RESUMEN

The elimination of ground reaction force (support withdrawal) vastly affects slow postural muscles in terms of their regulation and structure. One of the effects of support withdrawal in this study was an immediate postural muscle inactivation, followed by the daily gradual development of spontaneous activity of the slow postural soleus muscle in response to rat hindlimb suspension to mimic space flight. The origin of this activity is somewhat akin to muscle spasticity after spinal cord injuries and is the result of KCC2 content decline in the spinal cord's motor neurons. However, the physiological consequences of unloading-induced spontaneous activity remain unexplored. We have conducted an experiment with the administration of a highly specific KCC2 activator during 7-day unloading. For this experiment, 32 male Wistar rats were divided into 4 groups: C+placebo, C+CLP-290 (100 mg/kg b w), 7HS+placebo, and 7HS+CLP-hindlimb-suspended group with CLP-290 administration (100 mg/kg b w). The soleus muscles of the animals were dissected and analyzed for several proteostasis- and metabolism-related parameters. CLP-290 administration to the unloaded animals led to the upregulation of AMPK downstream (p-ACC) and mTOR targets (p-p70S6k and p-4E-BP) and an enhanced PGC1alpha decrease vs. the 7HS group, but neither prevented nor enhanced atrophy of the soleus muscle or myofiber CSA.

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RESUMEN

A gradual increase in rat soleus muscle electromyographic (EMG) activity is known to occur after 3-4 days of hindlimb suspension/unloading (HS). The physiological significance and mechanisms of such activity of motoneurons under unloading conditions are currently unclear. Since hyperactivity of motoneurons and muscle spasticity after spinal cord injury are associated with KCC2 downregulation, we hypothesized that a decrease in potassium (K+)/chloride (Cl-) co-transporter 2 (KCC2) in motoneurons would be responsible for an increase in soleus muscle EMG activity during HS. We aimed to investigate the effect of prochlorperazine (KCC2 activator) on the electrical activity of rat soleus muscle under HS. Wistar rats were divided into the following groups: (1) vivarium control (C), (2) 7-day HS group (7HS) and (3) 7-day HS group plus intraperitoneal injections of prochlorperazine (10 mg/kg, daily) (7HS + P). Expression of proteins in the motoneurons of the lumbar spinal cord was determined by Western blotting. An electromyogram of the rat soleus muscle was recorded using intramuscular electrodes. KCC2 content after 7-day HS significantly decreased by 34% relative to the control group. HS-induced decrease in KCC2 protein content was prevented by prochlorperazine administration. HS also induced a significant 80% decrease in KCC2 Ser940 phosphorylation; however prochlorperazine did not affect KCC2 phosphorylation. The treatment of the rats with prochlorperazine prevented a HS-induced increase in Na(+)/K(+)/(Cl-) co-transporter 1 (KCC2 antagonist) protein content. In parallel with the restoration of KCC2 content, prochlorperazine administration during HS partially prevented an increase in the soleus muscle tonic EMG activity. Thus, prochlorperazine administration during 7-day HS prevents a decrease in KCC2 protein expression in motoneurons and significantly reduces the level of HS-induced soleus muscle electrical activity.

RESUMEN

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.

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