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. 1981 Sep 15;198(3):551–560. doi: 10.1042/bj1980551

The rôle of mitochondrial pyruvate transport in the stimulation by glucagon and phenylephrine of gluconeogenesis from L-lactate in isolated rat hepatocytes.

A P Thomas, A P Halestrap
PMCID: PMC1163301  PMID: 7326022

Abstract

The sensitivity of glucose production from L-lactate by isolated liver cells from starved rats to inhibition by alpha-cyano-4-hydroxycinnamate was studied. A small percentage of the maximal rate of gluconeogenesis was insensitive to inhibition by alpha-cyano-4-hydroxycinnamate, and evidence is presented to show that this is due to pyruvate entry into the mitochondria as alanine. After subtraction of this rate, Dixon plots of the reciprocal of the rate of gluconeogenesis against inhibitor concentration were linear both in the absence and presence of glucagon, phenylephrine or valinomycin, each of which stimulated gluconeogenesis by 30-50%. Pyruvate kinase activity was decreased by glucagon, but not by phenylephrine or valinomycin. Inhibition of gluconeogenesis by quinolinate (inhibitor of phosphoenolpyruvate carboxykinase) or monochloroacetate (probably inhibiting pyruvate carboxylation) caused a significant deviation from linearity of the Dixon plot obtained with alpha-cyano-4-hydroxycinnamate. Amytal, however, inhibited gluconeogenesis without affecting the linearity of this plot. These data, coupled with a computer simulation study, suggest that pyruvate transport may control gluconeogenesis from L-lactate and that hormones may stimulate this process through an effect on the respiratory chain. An additional role for pyruvate kinase and pyruvate carboxylase is quite compatible with the data presented.

