Lactic Acid Efflux from White Skeletal Muscle Is Catalyzed by the Monocarboxylate Transporter Isoform MCT3

Abstract

The newly cloned proton-linked monocarboxylate transporter MCT3 was shown by Western blotting and immunofluorescence confocal microscopy to be expressed in all muscle fibers. In contrast, MCT1 is expressed most abundantly in oxidative fibers but is almost totally absent in fast-twitch glycolytic fibers. Thus MCT3 appears to be the major MCT isoform responsible for efflux of glycolytically derived lactic acid from white skeletal muscle. MCT3 is also expressed in several other tissues requiring rapid lactic acid efflux. The expression of both MCT3 and MCT1 was decreased by 40 -60% 3 weeks after denervation of rat hind limb muscles, whereas chronic stimulation of the muscles for 7 days increased expression of MCT1 2-3-fold but had no effect on MCT3 expression. The kinetics and substrate and inhibitor specificities of monocarboxylate transport into cell lines expressing only MCT3 or MCT1 have been determined. Differences in the properties of MCT1 and MCT3 are relatively modest, suggesting that the significance of the two isoforms may be related to their regulation rather than their intrinsic properties.Lactic acid is both a major fuel for skeletal muscle ("red" oxidative fibers) and a major metabolic end product ("white" glycolytic muscles). Even oxidative fibers become net lactic acid exporters when oxygen supply cannot meet demand, and glycolysis is stimulated to maintain ATP supplies. Fatigue occurs when lactic acid builds up within the myocyte. This causes intracellular pH (pH i ) to drop, inhibiting both glycolysis and contractile activity (1, 2). In the extreme case further muscle activity is totally prevented, a phenomenon used to advantage by anglers "playing" their fish to exhaustion. The transport of lactic acid out of skeletal muscle fibers is essential if such intracellular accumulation of lactic acid is to be prevented.Better removal of lactic acid from the muscle fibers might improve athletic performance during intense exercise and enable better muscle function and subsequent recovery under pathological conditions such as inherited mitochondrial diseases, hypoxia, and reperfusion following a period of ischemia.Transport of lactic acid into skeletal muscle fibers for oxidation is thought to be mediated by the proton-linked monocarboxylate transporter (MCT) 1 isoform MCT1 whose expression correlates with the oxidative capacity of muscle fibers and is increased following chronic muscle stimulation (3, 4). However, sarcolemmal membranes of muscle fibers that are primarily glycolytic do not contain significant amounts of MCT1 yet transport lactic acid by means of a saturable carrier that is inhibited by known inhibitors of MCT1 (3,5,6). These data imply the presence of another MCT isoform in such glycolytic fibers. MCT kinetics in heart (7-9) and liver (10) cells also imply the existence of other MCT isoforms, and this conclusion has been confirmed by cloning and sequencing studies.The first MCT isoform (MCT1) was cloned from Chinese hamster ovary cells (11) and has since been cloned and sequenced from huma...

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“…Further details of the protocol, including washing and blocking procedures, can be found elsewhere (21).…”

“…Both the supernatants were then mixed and centrifuged at 100,000 ϫ g for 60 min at 4°C and the pellet resuspended in 30 l of buffer and 30 l of SDS-PAGE sample buffer. Following separation by SDS-PAGE Western blotting was performed using mouse anti-FLAG monoclonal antibody (Sigma) with detection by enhanced chemiluminescence (ECL) as described previously (21).…”

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