Skip to main content
The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1987 Nov;80(5):1303–1310. doi: 10.1172/JCI113206

Determination of Krebs cycle metabolic carbon exchange in vivo and its use to estimate the individual contributions of gluconeogenesis and glycogenolysis to overall glucose output in man.

A Consoli 1, F Kennedy 1, J Miles 1, J Gerich 1
PMCID: PMC442384  PMID: 3680498

Abstract

Current isotopic approaches underestimate gluconeogenesis in vivo because of Krebs cycle carbon exchange and the inability to measure intramitochondrial precursor specific activity. We therefore applied a new isotopic approach that theoretically overcomes these limitations and permits quantification of Krebs cycle carbon exchange and the individual contributions of gluconeogenesis and glycogenolysis to overall glucose output. [6-3H]Glucose was infused to measure overall glucose output; [2-14C]acetate was infused to trace phosphoenolpyruvate gluconeogenesis and to calculate Krebs cycle carbon exchange as proposed by Katz. Plasma [14C]3-OH-butyrate specific activity was used to estimate intramitochondrial acetyl coenzyme A (CoA) specific activity, and finally the ratio between plasma glucose 14C-specific activity and the calculated intracellular phosphoenolpyruvate 14C-specific activity was used to determine the relative contributions of gluconeogenesis and glycogenolysis to overall glucose output. Using this approach, acetyl CoA was found to enter the Krebs cycle at twice (postabsorptive subjects) and three times (2 1/2-d fasted subjects) the rate of pyruvate, respectively. Gluconeogenesis in postabsorptive subjects (3.36 +/- 0.20 mumol/kg per min) accounted for 28 +/- 2% of overall glucose output and increased twofold in subjects fasted for 2 1/2-d (P less than 0.01), accounting for greater than 97% of overall glucose output. Glycogenolysis in postabsorptive subjects averaged 8.96 +/- 0.40 mumol/kg per min and decreased to 0.34 +/- 0.08 mumol/kg per min (P less than 0.01) after a 2 1/2-d fast. Since these results agree well with previously reported values for gluconeogenesis and glycogenolysis based on determinations of splanchnic substrate balance and glycogen content of serial liver biopsies, we conclude that the isotopic approach applied herein provides an accurate, noninvasive measurement of gluconeogenesis and glycogenolysis in vivo.

