Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 1977 Aug 15;166(2):225–235. doi: 10.1042/bj1660225

Effect of glucagon on metabolite compartmentation in isolated rat liver cells during gluconeogenesis from lactate

Elmar A Siess 1, Dietrich G Brocks 1, Herbert K Lattke 1, Otto H Wieland 1
PMCID: PMC1164999  PMID: 199159

Abstract

1. The subcellular distribution of adenine nucleotides, acetyl-CoA, CoA, glutamate, 2-oxoglutarate, malate, oxaloacetate, pyruvate, phosphoenolpyruvate, 3-phosphoglycerate, glucose 6-phosphate, aspartate and citrate was studied in isolated hepatocytes in the absence and presence of glucagon by using a modified digitonin procedure for cell fractionation. 2. In the absence of glucagon, the cytosol contains about two-thirds of cellular ATP, some 40–50% of ADP, acetyl-CoA, citrate and phosphoenolpyruvate, more than 75% of total 2-oxoglutarate, glutamate, malate, oxaloacetate, pyruvate, 3-phosphoglycerate and aspartate, and all of glucose 6-phosphate. 3. In the presence of glucagon the cytosolic space shows an increase in the content of malate, phosphoenolpyruvate and 3-phosphoglycerate by more than 60%, and those of aspartate and glucose 6-phosphate rise by about 25%. Other metabolites remain unchanged. After glucagon treatment, cytosolic pyruvate is decreased by 37%, whereas glutamate and 2-oxoglutarate decrease by 70%. The [NAD+]/[NADH] ratios calculated from the cytosolic concentrations of the reactants of lactate dehydrogenase and malate dehydrogenase were the same. Glucagon shifts this ratio and also that of the [NADP+]/[NADPH] couple towards a more reduced state. 4. In the mitochondrial space glucagon causes an increase in the acetyl-CoA and ATP contents by 25%, and an increase in [phosphoenolpyruvate] by 50%. Other metabolites are not changed by glucagon. Oxaloacetate in the matrix is only slightly decreased after glucagon, yet glutamate and 2-oxoglutarate fall to about 25% of the respective control values. The [NAD+]/[NADH] ratios as calculated from the [3-hydroxybutyrate]/[acetoacetate] ratio and from the matrix [malate]/[oxaloacetate] couple are lowered by glucagon, yet in the latter case the values are about tenfold higher than in the former. 5. Glucagon and oleate stimulate gluconeogenesis from lactate to nearly the same extent. Oleate, however, does not produce the changes in cellular 2-oxoglutarate and glutamate as observed with glucagon. 6. The changes of the subcellular metabolite distribution after glucagon are compatible with the proposal that the stimulation of gluconeogenesis results from as yet unknown action(s) of the hormone at the mitochondrial level in concert with its established effects on proteolysis and lipolysis.

Full text

PDF
228

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  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. Bewsher P. D., Ashmore J. Ketogenic and lipolytic effects of glucagon on liver. Biochem Biophys Res Commun. 1966 Aug 12;24(3):431–436. doi: 10.1016/0006-291x(66)90178-1. [DOI] [PubMed] [Google Scholar]
  3. Blair J. B., Cimbala M. A., Foster J. L., Morgan R. A. Hepatic pyruvate kinase. Regulation by glucagon, cyclic adenosine 3'-5'-monophosphate, and insulin in the perfused rat liver. J Biol Chem. 1976 Jun 25;251(12):3756–3762. [PubMed] [Google Scholar]
  4. Blair J. B., Cook D. E., Lardy H. A. Influence of glucagon on the metabolism of xylitol and dihydroxyacetone in the isolated perfused rat liver. J Biol Chem. 1973 May 25;248(10):3601–3607. [PubMed] [Google Scholar]
