The Krebs cycle plays a fundamental role in cardiac energy production and is often implicated in the energetic imbalance characteristic of heart disease. In this study, we measured Krebs cycle flux in real time in perfused rat hearts using hyperpolarized magnetic resonance spectroscopy (MRS). [2-(13)C]Pyruvate was hyperpolarized and infused into isolated perfused hearts in both healthy and postischemic metabolic states. We followed the enzymatic conversion of pyruvate to lactate, acetylcarnitine, citrate, and glutamate with 1 s temporal resolution. The appearance of (13)C-labeled glutamate was delayed compared with that of other metabolites, indicating that Krebs cycle flux can be measured directly. The production of (13)C-labeled citrate and glutamate was decreased postischemia, as opposed to lactate, which was significantly elevated. These results showed that the control and fluxes of the Krebs cycle in heart disease can be studied using hyperpolarized [2-(13)C]pyruvate.

SummaryThe citric acid cycle (CAC) metabolite fumarate has been proposed to be cardioprotective; however, its mechanisms of action remain to be determined. To augment cardiac fumarate levels and to assess fumarate's cardioprotective properties, we generated fumarate hydratase (Fh1) cardiac knockout (KO) mice. These fumarate-replete hearts were robustly protected from ischemia-reperfusion injury (I/R). To compensate for the loss of Fh1 activity, KO hearts maintain ATP levels in part by channeling amino acids into the CAC. In addition, by stabilizing the transcriptional regulator Nrf2, Fh1 KO hearts upregulate protective antioxidant response element genes. Supporting the importance of the latter mechanism, clinically relevant doses of dimethylfumarate upregulated Nrf2 and its target genes, hence protecting control hearts, but failed to similarly protect Nrf2-KO hearts in an in vivo model of myocardial infarction. We propose that clinically established fumarate derivatives activate the Nrf2 pathway and are readily testable cytoprotective agents.

PurposeButyrate, a short chain fatty acid, was studied as a novel hyperpolarized substrate for use in dynamic nuclear polarization enhanced magnetic resonance spectroscopy experiments, to define the pathways of short chain fatty acid and ketone body metabolism in real time.MethodsButyrate was polarized via the dynamic nuclear polarization process and subsequently dissolved to generate an injectable metabolic substrate. Metabolism was initially assessed in the isolated perfused rat heart, followed by evaluation in the in vivo rat heart.ResultsHyperpolarized butyrate was generated with a polarization level of 7% and was shown to have a T1 relaxation time of 20 s. These physical characteristics were sufficient to enable assessment of multiple steps in its metabolism, with the ketone body acetoacetate and several tricarboxylic acid cycle intermediates observed both in vitro and in vivo. Metabolite to butyrate ratios of 0.1–0.4% and 0.5–2% were observed in vitro and in vivo respectively, similar to levels previously observed with hyperpolarized [2-13C]pyruvate.ConclusionsIn this study, butyrate has been demonstrated to be a suitable hyperpolarized substrate capable of revealing multi-step metabolism in dynamic nuclear polarization experiments and providing information on the metabolism of fatty acids not currently achievable with other hyperpolarized substrates. Magn Reson Med 71:1663–1669, 2014. © 2013 Wiley Periodicals, Inc.

Functional and structural alterations in the SHR heart are consistent with the hypertrophied phenotype. Our in vivo work indicates a preference for glucose metabolism in the SHR heart, a move away from predominantly fatty acid oxidative metabolism. Interestingly, (13)C label flux into lactate was unchanged, indicating no switch to an anaerobic glycolytic phenotype, but rather an increased reliance on glucose oxidation in the SHR heart.

Background— Fatty acid and glucose transporters translocate between the sarcolemma and intracellular compartments to regulate substrate metabolism acutely. We hypothesised that during ischemia fatty acid translocase (FAT/CD36) would translocate away from the sarcolemma to limit fatty acid uptake when fatty acid oxidation is inhibited. Methods and Results— Wistar rat hearts were perfused during preischemia, low-flow ischemia, and reperfusion, using 3 H-substrates for measurement of metabolic rates, followed by metabolomic analysis and subcellular fractionation. During ischemia, there was a 32% decrease in sarcolemmal FAT/CD36 accompanied by a 95% decrease in fatty acid oxidation rates, with no change in intramyocardial lipids. Concomitantly, the sarcolemmal content of the glucose transporter, GLUT4, increased by 90% during ischemia, associated with an 86% increase in glycolytic rates, 45% decrease in glycogen content, and a 3-fold increase in phosphorylated AMP-activated protein kinase. Following reperfusion, decreased sarcolemmal FAT/CD36 persisted, but fatty acid oxidation rates returned to preischemic levels, resulting in a 35% decrease in myocardial triglyceride content. Elevated sarcolemmal GLUT4 persisted during reperfusion; in contrast, glycolytic rates decreased to 30% of preischemic rates, accompanied by a 5-fold increase in intracellular citrate levels and restoration of glycogen content. Conclusions— During ischemia, FAT/CD36 moved away from the sarcolemma as GLUT4 moved toward the sarcolemma, associated with a shift from fatty acid oxidation to glycolysis, while intramyocardial lipid accumulation was prevented. This relocation was maintained during reperfusion, which was associated with replenishing glycogen stores as a priority, occurring at the expense of glycolysis and mediated by an increase in citrate levels.