In vitroketone-supported mitochondrial respiration is minimal when other substrates are readily available in cardiac and skeletal muscle

Key points

Ketone bodies are proposed to represent an alternative fuel source driving energy production, particularly during exercise. Biologically, the extent to which mitochondria utilize ketone bodies compared to other substrates remains unknown. We demonstratein vitrothat maximal mitochondrial respiration supported by ketone bodies is low when compared to carbohydrate-derived substrates in the left ventricle and red gastrocnemius muscle from rodents, and in human skeletal muscle. When considering intramuscular concentrations of ketone bodies and the presence of other carbohydrate and lipid substrates, biological rates of mitochondrial respiration supported by ketone bodies are predicted to be minimal. At the mitochondrial level, it is therefore unlikely that ketone bodies are an important source for energy production in cardiac and skeletal muscle, particularly when other substrates are readily available. Ketone bodies (KB) have recently gained popularity as an alternative fuel source to support mitochondrial oxidative phosphorylation and enhance exercise performance. However, given the low activity of ketolytic enzymes and potential inhibition from carbohydrate oxidation, it remains unknown if KBs can contribute to energy production. We therefore determined the ability of KBs (sodiumdl-beta-hydroxybutyrate, beta-HB; lithium acetoacetate, AcAc) to stimulatein vitromitochondrial respiration in the left ventricle (LV) and red gastrocnemius (RG) of rats, and in human vastus lateralis. Compared to pyruvate, the ability of KBs to maximally drive respiration was low in isolated mitochondria and permeabilized fibres (PmFb) from the LV (similar to 30-35% of pyruvate), RG (similar to 10-30%), and human vastus lateralis (similar to 2-10%). In PmFb, the concentration of KBs required to half-maximally drive respiration (LV: 889 mu m beta-HB, 801 mu mAcAc; RG: 782 mu m beta-HB, 267 mu mAcAc) were greater than KB content representative of the muscle microenvironment (similar to 100 mu m). This would predict low rates (similar to 1-4% of pyruvate) of biological KB-supported respiration in the LV (8-14 pmol s(-1) mg(-1)) and RG (3-6 pmol s(-1) mg(-1)) at rest and following exercise. Moreover, KBs did not increase respiration in the presence of saturating pyruvate, submaximal pyruvate (100 mu m) reduced the ability of physiological beta-HB to drive respiration, and addition of other intracellular substrates (succinate + palmitoylcarnitine) decreased maximal KB-supported respiration. As a result, product inhibition is likely to limit KB oxidation. Altogether, the ability of KBs to drive mitochondrial respiration is minimal and they are likely to be outcompeted by other substrates, compromising their use as an important energy source.