One of the key events that occurs during skeletal muscle inactivation is a change in myosin phenotype, i.e. increased expression of fast isoforms and decreased expression of the slow isoform of myosin heavy chain (MyHC). It is known that calcineurin/nuclear factor of activated T-cells and AMP-activated protein kinase (AMPK) can regulate the expression of genes encoding MyHC slow isoform. Earlier, we found a significant decrease in phosphorylated AMPK in rat soleus after 24 h of hindlimb unloading (HU). We hypothesized that a decrease in AMPK phosphorylation and subsequent histone deacetylase (HDAC) nuclear translocation can be one of the triggering events leading to a reduced expression of slow MyHC. To test this hypothesis, Wistar rats were treated with AMPK activator (AICAR) for 6 days before HU as well as during 24 h of HU. We discovered that AICAR treatment prevented a decrease in pre-mRNA and mRNA expression of MyHC I as well as MyHC IIa mRNA expression. Twenty-four hours of hindlimb suspension resulted in HDAC4 accumulation in the nuclei of rat soleus but AICAR pretreatment prevented this accumulation. The results of the study indicate that AMPK dephosphorylation after 24 h of HU had a significant impact on the MyHC I and MyHC IIa mRNA expression in rat soleus. AMPK dephosphorylation also contributed to HDAC4 translocation to the nuclei of soleus muscle fibres, suggesting an important role of HDAC4 as an epigenetic regulator in the process of myosin phenotype transformation.

It is known that plantar mechanical stimulation (PMS) is able to attenuate unloading-induced skeletal muscle atrophy and impaired muscle function. However, molecular mechanisms underlying the effect of PMS on skeletal muscle during unloading remain undefined. The aim of the study was to evaluate the effects of PMS on anabolic and catabolic signaling pathways in rat soleus at the early stages of mechanical unloading. Wistar rats were randomly assigned to ambulatory control, hindlimb suspension (HS) for 1 or 3 days, and HS for 1 or 3 days with PMS. The key anabolic and catabolic markers were assessed by western blotting and RT-PCR. Protein synthesis (PS) rate was estimated using SUnSET technique. PMS attenuated a 1-day HS-induced decrease in 4E-BP1, GSK-3β, and AMPK phosphorylation. PMS also partially prevented a decrease in PS, phosphorylation of GSK-3β, nNOS, and an increase in eEF2 phosphorylation after 3-day HS. PMS during 1- and 3-day HS prevented MuRF-1, but not MAFbx, upregulation but did not affect markers of ribosome biogenesis (18S + 28S rRNA, c-myc) as well as AKT phosphorylation. Thus, PMS during 3-day HS partially prevented a decrease in the global rate of PS in rat soleus muscle, which was accompanied by attenuation of MuRF-1 mRNA expression as well as changes in GSK-3β, nNOS, and eEF2 phosphorylation.

Both research conducted under microgravity conditions and ground-based space analog studies have shown that air pump-based plantar mechanical stimulation (PMS) of cutaneous mechanoreceptors of the sole of the foot is able to increase neuromuscular activity in the musculature of the lower limbs. This type of stimulation is able to attenuate unloading-induced skeletal muscle atrophy and impaired muscle function. The aim of the present study was to evaluate the effects of PMS on anabolic signaling pathways in rat soleus muscle following 7-day hindlimb suspension (HS) and to elucidate if the effects of PMS on anabolic processes would be NO-dependent. The soles of the feet were stimulated with a frequency of 1-s inflation/1-s deflation with a total of 20 min followed by 10 min rest. This cycle was repeated for 4 h each day. We observed a decrease in the soleus muscle mass after 7-day HS, which was not prevented by PMS. We also observed a decrease in slow-type fiber cross-sectional area (CSA) by 56%, which significantly exceeded a decrease (–22%) in fast-type fiber CSA. PMS prevented a reduction in slow-twitch fiber CSA, but had no effect on fast-twitch fiber CSA. PMS prevented a 63% decrease in protein synthesis after 7-day HS as well as changes in several key anabolic signaling regulators, such as p70S6k, 4E-BP1, GSK3β, eEF-2, p90RSK. PMS also prevented a decrease in the markers of translational capacity (18S and 28S rRNA, c-myc, 45S pre-rRNA). Some effects of PMS on anabolic signaling were altered due to NO-synthase inhibitor (L-NAME) administration. Thus, PMS is able to partially prevent atrophic processes in rat soleus muscle during 7-day HS, affecting slow-type muscle fibers. This effect is mediated by alterations in anabolic signaling pathways and may depend on NO-synthase activity.

No abstract

It was observed that gravitational unloading during space missions and simulated microgravity in ground-based studies leads to both transformation of slow-twitch muscle fibers into fast-twitch fibers and to the elimination of support afferentation, leading to the “switching-off” of postural muscle motor units electrical activity. In recent years, plantar mechanical stimulation (PMS) has been found to maintain the neuromuscular activity of the hindlimb muscles. Nitric oxide (NO) was shown to be one of the mediators of muscle fiber activity, which can also promote slow-type myosin expression. We hypothesized that applying PMS during rat hindlimb unloading would lead to NO production upregulation and prevention of the unloading-induced slow-to-fast fiber-type shift in rat soleus muscles. To test this hypothesis, Wistar rats were hindlimb suspended and subjected to daily PMS, and one group of PMS-subjected animals was also treated with nitric oxide synthase inhibitor (L-NAME). We discovered that PMS led to sustained NO level in soleus muscles of the suspended animals, and NOS inhibitor administration blocked this effect, as well as the positive effects of PMS on myosin I and IIa mRNA transcription and slow-to-fast fiber-type ratio during rat hindlimb unloading. The results of the study indicate that NOS activity is necessary for the PMS-mediated prevention of slow-to-fast fiber-type shift and myosin I and IIa mRNA transcription decreases during rat hindlimb unloading.