Absence of humoral mediated 5'AMP-activated protein kinase activation in human skeletal muscle and adipose tissue during exercise

Abstract

5'AMP-activated protein kinase (AMPK) exists as a heterotrimer comprising a catalytic alpha subunit and regulatory beta and gamma subunits. The AMPK system is activated under conditions of cellular stress, indicated by an increase in the AMP/ATP ratio, as observed, e.g. in muscles during contractile activity. AMPK was originally thought to be activated only by local intracellular mechanisms. However, recently it has become apparent that AMPK in mammals is also regulated by humoral substances, e.g. catecholamines. We studied whether humoral factors released during exercise regulate AMPK activity in contracting and resting muscles as well as in abdominal subcutaneous adipose tissue in humans. In resting leg muscle and adipose tissue the AMPK activity was not up-regulated by humoral factors during one-legged knee extensor exercise even when arm cranking exercise, inducing a approximately 20-fold increase in plasma catecholamine level, was added simultaneously. In exercising leg muscle the AMPK activity was increased by one-legged knee extensor exercise eliciting a whole body respiratory load of only 30% .VO(2,peak) but was not further increased by adding arm cranking exercise. In conclusion, during exercise with combined leg kicking and arm cranking, the AMPK activity in human skeletal muscle is restricted to contracting muscle without influence of marked increased catecholamine levels. Also, with this type of exercise the catecholamines or other humoral factors do not seem to be physiological regulators of AMPK in the subcutaneous adipose tissue.

Figure 1. Plasma adrenaline (A) and noradrenaline (B) concentrations (nmol l⁻¹) at rest and after 10 min (A10) and 20 min (A20) of exercise in period A and 10 min (B10) and 20 min (B20) in exercise period B

Filled symbols show results from the main study (n = 11) and open symbols from the control study (n = 7). *Significant difference from rest (P < 0.05); †significant difference from the preceding value (P < 0.05); #significant difference from the control study. Values are means ± s.e.m.

Figure 2. Muscle cAMP concentration (μmol (kg d.w.)⁻¹) in muscle tissue from the main study (A, n = 11) and the control study (B, n = 7)

Filled bars show values for the resting leg and open bars show values for the exercising leg at rest and after exercise period A and B. (*)P = 0.057; ‡significant difference between the working and resting leg (P < 0.05).

Figure 3. α₂ and α₁ associated AMPK activity in muscle tissue lysates from the main study (A and C, n = 11) and the control study (B and D, n = 7)

Filled bars show values for the resting leg and open bars show values for the exercising leg at rest and after exercise period A and B. Values are related to the resting leg at rest. *Significant difference from rest (P < 0.05); (*)P = 0.06; ‡ significant difference between the working and resting leg (P < 0.05); (‡)P = 0.06. Values are means ± s.e.m.

Figure 4. αAMPK protein phosphorylation and ACCβ protein phosphorylation relative to αAMPK protein and ACCβ protein content, respectively, in muscle tissue lysates from the main study (A and C, n = 11) and the control study (B and D, n = 7)

Filled bars show values for the resting leg and open bars show values for the exercising leg at rest and after exercise period A and B. Values are related to the resting leg at rest. *Significant difference from rest (P < 0.05); (*)P = 0.057; ‡significant difference between the working and resting leg (P < 0.05); (‡)P = 0.057. Values are means ± s.e.m.

Figure 6. Representative immunoblots of protein expression and phosphorylation of invested proteins in the muscle (A) and adipose tissue (B) lysates

The muscle lysate immuno-blots are depicted with the resting (RL) and exercising (EL) leg at rest and after exercise period A (Ex A) and B (Ex.B). The adipose lysate blots are depicted at rest and after exercise A (Ex.A) and B (Ex.B), respectively.

Figure 5. A–D, αAMPK protein and ACC protein phosphorylation relative to αAMPK and ACC protein content, respectively, in adipose tissue lysates from the main study (A and C, n = 10) and the control study (B and D, n = 7) at rest and after exercise period A and B

*Significant difference from rest (P < 0.05); †significant difference from the preceding value (P < 0.05).