Brief ambulatory reloading elevates muscle protein synthesis but does not prevent disuse atrophy in hindlimb-unloaded rats

Abstract

Disuse muscle atrophy remains a major challenge in contexts such as prolonged bed rest or microgravity. Here, we investigated whether brief bouts of ambulatory reloading could attenuate skeletal muscle atrophy caused by five days of hindlimb unloading (HU) in rats. Using a deuterium oxide tracer, we measured integrative protein synthesis (fractional synthesis rate, FSR) in the soleus, plantaris, and gastrocnemius muscles, including distinct portions of the muscle that are composed mostly of red, white, and mixed fibers. HU significantly reduced both muscle mass and FSR in the predominantly slow-twitch soleus and in the predominantly fast gastrocnemius. Intermittent ambulatory reloading (HU + AR) partially restored FSR in the soleus and gastrocnemius but did not recover soleus or gastrocnemius mass to control levels. The plantaris muscle showed no differences in mass or FSR among groups, suggesting muscle-specific responses to unloading and reloading. Fiber-type analyses revealed that portions of the gastrocnemius that are mostly red fibers had higher baseline FSR than mixed or white portions, while HU consistently depressed protein synthesis across all fiber types. In conclusion, although intermittent ambulation increased protein synthesis during HU, it was not sufficient to prevent overall muscle mass loss. These findings emphasize the importance of both the duration and intensity of loading in preserving skeletal muscle during periods of disuse.

Keywords: Anabolic signaling; Countermeasures; Microgravity; Spaceflight.

Fig. 1

Soleus (A) muscle mass (g) and (B) fractional protein synthesis rate (FSR, % per day) in Control, HU, and HU + AR groups. In general, hindlimb unloading (HU) reduced both soleus mass and FSR relative to Control, and the addition of ambulatory reloading (HU + AR) partially rescued FSR but did not restore muscle mass to Control values. Bars represent mean ± SEM. Groups not sharing the same letter are significantly different one another.

Fig. 2

Plantaris (A) muscle mass (g) and (B) fractional protein synthesis rate (FSR, % per day) in the Control, HU, and HU + AR groups. Brief ambulatory reloading did not substantially affect plantaris muscle mass or FSR relative to hindlimb unloading alone, as no significant differences were detected among the three groups. Bars represent mean ± SEM.

Fig. 3

Gastrocnemius (A) muscle mass (g) in Control, HU, and HU + AR groups after five days. Hindlimb unloading (HU) significantly reduced muscle mass compared to Control, and intermittent ambulatory reloading (HU + AR) did not restore mass to Control levels. (B) Fractional protein synthesis rate (FSR, % per day) in mixed, red, and white portions for each group. Both fiber type and loading condition significantly affected FSR (main effects), but there was no interaction. Bars represent mean ± SEM; ∗ indicates that group is different from the other fiber regions within the factor of load. Pairwise comparisons are demonstrated where bars sharing the same letter are not significantly different (p > 0.05).