Skip to main content
NCBI home page
The Journal of General Physiology logoLink to The Journal of General Physiology
. 1977 Jun 1;69(6):681–704. doi: 10.1085/jgp.69.6.681

A reassessment of decreased amino acid accumulation by ehrlich ascites tumor cells in the presence of metabolic inhibitors

JA Schafer, BE Richey, AE Williams
PMCID: PMC2215337  PMID: 561160

Abstract

This study was undertaken to examine the mechanism by which metabolic inhibition reduces amino acid active transport in ehrlich ascites tumor cells. At 37 degrees C the metabolic inhibitor combination 0.1 mM 2,4-dinitrophenol (DNP) + 10 mM 2- deoxy-D-glucose (DOG) reduced the cell ATP concentration to 0.10- 0.15 mM in less than 5 min. This inhibition was associated with a 20.6 percent +/- 6.4 percent (SD) decrease in the initial influx of α-aminoisobutyric acid (AIB), and a two- to fourfold increase in the unidirectional efflux. These effects could be dissociated from changes in cell Na(+) or K(+) concentrations. Cells incubated to the steady state in 1.0-1.5 mM AIB showed an increased steady-state flux in the presence of DNP + DOG. Steady- state fluxes were consistent with trans-inhibition of AIB influx and trans-stimulation of efflux in control cells, but trans- stimulation of both fluxes in inhibited cells. In spite of the reduction of the cell ATP concentration to less than 0.15 mM and greatly reduced transmembrane concentration gradients of Na(+) and K(+), cells incubated to the steady state in the presence of the inhibitors still established an AIB distribution ration 13.8 +/- 2.6. The results are interpreted to indicate that a component of the reduction of AIB transport produced by metabolic inhibition is attributable to other actions in addition to the reduction of cation concentration gradients. Reduction of cell ATP alone is not responsible for the effects of metabolic inhibition, and both the transmembrane voltage and direct coupling to substrate oxidation via plasma-membrane-bound enzymes must be considered as possible energy sources for amino acid active transport.

Full Text

The Full Text of this article is available as a PDF (1.8 MB).

