| Spinal cord injury (SCI) is a kind of severely disabled disease. With theimprovement of standard of living and the rapid development of thetransportation industry, its incidence has also increased. Because of thedamaged central nervous tissue has extremely limited ability to regenerate, itbecome a difficult problem that repair itself after injury, consequentlyexploring a series of pathological physiology changes in mechanism after thecentral nervous system damaged and finding effective repair methods andways, and promoting the recovery of the brain and spinal cord’s function hasbecome a subject of neuroscience’s field which urgently need to be furtherdiscussed and researched.Clinical observation found that after spinal cord transverse injury,patients first experience shock stage in which any reflection significantlyreduced or disappear, and after5-10days,gradually evolves into varioushyper-reflexia cramps period. What is the morphological basis forstress-induced pathological physiology changes that occur in this criticalperiod after spinal cord injury? How the mechanism of molecular biology? Itis not very clear, and thus, in the clinical use of specific, targeted and effectivedrugs to treatment has become a thorny issue.Study found that, in the SCI model, the injection of serotonin precursor,5-hydroxytryptamin or tryptophan could make anterior horn motoneuroneenter the ultra-sensitive state. It suggested that spinal cord anterior hornmotoneurone have more sensitivity with5-HT may be one of the mainreason of spasms after SCI.AS is known at present,5-HT system have seventypes fourteen subtypes, of which5-HT2A receptor play an important role.However, in SCI, the changes of5-HT2A receptor and the function mechanism is not yet clear.This research use spinal cord complete transection injury rat model,in-depth and detailed study the changes of anterior horn motoneuronereceptors5-HT2A in the injury segment. in order to reveal the molecularmorphological basis and mechanisms of the pathological physiologychanges.To provide valuable reference material for effective prevention andtreatment of spinal cord injury in clinical, it is observated the changes ofneurone and its related structures after spinal cord transection. further to findout the neurons, neuronal and cell ultrastructural morphology changeprocesses following regional spinal cord injury at different time points. andthus to provide the basis for change of the pathophysiology of spinal cordfunction after SCI.PartⅠ Study onexpression of5-hydroxytryptamin2A receptorsof motoneuron in anterior horn of sacral regionafter spinal cord transection of the ratsObjective:To study the5-HT2A-IR density changes within spinal segment after SCI,the spinal sacral cord injury model of rats were used byimmunohistochemistry method. to reveal the basic of morphological afterspinal cord injury hyperreflexia of muscle, tetanic spasm in the molecularbiology mechanism and physiology of change, to provide the data for clinicaltreatment.Methods:One hundred and fourty adult male Wistar rats were randomly separatedinto seven groups:2,8hours and1,2,7,30,60days (n=20). The rats in differentgroups were also randomly separated into a spinal transection group and asham-operation group(n=10). In the spinal transection operation, the secondlumbar vertebra was removed, the dura was opened and a1~2mm piece ofspinal cord tissue at S2segment was gently removed with forceps and suction. In the sham-operation, only the skin and muscles in the lumbar vertebral levelwere cut open, but the dura was kept intact. The segments below injuredspinal cord were completely removed after perfusion-fixation with4%paraformaldehyde solution. Spinal cord were cut transversely into40μm thicksections with a freezing microtome at-25℃. Spinal cord slice was staining for5-HT2AR immunoreactivity. The pictures were obtained using thefluorescence confocal microscope and the changes of density of5-HT2ARimmunoreactivity between different groups were analyzed with Image J andAdobe Photoshop CS4.Results:1. In the gray matter,5-HT2AR immunoreactivity is mainly located inthe ventral horn（AH）motoneuron pool region and lateral funiculus.thoughsome labeling could also be observed in the dorsal horn, the intermediate zoneand the region surrounding the central canal.2. It was not significant statistically that5-HT2AR-IR density in theventral horn and lateral funiculus at spinal transection group andsham-operation group of2,8hours (P>0.05).3. Compared with the1day sham-operation group,5-HT2AR-IRdensity increased more than2times in the1day operation