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Selected References

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  1. Adam P. A., Haynes R. C., Jr Control of hepatic mitochondrial CO2 fixation by glucagon, epinephrine, and cortisol. J Biol Chem. 1969 Dec 10;244(23):6444–6450. [PubMed] [Google Scholar]
  2. Berry M. N., Friend D. S. High-yield preparation of isolated rat liver parenchymal cells: a biochemical and fine structural study. J Cell Biol. 1969 Dec;43(3):506–520. doi: 10.1083/jcb.43.3.506. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Blair J. B., James M. E., Foster J. L. Adrenergic control of glycolysis and pyruvate kinase activity in hepatocytes from young and old rats. J Biol Chem. 1979 Aug 25;254(16):7585–7590. [PubMed] [Google Scholar]
  4. Bryla J., Harris E. J., Plumb J. A. The stimulatory effect of glucagon and dibutyryl cyclic AMP on ureogenesis and gluconeogenesis in relation to the mitochondrial ATP content. FEBS Lett. 1977 Aug 15;80(2):443–448. doi: 10.1016/0014-5793(77)80494-8. [DOI] [PubMed] [Google Scholar]
  5. Chan T. M., Exton J. H. Studies on alpha-adrenergic activation of hepatic glucose output. J Biol Chem. 1978 Sep 25;253(18):6393–6400. [PubMed] [Google Scholar]
  6. Chen R. F. Removal of fatty acids from serum albumin by charcoal treatment. J Biol Chem. 1967 Jan 25;242(2):173–181. [PubMed] [Google Scholar]
  7. Cherrington A. D., Assimacopoulos F. D., Harper S. C., Corbin J. D., Park C. R., Exton J. H. Studies on the alpha-andrenergic activation of hepatic glucose output. II. Investigation of the roles of adenosine 3':5'-monophosphate and adenosine 3':5'-monophosphate-dependent protein kinase in the actions of phenylephrine in isolated hepatocytes. J Biol Chem. 1976 Sep 10;251(17):5209–5218. [PubMed] [Google Scholar]
  8. Cornell N. W., Lund P., Krebs H. A. The effect of lysine on gluconeogenesis from lactate in rat hepatocytes. Biochem J. 1974 Aug;142(2):327–337. doi: 10.1042/bj1420327. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Davidoff F. Effects of guanidine derivatives on mitochondrial function. 3. The mechanism of phenethylbiguanide accumulation and its relationship to in vitro respiratory inhibition. J Biol Chem. 1971 Jun 25;246(12):4017–4027. [PubMed] [Google Scholar]
  10. DeRosa G., Swick R. W. Metabolic implications of the distribution of the alanine aminotransferase isoenzymes. J Biol Chem. 1975 Oct 25;250(20):7961–7967. [PubMed] [Google Scholar]
  11. Demaugre F., Cepanec C., Leroux J. P. Characterization of oxalate as a catabolite of dichloroacetate responsible for the inhibition of gluconeogenesis and pyruvate carboxylation in rat liver cells. Biochem Biophys Res Commun. 1978 Dec 14;85(3):1180–1185. doi: 10.1016/0006-291x(78)90666-6. [DOI] [PubMed] [Google Scholar]
  12. Demaugre F., Leroux J. P., Cartier P. The effects of pyruvate concentration, dichloroacetate and alpha-cyano-4-hydroxycinnamate on gluconeogenesis, ketogenesis and [3-hydroxybutyrate]/[3-oxobutyrate] ratios in isolated rat hepatocytes. Biochem J. 1978 Apr 15;172(1):91–96. doi: 10.1042/bj1720091. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Denton R. M., Edgell N. J., Bridges B. J., Poole G. P. Acute regulation of pyruvate kinase activity in rat epididymal adipose tissue by insulin. Biochem J. 1979 Jun 15;180(3):523–531. doi: 10.1042/bj1800523. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Denton R. M., Halestrap A. P. Regulation of pyruvate metabolism in mammalian tissues. Essays Biochem. 1979;15:37–77. [PubMed] [Google Scholar]
  15. Dieterle P., Brawand F., Moser U. K., Walter P. Alanine metabolism in rat liver mitochondria. Eur J Biochem. 1978 Aug 1;88(2):467–473. doi: 10.1111/j.1432-1033.1978.tb12471.x. [DOI] [PubMed] [Google Scholar]
  16. Elliott K. R., Pogson C. I., Smith S. A. Permeability of the liver cell membrane to quinolinate. Biochem J. 1977 Apr 15;164(1):283–286. doi: 10.1042/bj1640283. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Exton J. H., Park C. R. Control of gluconeogenesis in liver. 3. Effects of L-lactate, pyruvate, fructose, glucagon, epinephrine, and adenosine 3',5'-monophosphate on gluconeogenic intermediates in the perfused rat liver. J Biol Chem. 1969 Mar 25;244(6):1424–1433. [PubMed] [Google Scholar]
  18. Foster J. L., Blair J. B. Acute hormonal control of pyruvate kinase and lactate formation in the isolated rat hepatocyte. Arch Biochem Biophys. 1978 Aug;189(2):263–276. doi: 10.1016/0003-9861(78)90212-6. [DOI] [PubMed] [Google Scholar]
