Abstract
Mitochondria were isolated from the liver, kidney and mixed hindlimb skeletal muscle of three vertebrate species; the laboratory rat Rattus norvegicus, the bearded dragon lizard Pogona vitticeps, and the cane toad Bufo marinus. These vertebrate species are approximately the same body mass and have similar body temperatures. The content of cytochromes B, C, C1, and A were measured in these isolated mitochondria by oxidised–reduced difference spectra. Adenine nucleotide translocase (ANT) was measured by titration of mitochondrial respiration with carboxyactractyloside and the protein and phospholipid content of isolated mitochondria were also measured. Fatty acid composition of mitochondrial phospholipids was measured. Mitochondrial respiration was measured at 37°C under states III and IV conditions as well as during oligomycin inhibition. Species differed in the ratios of different mitochondrial cytochromes. Muscle mitochondria differed from kidney and liver mitochondria by having a higher ANT content relative to cytochrome content. Respiration rates were compared relative to a number of denominators and found to be most variable when expressed relative to mitochondrial protein content and least variable when expressed relative to mitochondrial cytochrome A and ANT content. The turnover of cytochromes was calculated and found to vary between 1 and 94 electrons s−1. The molecular activity of mitochondrial cytochromes was found to be significantly positively correlated with the relative polyunsaturation of mitochondrial membrane lipids.
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Brand MD, Turner N, Ocloo A, Else PL, Hulbert AJ (2003) Proton conductance and fatty acyl composition of liver mitochondria correlates with body mass in birds. Biochem J 376:741–748
Brookes PS, Buckingham JA, Tenreiro AM, Hulbert AJ, Brand MD (1998) The proton permeability of the inner membrane of liver mitochondria from ectothermic and endothermic vertebrates and from obese rates: correlations with standard metabolic rate and phospholipid fatty acid composition. Comp Biochem Physiol 119B:325–334
Else PL, Wu BJ (1999) What role for membranes in determining the higher sodium pump molecular activity mammals compared to ectotherms? J Comp Physiol B 169:296–302
Estabrook RW (1967) Mitochondrial respiratory control and the polarographic measurement of ADP:O ratios. Methods Enzymol 10:41–47
Folch J, Lees M, Stanley GHS (1957) A simple method for the isolation and purification of total lipids from animal tissues. J Biol Chem 226:497–509
Guderley HE, Johnston IA (1996) Plasticity of fish muscle mitochondria with thermal acclimation. J Exp Biol 199:1311–1317
Hazel JR (1972a) The effect of temperature acclimation upon succinic dehydrogenase activity from the epaxial muscle of the common goldfish (Carassius auratus L.) -I. Properties of the enzyme and the effect of lipid extraction. Comp Biochem Physiol B 43:837–861
Hazel JR (1972b) The effect of temperature acclimation upon succinic dehydrogenase activity from the epaxial muscle of the common goldfish (Carassius auratus L.) -II. Lipid reactivation of the soluble enzyme. Comp Biochem Physiol B 43:863–882
Hazel JR, Williams EE (1990) The role of alterations in membrane lipid composition in enabling physiological adaptation of organisms to their physical environment. Prog Lipid Res 29:167–227
Hulbert AJ, Else PL (1999) Membranes as possible pacemakers of metabolism. J Theor Biol 199:257–274
Hulbert AJ, Else PL (2000) Mechanisms underlying the cost of living in animals. Ann Rev Physiol 62:207–235
Hulbert AJ, Else PL (2005) Membranes and the setting of energy demand. J Exp Biol 208:1593–1599
Leary SC, Lyons CN, Rosenberger AG, Ballantyne JS, Stillman J, Moyes CD (2003) Fiber-type differences in muscle mitochondrial profiles. Am J Physiol 285:R817–R826
Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193:265–275
Mills GL, Lane PA, Weech PK (1984) A guide book to lipoprotein technique. In: Laboratory techniques in biochemistry and molecular biology, vol. 14, Amsterdam, Elsevier, pp 240–241
