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BACKGROUND: Previous studies have shown that neurotrophin 3 (NT3)-chitosan can induce endogenous neurogenesis and axon regeneration in rats with spinal cord injury, and promote recovery of motor and sensory functions in rats. OBJECTIVE: To observe the effect of rehabilitation training combined with NT3-chitosan biomaterial scaffold on skeletal muscle morphological changes and functional recovery in rats with complete spinal cord injury. METHODS: Fifty adult female Wistar rats were randomly divided into five groups, 10 in each group. The sham group was not modeled; the remaining four groups were prepared with T7-T8 complete 5-mm spinal cord injury model, and the lesion control was not performed any intervention after modeling. The other three groups were given rehabilitation training, NT3-chitosan active biomaterial scaffold, NT3-chitosan active biomaterial scaffold combined with rehabilitation training intervention. Rehabilitation training started 2 weeks after modeling. Before operation, 2, 4, 6, 8, 10, and 12 weeks after operation, all of rats were subjected to double-blind open-field BBB scores. After 12 weeks, the skeletal muscles of the hind limbs (tibialis anterior muscle, gastrocnemius muscle, and soleus muscle) were taken for hematoxylin-eosin staining and acetylcholinesterase staining. The changes in muscle atrophy and motor endplates were assessed in each group. The experimental plan was approved by the Animal Experiment Committee of Capital Medical University (approval No. AEEI-2018-105). RESULTS AND CONCLUSION: (1) The BBB score at each time point in the sham group was higher than that in the other four groups (P < 0.05); and the scores at 8, 10, and 12 weeks after the NT3-chitosan combined rehabilitation training group were higher than the lesion control group, the lesion control combined rehabilitation training group, and NT3-chitosan group (P < 0.05). (2) At 12 weeks after operation, hematoxylin-eosin staining showed that the cross-sectional area and diameter of muscle fibers of each skeletal muscle were smaller in the other four groups than that in the sham group (P < 0.05). The cross-sectional area and diameter of muscle fibers of each skeletal muscle in the NT3-chitosan combined rehabilitation training group were higher than the lesion control group, the lesion control combined rehabilitation training group, and NT3-chitosan group (P < 0.05). (3) At 12 weeks after operation, the acetylcholinesterase staining showed that the average optical density of the acetylcholinesterase on motor endplate of the muscle was lower in the other four groups than that in the sham group (P < 0.05); the average optical density of the acetylcholinesterase of the motor endplate in the NT3-chitosan combined rehabilitation training was significantly higher than that in the lesion control, lesion control combined rehabilitation training, and NT3-chitosan groups (P < 0.05). (4) The results show that NT3-chitosan combined with rehabilitation training can effectively prevent muscular atrophy of hind limb skeletal muscles in rats with complete spinal cord injury, improve the average optical density of the acetylcholinesterase of the motor endplate, reduce neuromuscular joint degeneration, and improve rat hindlimb motor function.
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Upper limb performance is affected by diabetes mellitus (DM). Neuromuscular junction (NMJ) is a key structure to understand the relationship between performance and morphology in DM. The aim of the study was to analyze NMJ plasticity due to DM in an animal model and its relationship with the function of forelimbs in rats. Twelve Wistar rats were divided into control (C) and DM groups. Animals were trained to perform a grasping task, following procedures of habituation, shaping, and reaching task. DM was induced using streptozotocin. Forelimb neuromuscular performance for dexterity was evaluated one day before DM induction and five weeks following induction. After that, biceps, triceps, and finger flexors and extensors were removed. Connective tissue and muscle fiber cross-sectional area (CSA) were measured. NMJ was assessed by its morphometric characteristics (area, perimeter, and maximum diameter), using ImageJ software. Motor performance analyses were made using single pellet retrieval task performance test. Student's t-test was used for comparisons between groups. A significant decrease in all NMJ morphometric parameters was observed in the DM group compared with the C group. Results showed that DM generated NMJ retraction in muscles involved in a reaching task. These alterations are related to signs of muscular atrophy and to poor reaching task performance. In conclusion, induced DM caused NMJ retraction and muscular atrophy in muscles involved in reaching task performance. Induced DM caused significantly lower motor performance, especially in the final moments of evaluation, when DM compromised the tropism of the muscular tissue.
