Calorie restriction and exercise differentially regulate AMP-activated protein kinase across subcellular compartments...

AMP-activated protein kinase (AMPK) is a crucial energy sensor that regulates a wide range of important processes in skeletal muscle. AMPK is present in several subcellular compartments (including the cytosol, nucleus, and mitochondria). However, the influence of physiologically relevant interventions on AMPK's localization in skeletal muscle is not well understood, especially during older age. Accordingly, this study examined AMPK signaling in skeletal muscle from aged male rats (24-25-months-old) subjected to either eight-weeks of calorie restriction (CR; consuming 65% of ad libitum intake) or a single swim-exercise session. Phosphorylation of AMPK and its substrate acetyl-CoA carboxylase (ACC), as well as abundance of AMPK subunits (α1, α2, β1, β2, γ1, γ3), were assessed by immunoblotting in whole muscle lysates and cytosolic, nuclear, and mitochondrial-enriched fractions obtained by differential centrifugation. CR increased phosphorylation of AMPK and ACC in whole muscle lysates, but not in the three subcellular fractions that were tested, suggesting AMPK-activation occurs in other, currently unidentified compartments. In contrast, exercise significantly increased AMPK phosphorylation in the cytosolic fraction and ACC phosphorylation in whole lysates and all three subcellular fractions. AMPK-γ1 abundance was greater in the mitochondrial-enriched fraction of CR versus ad libitum muscles. These findings revealed strikingly different patterns of AMPK activation within key subcellular compartments in response to two important physiological interventions. This study substantially advances current knowledge and provides a foundation for future research on AMPK's compartment-specific roles in skeletal muscle physiology and aging.