Pan DA, Storlein LH (1993) Dietary lipid profile is a determinant of tissue phospholipid fatty acid composition and rate of weight gain in rats. J Nutr 123:512–519
Porter RK, Hulbert AJ, Brand MD (1996) Allometry of mitochondrial proton leak: influence of membrane surface area and fatty acid composition. Am J Physiol 271:R1550–R1556
Rolfe DF, Hulbert AJ, Brand MD (1994) Characteristics of mitochondrial proton leak and control of oxidative phosphorylation in the major oxygen consuming tissues of the rat. Biochim Biophys Acta 1118:405–416
Schneider H, Lemasters JJ, Höchli M, Hackenbrock CR (1980) Liposome-mitochondrial inner membrane fusion. J Biol Chem 255:3748–3756
Schwerzmann K, Cruz-Orive LM, Eggman R, Sänger A, Weibel ER (1986) Molecular architecture of the inner membrane of mitochondria from rat liver: a combined biochemical and stereological study. J Cell Biol 102:97–103
Schwerzmann K, Hoppeler H, Kayar SR, Weibel ER (1989) Oxidative capacity of muscle and mitochondria: correlation of physiological, biochemical, and morphometric characteristics. Proc Nat Acad Sci USA 86:1583–1587
Stillwell W, Jenski LJ, Crump T, Ehringer W (1997) Effect of DHA on mouse mitochondrial membrane properties. Lipids 32:497–506
St-Pierre J, Brand MD, Boutilier RG (2000) The effect of metabolic depression on proton leak rate in mitochondria from hibernating frogs. J Exp Biol 203:1469–1478
Suarez RK, Staples JF, Lighton JRB, Mathieu-Costello O (2000) Mitochondrial function in flying honeybees (Apis mellifera): respiratory chain enzymes and electron flow from complex III to oxygen. J Exp Biol 203:905–911
Turner N, Else PL, Hulbert AJ (2003) DHA content of membranes determines molecular activity of the sodium pump: implications for diseases states and metabolism. Naturwissenschaften 90:521–523
Turner N, Haga KL, Hulbert AJ, Else PL (2005) Relationship between body size, Na+-K+-ATPase activity, and membrane lipid composition in mammal and bird kidney. Am J Physiol Regul Integr Comp Physiol 288:301–310
Williams JN Jr (1964) A method for the simultaneous quantitative estimation of cytochromes a, b, c1 and c in mitochondria. Arch Biochem Biophys 107:537–543
Willis WT, Dallman PR (1989) Impaired control of respiration in iron-deficient muscle mitochondria. Am J Physiol 257:C1080–C1085
Wodtke E (1981a) Temperature adaptation of biological membranes. The effects of acclimation temperature on the unsaturation of the main neutral and charged phospholipids in mitochondrial membranes of the carp (Cyprinus carpio L.). Biochim Biophys Acta 698–709
Wodtke E (1981b) Temperature adaptation of biological membranes. Compensation of the molar activity of cytochrome c oxidase in the mitochondrial energy-transducing membrane during thermal acclimation of the carp (Cyprinus carpio L.). Biochim Biophys Acta 640:710–720
Wu BJ, Else PL, Storlein LH, Hulbert AJ (2001) Molecular activity of Na+/K+ ATPase from different sources is related to the packing of membrane lipids. J Exp Biol 204:4271–4280
Wu BJ, Hulbert AJ, Storlien LH, Else PL (2004) Membrane lipids and sodium pumps of cattle and crocodiles: an experimental test of the membrane pacemaker theory of metabolism. Am J Physiol 287:R633–R641
Acknowledgements
This study was supported by funds from the Australian Research Council to A.J. Hulbert and P. Else as well as by funds from NSERC (Canada) to H. Guderley. Sabbatical support funds from Univ. Laval (to H. Guderley) were instrumental in permitting this research to be carried out.
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Communicated by I.D. Hume
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Hulbert, A.J., Turner, N., Hinde, J. et al. How might you compare mitochondria from different tissues and different species?. J Comp Physiol B 176, 93–105 (2006). https://doi.org/10.1007/s00360-005-0025-z
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DOI: https://doi.org/10.1007/s00360-005-0025-z