Google Scholar 

  • Carey, N.H., Greville, G.D.: Mitochondria from embryonic tissues of the chick. 2. Metabolic activities. Biochem. J. 71, 166–176 (1959b).

    Google Scholar 

  • Chenoweth, M.B.: Monofluoroacetic acid and related compounds. J. Pharmacol. exp. Ther. Part II, 97, 383–424 (1949).

    Google Scholar 

  • Ernster, L., Kuylenstierna, B.: Outer membrane of mitochondria. In: Membranes of mitochondria and chloroplasts (E. Racker ed.), p. 172–212. New York: Van Nostrand Reinhold Co. 1970.

    Google Scholar 

  • Feinstein, A., Rowe, A.J.: Molecular mechanism of formation of an antigen-antibody complex. Nature (Lond.) 205, 147–149 (1965).

    Google Scholar 

  • Frédéric, J.: Recherches cytologiques sur le chondriome normal ou soumis à l'expérimentation dans des cellules vivantes cultivées in vitro. Arch. Biol. (Liège) 69, 167–349 (1958).

    Google Scholar 

  • Gavrilescu, N., Peters, R.A.: Biochemical lesions in vitamin B deficiency. Biochem. J. 25, 1397–1409 (1931a).

    Google Scholar 

  • Gavrilescu, N., Peters, R.A.: On the function of torulin. An in vitro effect of antineuritic vitamin concentrates. Biochem. J. 25, 2150–2161 (1931b).

    Google Scholar 

  • Godina, G., Buffa, P., Barasa, A.: Mitochondrial changes induced by fluoroacetate in living cells. Acta anat., (Basel) 47, 391 (1961a).

    Google Scholar 

  • Godina, G., Buffa, P., Barasa, A.: Transformations des mitochondries des cellules vivantes en culture soumis a l'action d'inhibiteurs enzymatiques. Bull. Association Anatomistes XLVIIe Réunion Naples, mars 1961 b, p. 366–369.

  • Godina, G., Buffa, P., Barasa, A.: I mitocondri di cellule viventi in coltura sotto l'azione di inibitori enzimatici. Arch. Sci. med. 112, 510–512 (1961c).

    Google Scholar 

  • Goyer, R.A.: The renal tubule in lead poisoning. I. Mitochondrial swelling and aminoaciduria. Lab. Invest. 19, 71–77 (1968).

    Google Scholar 

  • Green, D.E., Harris, R.A.: Conformational basis of energy transduction in mitochondria. FEBS Letters 5, 241–245 (1969).

    Google Scholar 

  • Gustafsson, R., Tata, J.R., Lindberg, O., Ernster, L.: The relationship between the structure and activity of rat skeletal muscle mitochondria after thyroidectomy and thyroid hormone treatment. J. Cell Biol. 26, 555–578 (1965).

    Google Scholar 

  • Hackenbrock, C.R.: Ultrastructural bases for metabolically linked mechanical activity in mitochondria. I. Reversible ultrastructural changes with change in metabolic steady state in isolated liver mitochondria. J. Cell Biol. 30, 269–297 (1966).

    Google Scholar 

  • Hackenbrock, C.R.: Ultrastructural bases for metabolically linked mechanical activity in mitochondria. II. Electron transport-linked ultrastructural transformations in mitochondria. J. Cell Biol. 37, 345–369 (1968).

    Google Scholar 

  • Hackenbrock, C.R., Rehn, T.G., Gamble, J.L. Jr., Weinbach, E.C., Lemasters, J.J.: Ultrastructural transformation in the mitochondrion: its relationship to the energy state of the mitochondrion and the energy state of the cell. In: Energy transduction in respiration and phosphorylation (E. Quagliariello, S. Papa, and C.S. Rossi eds.), p. 285–305. Bari: Adriatica Editrice 1971.

    Google Scholar 

  • Lehninger, A.L.: Water uptake and extrusion by mitochondria in relation to oxidative phosphorylation. Physiol. Rev. 42, 467–517 (1962).

