Direct observation of the phosphate acceptor and phosphagen pool sizes in vivo.
Earthworms were subjected to environmental anoxia (200 min) and electrical stimulation (5 min, 6 V, 50 Hz). The levels of high-energy phosphate compounds and the intracellular pH were monitored during anoxia, muscular contraction, and recovery by in vivo phosphorus-31 nuclear magnetic resonance (31P-NMR). Several key metabolites were determined by enzymatic analysis and reversed-phase high-performance liquid chromatography of perchlorate extracts. Because lombricine, the phosphagen phosphate acceptor of earthworms, is a phosphodiester, 31P-NMR permitted direct analysis of the extent of phosphorylation of the lombricine pool in vivo. Total lombricine, lombricine phosphate, ATP, and H+ were measured by NMR, and free ADP was calculated from the lombricine kinase equilibrium constant. The ADP concentration was not significantly changed by anoxia, but it rose threefold after stimulation. L-Lactate accumulated on stimulation, whereas multiple end products (L-lactate, alanine, succinate, and glutamine) were formed during environmental anaerobiosis.
Van Waarde
A
A
Department of Biology, University of Leiden, The Netherlands.
van den Thillart
G
G
Verhagen
M
M
Erkelens
C
C
Addink
A
A
Lugtenburg
J
J
D000244:Adenosine Diphosphate / Q000378:metabolism; D000818:Animals; D004558:Electric Stimulation; D004734:Energy Metabolism; D000860:Hypoxia / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D009835:Oligochaeta / Q000378:metabolism*; D010710:Phosphates / Q000378:metabolism*; D010766:Phosphorylation; D012694:Serine / Q000031:analogs & derivatives* / Q000378:metabolism
D010710:Phosphates; D012694:Serine; D000244:Adenosine Diphosphate; C008295:lombricine