Bed rest impairs skeletal muscle amino acid transporter expression, mTORC1 signaling, and protein synthesis in response to essential amino acids in older adults

Abstract

Skeletal muscle atrophy during bed rest is attributed, at least in part, to slower basal muscle protein synthesis (MPS). Essential amino acids (EAA) stimulate mammalian target of rapamycin (mTORC1) signaling, amino acid transporter expression, and MPS and are necessary for muscle mass maintenance, but there are no data on the effect of inactivity on this anabolic mechanism. We hypothesized that bed rest decreases muscle mass in older adults by blunting the EAA stimulation of MPS through reduced mTORC1 signaling and amino acid transporter expression in older adults. Six healthy older adults (67 ± 2 yr) participated in a 7-day bed rest study. We used stable isotope tracers, Western blotting, and real-time qPCR to determine the effect of bed rest on MPS, muscle mTORC1 signaling, and amino acid transporter expression and content in the postabsorptive state and after acute EAA ingestion. Bed rest decreased leg lean mass by ∼4% (P < 0.05) and increased postabsorptive mTOR protein (P < 0.05) levels while postabsorptive MPS was unchanged (P > 0.05). Before bed rest acute EAA ingestion increased MPS, mTOR (Ser(2448)), S6 kinase 1 (Thr(389), Thr(421)/Ser(424)), and ribosomal protein S6 (Ser(240/244)) phosphorylation, activating transcription factor 4, L-type amino acid transporter 1 and sodium-coupled amino acid transporter 2 protein content (P < 0.05). However, bed rest blunted the EAA-induced increase in MPS, mTORC1 signaling, and amino acid transporter protein content. We conclude that bed rest in older adults significantly attenuated the EAA-induced increase in MPS with a mechanism involving reduced mTORC1 signaling and amino acid transporter protein content. Together, our data suggest that a blunted EAA stimulation of MPS may contribute to muscle loss with inactivity in older persons.

Figures

Fig. 1.

Schematic of infusion protocol. Protocol was conducted before and after 7 days of bed rest. EAA, essential amino acids.

Fig. 2.

Data represent blood (A) and muscle phenylalanine (B) concentrations in the postabsorptive state (basal) and after ingestion of EAA in older adults (n = 6). Solid line, before bed rest (pre-bed rest); broken line, after 7 days of bed rest (post-bed rest). *Different from basal (P < 0.05). Values are presented as means ± SE.

Fig. 3.

Mixed-muscle protein synthesis in skeletal muscle of older adults (n = 6) in the postabsorptive state and during the EAA ingestion period (3 h; 0–3 h post-EAA). #Different from post-bed rest during the 3-h EAA period (P = 0.05). Values are presented as means ± SE.

Fig. 4.

Protein kinase B (Akt), mammalian target of rapamycin (mTOR), and downstream signaling. Data represent phosphorylation of Akt at Ser473 (A), mTOR at Ser2448 (B), S6 kinase 1 (S6K1) at Thr389 (C), S6K1 at Thr421/Ser424 (D), 4E binding protein 1 (4EBP1) at Thr37/46 (E), and ribosomal protein S6 (rpS6) at Ser240/244 (F) in the postabsorptive state (basal) and 1 and 3 h after ingestion of EAA in skeletal muscle of older adults (n = 6). *Different from basal (P < 0.05). #Different from postbed rest at corresponding time point (P < 0.05). Values are presented as means ± SE.

Fig. 5.

Activating transcription factor 4 (ATF4) and amino acid transporters. Data represent ATF4 protein content (A), L-type amino acid transporter 1 (LAT1) protein content (B), LAT1 mRNA expression (C), sodium-coupled neutral amino acid transporter 2 (SNAT2) protein content (D), and SNAT2 mRNA expression (E) in the postabsorptive state (basal) and 1 and 3 h after ingestion of EAA in skeletal muscle of older adults (n = 6). *Different from basal (P < 0.05). #Different from post-bed rest at corresponding time point (P < 0.05). Values are presented as means ± SE.

Fig. 6.

Representative phospho and total protein Western blot images in the postabsorptive state (basal) and after (1 and 3 h) the ingestion of EAA before (pre-bed rest) and after (post-bed rest) 7 days of bed rest.