Selected References

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

  1. BITTNER J., HEINZ E. DIE WIRKUNG VON G-STROPHANTIN AUF DEN GLYZINTRANSPORT IN EHRLICH-ASCITES-TUMORZELLEN. Biochim Biophys Acta. 1963 Aug 13;74:392–400. doi: 10.1016/0006-3002(63)91383-0. [DOI] [PubMed] [Google Scholar]
  2. CHRISTENSEN H. N., RIGGS T. R. Concentrative uptake of amino acids by the Ehrlich mouse ascites carcinoma cell. J Biol Chem. 1952 Jan;194(1):57–68. [PubMed] [Google Scholar]
  3. Chez R. A., Palmer R. R., Schultz S. G., Curran P. F. Effect of inhibitors on alanine transport in isolated rabbit ileum. J Gen Physiol. 1967 Nov;50(10):2357–2375. doi: 10.1085/jgp.50.10.2357. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Christensen H. N., Handlogten M. E. Modes of mediated exodus of amino acids from the Ehrlich ascites tumor cell. J Biol Chem. 1968 Oct 25;243(20):5428–5438. [PubMed] [Google Scholar]
  5. Christensen H. N. On the meaning of effects of substrate structure on biological transport. J Bioenerg. 1973 Jan;4(1):31–61. doi: 10.1007/BF01516050. [DOI] [PubMed] [Google Scholar]
  6. Christensen H. N. Towards a sharper definition of energetic coupling through integration of membrane transport into bioenergetics. J Theor Biol. 1976 Apr;57(2):419–431. doi: 10.1016/0022-5193(76)90013-8. [DOI] [PubMed] [Google Scholar]
  7. Christensen H. N., de Cespedes C., Handlogten M. E., Ronquist G. Energization of amino acid transport, studied for the Ehrlich ascites tumor cell. Biochim Biophys Acta. 1973 Dec 28;300(4):487–522. doi: 10.1016/0304-4157(73)90017-8. [DOI] [PubMed] [Google Scholar]
  8. De Cespedes C., Christensen H. N. Complexity in valinomycin effects on amino acid transport. Biochim Biophys Acta. 1974 Feb 26;339(1):139–145. doi: 10.1016/0005-2736(74)90339-3. [DOI] [PubMed] [Google Scholar]
  9. Eddy A. A., Mulcahy M. F., Thomson P. J. The effects of sodium ions and potassium ions on glycine uptake by mouse ascites-tumour cells in the presence and absence of selected metabolic inhibitors. Biochem J. 1967 Jun;103(3):863–876. doi: 10.1042/bj1030863. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Eddy A. A. The effects of varying the cellular and extracellular concentrations of sodium and potassium ions on the uptake of glycine by mouse ascites-tumour cells in the presence and absence of sodium cyanide. Biochem J. 1968 Jul;108(3):489–498. doi: 10.1042/bj1080489. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Geck P., Heinz E., Pfeiffer B. Evidence against direct coupling between amino acid transport and ATP hydrolysis. Biochim Biophys Acta. 1974 Mar 29;339(3):419–425. doi: 10.1016/0005-2736(74)90169-2. [DOI] [PubMed] [Google Scholar]
  12. Glick N., Gillespie E., Scholefield P. G. Oxidation of glucose and other substrates by Ehrlich ascites tumor cells. Can J Biochem. 1967 Sep;45(9):1401–1412. doi: 10.1139/o67-165. [DOI] [PubMed] [Google Scholar]
  13. HEINZ E., MARIANI H. A. Concentration work and energy dissipation in active transport of glycine into carcinoma cells. J Biol Chem. 1957 Sep;228(1):97–111. [PubMed] [Google Scholar]
  14. HEINZ E. The exchangeability of glycine accumulated by carcinoma cells. J Biol Chem. 1957 Mar;225(1):305–315. [PubMed] [Google Scholar]
  15. HEINZ E., WALSH P. M. Exchange diffusion, transport, and intracellular level of amino acids in Ehrlich carcinoma cells. J Biol Chem. 1958 Dec;233(6):1488–1493. [PubMed] [Google Scholar]
  16. Heinz E. Energetische Probleme beim aktiven Transport. Biophysik. 1973 Jul 27;9(4):291–298. doi: 10.1007/BF01189845. [DOI] [PubMed] [Google Scholar]
  17. Heinz E., Geck P., Pietrzyk C. Driving forces of amino acid transport in animal cells. Ann N Y Acad Sci. 1975 Dec 30;264:428–441. doi: 10.1111/j.1749-6632.1975.tb31501.x. [DOI] [PubMed] [Google Scholar]
  18. Heinz E., Geck P. The efficiency of energetic couping between Na+ flow and amino acid transport in Ehrlich cells-a revised assessment. Biochim Biophys Acta. 1974 Mar 29;339(3):426–431. doi: 10.1016/0005-2736(74)90170-9. [DOI] [PubMed] [Google Scholar]