  5. Blair J. B., Cook D. E., Lardy H. A. Interaction of propionate and lactate in the perfused rat liver. Effects of glucagon and oleate. J Biol Chem. 1973 May 25;248(10):3608–3614. [PubMed] [Google Scholar]
  6. Carlson C. W., Baxter R. C., Ulm E. H., Pogell B. M. Role of oleate in the regulation of "neutral" rabbit liver fructose 1,6-diphosphatase activity. J Biol Chem. 1973 Aug 25;248(16):5555–5561. [PubMed] [Google Scholar]
  7. 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]
  8. Clark M. G., Kneer N. M., Bosch A. L., Lardy H. A. The fructose 1,6-diphosphatase-phosphofructokinase substrate cycle. A site of regulation of hepatic gluconeogenesis by glucagon. J Biol Chem. 1974 Sep 25;249(18):5695–5703. [PubMed] [Google Scholar]
  9. Claycomb W. C., Kilsheimer G. S. Effect of glucagon, adenosine-3',5'-monophosphate and theophylline on free fatty acid release by rat liver slices and on tissue levels of coenzyme A esters. Endocrinology. 1969 May;84(5):1179–1183. doi: 10.1210/endo-84-5-1179. [DOI] [PubMed] [Google Scholar]
  10. Cornell N. W., Lund P., Hems R., Krebs H. A. Acceleration of gluconeogenesis from lactate by lysine (Short Communication). Biochem J. 1973 Jun;134(2):671–672. doi: 10.1042/bj1340671. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. 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]
  12. Exton J. H., Corbin J. G., Park C. R. Control of gluconeogenesis in liver. IV. Differential effects of fatty acids and glucagon on ketogenesis and gluconeogenesis in the perfused rat liver. J Biol Chem. 1969 Aug 10;244(15):4095–4102. [PubMed] [Google Scholar]
  13. 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]
  14. Feliú J. E., Hue L., Hers H. G. Hormonal control of pyruvate kinase activity and of gluconeogenesis in isolated hepatocytes. Proc Natl Acad Sci U S A. 1976 Aug;73(8):2762–2766. doi: 10.1073/pnas.73.8.2762. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Fröhlich J., Wieland O. Glucagon and the permissive action of fatty acids in hepatic gluconeogenesis. Eur J Biochem. 1971 Apr 30;19(4):557–562. doi: 10.1111/j.1432-1033.1971.tb01349.x. [DOI] [PubMed] [Google Scholar]
  16. 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]
  17. Greenbaum A. L., Gumaa K. A., McLean P. The distribution of hepatic metabolites and the control of the pathways of carbohydrate metabolism in animals of different dietary and hormonal status. Arch Biochem Biophys. 1971 Apr;143(2):617–663. doi: 10.1016/0003-9861(71)90247-5. [DOI] [PubMed] [Google Scholar]
  18. Johnson M. E., Das N. M., Butcher F. R., Fain J. N. The regulation of gluconeogenesis in isolated rat liver cells by glucagon, insulin, dibutyryl cyclic adenosine monophosphate, and fatty acids. J Biol Chem. 1972 May 25;247(10):3229–3235. [PubMed] [Google Scholar]
  19. KEECH D. B., UTTER M. F. PYRUVATE CARBOXYLASE. II. PROPERTIES. J Biol Chem. 1963 Aug;238:2609–2614. [PubMed] [Google Scholar]
  20. Lardy H. A., Paetkau V., Walter P. Paths of carbon in gluconeogenesis and lipogenesis: the role of mitochondria in supplying precursors of phosphoenolpyruvate. Proc Natl Acad Sci U S A. 1965 Jun;53(6):1410–1415. doi: 10.1073/pnas.53.6.1410. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Ljunström O., Berglund L., Engström L. Studies on the kinetic effects of adenosine-3':5'-monophosphate-dependent phosphorylation of purified pig-liver pyruvate kinase type L. Eur J Biochem. 1976 Sep 15;68(2):497–506. doi: 10.1111/j.1432-1033.1976.tb10837.x. [DOI] [PubMed] [Google Scholar]
  22. Llorente P., Marco R., Sols A. Regulation of liver pyruvate kinase and the phosphoenolpyruvate crossroads. Eur J Biochem. 1970 Mar 1;13(1):45–54. doi: 10.1111/j.1432-1033.1970.tb00897.x. [DOI] [PubMed] [Google Scholar]
