Regulation of glutamine synthetase and glutaminase activities in cultured skeletal muscle cells

Abstract

Glutamine is synthesized in skeletal muscle, released to the circulation, and transported to other tissues, where it may provide important substrate for gluconeogenesis, ammoniagenesis, and energy-yielding pathways. With the ultimate goal of delineating the factors that control glutamine production and release by skeletal muscle, we have studied the regulation of two key enzymes, glutamine synthetase and glutaminase, in the L6 line of rat skeletal muscle cells grown in monolayer culture. The cultured myotubes were found to have glutamine synthetase and phosphate-dependent glutaminase activities. Glutamine synthetase activity was increased following incubation (1) in glutamine-free medium (threefold); (2) in medium containing high glutamic acid concentrations (fourfold); and (3) in medium supplemented with dexamethasone (threefold). In each case the increase in glutamine synthetase activity required several hours to reach a maximum and was prevented by cycloheximide, suggesting that the change occurred through increased enzyme biosynthesis. No substances tested were found to affect glutaminase activity. We conclude that glutamine synthetase in cultured skeletal muscle is responsive to substrate, product, and hormonal regulation.

Glutamine, insulin and glucocorticoids regulate glutamine synthetase expression in C2C12 myotubes, Hep G2 hepatoma cells and 3T3 L1 adipocytes

“…DeMars [24] was first to demonstrate the effect in HeLa cells and others have shown it in chick embryo retinal cells, V79 and L2 lung cells, hepatomas, L cells, astrocytes, IEC intestinal cells, and L6 muscle cells [25][26][27][28][29][30][31][32][33][34][35][36][37][38][39]. Where examined, the effect has usually been found to be independent of changes in mRNA abundance or enzyme protein synthesis, and the primary mechanism involves an acceleration of glutamine synthetase protein degradation in the presence of glutamine.…”

“…In addition to hormonal regulation, it has been known since the 1950s that glutamine synthetase activity in a variety of cell lines in culture is subject to down-regulation in the presence of glutamine [24][25][26][27][28][29][30][31][32][33][34][35][36][37][38][39]. The principal mechanism involved has been established as glutamine causing a rapid acceleration of the degradation of the glutamine synthetase protein with little change in transcription or translation in most cells.…”

L6 cells as a tissue culture model for thyroid hormone effects on skeletal muscle metabolism.

“…The L6 cells used in the present studies were provided by Dr. Peter Nissley (Metabolism Branch, National Cancer Institute, Bethesda, MD), and were grown in 100-mm petri dishes as described previously (5). Cells were harvested on the 15th or 16th d after plating, at least 6 d after treatment with cytosine arabinoside, at which time they consisted of fully differentiated myotubes (5).…”

Augmentation of the effects of insulin and insulin-like growth factors I and II on glucose uptake in cultured rat skeletal muscle cells by sulfonylureas

“…The data represent mean + SD from four experiments in duplicate post-mitotic, multinucleated myotubes with many characteristics of skeletal muscle [23,28]. By treating cultures with cytosine arabinoside at the time of fusion, it is possible to selectively eliminate residual myoblasts and obtain a homogeneous population of differentiated skeletal muscle myotubes [29]. Insulin stimulates glucose uptake in differentiated L6 skeletal muscle cells, with a maximal 40% increase above basal under these conditions at an insulin concentration of 10 .7 mol/1.…”