FAT/CD36-null mice reveal that mitochondrial FAT/CD36 is required to upregulate mitochondrial fatty acid oxidation in contracting muscle

Abstract

The plasma membrane fatty acid transport protein FAT/CD36 is also present at the mitochondria, where it may contribute to the regulation of fatty acid oxidation, although this has been challenged. Therefore, we have compared enzyme activities and rates of mitochondrial palmitate oxidation in muscles of wild-type (WT) and FAT/CD36 knockout (KO) mice, at rest and after muscle contraction. In WT and KO mice, carnitine palmitoyltransferase-I, citrate synthase, and beta-hydroxyacyl-CoA dehydrogenase activities did not differ in subsarcolemmal (SS) and intermyofibrillar (IMF) mitochondria of WT and FAT/CD36 KO mice. Basal palmitate oxidation rates were lower (P < 0.05) in KO mice (SS -18%; IMF -13%). Muscle contraction increased fatty acid oxidation (+18%) and mitochondrial FAT/CD36 protein (+16%) in WT IMF but not in WT SS, or in either mitochondrial subpopulation in KO mice. This revealed that the difference in IMF mitochondrial fatty acid oxidation between WT and KO mice can be increased approximately 2.5-fold from 13% under basal conditions to 35% during muscle contraction. The FAT/CD36 inhibitor sulfo-N-succinimidyl oleate (SSO), inhibited palmitate transport across the plasma membrane in WT, but not in KO mice. In contrast, SSO bound to mitochondrial membranes and reduced palmitate oxidation rates to a similar extent in both WT and KO mitochondria ( approximately 80%; P < 0.05). In addition, SSO reduced state III respiration with succinate as a substrate, without altering mitochondrial coupling (P/O ratios). Thus, while SSO inhibits FAT/CD36-mediated palmitate transport at the plasma membrane, SSO has undefined effects on mitochondria. Nevertheless, the KO animals reveal that FAT/CD36 contributes to the regulation of mitochondrial fatty acid oxidation, which is especially important for meeting the increased metabolic demands during muscle contraction.

AMPKα2deficiency uncovers time dependency in the regulation of contraction-induced palmitate and glucose uptake in mouse muscle

“…Because we could not isolate mitochondrial membrane fractions on top of nuclear fractions from the small amount of hindlimb muscles that we had, we could not determine whether CD36 protein content was altered in mitochondrial membranes. Indeed, it has been suggested by some (18,19), but not others (23), that mitochondrial membrane CD36 content is an important factor in the regulation of FA oxidation. Because of the methodological limitations described above, we cannot contribute data to this debate.…”

Skeletal muscle lipid flux: running water carries no poison

“…However, skeletal muscle from the whole body ATGL knockout mouse does not have decreased expression of genes relevant to fatty acid oxidation (51), an effect that may be different in cardiac muscle (34). Skeletal muscle CD36 deficiency decreased PPAR␤/␦ gene expression and mitochondrial fat oxidation (46,82). In nonmuscle tissues, including liver, brain, and macrophages, the lipogenic enzyme FAS regulates PPAR␣-dependent gene expression (12,13,100).…”

Cardiac and skeletal muscle fatty acid transport and transporters and triacylglycerol and fatty acid oxidation in lean and Zucker diabetic fatty rats

“…Cardiac and intramuscular triacylglycerol content was determined as we previously reported (3,30). Briefly, tissues (40 -50 mg) were homogenized (Polytron, Kinematica, Brinkmann, Littau-Lucerne, Switzerland) for 15 s in 2 ml of 1:1 chloroform-methanol on ice at a speed setting of 8, allowed to rest for 15 s, and homogenized again under the same conditions.…”

“…Palmitate oxidation was measured in a sealed system, as we described previously (3,30). Briefly, mitochondria were added to a pregassed modified Krebs-Ringer buffer at 37°C.…”

“…Oxidation was determined from trapping of the 14 CO2 produced and the 14 C label released from acidified buffer, as we reported previously (3,11,30,60).…”