  23. MILLER L. L. DIRECT ACTIONS OF INSULIN, GLUCAGON, AND EPINEPHRINE ON THE ISOLATED PERFUSED RAT LIVER. Fed Proc. 1965 May-Jun;24:737–744. [PubMed] [Google Scholar]
  24. McClure W. R., Lardy H. A. Rat liver pyruvate carboxylase. IV. Factors affeing the regulation in vivo. J Biol Chem. 1971 Jun 10;246(11):3591–3596. [PubMed] [Google Scholar]
  25. Menahan L. A., Wieland O. Interactions of glucagon and insulin on the metabolism of perfused livers from fasted rats. Eur J Biochem. 1969 May 1;9(1):55–62. doi: 10.1111/j.1432-1033.1969.tb00575.x. [DOI] [PubMed] [Google Scholar]
  26. Müllhofer G., Loy E. A possible role of the glycerol phosphate cycle in cyclic AMP-stimulated gluconeogenesis from lactate in perfused rat livers. Hoppe Seylers Z Physiol Chem. 1974 Mar;355(3):239–254. doi: 10.1515/bchm2.1974.355.1.239. [DOI] [PubMed] [Google Scholar]
  27. Parrilla R., Jimenez I., Ayuso-Parrilla M. S. Glucagon and insulin control of gluconeogenesis in the perfused isolated rat liver. Effects on cellular metabolite distribution. Eur J Biochem. 1975 Aug 15;56(2):375–383. doi: 10.1111/j.1432-1033.1975.tb02243.x. [DOI] [PubMed] [Google Scholar]
  28. Parrilla R., Jimenez M. I., Ayuso-Parrilla M. S. Cellular redistribution of metabolites during glucagon and insulin control of gluconeogenesis in the isolated perfused rat liver. Arch Biochem Biophys. 1976 May;174(1):1–12. doi: 10.1016/0003-9861(76)90317-9. [DOI] [PubMed] [Google Scholar]
  29. Penhos J. C., Wu C. H., Daunas J., Reitman M., Levine R. Effect of glucagon on the metabolism of lipids and on urea formation by the perfused rat liver. Diabetes. 1966 Oct;15(10):740–748. doi: 10.2337/diab.15.10.740. [DOI] [PubMed] [Google Scholar]
  30. Pilkis S. J., Claus T. H., Riou J. P., Park C. R. Possible role of pyruvate kinase in the hormonal control of dihydroxyacetone gluconeogenesis in isolated heptatocytes. Metabolism. 1976 Nov;25(11 Suppl 1):1355–1360. doi: 10.1016/s0026-0495(76)80141-2. [DOI] [PubMed] [Google Scholar]
  31. Pogell B. M., Taketa K., Sarngadharan M. G. Reversal of the adenosine triphosphate and adenosine diphosphate inactivation of liver fructose 1,6-diphosphatase by 3-phosphoglycerate. J Biol Chem. 1971 Mar 25;246(6):1947–1948. [PubMed] [Google Scholar]
  32. Proffitt R. T., Sankaran L. Specific, reversible inactivation of phosphofructokinase by fructose-1,6-bisphosphatase. Involvement of adenosine 5'-triphosphate, oleate, and 3-phosphoglycerate. Biochemistry. 1976 Jun 29;15(13):2918–2925. doi: 10.1021/bi00658a034. [DOI] [PubMed] [Google Scholar]
  33. 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]
  34. Ross B. D., Hems R., Krebs H. A. The rate of gluconeogenesis from various precursors in the perfused rat liver. Biochem J. 1967 Mar;102(3):942–951. doi: 10.1042/bj1020942. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Rozengurt E., Jiménez de Asúa L., Carminatti H. Some kinetic properties of liver pyruvate kinase (type L). II. Effect of pH on its allosteric behavior. J Biol Chem. 1969 Jun 25;244(12):3142–3147. [PubMed] [Google Scholar]
  36. SCHIMASSEK H., MITZKAT H. J. UBER EINE SPEZIFISCHE WIRKUNG DES GLUCAGON AUF DIE EMBDEN-MEYERHOF-KETTE IN DER LEBER. VERSUCHE AN DER ISOLIERT PERFUNDIERTEN RATTENLEBER. Biochem Z. 1963 Aug 14;337:510–518. [PubMed] [Google Scholar]