  19. Friedmann N., Dambach G. Antagonistic effect of insulin on glucagon-evoked hyperpolarization. A correlation between changes in membrane potential and gluconeogenesis. Biochim Biophys Acta. 1980 Feb 28;596(2):180–185. doi: 10.1016/0005-2736(80)90352-1. [DOI] [PubMed] [Google Scholar]
  20. Garrison J. C., Borland M. K. Regulation of mitochondrial pyruvate carboxylation and gluconeogenesis in rat hepatocytes via an alpha-adrenergic, adenosine 3':5'-monophosphate-independent mechanism. J Biol Chem. 1979 Feb 25;254(4):1129–1133. [PubMed] [Google Scholar]
  21. Garrison J. C., Haynes R. C., Jr The hormonal control of gluconeogenesis by regulation of mitochondrial pyruvate carboxylation in isolated rat liver cells. J Biol Chem. 1975 Apr 25;250(8):2769–2777. [PubMed] [Google Scholar]
  22. HOPPER S., SEGAL H. L. Kinetic studies of rat liver glutamicalanine transaminase. J Biol Chem. 1962 Oct;237:3189–3195. [PubMed] [Google Scholar]
  23. Halestrap A. P., Denton R. M. The specificity and metabolic implications of the inhibition of pyruvate transport in isolated mitochondria and intact tissue preparations by alpha-Cyano-4-hydroxycinnamate and related compounds. Biochem J. 1975 Apr;148(1):97–106. doi: 10.1042/bj1480097. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Halestrap A. P. Pyruvate and ketone-body transport across the mitochondrial membrane. Exchange properties, pH-dependence and mechanism of the carrier. Biochem J. 1978 Jun 15;172(3):377–387. doi: 10.1042/bj1720377. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Halestrap A. P., Scott R. D., Thomas A. P. Mitochondrial pyruvate transport and its hormonal regulation. Int J Biochem. 1980;11(2):97–105. doi: 10.1016/0020-711x(80)90241-4. [DOI] [PubMed] [Google Scholar]
  26. Halestrap A. P. Stimulation of pyruvate transport in metabolizing mitochondria through changes in the transmembrane pH gradient induced by glucagon treatment of rats. Biochem J. 1978 Jun 15;172(3):389–398. doi: 10.1042/bj1720389. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Halestrap A. P. Stimulation of the respiratory chain of rat liver mitochondria between cytochrome c1 and cytochrome c by glucagon treatment of rats. Biochem J. 1978 Jun 15;172(3):399–405. doi: 10.1042/bj1720399. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Halestrap A. P. The mechanism of the inhibition of the mitochondrial pyruvate transportater by alpha-cyanocinnamate derivatives. Biochem J. 1976 Apr 15;156(1):181–183. doi: 10.1042/bj1560181. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Halestrap A. P. The mechanism of the stimulation of pyruvate transport into rat liver mitochondria by glucagon. Biochem Soc Trans. 1977;5(1):216–219. doi: 10.1042/bst0050216. [DOI] [PubMed] [Google Scholar]
  30. Halestrap A. P. The mitochondrial pyruvate carrier. Kinetics and specificity for substrates and inhibitors. Biochem J. 1975 Apr;148(1):85–96. doi: 10.1042/bj1480085. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Haynes R. C., Jr, Garrison J. C., Yamazaki R. K. Comparison of effects of glucagon and valinomycin on rat liver mitochondria and cells. Mol Pharmacol. 1974 May;10(3):381–388. [PubMed] [Google Scholar]
  32. Holland P. C., Clark M. G., Bloxham D. P., Lardy H. A. Mechanism of action of the hypoglycemic agent diphenyleneiodonium. J Biol Chem. 1973 Sep 10;248(17):6050–6056. [PubMed] [Google Scholar]
  33. Hue L., Felíu J. E., Hers H. G. Control of gluconeogenesis and of enzymes of glycogen metabolism in isolated rat hepatocytes. A parallel study of the effect of phenylephrine and of glucagon. Biochem J. 1978 Dec 15;176(3):791–797. doi: 10.1042/bj1760791. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Longshaw I. D., Bowen N. L., Pogson C. I. The pathway of gluconeogenesis in the cortex of guinea-pig kidney. Use of aminooxyacetate as a transaminase inhibitor. Eur J Biochem. 1972 Feb 15;25(2):366–371. doi: 10.1111/j.1432-1033.1972.tb01705.x. [DOI] [PubMed] [Google Scholar]
  35. McDaniel H. G., Reddy W. J., Boshell B. R. The mechanism of inhibition of phosphoenolpyruvate carboxylase by quinolinic acid. Biochim Biophys Acta. 1972 Aug 28;276(2):543–550. doi: 10.1016/0005-2744(72)91015-7. [DOI] [PubMed] [Google Scholar]