Full text

PDF
1303

Selected References

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

  1. Bortz W. M., Paul P., Haff A. C., Holmes W. L. Glycerol turnover and oxidation in man. J Clin Invest. 1972 Jun;51(6):1537–1546. doi: 10.1172/JCI106950. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Brady P. S., Scofield R. F., Ohgaku S., Schumann W. C., Bartsch G. E., Margolis J. M., Kumaran K., Horvat A., Mann S., Landau B. R. Pathways of acetoacetate's formation in liver and kidney. J Biol Chem. 1982 Aug 25;257(16):9290–9293. [PubMed] [Google Scholar]
  3. Dietze G., Wicklmayr M., Hepp K. D., Bogner W., Mehnert H., Czempiel H., Henftling H. G. On gluconeogenesis of human liver. Accelerated hepatic glucose formation induced by increased precursor supply. Diabetologia. 1976 Dec;12(6):555–561. doi: 10.1007/BF01220631. [DOI] [PubMed] [Google Scholar]
  4. Garber A. J., Cryer P. E., Santiago J. V., Haymond M. W., Pagliara A. S., Kipnis D. M. The role of adrenergic mechanisms in the substrate and hormonal response to insulin-induced hypoglycemia in man. J Clin Invest. 1976 Jul;58(1):7–15. doi: 10.1172/JCI108460. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Gauthier C., Vranic M., Hetenyi G., Jr Nonhypoglycemic glucoregulation: role of glycerol and glucoregulatory hormones. Am J Physiol. 1983 Apr;244(4):E373–E379. doi: 10.1152/ajpendo.1983.244.4.E373. [DOI] [PubMed] [Google Scholar]
  6. Grunnet N., Katz J. Effects of ammonia and norvaline on lactate metabolism by hepatocytes from starved rats. The use of 14C-labelled lactate in studies of hepatic gluconeogenesis. Biochem J. 1978 Jun 15;172(3):595–603. doi: 10.1042/bj1720595. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hetenyi G., Jr Correction factor for the estimation of plasma glucose synthesis from the transfer of 14C-atoms from labelled substrate in vivo: A preliminary report. Can J Physiol Pharmacol. 1979 Jul;57(7):767–770. doi: 10.1139/y79-119. [DOI] [PubMed] [Google Scholar]
  8. Hetenyi G., Jr, Ferrarotto C. Correction for metabolic exchange in the calculation of the rate of gluconeogenesis in rats. Biochem Med. 1983 Jun;29(3):372–378. doi: 10.1016/0006-2944(83)90073-x. [DOI] [PubMed] [Google Scholar]
  9. Hetenyi G., Jr, Lussier B., Ferrarotto C., Radziuk J. Calculation of the rate of gluconeogenesis from the incorporation of 14C atoms from labelled bicarbonate or acetate. Can J Physiol Pharmacol. 1982 Dec;60(12):1603–1609. doi: 10.1139/y82-237. [DOI] [PubMed] [Google Scholar]
  10. Katz J. Determination of gluconeogenesis in vivo with 14C-labeled substrates. Am J Physiol. 1985 Apr;248(4 Pt 2):R391–R399. doi: 10.1152/ajpregu.1985.248.4.R391. [DOI] [PubMed] [Google Scholar]
  11. Katz L. D., Glickman M. G., Rapoport S., Ferrannini E., DeFronzo R. A. Splanchnic and peripheral disposal of oral glucose in man. Diabetes. 1983 Jul;32(7):675–679. doi: 10.2337/diab.32.7.675. [DOI] [PubMed] [Google Scholar]
  12. Kelleher J. K. Gluconeogenesis from labeled carbon: estimating isotope dilution. Am J Physiol. 1986 Mar;250(3 Pt 1):E296–E305. doi: 10.1152/ajpendo.1986.250.3.E296. [DOI] [PubMed] [Google Scholar]
  13. Kosugi K., Scofield R. F., Chandramouli V., Kumaran K., Schumann W. C., Landau B. R. Pathways of acetone's metabolism in the rat. J Biol Chem. 1986 Mar 25;261(9):3952–3957. [PubMed] [Google Scholar]
  14. Krebs H. A., Hems R., Weidemann M. J., Speake R. N. The fate of isotopic carbon in kidney cortex synthesizing glucose from lactate. Biochem J. 1966 Oct;101(1):242–249. doi: 10.1042/bj1010242. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kreisberg R. A., Siegal A. M., Owen W. C. Alanine and gluconeogenesis in man: effect of ethanol. J Clin Endocrinol Metab. 1972 May;34(5):876–883. doi: 10.1210/jcem-34-5-876. [DOI] [PubMed] [Google Scholar]
  16. Nilsson L. H., Hultman E. Liver glycogen in man--the effect of total starvation or a carbohydrate-poor diet followed by carbohydrate refeeding. Scand J Clin Lab Invest. 1973 Dec;32(4):325–330. doi: 10.3109/00365517309084355. [DOI] [PubMed] [Google Scholar]
  17. Nilsson L. H., Hultman E. Liver glycogen in man--the effect of total starvation or a carbohydrate-poor diet followed by carbohydrate refeeding. Scand J Clin Lab Invest. 1973 Dec;32(4):325–330. doi: 10.3109/00365517309084355. [DOI] [PubMed] [Google Scholar]
  18. Nurjhan N., Campbell P. J., Kennedy F. P., Miles J. M., Gerich J. E. Insulin dose-response characteristics for suppression of glycerol release and conversion to glucose in humans. Diabetes. 1986 Dec;35(12):1326–1331. doi: 10.2337/diab.35.12.1326. [DOI] [PubMed] [Google Scholar]
  19. Reichard G. A., Jr, Haff A. C., Skutches C. L., Paul P., Holroyde C. P., Owen O. E. Plasma acetone metabolism in the fasting human. J Clin Invest. 1979 Apr;63(4):619–626. doi: 10.1172/JCI109344. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Rizza R. A., Mandarino L. J., Gerich J. E. Dose-response characteristics for effects of insulin on production and utilization of glucose in man. Am J Physiol. 1981 Jun;240(6):E630–E639. doi: 10.1152/ajpendo.1981.240.6.E630. [DOI] [PubMed] [Google Scholar]
  21. Rizza R., Verdonk C., Miles J., Service F. J., Gerich J. Effect of intermittent endogenous hyperglucagonemia on glucose homeostasis in normal and diabetic man. J Clin Invest. 1979 Jun;63(6):1119–1123. doi: 10.1172/JCI109404. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Rognstad R., Katz J. Gluconeogenesis in the kidney cortex. Quantitative estimation of carbon flow. J Biol Chem. 1972 Oct 10;247(19):6047–6054. [PubMed] [Google Scholar]
  23. Rognstad R., Woronsberg J. An enzymic-chemical degradation of glucose. Anal Biochem. 1968 Oct 24;25(1):448–451. doi: 10.1016/0003-2697(68)90121-8. [DOI] [PubMed] [Google Scholar]
  24. Ruderman N. B. Muscle amino acid metabolism and gluconeogenesis. Annu Rev Med. 1975;26:245–258. doi: 10.1146/annurev.me.26.020175.001333. [DOI] [PubMed] [Google Scholar]
  25. STEELE R. Influences of glucose loading and of injected insulin on hepatic glucose output. Ann N Y Acad Sci. 1959 Sep 25;82:420–430. doi: 10.1111/j.1749-6632.1959.tb44923.x. [DOI] [PubMed] [Google Scholar]
  26. STRISOWER E. H., KOHLER G. D., CHAIKOFF I. L. Incorporation of acetate carbon into glucose by liver slices from normal and alloxan-diabetic rats. J Biol Chem. 1952 Sep;198(1):115–126. [PubMed] [Google Scholar]
  27. Skutches C. L., Holroyde C. P., Myers R. N., Paul P., Reichard G. A. Plasma acetate turnover and oxidation. J Clin Invest. 1979 Sep;64(3):708–713. doi: 10.1172/JCI109513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. WEINMAN E. O., STRISOWER E. H., CHAIKOFF I. L. Conversion of fatty acids to carbohydrate; application of isotopes to this problem and role of the Krebs cycle as a synthetic pathway. Physiol Rev. 1957 Apr;37(2):252–272. doi: 10.1152/physrev.1957.37.2.252. [DOI] [PubMed] [Google Scholar]
  29. Wahren J., Felig P., Cerasi E., Luft R. Splanchnic and peripheral glucose and amino acid metabolism in diabetes mellitus. J Clin Invest. 1972 Jul;51(7):1870–1878. doi: 10.1172/JCI106989. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Clinical Investigation are provided here courtesy of American Society for Clinical Investigation

RESOURCES