  37. Saggerson D., Evans C. J. The activities and intracellular distribution of nicotinamide-adenine dinucleotide phosphate-malate dehydrogenase, phosphoenolpyruvate carboxykinase and pyruvate carboxylase in rat, guinea-pig and rabbit tissues. Biochem J. 1975 Feb;146(2):329–332. doi: 10.1042/bj1460329. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Scrutton M. C., White M. D. Pyruvate carboxylase. Inhibition of the mammalian and avian liver enzymes by alpha-ketoglutarate and L-glutamate. J Biol Chem. 1974 Sep 10;249(17):5405–5415. [PubMed] [Google Scholar]
  39. Seufert D., Herlemann E. M., Albrecht E., Seubert W. On the mechanism of gluconeogenesis and its regulation. VII. Purification and properties of pyruvate carboxylase from rat liver. Hoppe Seylers Z Physiol Chem. 1971 Mar;352(3):459–478. doi: 10.1515/bchm2.1971.352.1.459. [DOI] [PubMed] [Google Scholar]
  40. Siess E. A., Brocks D. G., Wieland O. H. A sensitive and simple method for the study of oxaloacetate compartmentation in isolated hepatocytes. FEBS Lett. 1976 Nov;70(1):51–55. doi: 10.1016/0014-5793(76)80724-7. [DOI] [PubMed] [Google Scholar]
  41. Siess E. A., Brocks D. G., Wieland O. H. Subcellular distribution of key metabolites in isolated liver cells from fasted rats. FEBS Lett. 1976 Oct 15;69(1):265–271. doi: 10.1016/0014-5793(76)80701-6. [DOI] [PubMed] [Google Scholar]
  42. Siess E. A., Wieland O. H. Improved response of isolated liver cells to glucagon in the presence of rat serum. Biochem Biophys Res Commun. 1975 May 5;64(1):323–330. doi: 10.1016/0006-291x(75)90256-9. [DOI] [PubMed] [Google Scholar]
  43. Siess E. A., Wieland O. H. Phosphorylation state of cytosolic and mitochondrial adenine nucleotides and of pyruvate dehydrogenase in isolated rat liver cells. Biochem J. 1976 Apr 15;156(1):91–102. doi: 10.1042/bj1560091. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Struck E., Ashmore J., Wieland O. Effects of glucagon and long chain fatty acids on glucose production by isolated perfused rat liver. Adv Enzyme Regul. 1966;4:219–224. doi: 10.1016/0065-2571(66)90016-1. [DOI] [PubMed] [Google Scholar]
  45. Struck E., Ashmore J., Wieland O. Stimulierung der Gluconeogenese durch langkettige Fettsäuren und Glucagon. Biochem Z. 1965 Nov 5;343(1):107–110. [PubMed] [Google Scholar]
  46. Stucki J. W., Brawand F., Walter P. Regulation of pyruvate metabolim in rat-liver mitochondria by adenine nucleotides and fatty acids. Eur J Biochem. 1972 May;27(1):181–191. doi: 10.1111/j.1432-1033.1972.tb01824.x. [DOI] [PubMed] [Google Scholar]
  47. Stucki J. W., Walter P. Pyruvate metabolism in mitochondria from rat liver. Measured and computer-simulated fluxes. Eur J Biochem. 1972 Oct 17;30(1):60–72. doi: 10.1111/j.1432-1033.1972.tb02072.x. [DOI] [PubMed] [Google Scholar]
  48. Söling H. D., Willms B., Friedrichs D., Kleineke J. Regulation of gluconeogenesis by fatty acid oxidation in isolated perfused livers of non-starved rats. Eur J Biochem. 1968 Apr;4(3):364–372. doi: 10.1111/j.1432-1033.1968.tb00220.x. [DOI] [PubMed] [Google Scholar]
  49. Söling H. D., Willms B., Kleineke J., Gehlhoff M. Regulation of gluconeogenesis in the guinea pig liver. Eur J Biochem. 1970 Oct;16(2):289–302. doi: 10.1111/j.1432-1033.1970.tb01084.x. [DOI] [PubMed] [Google Scholar]
  50. Teufel H., Menahan L. A., Shipp J. C., Böning S., Wieland O. Effect of oleic acid on the oxidation and gluconeogenesis from [1-14C]pyruvate in the perfused rat liver. Eur J Biochem. 1967 Sep;2(2):182–186. doi: 10.1111/j.1432-1033.1967.tb00124.x. [DOI] [PubMed] [Google Scholar]
  51. 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]
  52. 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]
  53. 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]
  54. Ui M., Exton J. H., Park C. R. Effects of glucagon on glutamate metabolism in the perfused rat liver. J Biol Chem. 1973 Aug 10;248(15):5350–5359. [PubMed] [Google Scholar]