  36. McGarry J. D., Foster D. W. Regulation of hepatic fatty acid oxidation and ketone body production. Annu Rev Biochem. 1980;49:395–420. doi: 10.1146/annurev.bi.49.070180.002143. [DOI] [PubMed] [Google Scholar]
  37. McGivan J. D. Mechanism of the stimulation of serine and alanine transport into isolated rat liver cells by bicarbonate ions. Biochem J. 1979 Sep 15;182(3):697–705. doi: 10.1042/bj1820697. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Mendes-Mourāo J., Halestrap A. P., Crisp D. M., Pogson C. I. The involvement of mitochondrial pyruvate transport in the pathways of gluconeogenesis from serine and alanine in isolated rat and mouse liver cells. FEBS Lett. 1975 Apr 15;53(1):29–32. doi: 10.1016/0014-5793(75)80674-0. [DOI] [PubMed] [Google Scholar]
  39. Nagano M., Ishibashi H., McCully V., Cottam G. L. Epinephrine-stimulated phosphorylation of pyruvate kinase in hepatocytes. Arch Biochem Biophys. 1980 Aug;203(1):271–281. doi: 10.1016/0003-9861(80)90177-0. [DOI] [PubMed] [Google Scholar]
  40. Pande S. V., Parvin R. Pyruvate and acetoacetate transport in mitochondria. A reappraisal. J Biol Chem. 1978 Mar 10;253(5):1565–1573. [PubMed] [Google Scholar]
  41. Rognstad R. Cyclic AMP induced inhibition of pyruvate kinase flux in the intact liver cell. Biochem Biophys Res Commun. 1975 Apr 21;63(4):900–905. doi: 10.1016/0006-291x(75)90653-1. [DOI] [PubMed] [Google Scholar]
  42. Rognstad R. Effects of ethyl hydrazinoacetate on gluconeogenesis and on ethanol oxidation in rat hepatocytes. Biochim Biophys Acta. 1980 Feb 21;628(1):116–118. doi: 10.1016/0304-4165(80)90357-8. [DOI] [PubMed] [Google Scholar]
  43. Rognstad R., Katz J. Gluconeogenesis in the kidney cortex. Effects of D-malate and amino-oxyacetate. Biochem J. 1970 Feb;116(3):483–491. doi: 10.1042/bj1160483. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Rognstad R. Rate-limiting steps in metabolic pathways. J Biol Chem. 1979 Mar 25;254(6):1875–1878. [PubMed] [Google Scholar]
  45. Ross B. D., Hems R., Freedland R. A., Krebs H. A. Carbohydrate metabolism of the perfused rat liver. Biochem J. 1967 Nov;105(2):869–875. doi: 10.1042/bj1050869. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Siess E. A., Brocks D. G., Lattke H. K., Wieland O. H. Effect of glucagon on metabolite compartmentation in isolated rat liver cells during gluconeogenesis from lactate. Biochem J. 1977 Aug 15;166(2):225–235. doi: 10.1042/bj1660225. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Stanley P. E., Williams S. G. Use of the liquid scintillation spectrometer for determining adenosine triphosphate by the luciferase enzyme. Anal Biochem. 1969 Jun;29(3):381–392. doi: 10.1016/0003-2697(69)90323-6. [DOI] [PubMed] [Google Scholar]
  48. Steiner K. E., Chan T. M., Claus T. H., Exton J. H., Pilkis S. J. The role of phosphorylation in the alpha-adrenergic-mediated inhibition of rat hepatic pyruvate kinase. Biochim Biophys Acta. 1980 Oct 15;632(3):366–374. doi: 10.1016/0304-4165(80)90232-9. [DOI] [PubMed] [Google Scholar]
  49. Titheradge M. A., Binder S. B., Yamazaki R. K., Haynes R. C., Jr Glucagon treatment stimulates the metabolism of hepatic submitochondrial particles. J Biol Chem. 1978 May 25;253(10):3357–3360. [PubMed] [Google Scholar]
  50. Titheradge M. A., Coore H. G. Hormonal regulation of liver mitochondrial pyruvate carrier in relation to gluconeogenesis and lipogenesis. FEBS Lett. 1976 Nov 15;72(1):73–78. doi: 10.1016/0014-5793(76)80901-5. [DOI] [PubMed] [Google Scholar]
  51. Titheradge M. A., Coore H. G. The mitochondrial pyruvate carrier, its exchange properties and its regulation by glucagon. FEBS Lett. 1976 Mar 15;63(1):45–50. doi: 10.1016/0014-5793(76)80191-3. [DOI] [PubMed] [Google Scholar]
  52. Titheradge M. A., Stringer J. L., Haynes R. C., Jr The stimulation of the mitochondrial uncoupler-dependent ATPase in isolated hepatocytes by catecholamines and glucagon and its relationship to gluconeogenesis. Eur J Biochem. 1979 Dec;102(1):117–124. doi: 10.1111/j.1432-1033.1979.tb06271.x. [DOI] [PubMed] [Google Scholar]
  53. Tolbert M. E., Fain J. N. Studies on the regulation of gluconeogenesis in isolated rat liver cells by epinephrine and glucagon. J Biol Chem. 1974 Feb 25;249(4):1162–1166. [PubMed] [Google Scholar]
  54. Ui M., Claus T. H., Exton J. H., Park C. R. Studies on the mechanism of action of glucagon on gluconeogenesis. J Biol Chem. 1973 Aug 10;248(15):5344–5349. [PubMed] [Google Scholar]

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