title: ¹³C magnetic resonance spectroscopy detection of changes in serine isotopomers reflects changes in mitochondrial redox status.
abstract: The glycine cleavage system (GCS), the major pathway of glycine catabolism in liver, is found only in the mitochondria matrix and is regulated by the oxidized nicotinamide adenine dinucleotide (NAD(+) )/reduced nicotinamide adenine dinucleotide (NADH) ratio. In conjunction with serine hydroxymethyltransferase, glycine forms the 1 and 2 positions of serine, while the 3 position is formed exclusively by GCS. Therefore, we sought to exploit this pathway to show that quantitative measurements of serine isotopomers in liver can be used to monitor the NAD(+) /NADH ratio using (13) C NMR spectroscopy. Rat hepatocytes were treated with modulators of GCS activity followed by addition of 2-(13) C-glycine, and the changes in the proportions of newly synthesized serine isotopomers were compared to controls. Cysteamine, a competitive inhibitor of GCS, prevented formation of mitochondrial 3-(13) C-serine and 2,3-(13) C-serine isotopomers while reducing 2-(13) C-serine by 55%, demonstrating that ca. 20% of glycine-derived serine is produced in the cytosol. Glucagon, which activates GCS activity, and the mitochondrial uncoupler carbonyl cyanide-3-chlorophenylhydrazone both increased serine isotopomers, whereas rotenone, an inhibitor of complex I, had the opposite effect. These results demonstrate that (13) C magnetic resonance spectroscopy monitoring of the formation of serine isotopomers in isolated rat hepatocytes given 2-(13) C-glycine reflects the changes of mitochondrial redox status.
authors.affiliation: Department of Biomedical Engineering, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA.
authors.forename: C Bryce
authors.initials: CB
authors.lastname: Johnson
authors.forename: Andrey P
authors.initials: AP
authors.lastname: Tikunov
authors.forename: Haakil
authors.initials: H
authors.lastname: Lee
authors.forename: Justyna E
authors.initials: JE
authors.lastname: Wolak
authors.forename: Peter
authors.initials: P
authors.lastname: Pediaditakis
authors.forename: Doug A
authors.initials: DA
authors.lastname: Romney
authors.forename: Ekhson
authors.initials: E
authors.lastname: Holmuhamedov
authors.forename: Michael P
authors.initials: MP
authors.lastname: Gamcsik
authors.forename: Jeffrey M
authors.initials: JM
authors.lastname: Macdonald
mesh_terms: D000818:Animals; D002247:Carbon Isotopes; D002478:Cells, Cultured; D022781:Hepatocytes / Q000378:metabolism* / Q000648:ultrastructure; D008297:Male; D008930:Mitochondria, Liver / Q000378:metabolism*; D009243:NAD / Q000032:analysis*; D010084:Oxidation-Reduction; D019275:Radiopharmaceuticals; D051381:Rats; D017207:Rats, Sprague-Dawley; D012694:Serine / Q000032:analysis*
chemical_list: D002247:Carbon Isotopes; D019275:Radiopharmaceuticals; D009243:NAD; D012694:Serine