  55. Usatenko M. S. Hormonal regulation of phosphoenolpyruvate carboxykinase activity in liver and kidney of adult animals and formation of this enzyme in developing rabbit liver. Biochem Med. 1970 Feb;3(4):298–310. doi: 10.1016/0006-2944(70)90030-x. [DOI] [PubMed] [Google Scholar]
  56. Utter M. F., Keech D. B., Scrutton M. C. A possible role for acetyl CoA in the control of gluconeogenesis. Adv Enzyme Regul. 1964;2:49–68. doi: 10.1016/s0065-2571(64)80005-4. [DOI] [PubMed] [Google Scholar]
  57. Veech R. L., Eggleston L. V., Krebs H. A. The redox state of free nicotinamide-adenine dinucleotide phosphate in the cytoplasm of rat liver. Biochem J. 1969 Dec;115(4):609–619. doi: 10.1042/bj1150609a. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Veneziale C. M., Gabrielli F., Lardy H. A. Gluconeogenesis from pyruvate in isolated perfused rat liver. Biochemistry. 1970 Sep 29;9(20):3960–3970. doi: 10.1021/bi00822a014. [DOI] [PubMed] [Google Scholar]
  59. Veneziale C. M. Gluconeogenesis from D-glyceraldehyde and dihydroxyacetone in isolated rat liver. Stimulation by glucagon. Biochemistry. 1972 Aug 15;11(17):3286–3289. doi: 10.1021/bi00767a025. [DOI] [PubMed] [Google Scholar]
  60. Veneziale C. M. Gluconeogenesis in the isolated rat liver. Studies with bicarbonate-14C. Biochemistry. 1971 Jul 6;10(14):2793–2798. doi: 10.1021/bi00790a022. [DOI] [PubMed] [Google Scholar]
  61. Williamson J. R., Browning E. T., Scholz R. Control mechanisms of gluconeogenesis and ketogenesis. I. Effects of oleate on gluconeogenesis in perfused rat liver. J Biol Chem. 1969 Sep 10;244(17):4607–4616. [PubMed] [Google Scholar]
  62. Williamson J. R., Browning E. T., Thurman R. G., Scholz R. Inhibition of glucagon effects in perfused rat liver by (+)decanoylcarnitine. J Biol Chem. 1969 Sep 25;244(18):5055–5064. [PubMed] [Google Scholar]
  63. Williamson J. R., Garcia A., Renold A. E., Cahill G. F., Jr Studies on the perfused rat liver. I. Effects of glucagon and insulin on glucose metabolism. Diabetes. 1966 Mar;15(3):183–187. doi: 10.2337/diab.15.3.183. [DOI] [PubMed] [Google Scholar]
  64. Williamson J. R., Herczeg B., Coles H., Danish R. Studies on the ketogenic effect of glucagon in intact rat liver. Biochem Biophys Res Commun. 1966 Aug 12;24(3):437–442. doi: 10.1016/0006-291x(66)90179-3. [DOI] [PubMed] [Google Scholar]
  65. Williamson J. R., Kreisberg R. A., Felts P. W. Mechanism for the stimulation of gluconeogenesis by fatty acids in perfused rat liver. Proc Natl Acad Sci U S A. 1966 Jul;56(1):247–254. doi: 10.1073/pnas.56.1.247. [DOI] [PMC free article] [PubMed] [Google Scholar]
  66. Williamson J. R., Scholz R., Browning E. T. Control mechanisms of gluconeogenesis and ketogenesis. II. Interactions between fatty acid oxidation and the citric acid cycle in perfused rat liver. J Biol Chem. 1969 Sep 10;244(17):4617–4627. [PubMed] [Google Scholar]
  67. Yamazaki R. K. Glucagon stimulation of mitochondrial respiration. J Biol Chem. 1975 Oct 10;250(19):7924–7930. [PubMed] [Google Scholar]
  68. Yamazaki R. K., Haynes R. C., Jr Dissociation of pyruvate dehydrogenase from the glucagon stimulation of pyruvate carboxylation in rat liver mitochondria. Arch Biochem Biophys. 1975 Feb;166(2):575–583. doi: 10.1016/0003-9861(75)90422-1. [DOI] [PubMed] [Google Scholar]
  69. von Glutz G., Walter P. Regulation of pyruvate carboxylation by acetyl-CoA in rat liver mitochondria. FEBS Lett. 1976 Dec 31;72(2):299–303. doi: 10.1016/0014-5793(76)80991-x. [DOI] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES