4-acetamido-4--isothiocyanatostilbene-2-2--disulfonic-acid and 2-2-dimethyl-beta-alanine

1. In order to investigate the relationship between cellular hydration state and the rate of glutamine transport, tracer glutamine uptake into primary rat myotubes was studied at external osmolalities of 170, 320 or 430 mosmol kg-1. 2. Incubation of myotubes with glutamine (2 mM; 30 min) at 320 mosmol kg-1 increased cell volume and glutamine transport (by 35 and 36%, respectively); insulin (66 nM; 30 min) also increased cell volume and glutamine transport (by 22 and 40%, respectively) and the effects of insulin and glutamine combined were additive. The increase in glutamine uptake following glutamine pre-incubation represented an increase in Vmax of Na(+)-dependent glutamine transport. 3. There was an inverse relationship between myotube glutamine transport and external osmolality after 30 min exposure. 4. During hyposmotic (170 mosmol kg-1) exposure there were large, rapid increases of cell volume and glutamine transport; the latter increased transiently (during the cell swelling phase) by a maximum of approximately 80% at 2 min, (due to an increased Vmax for Na(+)-dependent glutamine transport) then decayed to a new elevated steady state after 30 min exposure. 5. During hyperosmotic (430 mosmol kg-1) exposure there were rapid decreases in glutamine transport and myotube cell volume (both by approximately 30%) to values which were maintained for at least 15 min. 6. The volume-sensitive glutamine transport process features characteristics of the insulin-sensitive system Nm transporter. 7. Modulation of Na(+)-dependent glutamine transport by insulin and cell volume changes may contribute towards regulation of muscle metabolism.

Topics: 4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; Amino Acid Transport Systems, Basic; Animals; beta-Alanine; Carrier Proteins; Cell Size; Cells, Cultured; Glucose; Glutamine; Insulin; Muscle, Skeletal; Nitrobenzenes; Osmolar Concentration; Rats; Water

Studies were carried out in human lung fibroblasts (IMR-90) to investigate 1) the relative contribution of two extracellular pools, inorganic sulfate and sulfur-containing amino acids, to the intracellular fraction precipitable by trichloroacetic acid and 2) the possibility that the transport of these sulfur-containing substrates at the plasma membrane may be a limiting step for macromolecular sulfation. Our studies indicate that the ability to use SO4(2-) released by intracellular catabolism of the sulfur-containing amino acid L-cysteine differs from one cell system to another. In contrast to smooth muscle cells, in the human lung fibroblast, L-cysteine contributes significantly to the intercellular pool of SO4(2-) used for sulfation at extracellular [SO4(2-)] less than 100 microM. However, under physiological conditions with respect to SO4(2-) ([SO4(2-)]0 = 300 microM), L-cysteine does not contribute greater than 30% of the sulfate incorporated into the cellular fraction. Taurine (2-aminoethanesulfonic acid) inhibits SO4(2-) incorporation into the cell-associated macromolecular fraction. However, results suggest that the effect is not due to either SO4(2-) released by its catabolism or to an effect on SO4(2-) transport into the cell. The fact that the transport inhibitor 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid inhibits sulfate incorporation indicates that carrier-mediated sulfate transport at the cellular plasma membrane may be a limiting step for sulfate incorporation. In conclusion, under physiological conditions with respect to SO4(2-), inorganic sulfate is a major source of sulfate for sulfation in human lung fibroblasts and macromolecular sulfation may be limited by its transport into the cells.

Topics: 4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; beta-Alanine; Biological Transport; Cell Line; Cysteine; Fibroblasts; Glucosamine; Glycosaminoglycans; Humans; Kinetics; Lung; Sulfates; Taurine