  19. JOHNSTONE R. M., SCHOLEFIELD P. G. THE NEED FOR IONS DURING TRANSPORT AND EXCHANGE DIFFUSION OF AMINO ACIDS INTO EHRLICH ASCITES CARCINOMA CELLS. Biochim Biophys Acta. 1965 Jan 25;94:130–135. doi: 10.1016/0926-6585(65)90016-6. [DOI] [PubMed] [Google Scholar]
  20. JOHNSTONE R. M., SCHOLEFIELD P. G. The influence of amino acids and antimetabolities on glycine retention by Ehrlich ascites carcinoma cells. Cancer Res. 1959 Dec;19:1140–1149. [PubMed] [Google Scholar]
  21. Jacquez J. A. One-way fluxes of alpha-aminoisobutyric acid in Ehrlich ascites tumor cells. Trans effects and effects of sodium and potassium. J Gen Physiol. 1975 Jan;65(1):57–83. doi: 10.1085/jgp.65.1.57. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Jacquez J. A., Schafer J. A. Na+ and K+ electrochemical potential gradients and the transport of alpha-aminoisobutyric acid in Ehrlich ascites tumor cells. Biochim Biophys Acta. 1969;193(2):368–383. doi: 10.1016/0005-2736(69)90197-7. [DOI] [PubMed] [Google Scholar]
  23. Johnstone R. M. Role of ATP on the initial rate of amino acid uptake in Ehrlich ascites cells. Biochim Biophys Acta. 1974 Aug 9;356(3):319–330. doi: 10.1016/0005-2736(74)90272-7. [DOI] [PubMed] [Google Scholar]
  24. KALTENBACH J. P., KALTENBACH M. H., LYONS W. B. Nigrosin as a dye for differentiating live and dead ascites cells. Exp Cell Res. 1958 Aug;15(1):112–117. doi: 10.1016/0014-4827(58)90067-3. [DOI] [PubMed] [Google Scholar]
  25. Laris P. C., Pershadsingh H. A., Johnstone R. M. Monitoring membrane potentials in Ehrlich ascites tumor cells by means of a fluorescent dye. Biochim Biophys Acta. 1976 Jun 17;436(2):475–488. doi: 10.1016/0005-2736(76)90209-1. [DOI] [PubMed] [Google Scholar]
  26. Lin K. T., Johnstone R. M. Active transport of glycine by mouse pancreas. Evidence against the Na + gradient hypothesis. Biochim Biophys Acta. 1971 Oct 12;249(1):144–158. doi: 10.1016/0005-2736(71)90091-5. [DOI] [PubMed] [Google Scholar]
  27. McClellan W. M., Schafer J. A. Transport of the amino acid analog, 2-aminobicyclo(2,2,1)-heptane-2-carboxylic acid, by Ehrlich ascites tumor cells. Biochim Biophys Acta. 1973 Jul 6;311(3):462–475. doi: 10.1016/0005-2736(73)90326-x. [DOI] [PubMed] [Google Scholar]
  28. Pietrzyk C., Heinz E. The sequestration of Na+, K+ and Cl- in the cellular nucleus and its energetic consequences for the gradient hypothesis of amino acid transport in Ehrlich cells. Biochim Biophys Acta. 1974 Jun 29;352(3):397–411. doi: 10.1016/0005-2736(74)90231-4. [DOI] [PubMed] [Google Scholar]
  29. Poole D. T., Butler T. C., Williams M. E. The effects of nigericin, valinomycin, and 2,4-dinitrophenol on intracellular pH, glycolysis, and K + concentration of Ehrlich ascites tumor cells. Biochim Biophys Acta. 1972 May 9;266(2):463–470. doi: 10.1016/0005-2736(72)90102-2. [DOI] [PubMed] [Google Scholar]
  30. RIGGS T. R., WALKER L. M., CHRISTENSEN H. N. Potassium migration and amino acid transport. J Biol Chem. 1958 Dec;233(6):1479–1484. [PubMed] [Google Scholar]
  31. Reid M., Gibb L. E., Eddy A. A. Ionophore-mediated coupling between ion fluxes and amino acid absorption in mouse ascites-tumour cells. Restoration of the physiological gradients of methionine by valinomycin in the absence of adenosine triphosphate. Biochem J. 1974 Jun;140(3):383–393. doi: 10.1042/bj1400383. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Schafer J. A., Heinz E. The effect of reversal on Na + and K + electrochemical potential gradients on the active transport of amino acids in Ehrlich ascites tumor cells. Biochim Biophys Acta. 1971 Oct 12;249(1):15–33. doi: 10.1016/0005-2736(71)90079-4. [DOI] [PubMed] [Google Scholar]
  33. Schultz S. G., Curran P. F. Coupled transport of sodium and organic solutes. Physiol Rev. 1970 Oct;50(4):637–718. doi: 10.1152/physrev.1970.50.4.637. [DOI] [PubMed] [Google Scholar]
  34. 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]

Articles from The Journal of General Physiology are provided here courtesy of The Rockefeller University Press

RESOURCES