Premium
Glutamine and glutamate nitrogen exchangeable pools in cultured fibroblasts: A stable isotope study
Author(s) -
Darmaun Dominique,
Matthews Dwight E.,
Desjeux JehanFrancois,
Bier Dennis M.
Publication year - 1988
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.1041340118
Subject(s) - glutamine , intracellular , extracellular , glutamate receptor , incubation , metabolism , biochemistry , biology , amino acid , receptor
Abstract Glutamine's role as an energetic fuel has been extensively studied in the past using 14 C‐ and 3 H‐labeled tracers in cultured human cells. Yet another prominent role of glutamine, that of a nitrogen shuttle, cannot be approached without an N‐tracer. We therefore used 15 N‐labeled glutamine and glutamate to address the following questions: (1) is it possible to study the exchangeable pools of intracellular free glutamine and glutamate nitrogen with stable isotope methods? and (2) to what extent is intracellular glutamine pool regulated by extracellular glutamine? We observed that: (1) intracellular [ 15 N]‐glutamine enrichment reached a plateau at 80% within 20 min of incubation in a buffer containing 0.7 mM pure 15 N‐glutamine and no glutamate; in contrast, intracellular 15 N‐glutamate enrichment rose only to 40% after 4 hours of incubation in a buffer containing 0.5 mM pure 15 N‐glutamate and no glutamine; (2) the cell‐free glutamine content was tightly dependent on extracellular glutamine level, while the cell‐free glutamate remained steady irrespective of the extracellular glutamate level; (3) the cells took up glutamine and glutamate against a concentration gradient; the rate of glutamine uptake accounted for 90% of the cell glutamine turnover rate; and (4) when cells were confronted with a glutamine‐free medium, only one fourth of intracellular glutamine was derived from the exchangeable glutamate. We conclude that: (1) The size and turnover rate of the intracellular pool of free glutamine nitrogen are measurable using stable isotope methodology; (2) glutamine uptake from the extracellular medium accounts for most of glutamine turnover rate in cultured fibroblasts; and (3) intracellular free glutamate is divided up between several pools in cultured human fibroblasts.