Extracellular flux analysis in intact cardiac tissue slices—A novel tool to investigate cardiac substrate metabolism in mouse myocardium

Abstract

Aim: Cardiac pathologies are accompanied by alterations in substrate metabolism, and extracellular flux analysis is a standard tool to investigate metabolic disturbances, especially in immortalized cell lines. However, preparations of primary cells, such as adult cardiomyocytes require enzymatic dissociation and cultivation affecting metabolism. Therefore, we developed a flux analyzer-based method for the assessment of substrate metabolism in intact vibratome-sliced mouse heart tissue. Methods: Oxygen consumption rates were determined using a Seahorse XFe24-analyzer and “islet capture plates.” We demonstrate that tissue slices are suitable for extracellular flux analysis and metabolize both free fatty acids (FFA) and glucose/glutamine. Functional integrity of tissue slices was proven by optical mapping-based assessment of action potentials. In a proof-of-principle approach, the sensitivity of the method was tested by analyzing substrate metabolism in the remote myocardium after myocardial infarction (I/R). Results: Here, I/R increased uncoupled OCR compared with sham animals indicating a stimulated metabolic capacity. This increase was caused by a higher glucose/glutamine metabolism, whereas FFA oxidation was unchanged. Conclusion: In conclusion, we describe a novel method to analyze cardiac substrate metabolism in intact cardiac tissue slices by extracellular flux analysis. The proof-of-principle experiment demonstrated that this approach has a sensitivity allowing the investigation of pathophysiologically relevant disturbances in cardiac substrate metabolism.