    Google Scholar 

  • Lehninger, A.L.: The mitochondrion. Molecular basis of structure and function, p. 10. New York: Benjamin Inc. 1964.

    Google Scholar 

  • Lewis, M.R., Lewis, W.H.: Mitochondria (and other cytoplasmic structures) in tissue cultures. Amer. J. Anat. 17, 339–401 (1914–15).

    Google Scholar 

  • McDowell, E.M.: Light and electron microscopic studies of the rat kidney after administration of inhibitors of the citric acid cycle in vivo: I. Effects of sodium fluoroacetate on the proximal convoluted tubule. Amer. J. Path. 66, 513–542 (1972a).

    Google Scholar 

  • McDowell, E.M.: Light and electron microscopic studies of the rat kidney after administration of inhibitors of the citric acid cycle in vivo: II. Effects of sodium malonate on proximal convoluted tubule. Amer. J. Path. 67, 141–158 (1972b).

    Google Scholar 

  • Muirhead, H., Perutz, M.F.: Structure of haemoglobin. A threedimensional Fourier synthesis of reduced human haemoglobin at 5.5 Å resolution. Nature (Lond.) 199, 633–638 (1963).

    Google Scholar 

  • Muscatello, U., Guarriera-Bobyleva, V., Buffa, P.: Configurational changes in isolated rat liver mitochondria as revealed by negative staining. I. Modifications caused by osmotic and other factors. J. Ultrastruct. Res. 40, 215–234 (1972a).

    Google Scholar 

  • Muscatello, U., Guarriera-Bobyleva, V., Buffa, P.: Configurational changes in isolated rat liver mitochondria as revealed by negative staining. II. Modifications caused by changes in respiratory states. J. Ultrastruct. Res. 40, 235–260 (1972b).

    Google Scholar 

  • Muscatello, U., Pasquali-Ronchetti, I., Barasa, A.: An electron microscopic study of myoblasts from chick embryo heart cultured in vitro. J. Ultrastruct. Res. 23, 44–59 (1968).

    Google Scholar 

  • Packer, L.: Relation of structure to energy coupling in rat liver mitochondria. Fed. Proc. 29, 1533–1540 (1970).

    Google Scholar 

  • Paradisi, F.: Effect of some substances on mitochondrial swelling induced by diphtheria toxin in chicken embryo heart cell cultures. Experientia (Basel) 23, 742–743 (1967a).

    Google Scholar 

  • Paradisi, F.: Experiments on mitochondrial swelling in vivo. Experientia (Basel) 23, 752–753 (1967b).

    Google Scholar 

  • Paradisi, F.: Mitochondrial swelling induced by diphtheria toxin in cell cultures. Path. Microbiol. 30, 481–494 (1967c).

    Google Scholar 

  • Parsons, D.F., Williams, G.R., Thompson, W., Wilson, D., Chance, B.: Improvements in the procedure for purification of mitochondrial outer and inner membrane. Comparison of the outer membrane with smooth endoplasmic reticulum. In: Mitochondrial structure and compartmentation (E. Quagliariello, S. Papa, E.C. Slater and J.M. Tager eds.), p. 29–70. Bari: Adriatica Editrice. 1967.

    Google Scholar 

  • Pasquali-Ronchetti, I., Buffa, P.: Correlazione fra ultrastruttura e biochimica respiratoria dei mitocondri in vivo. Atti VII Congr. Ital. Microsc. Elettr., Modena, 1969, p. 113–115.

  • Penniston, J.T., Harris, R.A., Asai, J., Green, D.E.: The conformational basis of energy transformations in membrane systems, I. Conformational changes in mitochondria. Proc. nat. Acad. Sci. (Wash.) 59, 624–631 (1968).

    Google Scholar 

  • Peters, R. A.: Lethal synthesis. Proc. roy. Soc. B 139, 143–170 (1952).

    Google Scholar 

  • Peters, R.A.: Mechanism of the toxicity of the active constituent of Dichapetalum cymosum and related compounds. Advanc. Enzymol. 18, 113–159 (1957).

