Prolonged disuse of skeletal muscle causes significant loss of myofibrillar contents, muscle tension, and locomotory capacity. However, hibernating mammals like bats appear to deviate from this trend. Although low functional demands during winter dormancy has been implicated as a factor contributing to reduced muscle loss, the precise mechanism that actively prevents muscle atrophy remains unclear. We explored proteomic and molecular assessments of bat muscle to test a hypothesis that expression levels of major myofibrillar proteins are retained during hibernation, with periodic arousals utilized as a potential mechanism to prevent disuse atrophy. We examined changes in myofibrillar contents and contractile properties of the pectoral or biceps brachii muscles of the bat Murina leucogaster in summer active (SA), hibernation (HB) and early phase of arousal (AR) states. We found the bat muscles did not show any sign of atrophy or tension reduction over the 3-month winter dormancy. Levels of most sarcomeric and metabolic proteins examined were maintained through hibernation, with some proteins (e.g., actin and voltage dependent anion channel 1) 1.6- to 1.8-fold upregulated in HB and AR compared to SA. Moreover, expression levels of six heat shock proteins (HSPs) including glucose-regulated protein 75 precursor were similar among groups, while the level of HSP70 was even 1.7-fold higher in HB and AR than in SA. Thus, considering the nature of arousal with strenuous muscle shivering and heat stress, upregulation or at least balanced regulation of the chaperones (HSPs) would contribute to retaining muscle properties during prolonged disuse of the bat.

Hibernators like bats show only marginal muscle atrophy during prolonged hibernation. The current study was designed to test the hypothesis that hibernators use periodic arousal to increase protein anabolism that compensates for the continuous muscle proteolysis during disuse. To test this hypothesis, we investigated the effects of 3-month hibernation (HB) and 7-day post-arousal torpor (TP) followed by re-arousal (RA) on signaling activities in the pectoral muscles of summer-active (SA) and dormant Murina leucogaster bats. The bats did not lose muscle mass relative to body mass during the HB or TP-to-RA period. For the first 30-min following arousal, the peak amplitude and frequency of electromyographic spikes increased 3.1- and 1.4-fold, respectively, indicating massive myofiber recruitment and elevated motor signaling during shivering. Immunoblot analyses of whole-tissue lysates revealed several principal outcomes: (1) for the 3-month HB, the phosphorylation levels of Akt1 (p-Akt1) and p-mTOR decreased significantly compared to SA bats, but p-FoxO1 levels remained unaltered; (2) for the TP-to-RA period, p-Akt1 and p-FoxO1 varied little, while p-mTOR showed biphasic oscillation; (3) proteolytic signals (i.e., atrogin-1, MuRF1, Skp2 and calpain-1) remained constant during the HB and TP-to-RA period. These results suggest that the resistive properties of torpid bat muscle against atrophy might be attained primarily by relatively constant proteolysis in combination with oscillatory anabolic activity (e.g., p-mTOR) corresponding to the frequency of arousals occurring throughout hibernation.

At times, exercise accompanied by its anabolic effects is not a tractable countermeasure to muscle atrophy. Instead, training is often attempted after the affected muscle has atrophied greatly as a result of unloading. This study was designed to elucidate stress and signaling mechanisms underlying a process of muscle catch-up growth as a result of transitory exercise during unloading. Rats were exercised daily with a routine of 20- or 40-minute treadmill running (at 60% of maximum oxygen uptake) during the second week of a two-week hindlimb suspension. We examined the expression and activation of heat shock proteins and anabolic and proteolytic markers in the rat soleus muscle. Muscle mass relative to body mass decreased 2.4-fold in the unloaded group (HU) with respect to controls but decreased only 1.7-fold in the 40-min trained group (HT40) (P < 0.05) - equivalent to a 1.4-fold increase in the relative muscle mass over HU. Immunoblotting analyses on whole-tissue lysates demonstrated the following: (1) HSP72 and αB-crystallin were upregulated 7- and 2.5-fold, respectively, in HT40 versus HU; (2) phosphorylation of Akt1 and p70/S6K decreased only slightly in HU; (3) when compared to HU, HT40 phosphorylation of Akt1, S6K, and FoxO1 increased 1.4- to 3.0-fold while phosphorylation of FoxO3 was unchanged; and (4) activities of the ubiquitin E3 ligases, calpain 1 and caspase-3 increased 2- to 4-fold in the unloaded groups regardless of exercise duration. These results suggest that the significant upregulation of chaperones and anabolic markers (e.g., HSP72, p-Akt1, p-S6K) in HT40, along with the lack of the training effect on proteolytic activity, is likely crucial for muscle mass catch-up in the unloaded muscle.

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