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Animals , Male , Rabbits , Rats , Task Performance and Analysis , Adaptation, Physiological/physiology , Diabetes Mellitus, Experimental/pathology , Neuromuscular Junction/pathology , Rats, Wistar , Diabetes Mellitus, Experimental/physiopathology , Neuromuscular Junction/physiopathologyABSTRACT
Objective To investigate the distribution characteristics of motor endplates in musculuslevatorani of dif-ferent gestational ages. Methods 20~24 weeks female fetuses were categorized as Group A, and 24~28 weeks female fetuses as group B. Karnovsky-Roots iron cyanide copper staining ( hematoxylin) was used to study the aver-age weight, the morphology, distribution and changes in number of motor end plate. Results The motor endplates morphology and distribution of consistent oval of 20 ~28 weeks female fetuses showed a curve distribution in the musculuslevatorani fibers foreign 1/3 . The weight and numbers of motor endplates in Group A were less than those of group B. Conclusion 20 ~28 weeks female fetuses' musculuslevatorani are visible on the motor end plates, and no significant difference is found in morphology and distribution of the curve along the muscle fibers. The num-ber of motor endplates increases with the increase of anal muscle weight mentioned.
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@#ObjectiveTo observe the hindlimb motor end-plate morphology and activity of acetyl cholinesterase (AChE) in different types of muscle fiber end-plate areas after spinal cord injury and spinal cord reconstruction tubes inducing spinal cord regeneration. Methods43 adult Wistar rats were randomly divided into spinal cord transected group at T8 (Cx group), spinal cord transected with transplantation of reconstruction tubes group (CxTp group) and control group (Co group). 2 weeks, 4 weeks, 3 months, 6 months and 12 months after operation, gastrocnemius, soleus and extensor digitorum longus were dissected respectively, and stained with gold chloride to observe the motor end-plate and stained with Karnovsky-Roots to detect AChE. ResultsIn Cx group, the end-plates degenerated since 3 months after operation, while the AChE activity declined. In CxTp group, end-plate structure and morphology were relatively stable and there were no signs of degeneration. ConclusionAfter spinal cord injury, motor end-plates undergo degeneration. The spinal cord reconstruction tubes graft can prevent end-plate degeneration and benefit for AChE reactivation and motor end-plate morphological and structural plasticity towards the direction of neurological rehabilitation.
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@#Objective To observe the effect of Xiaoyingqiangjitang on the ultrastructure of motor end-plate of skeletal muscle in rats with hyperthyroid myopathy, and changes of acetylcholin esterase (AchE).Methods The 36 healthy female Wistar rats were randomly divided into the normal control group (n=6), and model group, Chinese medicine treated group and saline control group (all n=10). All animals in the model group, Chinese medicine treated group and saline control group were treated with group given L-thyroxine intraperitoneal injection for 42 days to establish the animal model of hyperthyroid myopathy. The animals of the Chinese medicine treated group were treated with Xiaoyngqiangjitang for 30 days after L-thyroxine intraperitoneal injection. The enzyme activity of AchE and ultrastructure of motor end-plate were observed by light and electron microscopes.Results The enzyme activity and content in the group of hyperthyroid myopathy was decreased, the reaction products of AchE located in synapse became weaker and distributed heterogeneously, the junction infolding of motor end-plate became smaller and shorter, the mitochondria in axon terminal became vacuolated. After given Chinese medicine, the enzyme activity and content increased, the junction folding, the mitochondria showed normal ultrastructural appearance.Conclusion Xiaoyingqiangjitang can improve the function of skeletal muscle in hyperthyroid myopathy by recovering the enzyme activity of AchE and ultrastructures of motor end-plate.