    Google Scholar 

  • Peters, R.A., Wakelin, R.W., Buffa, P., Thomas, L.C.: Biochemistry of fluoroacetate poisoning. The isolation and some properties of the fluorotricarboxylic acid inhibitor of citrate metabolism. Proc. roy. Soc. B 140, 497–507 (1953).

    Google Scholar 

  • Pucci, I.: Mitochondrial changes induced by fluoroacetate in chick embryo myocardium in vivo. Exp. Cell Res. 35, 412–414 (1964).

    Google Scholar 

  • Racker, E.: Function and structure of the inner membrane of mitochondria and chloroplasts. In: Membranes of mitochondria and chloroplasts (E. Racker ed.), p. 127–171. New York: Van Nostrand Reinhold Co. 1970.

    Google Scholar 

  • Reynolds, E.S.: Liver parenchymal cell injury. I. Initial alterations of the cell following poisoning with carbon tetrachloride. J. Cell Biol. 19, 139–157 (1963).

    Google Scholar 

  • Ruggiero, G., Paradisi, F., Giusti, G.: Mitochondrial swelling induced by diphtheria toxin. The behaviour of citrate in intoxicated cells. Enzym. biol. clin. 9, 168–174 (1968).

    Google Scholar 

  • Schulz, H., Löw, H., Ernster, L., Sjöstrand, F.S.: Elektronenmikroskopische Studien an Leberschnitten von Thyroxin-behandelten Ratten. Proc. Stockholm Conf. Electron Microsc. 1956, p. 133–137.

  • Seno, S., Utsumi, K., Yokomura, E.: Swelling and contraction of mitochondria related to metabolism. In: Intracellular membranous structure (S. Seno and E.V. Cowdry eds.), p. 155–166. Okayama: Japan Society for Cell Biology, 1965.

    Google Scholar 

  • Stoeckenius, W.: Electron microscopy of mitochondrial and model membranes. In.: Membranes of mitochondria and chloroplasts (E. Racker ed.), p. 53–90. New York: Van Nostrand Reinhold Co. 1970.

    Google Scholar 

  • Suzuki, K.: Giant hepatic mitochondria. Production in mice fed with cuprizone. Science 163, 81–82 (1969).

    Google Scholar 

  • Tandler, B., Erlandson, R.A., Smith, A.L., Wynder, E.L.: Riboflavin and mouse hepatic cell structure and function. II. Division of mitochondria during recovery from simple deficiency. J. Cell Biol. 41, 477–493 (1969).

    Google Scholar 

  • Ward, P.F.V., Peters, R.A.: The chemical and biochemical properties of fluorocitric acid. Biochem. J. 78, 661–668 (1961).

    Google Scholar 

  • Wichertjes, T., Schepman, A.M.H., Mellema, J.E., Bruggen, E.F.J. van: Electron microscopy of glutamate dehydrogenase and hemocyanin crystals: structural analysis by optical diffraction. 7th Internat. Congr. Electr. Micr., Grenoble, 1970, Vol. II, 625–626.

    Google Scholar 

  • Williams, C.H., Vail, W.J., Harris, R.A., Caldwell, M., Green, D.E.: Conformational basis of energy transduction in membrane systems. VIII. Configurational changes of mitochondria in situ and in vitro. Bioenergetics 1, 147–180 (1970).

    Google Scholar 

  • Wollenberger, A., Schulze, W.: Mitochondrial alterations in the myocardium of dogs with aortic stenosis. J. biophys. biochem. Cytol. 10, 285–288 (1961).

    Google Scholar 

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This work was supported by grants of the Consiglio Nazionale delle Richerce of Italy to both Institutes.

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Barasa, A., Godina, G., Buffa, P. et al. Biochemical lesions of respiratory enzymes and configurational changes of mitochondria in vivo . Z.Zellforsch 138, 187–210 (1973). https://doi.org/10.1007/BF00306607

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