Resistance exercise increases AMPK activity and reduces 4E-BP1 phosphorylation and protein synthesis in human skeletal muscle

Abstract

Resistance exercise is a potent stimulator of muscle protein synthesis and muscle cell growth, with the increase in protein synthesis being detected within 2-3 h post-exercise and remaining elevated for up to 48 h. However, during exercise, muscle protein synthesis is inhibited. An increase in AMP-activated protein kinase (AMPK) activity has recently been shown to decrease mammalian target of rapamycin (mTOR) signalling to key regulators of translation initiation. We hypothesized that the cellular mechanism for the inhibition of muscle protein synthesis during an acute bout of resistance exercise in humans would be associated with an activation of AMPK and an inhibition of downstream components of the mTOR pathway (4E-BP1 and S6K1). We studied 11 subjects (seven men, four women) before, during, and for 2 h following a bout of resistance exercise. Muscle biopsy specimens were collected at each time point from the vastus lateralis. We utilized immunoprecipitation and immunoblotting methods to measure muscle AMPKalpha2 activity, and mTOR-associated upstream and downstream signalling proteins, and stable isotope techniques to measure muscle fractional protein synthetic rate (FSR). AMPKalpha2 activity (pmol min(-1) (mg protein)(-1)) at baseline was 1.7 +/- 0.3, increased immediately post-exercise (3.0 +/- 0.6), and remained elevated at 1 h post-exercise (P < 0.05). Muscle FSR decreased during exercise and was significantly increased at 1 and 2 h post-exercise (P < 0.05). Phosphorylation of 4E-BP1 at Thr37/46 was significantly reduced immediately post-exercise (P < 0.05). We conclude that AMPK activation and a reduced phosphorylation of 4E-BP1 may contribute to the inhibition of muscle protein synthesis during resistance exercise. However, by 1-2 h post-exercise, muscle protein synthesis increased in association with an activation of protein kinase B, mTOR, S6K1 and eEF2.

Figure 1. Schematic displaying the study design used to measure the effect of resistance exercise on the regulation of muscle protein synthesis in human subjects

The study design consisted of a basal period (hours 2–3), an exercise period (hours 3–4), and two post-exercise periods (hours 4–5, and 5–6). ICG, indocyanine green was infused to measure blood flow in each of the periods. Blood samples were collected to measure blood glucose uptake, lactate concentration and pH. Muscle biopsy samples were used to measure muscle protein synthesis, AMP-activated protein kinase (AMPK) enzyme activity, and signalling pathways involved in translation initiation and elongation.

Figure 2. Muscle protein synthesis as expressed by the mixed muscle fractional synthetic rate (FSR) before, during, and after a bout of resistance exercise

Data are expressed as means ±s.e.m., n = 11. *Significantly different from basal (P < 0.05).

Figure 3. Muscle AMPKα2 activity before, during and after a bout of resistance exercise

Data are expressed as means ±s.e.m., n = 11. *Significantly different from basal (P < 0.05).

Figure 4. Phosphorylation of TSC2 and PKB: upstream regulators of mTOR signalling before, during and after a bout of resistance exercise

Data are expressed as means ±s.e.m.*Significantly different from basal (P < 0.05). Inset shows duplicate samples for each time point.

Figure 5. Phosphorylation of mTOR and downstream indicators of mTOR signalling (4E-BP1 and S6K1) before, during and after a bout of resistance exercise

Data are expressed as means ±s.e.m.*Significantly different from basal (P < 0.05). Inset shows duplicate samples for each time point.

Figure 6. Phosphorylation of eEF2: indicator of translation elongation status before, during and after a bout of resistance exercise

Data are expressed as means ±s.e.m.*Significantly different from basal (P < 0.05). Inset shows duplicate samples for each time point.