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 hyperthermic stresses upon the activity of the System A amino acid transporter in L6 rat muscle cells. Uptake of alpha-methyl-aminoisobutyric acid (Me-AIB), a non-metabolisable System A substrate, was increased by between 50% and 80% when muscle cells were exposed to a maximally effective concentration of insulin (100 nM), sodium arsenite (ARS, 0.5 mM) or a 42 degrees C heat shock (HS). The observed activation in System A in response to all three stimuli was maximal within 20 min and in the case of insulin and ARS primarily involved an increase in the Vmax of System A transport. In contrast, HS induced significant increases in both Vmax and Km of System A transport suggesting that hyperthermic stress may activate System A by a mechanism distinct from that mediating the effects of insulin and ARS. The hormonal stimulation of System A was blocked by the phosphoinositide 3-kinase (PI3k) inhibitor, wortmannin, but not by rapamycin or PD 98059 which respectively inhibit the mTOR and classical MAP kinase pathways. Exposure of L6 cells to ARS, but not HS, caused a 4.7-fold stimulation in MAPKAP-K2 activity that was blocked by SB 203580, a specific inhibitor of the stress activated protein kinase SAPK2/p38. However, neither SB 203580, rapamycin nor wortmannin were able 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 through changes in the transport kinetics of the System A carrier. Our data show that whilst the hormonal response is PI3k dependent, the signalling mechanisms which instigate changes in System A activity in response to chemical or hyperthermic stress do not appear to involve PI3k or components of the mTOR, p42/p44 MAP kinase or SAPK2/p38 signalling pathways.
McDowell
H E
HE
Department of Anatomy and Physiology, University of Dundee, UK.
Eyers
P A
PA
Hundal
H S
HS
D000596:Amino Acids / Q000378:metabolism*; D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D018053:Arsenites / Q000494:pharmacology*; D001692:Biological Transport / Q000187:drug effects; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D002460:Cell Line; D003513:Cycloheximide / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005419:Flavonoids / Q000494:pharmacology; D006358:Hot Temperature; D007328:Insulin / Q000494:pharmacology* / Q000502:physiology; D007700:Kinetics; D018482:Muscle, Skeletal / Q000187:drug effects / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D020123:Sirolimus / Q000494:pharmacology; D017670:Sodium Compounds / Q000494:pharmacology*; D000077191:Wortmannin
D000596:Amino Acids; D000730:Androstadienes; D018053:Arsenites; D004791:Enzyme Inhibitors; D005419:Flavonoids; D007328:Insulin; D000081082:Phosphoinositide-3 Kinase Inhibitors; D017670:Sodium Compounds; C017947:sodium arsenite; D003513:Cycloheximide; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one; D020123:Sirolimus; D000077191:Wortmannin