Regulation of System A amino acid transport in L6 rat skeletal muscle cells by insulin, chemical and hyperthermic stress

In this study we have investigated the effects of insulin, chemical and hyp erthermic stresses upon the activity of the System A amino acid transporter in L6 rat muscle cells. Uptake of a-methyl-aminoisobutyric acid (Me-AIB), a non-metabolisable System A substrate, was increased by between 50% and 80 % when muscle cells mere exposed to a maximally effective concentration of insulin (100 nM), sodium arsenite (ARS, 0.5 mM) or a 42 degrees C heat shoc k (HS). The observed activation in System A in response to all three stimul i was maximal within 20 min and in the case of insulin and ARS primarily in volved an increase in the V-max of System A transport. In contrast, HS indu ced significant increases in both V-max and K-m of System A transport sugge sting that hyperthermic stress may activate System A by a mechanism distinc t from that mediating the effects of insulin and ARS. The hormonal stimulat ion of System A was blocked by the phosphoinositide 3-kinase (PI3k) inhibit or, wortmannin, but not by rapamycin or PD 98059 which respectively inhibit the mTOR and classical MAP kinase pathways. Exposure of L6 cells to ARS, b ut not HS, caused a 4.7-fold stimulation in MAPKAP-K2 activity that was blo cked by SE 203580, a specific inhibitor of the stress activated protein kin ase SAPK2/p38. However, neither SE 203580, rapamycin nor wortmannin were ab le to suppress the ARS- or HS-induced stimulation in System A transport, In summary, our results demonstrate that activity of the System A transporter can be rapidly upregulated in response to hormonal and stress stimuli thro ugh changes in the transport kinetics of the System A carrier, Our data sho w that whilst the hormonal response is PI3k dependent, the signalling mecha nisms which instigate changes in System A activity in response to chemical or hyperthermic stress do not appear to involve PI3k or components of the m TOR, p42/p44 MAP kinase or SAPK2/p38 signalling pathways. (C) 1998 Federati on of European Biochemical Societies.