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[Objective]To study the effect of End-to-side neurorrhaphy for protection target muscle.[Method]Twenty-six wistar rats were divided into two groups,with 13 in each group,peroneal nerve was transacted on the right side of all the animals,in one group,the proximal end of peroneal nerve was turned back and ligated,the distal end was sutured to the side of tibial nerve with perineurial window,in another group,both ends of the transacted peroneal nerve were turned back and ligated as contrast,no operation was done on the left side.Three months after operation,the peroneal nerve and the tibial muscle were taken for histologic exmination,and the muscle's wet weight was examed.[Result]Significant difference was seen between the two groups in the muscle wet weight,the muscle fiber cross sectional area,the moter endplate area and color(P
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@#ObjectiveTo observe changes of calcitonin gene related peptide (CGRP) and aetylcholinesterase (AChE) in motor endplate (MEP) after complete spinal cord injury (SCI) in the rat.MethodsSCI model of Wistar rats were established by transecting T10 spinal cord completely. The venters of the tibial anterior muscle were taken at 1, 2, 4 and 8 weeks postoperatively to carry out the experiment.The changes of CGRP and AChE in the MEP were investigated with immunohistochemical test.ResultsAfter complete SCI, the quantity and distribution of CGRP decreased in MEP of 1-week sample, and the changes of AChE were observed until 4-week sample, and although the staining was light and sparse, CGRP and AChE existed in MEP during 2 months postoperatively.ConclusionThere is degeneration of MEP in skeletal muscle after upper motor neuron injury; CGRP and AChE are related to the degeneration of MEP; CGRP test can show the changes of MEP earlier.
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Objective To investigate the changes at the motor end-plate (i. e. neuromuscular junction) following the free neurovascular muscle transfer. Methods Light and electron microscopy were used to study serially during 30 weeks the changes at the neuromuscular junction after free muscle transfer of the gracilis muscle in the adult Wistar rat. Results Under light microscopy, after staining with acetylthocholine the neuromuscular junction showed changes of degeneration with withdrawal of the innervating axon terminal followed by regeneration and reconstitution of the neuromuscular junction. With the electron microscope, mitochondrial sweelling and clumping of the synaptic vesicles were followed by withdrawal of the axon terminal from the muscle membrane on denervation. The infolding of the muscle membrane at the neuromuscular junction became less prominent. With reinnervation the ulatrastructure of the junction was only partially reestablishied with poorly reconstituted primary and secondary folds of the muscle membrane. The newly formed neuromuscular junction still lacked the structural detail seen in the control neuromuscular junction, even after 30 weeks. Conclusion The persistent weakness following free neurovascular muscle transfer may be attributed to the failure of full recovery of the intricate ultrastructure of the neuromuscular junction.
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Recently, botulinum toxin has been widely used for the management of spasticity. However it's mechanism of action in the skeletal muscle has not been well clarified. This study was performed to investigate the histopathologic changes in the skeletal muscle after botulinum toxin injection, and to determine the clinical standards of muscle fiber conduction study as an objective indicator for the changes of muscle fiber. As a study group, 35 Sprague Dawley rats were injected intra-muscularly with the botulinum toxin type A around two heads of right gastrocnemius muscle. After the injection of botulinum toxin, histopathologic studies and muscle fiber conduction studies were performed in 5 rats of the study group at 0, 1, 3, 5, 7, 14, and 28th day respectively. Based on the morphologic studies, the mechanisms of paralysis following the botulinum toxin injection were found to be both myogenic and neurogenic, and the motor function recovered through the formation of new motor end-plate and proliferation of Schwann cells. The muscle fiber conduction study revealed that the mean latencies of study group at 1, 3, 5, 7, and 14th day after the injection of botulinum toxin were significantly prolonged than those of the control group(p<0.05). The prolongation and slow recovery of latencies in a muscle fiber conduction study after the injection of botulinum toxin significantly reflect the morphologic changes of paralized skeletal muscles.