group in the ventralhorn. There was significant differences statistically (P <0.05).4.5-HT2AR-IR density in2,7,30,60days operation groups weresignificantly increased than in the sham operation group in the ventral horn.There was significant differences statistically (P <0.01).Conclusion:1. On one day after spinal sacral cord transection,5-HT2AR-IR began toupregulation, after second days significantly up-regulated and the time ofupregulation is durated up to60days.2.5-HT2AR immunoreactivity is mainly located in the ventral hornmotoneuron pool region and lateral funiculus.3.5-HT2AR up-regulation with grown time and5-HT denervation in theextension of time to have a very closely relationship of time and schedule. 4.5-HT2AR up-regulation and5-HT denervation hypersensitivity areclosely related after spinal sacral cord transection of rats, upregulation of5-HT2AR at least partly underlies supersensitivity of the5-HT denervation.Part Ⅱ Study on the ultrastructure in anterior horn of sacralregion after spinal cord transection of the ratsObjective:To observate the changes of neurons and myeline sheath of ultrastructuralon spinal cord segments in different period of SCI, SCI model of the ratsdifferent postoperative survival time and electron microscope were used. Itprovided a theoretical basis of pathomechanism and rahabilitation of function.Methods:Forty-eight adult male Wistar rats were randomly separated into normalcontrol group and2,8hours and1,2,7,30,60days of spinal transectiongroup(n=6). In the spinal transection operation, the second lumbar vertebrawas removed, the dura was opened and a1~2mm piece of spinal cord tissue atS2segment was gently removed with forceps and suction. AfterPerfusion-fixation with4%+2%glutaraldehyde (0.1mol/L phosphate buffer)paraformaldehyde solution, the segments below injury of spinal cord wascompletely removed after2hours.Electron microscope was used to observeand photograph.Results:1.Spinal cord neuron cell bodies were regular. The karyotheca was clear.The nuclear chromatin was uniform. The nucleoli was prominent, large andround, locating in the cell bodies of the central. In the cytoplasm, the roughendoplasmic reticulum ribosomes were arranged regularly. The mitochondrialwas oval with clear crest. The myelin lamellar structure were arrangedregularly.2. It was observated that the neural cells were swelling and myelin sheathwas scattered and twisted at2hours after SCI by electron microscopic.Eight hours after SCI, most of the neurons were necrosis, the structure of membranewas ambiguity, nucleus was swelling, numbers of lysosomes were increased,the structure of myelin sheath was disordered, the cystic space appeared.3. One day after SCI, neuronal cytoplasmic extensive edema, rupture anddissolve of the cell membrane, nuclear chromatin condensation, mitochondriacristae was broke, vacuolar degeneration, loose of rough endoplasmicreticulum, ribosomes was increased, lamellar of myelin sheath furtherseparated and ruptured.4. Two days after SCI, cell structure was serious damaged, nuclearfragmentation, Golgi apparatus dissolved, swelling of myelin sheath inwhichvisible swelling of mitochondria.5. Seven days after SCI, partly of neuronal cell nuclear have somerecovered, neuronal chromatin margination, nucleolus disappear, structure ofcytoplasmic organelles was damaged, cell membrane boundary was not clear,separated, fractured and distorted of myelin sheath lamellar. mitochondriaswelling and hyperplasia, ribosomes was increased.6. Tirthty days after SCI, neuronal nuclei is close to normal, nuclearchromatin slightly concentrated, mitochondria was swelling. the proliferationof glial cells, wrapped by a small amount of myelin sheath, axonal and myelinsheath disintegration, and became to vacuolated, degree of edema andseparation of myelin sheath lamellar was relieved.7. Sixty days after SCI, partly of myelin sheath was loss, there was filledby collagen fiber, the spinal cord lost its structure, and was replaced byneuroglia and fiber.Conclusion:1. Two hours after spinal cord injury it is observated motoneuron andmyelin has changes of ultramicro-pathological.2. Form eight hours to seven days after spinal cord injury, it is observatedmotoneuron and myelin has changes gradually severely ofultramico-pathological. the most organelle were necrosis3. Form thirty days to sixty days after SCI, the most organelle were necrosis and in the late-stage, the spinal cord has lost neural structure and wasreplaced by neuroglia and fiber, The cell nucleus has been restored. |
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