Skeletal Muscle AMP-activated Protein Kinase Is Essential for the Metabolic Response to Exercise in Vivo

AMP-activated protein kinase (AMPK) has been postulated as a super-metabolic regulator, thought to exert numerous effects on skeletal muscle function, metabolism, and enzymatic signaling. Despite these assertions, little is known regarding the direct role(s) of AMPK in vivo, and results obtained in vitro or in situ are conflicting. Using a chronically catheterized mouse model (carotid artery and jugular vein), we show that AMPK regulates skeletal muscle metabolism in vivo at several levels, with the result that a deficit in AMPK activity markedly impairs exercise tolerance. Compared with wild-type littermates at the same relative exercise capacity, vascular glucose delivery and skeletal muscle glucose uptake were impaired; skeletal muscle ATP degradation was accelerated, and arterial lactate concentrations were increased in mice expressing a kinase-dead AMPKα2 subunit (α2-KD) in skeletal muscle. Nitric-oxide synthase (NOS) activity was significantly impaired at rest and in response to exercise in α2-KD mice; expression of neuronal NOS (NOSμ) was also reduced. Moreover, complex I and IV activities of the electron transport chain were impaired 32 ± 8 and 50 ± 7%, respectively, in skeletal muscle of α2-KD mice (p < 0.05 versus wild type), indicative of impaired mitochondrial function. Thus, AMPK regulates neuronal NOSμ expression, NOS activity, and mitochondrial function in skeletal muscle. In addition, these results clarify the role of AMPK in the control of muscle glucose uptake during exercise. Collectively, these findings demonstrate that AMPK is central to substrate metabolism in vivo, which has important implications for exercise tolerance in health and certain disease states characterized by impaired AMPK activation in skeletal muscle.The ubiquitously expressed serine/threonine AMP-activated protein kinase (AMPK)² is an αβγ heterotrimer postulated to play a key role in the response to energetic stress (1, 2), because of its sensitivity to increased cellular AMP levels (3). Pharmacological activation of AMPK (primarily via the AMP analogue ZMP) increases catabolic processes such as GLUT4 translocation (4, 5), glucose uptake (6, 7), long chain fatty acid (LCFA) uptake (8), and substrate oxidation (6). Concomitantly, pharmacological activation of AMPK inhibits anabolic processes, and in skeletal muscle genetic reduction of the catalytic AMPKα2 subunit eliminates these pharmacological effects (9–12). Thus, AMPK has been proposed to act as a metabolic master switch (2, 13, 14). Physiologically, exercise at intensities sufficient to increase free cytosolic AMP (AMP_free) levels is a potent stimulus of AMPK, preferentially activating AMPKα2 in skeletal muscle (15–17). The metabolic profile of skeletal muscle during moderate to high intensity exercise is remarkably similar to skeletal muscle in which AMPK has been pharmacologically activated (i.e. increases in catabolic processes). This is consistent with the hypothesis that AMPK activation is required for the metabolic response to increased cellular stress. Given this, it is surprising that the direct role(s) of skeletal muscle AMPK during exercise under physiological in vivo conditions is unknown.A number of studies have tried to attribute causality to the AMPK and metabolic responses to exercise using transgenic models. In mouse models in which AMPKα2 protein expression and/or activity has been impaired, contractions performed in isolated skeletal muscle in vitro, ex vivo, or in situ have demonstrated that skeletal muscle glucose uptake (MGU) is normal (9, 10), partially impaired (11, 18), or ablated (19). Furthermore, ex vivo skeletal muscle LCFA uptake and oxidation in response to contraction appears to be AMPK-independent (20, 21). A key limitation of these studies is that the experimental models were not physiological. Under in vivo conditions, mice expressing a kinase-dead (18) or inactive (22) AMPKα2 subunit in cardiac and skeletal muscle have impaired voluntary and maximal physical activity, respectively, indicative of a physiological role for AMPK during exercise. In this context, obese non-diabetic and diabetic individuals have impaired skeletal muscle AMPK activation during moderate intensity exercise (23) as well as during the post-exercise period (24), yet the contribution of this impairment to the disease state is unclear. Thus, in vivo studies are essential to define the role of AMPK in skeletal muscle during exercise.Physical exercise of a moderate intensity is an effective adjunct treatment for chronic metabolic diseases such as obesity and type 2 diabetes (25). Given the importance of elucidating the molecular mechanism(s) regulating skeletal muscle substrate metabolism during exercise and the putative role of AMPK as a critical mediator in this process, we tested the hypothesis that AMPKα2 is functionally linked to substrate metabolism in vivo.