JNK activation in TA and EDL muscle is load-dependent in rats receiving identical excitation patterns

Abstract

As the excitation-contraction coupling is inseparable during voluntary exercise, the relative contribution of the mechanical and neural input on hypertrophy-related molecular signalling is still poorly understood. Herein, we use a rat in-vivo strength exercise model with an electrically-induced standardized excitation pattern, previously shown to induce a load-dependent increase in myonuclear number and hypertrophy, to study acute effects of load on molecular signalling. We assessed protein abundance and specific phosphorylation of the four protein kinases FAK, mTOR, p70S6K and JNK after 2, 10 and 28 min of a low- or high-load contraction, in order to assess the effects of load, exercise duration and muscle-type on their response to exercise. Specific phosphorylation of mTOR, p70S6K and JNK was increased after 28 min of exercise under the low- and high-load protocol. Elevated phosphorylation of mTOR and JNK was detectable already after 2 and 10 min of exercise, respectively, but greatest after 28 min of exercise, and JNK phosphorylation was highly load-dependent. The abundance of all four kinases was higher in TA compared to EDL muscle, p70S6K abundance was increased after exercise in a load-independent manner, and FAK and JNK abundance was reduced after 28 min of exercise in both the exercised and control muscles. In conclusion, the current study shows that JNK activation after a single resistance exercise is load-specific, resembling the previously reported degree of myonuclear accrual and muscle hypertrophy with repetition of the exercise stimulus.

Figure 1

Exercise-induced JNK phosphorylation is load-dependent, while mTOR and p70S6K phosphorylation is muscle-type dependent. Specific phosphorylation (phosphorylated protein / total protein) of Y397-FAK (a), pS2448-mTOR (b), pT421/S424-p70S6K (c), and pT183/Y185-JNK (d) as measured in 33 µg soluble protein from exercised and contralateral control TA and EDL muscles after a 28-min exercise stimulus with a high or a low load. Post-hoc significant differences denoted with *, ** p < 0.05 and 0.01 versus control leg; + ,  ++ p < 0.05 and 0.01 vs EDL within same exercise group; $$ p < 0.01 versus low load exercised group within the same muscle. Data are presented as individual muscle values (circles), contralateral comparisons (lines) and group means (bars) (n = 8 per group).

Figure 2

Kinase abundance is dependent on muscle type, and p70S6K abundance is increased by exercise in TA, while JNK abundance is decreased by exercise in a load-dependent manner in TA. Kinase abundance of FAK (A), mTOR (B), p70S6K (C), and JNK (D) as measured in 33 µg soluble protein from exercised and contralateral control TA and EDL muscles after a 28-min exercise stimulus with a high or a low load. Post-hoc significant differences denoted with *, ** p < 0.05 and 0.01 vs control leg; ++ p < 0.01 vs EDL within same exercise group. Data are presented as individual muscle values (circles), contralateral comparisons (lines) and group means (bars) (n = 8 per group).

Figure 3

mTOR and JNK phosphorylation is increased with exercise duration, and for JNK also in a load-dependent manner. Specific phosphorylation of Y397-FAK (a), pS2448-mTOR (b) and pT183/Y185-JNK (c) shown as fold change in exercised relative to their respective contralateral control TA muscles after 2, 10 and 28 min of exercise with a high or a low load. Post-hoc significant differences denoted with *,** p < 0.05 and 0.01 versus control leg; + ,  ++ p < 0.05 and 0.01 versus 28 min exercise within loading groups; $$ p < 0.01 versus low load exercised group after 28 min of exercise. Data are presented as individual muscle values (circles) and group means with 95% CI (n = 8 per group for the 2 min and 28 min protocol, n = 9 per group for the 10 min protocol). Dotted line at y = 1 representing no difference between exercised leg and contralateral control leg.

Figure 4

FAK and JNK abundance is decreased with exercise duration in both exercised and contralateral control leg, but not by exercise nor load per se. Kinase abundance of FAK (a), mTOR (b) and JNK (c) as measured in 33 µg soluble protein from exercised and contralateral control TA muscles after 2, 10 and 28 min of exercise with a high or a low load. Post-hoc significant differences denoted with ++ p < 0.01 versus 2- and 10-min exercise irrespective of group. Data are presented as individual muscle values (circles), contralateral comparisons (lines) and group means (bars) (n = 8 per group for the 2 min and 28 min protocol, n = 9 per group for the 10 min protocol).