The changes in the content of giant proteins of the sarcomeric cytoskeleton titin (3000-3700 kDa) and nebulin (770 kDa) in skeletal muscles (m. soleus, m. gastrocnemius), titin in the myocardium (left ventricle), as well as membrane cytoskeletal protein dystrophin (427 kDa) in m. soleus and m. extensor digitorum longus of edible dormouse Glis glis during hibernation have been studied. The animals were of two experimental groups: “Summer activity” and “Hypothermia”. It was found that the development of atrophic changes in the skeletal muscles of the animal during hibernation is accompanied by a decrease in the content of dystrophin. In particular, the intensity of fluorescence on cross-sections of skeletal muscles stained with primary antibodies to dystrophin and secondary antibodies conjugated to the Alexa488 fluorochrome decreased 2.7 times (p < 0.05) and 2.0 times (p < 0.05) in m. soleus and m. EDL, respectively, in animals of the “Hypothermia” group. SDS electrophoresis of proteins in agarose-strengthened macroporous 2.2%-polyacrylamide gel revealed a slight decrease (by 15%, p ≤ 0.01) in the titin content in relation to the content of myosin heavy chains in m. gastrocnemius of animals of the “Hypothermia” group. Titin content in m. soleus and cardiac muscle, as well as the nebulin content in the studied skeletal muscles did not decrease during hibernation. These results are consistent with our earlier data for other hibernates: long-tailed ground squirrel, brown and Himalayan black bears. It can be assumed that in the process of evolution in hibernating animals, molecular mechanisms were developed that are responsible for maintaining a stable level of giant proteins of the sarcomeric cytoskeleton during hibernation.

Exposure to microgravity has been shown to result in damaging alterations to skeletal muscle, bones, and inner organs. In this study, we investigated the effects of microgravity by using a hindlimb unloading model (HUM) in mice. The characteristics of the lumbar spinal cords of HUM mice 30 days after hindlimb unloading were examined. Morphometric analysis showed reductions of the total area, gray matter, and white matter by 17%, 20%, and 12%, respectively. Myelinated fibers in the white matter showed prominent myelin destruction. Analysis of the number of glial fibrillary acidic protein (GFAP+)/S100 calcium-binding protein B (S100B), GFAP+/S100B+, and GFAP-/S100B+ astrocytes in the ventral horn (VH), central channel area (CC), dorsal root entry zone (DREZ), main corticospinal tract (CST), and ventral funiculi (VF) showed that the number of GFAP+/S100B- astrocytes was increased in the DREZ and CST of HUM mice. Additionally, GFAP+/S100B+ cell numbers were significantly decreased in the VH and CST but did not differ in the CC or DREZ of HUM mice, as compared with the control. The numbers of GFAP-/S100B+ cells were significantly reduced only in the VH of HUM mice. Moreover, the number of ionized calcium-binding adaptor molecule 1 (Iba1+) microglia cells was significantly increased in the CC and DREZ of HUM mice. In control mice, homeobox protein HoxB8 (HoxB8+) cells were found only in the CC; in contrast, HoxB8+ cells were observed in all studied areas in HUM mice, with the greatest number found in the CC. Genome-wide transcriptome analysis of the lumbar spinal cords of HUM mice showed decreased expression of genes encoding myelin, extracellular matrix, cytoskeleton, and cell adhesion proteins. Real-time polymerase chain reaction (PCR) confirmed reductions in the expression of mpz, pmp2, pmp22, and prx genes, which are involved in myelination, as well as decreases in the levels of genes encoding extracellular matrix molecules, including glycoproteins (matrix gla protein (MGP), osteoglycin (OGN), microfibrillar associated protein 5 (MFAP), and collagen, type IV, alpha 1 (COL4A)), proteoglycans (perlecan (heparan sulfate proteoglycan) (HSPG)), and metalloproteinases (lysyl oxidase (LOX)). Thus, our results showed that hindlimb unloading caused decreases in gray and white matter areas, changes in gene expression, alterations in myelination, and phenotypic modifications in glial cells in the lumbar spinal cords of mice. (C) 2014 IBRO. Published by Elsevier Ltd. All rights reserved.

We made immunohistochemical study of the lumbar spinal cord of c57black/6 mice after a 30-days space flight using antibodies against proteins which involved in providing of synaptic transmission (synaptophysin and PSD95). Also we use antibodies against proteins of intracellular protection (Hsp25 and Hsp70), and neuroprotective protein (VEGF) and its receptor (Flt-1). We found, that the levels of synaptophysin immune expression decreased by 21%, PSD – by 55%, Hsp25 – by 15% and Hsp70 – by 9% in mice lumbar spinal cord motoneurons after space flight. Thus, there were no significant changes in the immune expression of VEGF and its receptor Flt-1. Obtained data reveal the changes in the functional state (synaptic activity and resistance to stress) of mice lumbar spinal cord motoneurons after space flight. So, we have shown a new data about spinal cord motoneurons role in the development of hypogravity motor syndrome.

Studies results of gravity unloading influence on spinal control system of muscle structure and functions are summarized. It was shown that demyelization of axons due to reduction of genes expression responsible for myelin proteins synthesis, decrease in one of the key enzymes of cholinergic system--cholineacetyltransferase activity, alteration of normal kinetics of quantal and non-quantal neurotransmitter secretion, impaired autoregulation of acetylcholine secretion from motor nerve endings through presynaptic cholinergic receptors, slowing of axonal transport of substances in motor neurons that innervate postural muscles played the important role in the development of hypogravitational motor symptoms. At the same time, the evidences of neuroprotective mechanisms enclosing (increase in heat shock proteins Hsp25 and Hsp70 expression), that hinder apoptosis development in motor neurons and glial cells in the spinal cord under conditions of model hypogravity, were revealed.

The spontaneous quantum secretion of neurotransmitter and its regulation through the system of presynaptic acetylcholine receptors have been studied on a neuromuscular preparation of rat m. soleus of intact animals and animals in which the axonal transport was blocked via the application of colchicine to the sciatic nerve. It was shown that, after six days of colchicine application, the spontaneous quantum secretion, the reaction of presynaptic membrane, and the reaction of neurosecretory apparatus to the depolarization of nerve endings via increase of the content of potassium ions in the environment and to the activation of presynaptic receptors by carbachol are not disturbed. Keeping in mind a rather short half-life of proteins that take part in the exocytosis and its regulation, it may be concluded that their functioning does not depend on the state of the axonal transport. These data correspond to the hypothesis put forward earlier that the synthesis of some proteins performing their function in nerve terminals occurs directly at the site of their utilization but not in the perikaryon, as it has been traditionally assumed.

Some of the electrophysiological parameters of m. soleus of rat and Mongolian gerbil, and Ca ions content in fiber myoplasm were compared in different periods of gravitational unloading simulated by tail-suspension. No difference was found between the control animals as for membrane potential at rest, electrogenic activities of Na-K-ATPase and its isoforms, and input resistance of m. soleus fibers. At the same time, unlike rats, gerbils exhibited a substantial Ca decrease in myoplasm. From day one to 14 of gravitational unloading the pace of electrophysiological changes in gerbil's m. soleus was noticeably slower than of rat's, whereas Ca ions depositing in myoplasm was observed in both species already at the beginning ofsuspension. Analysis of the results suggests that adaptive changes in m. soleus of Mongolian gerbil and rat during simulated gravitational unloading are fundamentally different due to, probably, peculiar water-electrolyte metabolism, type of locomotion, and other factors which are still unclear.