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Open AccessArticle

In Vivo Microdialysis of Endogenous and 13C-labeled TCA Metabolites in Rat Brain: Reversible and Persistent Effects of Mitochondrial Inhibition and Transient Cerebral Ischemia

1
VILLUM Center for Bioanalytical Sciences, Department of Biochemistry and Molecular Biology, University of Southern Denmark, Campusvej 55, 5230 Odense M, Denmark
2
Department of Neurosurgery, Odense University Hospital, University of Southern Denmark, Sdr. Boulevard 29, 5000 Odense C, Denmark
3
BRIDGE—Brain ResearchE—Inter-Disciplinary Guided Excellence, Institute of Clinical Research, University of Southern Denmark, Winsløwparken 19, 5000 Odense C, Denmark
4
Institute of Molecular Medicine, University of Southern Denmark, 55, 5230 Odense C, Denmark
*
Author to whom correspondence should be addressed.
These authors contributed equally to this work.
Metabolites 2019, 9(10), 204; https://doi.org/10.3390/metabo9100204
Received: 31 July 2019 / Revised: 24 September 2019 / Accepted: 25 September 2019 / Published: 27 September 2019
(This article belongs to the Special Issue Metabolomics in Neurodegenerative Disease)
Cerebral micro-dialysis allows continuous sampling of extracellular metabolites, including glucose, lactate and pyruvate. Transient ischemic events cause a rapid drop in glucose and a rise in lactate levels. Following such events, the lactate/pyruvate (L/P) ratio may remain elevated for a prolonged period of time. In neurointensive care clinics, this ratio is considered a metabolic marker of ischemia and/or mitochondrial dysfunction. Here we propose a novel, sensitive microdialysis liquid chromatography-mass spectrometry (LC-MS) approach to monitor mitochondrial dysfunction in living brain using perfusion with 13C-labeled succinate and analysis of 13C-labeled tricarboxylic acid cycle (TCA) intermediates. This approach was evaluated in rat brain using malonate-perfusion (10–50 mM) and endothelin-1 (ET-1)-induced transient cerebral ischemia. In the malonate model, the expected changes upon inhibition of succinate dehydrogenase (SDH) were observed, i.e., an increase in endogenous succinate and decreases in fumaric acid and malic acid. The inhibition was further elaborated by incorporation of 13C into specific TCA intermediates from 13C-labeled succinate. In the ET-1 model, increases in non-labeled TCA metabolites (reflecting release of intracellular compounds) and decreases in 13C-labeled TCA metabolites (reflecting inhibition of de novo synthesis) were observed. The analysis of 13C incorporation provides further layers of information to identify metabolic disturbances in experimental models and neuro-intensive care patients. View Full-Text
Keywords: 13C-labeled succinate; cerebral ischemia; energy metabolism; endothelin-1; LC-MS; malonate; micro-dialysis; mitochondrial dysfunction; reperfusion; tricarboxylic acid cycle 13C-labeled succinate; cerebral ischemia; energy metabolism; endothelin-1; LC-MS; malonate; micro-dialysis; mitochondrial dysfunction; reperfusion; tricarboxylic acid cycle
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Havelund, J.F.; Nygaard, K.H.; Nielsen, T.H.; Nordström, C.-H.; Poulsen, F.R.; Færgeman, N.J.; Forsse, A.; Gramsbergen, J.B. In Vivo Microdialysis of Endogenous and 13C-labeled TCA Metabolites in Rat Brain: Reversible and Persistent Effects of Mitochondrial Inhibition and Transient Cerebral Ischemia. Metabolites 2019, 9, 204.

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