Impaired system A amino acid transport mimics the catabolic effects of acid in L6 cells

Background Metabolic acidaemia stimulates protein catabolism in skeletal muscle cells, leading to muscle wasting. As this occurs without decreasing cytosolic pH, the initial signal is unclear. A possible explanation is that extracellular pH acts on solute transporters at the cell surface, inhibiting nutrient influx. Design Influx through glucose and Pi transporters and System A amino acid transporters into L6 skeletal muscle cells was assessed using (3) H-2-deoxyglucose (2-DG), (33) Pi and (14) C-methylaminoisobutyrate (MeAIB), respectively. Protein degradation (PD) was assessed from (14) C efflux from cells prelabelled with (14) C-Phe. Branched-chain amino acids and Phe were assayed by selective fluorimetric assays. Results While acid (pH 7.1) had little immediate effect on 2-DG or (33) Pi influx, exposure to pH 7.1 rapidly inhibited MeAIB influx. To determine whether System A inhibition was sufficient to trigger PD, it was blocked at pH 7.5 by a saturating dose (10 mmol L-1 ) of nonmetabolisable substrate (MeAIB). Like acid, this increased PD and decreased total protein. It also mimicked the decreases in protein synthesis, DNA synthesis, glucose transport and glycolysis, and depletion of branched-chain amino acids and Phe, which are induced in L6 by acid. The onset of inhibition of PD by an extracellular Gln load was retarded at pH 7.1, and stimulation of PD by acid was negligible if PD had already been stimulated by Gln depletion. The stimulatory effect of MeAIB on PD was selectively blunted by an excess of Gln, whereas the inhibitory effect of Gln on PD was blocked by excess MeAIB. Conclusions The similarity of changes in response to MeAIB and acid implies that these share a common intracellular signalling pathway triggered by inhibition of System A. Even though System A is only a minor contributor to total Gln influx in L6 cells, it is suggested that blockade of System A with acid or MeAIB induces a catabolic state by denying Gln access to a key intracellular regulatory site.