Hypoxia and AMP independently regulate AMP-activated protein kinase activity in heart.
The hypothesis was tested that hypoxia increases AMP-activated protein kinase (AMPK) activity independently of AMP concentration ([AMP]) in heart. In isolated perfused rat hearts, cytosolic [AMP] was changed from 0.2 to 16 microM using metabolic inhibitors during both normal oxygenation (95% O2-5% CO2, normoxia) and limited oxygenation (95% N2-5% CO2, hypoxia). Total AMPK activity measured in vitro ranged from 2 to 40 pmol.min(-1).mg protein(-1) in normoxic hearts and from 5 to 55 pmol.min(-1).mg protein(-1) in hypoxic hearts. The dependence of the in vitro total AMPK activity on the in vivo cytosolic [AMP] was determined by fitting the measurements from individual hearts to a hyperbolic equation. The [AMP] resulting in half-maximal total AMPK activity (A0.5) was 3 +/- 1 microM for hypoxic hearts and 28 +/- 13 microM for normoxic hearts. The A0.5 for alpha2-isoform AMPK activity was 2 +/- 1 microM for hypoxic hearts and 13 +/- 8 microM for normoxic hearts. Total AMPK activity correlated with the phosphorylation of the Thr172 residue of the AMPK alpha-subunit. In potassium-arrested hearts perfused with variable O2 content, alpha-subunit Thr172 phosphorylation increased at O2 < or = 21% even though [AMP] was <0.3 microM. Thus hypoxia or O2 < or = 21% increased AMPK phosphorylation and activity independently of cytosolic [AMP]. The hypoxic increase in AMPK activity may result from either direct phosphorylation of Thr172 by an upstream kinase or reduction in the A0.5 for [AMP].
Frederich
Markus
M
Nuclear Magnetic Resonance Laboratory for Physiological Chemistry, Division of Cardiovascular Medicine, Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, USA.
Zhang
Li
L
Balschi
James A
JA
D055372:AMP-Activated Protein Kinases; D000103:Acetyl-CoA Carboxylase / Q000378:metabolism; D000249:Adenosine Monophosphate / Q000378:metabolism*; D000818:Animals; D003600:Cytosol / Q000378:metabolism; D004789:Enzyme Activation / Q000502:physiology; D000860:Hypoxia / Q000378:metabolism*; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D008297:Male; D009097:Multienzyme Complexes / Q000378:metabolism*; D009206:Myocardium / Q000201:enzymology*; D010759:Phosphorus Isotopes; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012694:Serine / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D013912:Threonine / Q000378:metabolism
D007527:Isoenzymes; D009097:Multienzyme Complexes; D010759:Phosphorus Isotopes; D013912:Threonine; D000249:Adenosine Monophosphate; D012694:Serine; D017346:Protein Serine-Threonine Kinases; D055372:AMP-Activated Protein Kinases; D000103:Acetyl-CoA Carboxylase