Cardiac performance is limited by oxygen delivery to the mitochondria in the crystalloid-perfused working heart

Abstract

The left ventricular working, crystalloid-perfused heart is used extensively to evaluate basic cardiac function, pathophysiology, and pharmacology. Crystalloid-perfused hearts may be limited by oxygen delivery, as adding oxygen carriers increases myoglobin oxygenation and improves myocardial function. However, whether decreased myoglobin oxygen saturation impacts oxidative phosphorylation (OxPhos) is unresolved, since myoglobin has a much lower affinity for oxygen than cytochrome c oxidase (COX). In the present study, a laboratory-based synthesis of an affordable perfluorocarbon (PFC) emulsion was developed to increase perfusate oxygen carrying capacity without impeding optical absorbance assessments. In left ventricular working hearts, along with conventional measurements of cardiac function and metabolic rate, myoglobin oxygenation and cytochrome redox state were monitored using a novel transmural illumination approach. Hearts were perfused with Krebs-Henseleit (KH) or KH supplemented with PFC, increasing perfusate oxygen carrying capacity by 3.6-fold. In KH-perfused hearts, myoglobin was deoxygenated, consistent with cytoplasmic hypoxia, and the mitochondrial cytochromes, including COX, exhibited a high reduction state, consistent with OxPhos hypoxia. PFC perfusate increased aortic output from 76 ± 6 to 142 ± 4 ml/min and increased oxygen consumption while also increasing myoglobin oxygenation and oxidizing the mitochondrial cytochromes. These results are consistent with limited delivery of oxygen to OxPhos resulting in an adapted lower cardiac performance with KH. Consistent with this, PFCs increased myocardial oxygenation, and cardiac work was higher over a wider range of perfusate Po. In summary, heart mitochondria are limited by oxygen delivery with KH; supplementation of KH with PFC reverses mitochondrial hypoxia and improves cardiac performance, creating a more physiological tissue oxygen delivery. NEW & NOTEWORTHY Optical absorbance spectroscopy of intrinsic chromophores reveals that the commonly used crystalloid-perfused working heart is oxygen limited for oxidative phosphorylation and associated cardiac work. Oxygen-carrying perfluorocarbons increase myocardial oxygen delivery and improve cardiac function, providing a more physiological mitochondrial redox state and emphasizing cardiac work is modulated by myocardial oxygen delivery.

Intra-cardiac Side-Firing Light Catheter for Monitoring Cellular Metabolism using Transmural Absorbance Spectroscopy of Perfused Mammalian Hearts

“…Using the cyanide data as an estimate of total myoglobin oxygenation and the ischemia data for fully deoxygenated myoglobin, we estimate that under control conditions myoglobin was only 88.2% ± 1.0% (n = 10) oxygenated, consistent with prior studies 20,21,25 . .…”

“…The limitations of reflectance absorption spectroscopy presented above were resolved by introducing an optical catheter into the left ventricle cavity, permitting the collection of transmitted light across the left ventricle free wall 20 . The benefits of transmission spectroscopy for this type of study was appreciated in early invasive studies by Tamura et al 9 The current implementation provides a very robust transmission absorption spectroscopy analysis of the intact heart with regards to cytosolic oxygenation and mitochondria redox state under a variety of conditions 21 . These initial studies used a specially fabricated catheter with a powered LED on the tip oriented to generate a side-firing pattern of white light through the myocardium.…”

Artificial Oxygen Carriers—Past, Present, and Future—a Review of the Most Innovative and Clinically Relevant Concepts

“…Albumin-derived perfluorodecalin-filled nanocapsules showed promising results in a first in vivo toxicity study (Wrobeln et al, 2017a) and protected a Langendorff heart (rat) during massive ischemia (Wrobeln et al, 2017b). Similarly, a novel PFC emulsion increased myocardial O 2 delivery, improved cardiac function, and generated a more physiologic redox state in a Langendorff heart (rabbit) compared with perfusion without the PFC emulsion (Kuzmiak-Glancy et al, 2018).…”

Optocardiography and Electrophysiology Studies of Ex Vivo Langendorff-perfused Hearts

“…39,40 . (-/-)Blebbistatin (5 µM circulating concentration) may be added slowly as a bolus dose of 0.5 mM in 5 mL of perfusate (100x of final concentration)41 .Alternatively, BDM may be initially included in the perfusate media at a circulating concentration of 20 mM.1.…”