David L. Williamson

Immediate Response of Mammalian Target of Rapamycin (mTOR)‐Mediated Signalling Following Acute Resistance Exercise in Rat Skeletal Muscle

The purpose of the present investigation was to determine whether mammalian target of rapamycin (mTOR)‐mediated signalling and some key regulatory proteins of translation initiation are altered in skeletal muscle during the immediate phase of recovery following acute resistance exercise. Rats were operantly conditioned to reach an illuminated bar located high on a Plexiglass cage, such that the animals completed concentric and eccentric contractions involving the hindlimb musculature. Gastrocnemius muscle was extracted immediately after acute exercise and 5, 10, 15, 30 and 60 min of recovery. Phosphorylation of protein kinase B (PKB) on Ser‐473 peaked at 10 min of recovery (282 % of control, P < 0.05) with no significant changes noted for mTOR phosphorylation on Ser‐2448. Eukaryotic initiation factor (eIF) 4E‐binding protein‐1 (4E‐BP1) and S6 kinase‐1 (S6K1), both downstream effectors of mTOR, were altered during recovery as well. 4E‐BP1 phosphorylation was significantly elevated at 10 min (292 %, P < 0.01) of recovery. S6K1 phosphorylation on Thr‐389 demonstrated a trend for peak activation at 10 min following exercise (336 %, P= 0.06) with ribosomal protein S6 phosphorylation being maximally activated at 15 min of recovery (647 %, P < 0.05). Components of the eIF4F complex were enhanced during recovery as eIF4E association with eIF4G peaked at 10 min (292 %, P < 0.05). Events regulating the binding of initiator methionyl‐tRNA to the 40S ribosomal subunit were assessed through eIF2B activity and eIF2α phosphorylation on Ser‐51. No differences were noted with either eIF2B or eIF2α. Collectively, these results provide strong evidence that mTOR‐mediating signalling is transiently upregulated during the immediate period following resistance exercise and this response may constitute the most proximal growth response of the cell.

Exercise-induced alterations in extracellular signal-regulated kinase 1/2 and mammalian target of rapamycin (mTOR) signalling to regulatory mechanisms of mRNA translation in mouse muscle

The present study examined the effects of an acute bout of treadmill exercise on signalling through the extracellular signal‐regulated kinase (ERK)1/2 and mammalian target of rapamycin (mTOR) pathways to regulatory mechanisms involved in mRNA translation in mouse gastrocnemius muscle. Briefly, C57BL/6 male mice were run at 26 m min−1 on a treadmill for periods of 10, 20 or 30 min, then the gastrocnemius was rapidly removed and analysed for phosphorylation and/or association of protein components of signalling pathways and mRNA translation regulatory mechanisms. Repression of global mRNA translation was suggested by disaggregation of polysomes into free ribosomes, which occurred by 10 min and was sustained throughout the time course. Exercise repressed the mTOR signalling pathway, as shown by dephosphorylation of the eukaryotic initiation factor (eIF)4E‐binding protein‐1 (4E‐BP1), enhanced association of the regulatory‐associated protein of mTOR with mTOR, and increased assembly of the tuberin–hamartin complex. In contrast, exercise caused no change in phosphorylation of either Akt/PKB or tuberin. Upstream of mTOR, exercise was associated with an increase in cAMP, protein kinase A activity, and AMP‐activated protein kinase phosphorylation. Simultaneously, exercise caused a rapid and sustained activation of the MEK1/2–ERK1/2–p90RSK pathway, resulting in increased phosphorylation of downstream targets including eIF4E and the ribosomal protein (rp)S6 on S235/S236. Overall, the data are consistent with exercise‐induced repression of mTOR signalling and global rates of mRNA translation, accompanied perhaps by up‐regulated translation of selected mRNAs through regulatory mechanisms such as eIF4E and rpS6 phosphorylation, mediated by activation of the ERK1/2 pathway.

Age-related differences in the pancreatic β-cell response to hyperglycemia after eccentric exercise

Eccentric exercise (ECC) causes muscle damage, insulin resistance, and increased pancreatic β-cell secretion in young individuals. However, the effects of age on the pancreatic β-cell response to glucose after ECC are unknown. Hyperglycemic clamps (180 min, 10.0 mM) were performed on eight young (age 22 ± 1 yr) and eight older (age 66 ± 2 yr) healthy sedentary males without exercise (CONT) and 48 h after ECC. ECC increased ( P < 0.02) muscle soreness ratings and plasma creatine kinase concentrations in both groups. Insulin and C-peptide secretions were similar between young and older subjects during CONT clamps. ECC increased ( P < 0.05) first-phase (0-10 min) C-peptide area under the curve in young (4.2 ± 0.4 vs. 3.7 ± 0.6 nM ⋅ min; ECC vs. CONT, respectively) but not in older subjects (3.2 ± 0.7 vs. 3.5 ± 0.7 nM ⋅ min; ECC vs. CONT), with significant group differences ( P < 0.02). Indeed, ECC repressed ( P < 0.05) first-phase peak C-peptide concentrations in older subjects (0.93 ± 0.16 vs. 1.12 ± 0.11 nM; ECC vs. CONT). Moreover, first-phase C-peptide-to-insulin molar ratios suggest age-related differences ( P < 0.05) in insulin/C-peptide clearance after ECC. Furthermore, the observed C-peptide response after ECC was related to abdominal adiposity [ r = -0.62, P < 0.02, and r = -0.66, P < 0.006, for first and second (10-180 min) phases, respectively]. In conclusion, older individuals did not exhibit the compensatory increase in β-cell secretion observed among young individuals after ECC. Thus, with increasing age, the pancreatic β-cell may be less responsive to the physiological stress associated with ECC.

REDD1 Induction Regulates the Skeletal Muscle Gene Expression Signature following Acute Aerobic Exercise

The metabolic stress placed on skeletal muscle by aerobic exercise promotes acute and long-term health benefits in part through changes in gene expression. However, the transducers that mediate altered gene expression signatures have not been completely elucidated. Regulated in development and DNA damage 1 (REDD1) is a stress-induced protein whose expression is transiently increased in skeletal muscle following acute aerobic exercise. However, the role of this induction remains unclear. Because REDD1 altered gene expression in other model systems, we sought to determine whether REDD1 induction following acute exercise altered the gene expression signature in muscle. To do this, wild-type and REDD1-null mice were randomized to remain sedentary or undergo a bout of acute treadmill exercise. Exercised mice recovered for 1, 3, or 6 h before euthanization. Acute exercise induced a transient increase in REDD1 protein expression within the plantaris only at 1 h postexercise, and the induction occurred in both cytosolic and nuclear fractions. At this time point, global changes in gene expression were surveyed using microarray. REDD1 induction was required for the exercise-induced change in expression of 24 genes. Validation by RT-PCR confirmed that the exercise-mediated changes in genes related to exercise capacity, muscle protein metabolism, neuromuscular junction remodeling, and Metformin action were negated in REDD1-null mice. Finally, the exercise-mediated induction of REDD1 was partially dependent upon glucocorticoid receptor activation. In all, these data show that REDD1 induction regulates the exercise-mediated change in a distinct set of genes within skeletal muscle.