Subject(s)
Animals , Rats , Botulinum Toxins , Botulinum Toxins, Type A , Head , Motor Endplate , Muscle Spasticity , Muscle, Skeletal , Paralysis , Rats, Sprague-Dawley , Schwann CellsABSTRACT
The effect of concanavalin A (Con A) on nerve regeneration in heterograft was done in the present study. A segment about 5mm long was removed from the right sciatic nerve of adult rats. A 8mm of the heterograft nerve segment from the tibial nerve of the rabbit and pretreated by Con A was transplanted into the gap of the severed rat nerve. Regenerating nerve fibers were found from the proximal part of the sciatic nerve into the graft, and also from the graft into the distal part of the sciatic nerve at the 4th,8th and 12th week after transplantation. Regenerating nerve fibers existed separately or clustered into fascicles in the graft,as well as in the distal part of sciatic nerve. Some unmyeli-nated nerve fibers were scattered among the regenerating myelinated fibers. The gastrocnemius muscle was AChE-positive at 8th and 12th week after transplantation. In silver-combined AChE staining section, the regenerating nerve fibers could be seen connecting with the motor endplates. There were regenerating free nerve endings and nerve fascicles in the dermis and epidermis at 8th and 12th week after transplantation. The present study showed that the pretreatment of Con A had benefit to that the regenerating nerve fibers to pass through the heterograft and reinnervated to the targetes.ADepartment of Anatomy,Jiangxi Medical College,Nanchang 330006,China
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Neuromuscular junctions were formed in vitro between spatially separated explants of the fetal mouse spinal cord and the skeletal muscle. The interdependence between nerve and muscle and the changes of pre- and postsynaptic structures were observed during the formation of junctions in living and stained specimens. On the 7th day in vitro, the myotube was innervated by the neurite to form a small bulblike ending, and the cholinesterase activity was not observed at the contact region between nerve and myotube. On the 14th day in vitro, the nerve fiber sent out its branches, and the myofibers were innervated by these branches. Except some bulblike endings, some simple arborized nerve endings were also observed on the myofibers. At this stage, the cholinesterase activity began to appear at the contact region between the ending and the myofiber, and the multiple innervation of myofiber also occurred. Sometimes there were more than one nerve-muscle contact on a single myofiber, and sometimes a myofiber was innervated by several axons to form a "mixed-contact" at same place. The cholinesterase activity can only be examined in one of the multiple contacts but it is absent in the "mixed-contact". On the 21st day, both the number and the cholinesterase activity of the endplate were increased markedly. After 30 days in culture, there were intense cholinesterase activities on some endplates which were similar to young endplates in vivo, these endplates were innervated separately by single axons. When the spinal cord and muscle were co-cultured, the differentiation of muscle fiber depended on the innervation of motor nerve, and not on sensory nerve. On the other hand, the growth and maintenance of the neuron is closely related to the presence of the muscle fiber.
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Objective The experiment was designed to investigate the effect of acellular nerve allografts on the functional recovery and reconstruction of the nerve-muscle structure of the sciatic nerve defect in rats. Methods Acellular nerve allograft was transferred into the defected rat sciatic nerve with 10mm long.The wet weight of tibialis anterior was weighed at 12 and 24 weeks postoperatively compared with control group.The conducted velocity of regenerated nerve and the effect of regenerated nerve on tibialis anterior were investigated by electrophysiologic test,and silver staining combined with AChE histochemical methods were used in the experiment separately. Results The wet weight of tibialis anterior and the conducted velocity of regenerated nerve in experimental group were similar to those in control group in 12 and 24 weeks after transplantation.The positive acetylcholinesterase(AChE)histochemical reaction was observed in the tibialis anterior at 12 weeks with deeper staining and located in the middle of tibialis anterior tidily at 24 weeks after operation.The regenerated nerve bundles and nerve terminals were found to grow into the motor end-plate of the tibialis anterior in silver staining combined with AChE staining in experiment group.Electromyogram showed that the regenerated nerve has innervated tibialis anterior already.Conclusion The results indicated that extracted nerve allografts as a bridge can promote the motor functional recovery and reconstruction of the nerve-muscle structure of the defected rat sciatic nerve.