Research On The Plasticity Of The PPN And Gi For Body Weight Supported Treadmill Training In Incomplete Spinal Cord Injury Rats

Background and objectsSpinal cord injury(SCI)is a kind of highly disabling disease,which can can cause the dysfunction of motor,sensory,autonomic nerve and sphincter.Not only the patients and their families have a disastrous impact,but also increased the economic burden of society.Therefore,scholars around the world are actively looking for effective treatment to help patients with spinal cord injury at an early date.At present,it has not been able to find a fully restorative cure for the patients with spinal cord injury.More than 50%of the patients with spinal cord injury had motor dysfunction,especially in patients with locomotion.Body Weight Support Treadmill Training(BWSTT)is a kind of therapy aims at ameliorating lower limb dysfunction and improving the walking ability,because it has found the plasticity of locomotor central pattern generators(CPGs)in the spinal cord after this rehabilitation therapy in animal experiments.However,there are rarely literatures about supraspinal locomotor control systems of rats received BWSTT,the present study will research on rats pedunculopontine tegmental nucleus and gigantocellular reticular nucleus,which had been verified involving in locomotion,to identify the task-dependent plasticity of these two nucleus in incomplete spinal cord injury rats that received BWSTT.Afterwards,the plasticity of these two nuclei will be described,and the relationship between the plasticity and the improvement of the rats BBB scores will be analyzed.MethodsThis research includes two parts of experiments:1.the first experiment:30 three-month-old female SD rats,weight 250-280g,were randomly divided into BWSTT group,untrained group and sham operation group,10 rats in each group.For the BWSTT group and untraind group,we use the NYU-MASCIS impactor to make a incomplete spinal cord injury rat model at T10.PET/CT,immunofluorescence staining and BBB score were applied to observe the plasticity of pedunculopontine tegmental nucleus,and assess the motor function of the hind limbs in rats with incomplete spinal cord injury.2.the second parts:a total of 60 three-month-old female SD rats weighing 250-280g were divided into sham operation group,BWSTT group and untrained group.Utilizing the same incomplete spinal cord injury rat model,and immunofluorescence staining,HE staining,Nissl staining to observe the plasticity of the gigantocellular reticular nucleus in incomplete spinal cord injury rat.We measure their motor function with Basso Beattie Bresnahan Locomotor Rating Scale(BBB),and do correlation analysis between the motor function and the plasticity.ResultsThe first part of our research showed:1.BBB Scores The scores of locomotor function of the rats hindlimbs in BWSTT group was higher than that in untrained groups at the time of the 4 and the 7 week(P<0.05).At the 1 week,the 4 week and the 7 week after spinal cord injury,BBB scores of sham operated group were respectively and significantly higher than those in the training group and the untrained group(P<0.01).2.Metabolism of Pedunculopontine Tegmental Nucleus Although the mean value of 18F-fluorodeoxyglucose(18F-FDG)standard uptake value(SUV)in BWSTT group is higher than that in untrained group,while sham operation group is the highest,we found no statistically significant difference to the SUV mean values among all groups(P>0.05).3.Immunofluorescence Neuron-specific nuclear protein(NeuN)immunofluorescence staining showed that the number of rats pedunculopontine nucleus neurons in BWSTT group,untrained group and sham operation group also have no statistically significant differences(P>0.05).But comparisons of the integral optical density value(IOD)of pedunculopontine nucleus type 2 vesicular glutamate transporters(VGLUT2)among the three groups showed significant differences.Sham operation group gets the highest expression of IOD,meanwhile IOD of BWSTT group is higher than that in untrained group(P<0.01).4.Correlation Analysis There was a significant correlation between the IOD of pedunculopontine tegmental nucleus and the BBB scores of rats with incomplete spinal cord injury(r=0.989,P<0.01).The results of second part of research:1.BBB Scores 28days after being modeled,BBB scores of BWSTT group were significant higher compared to untrained group(P<0.05).Hindlimbs motor function in BWSTT group was better than that in the untrained group.In sham operation group,the rats trunks,tails and limbs movement are fully functional.Motor function recovery of incomplete spinal cord injury rats in both part 1 and 2 are similar.2.HE Staining of Spinal Cord Spinal cord slices stained with Hematoxylin-eosin(HE)showed that the dorsal side and ventral side anatomical structure of spinal cord of sham operation group are integral,and cellular morphology of spinal cord gray matter and white matter are normal,which without neuron edema and atrophy.In the untrained group,dorsal spinal cord anatomical structure is disorganized,along with hemorrhage and cavitation.Meanwhile,neurons in dorsal and ventral spinal cord gray matter being atrophy and edema,and the dorsal side is worse.Local hemorrhage and cavitation in spinal cord slices of BWSTT group is still visible,but the level is milder than untrained group.3.Nissl staining of Gigantocellular Reticular Nucleus According to the gigantocellular reticular nucleus with Nissl staining,there is no statistical differences in the number of neurons among the three groups(P>0.05),but also the morphology of neurons of the gigantocellular reticular nucleus was no significantly change.4.Immunofluorescence The gigantocellular reticular nucleus immunofluorescence staining with VGLUT2 showed that sham group had the highest IOD,and IOD of BWSTT group is higher than untrained group.The comparison of IOD among three groups had statistically significant differences(P<0.05).5.Correlation Analysis There was a significant correlation between the IOD of gigantocellular reticular nucleus and the BBB scores of rats with incomplete spinal cord injury(r=0.975,P<0.01).Conclusion1.The plasticity of neurons in the pedunculopontine nucleus and gigantocellular reticular nucleus are decreased after incomplete spinal cord injury in rats.2.BWSTT is able to promote the recovery of hindlimbs locomotor function in rats with incomplete spinal cord injury.3.BWSTT can raise the expression of VGLUT2 which represents the plasticity of the neurons in the pedunculopontine nucleus and gigantocellular reticular nucleus.4.The mechanism of hindlimbs locomotor function recovery in incomplete spinal cord rats receiving BWSTT is about the plasticity of neurons in the pedunculopontine nucleus and gigantocellular reticular nucleus,which may be related to increasing CPG regulation.