| PART I Therapeutic efficacy and neuroprotective effect study of deep brain stimulation of the subthalamic nucleus in Parkinsonian RatsObjective:As a progressive neurodegenerative disease,the main clinical manifestations of Parkinson's disease are static tremor,muscle rigidity,bradykinesia,and abnormal posture gait.There is no effective cure at present.For Parkinson's patients with long-term medical treatment but poor therapeutic efficacy,deep brain stimulation(DBS)of the subthalamic nucleus(STN)could be an alternative effective treatment.However,the effects and mechanisms of deep brain stimulation at different frequencies are still unclear.In this study,diffusion tensor imaging(DTI)was used to detect the parameter changes in different regions of brain after DBS at different frequencies.Besides,the relationship between DTI parameters changes and exercise performance as well as neuroprotective effect was analyzed,in order to provide a theoretical basis for clinical treatment and assessment of Parkinson's disease.Methods:The 6-OHDA-induced rat models of hemiparkinsonian were made and divided into four groups:control group,sham operation group,low-frequency group,and high-frequency group.The portable rat brain deep electrical stimulation system was used for implantation.Low-frequency(30Hz)and high-frequency(130Hz)deep brain stimulations were given to the subthalamic nucleus in rats.First,an open field experiment was used to evaluate the changes in exercise performance before the rats were modeled,2 weeks after modeling,4 weeks after modeling,and 1 week after the electrodes were implanted.Then 1 week after deep brain stimulation,the DTI imaging technique was used to measure the parameter changes of anisotropy(FA)and apparent diffusion coefficient(ADC)in the substantia nigra,striatum,and hippocampus in rats.Finally,the expression of TH,NeuN and a-syn in substantia nigra,striatum and hippocampus were analyzed by immunohistochemistry.The effects of deep brain stimulation at different frequencies in the subthalamic nucleus and the effectiveness of DTI imaging technique evaluating the therapeutic efficacy and severity of Parkinson's disease were studied.Results:After 1 week of stimulation electrode implantation in subthalamic nucleus,the total movement distance increased in the high-frequency group and the sham operation group,while decreased in the low-frequency group and the control group.There were significant statistical differences in movement distance changes between the high-frequency group and the low-frequency group,the high-frequency group and the control group,and also the sham operation group and the control group(all P<0.05).In the high-frequency group rats,FA value of substantia nigra evaluated by DTI was significantly high than the other three groups(P<0.05),ADC and ?trans values of substantia nigra were significantly lower than the other three groups(P<0.01)whereas there was no significant difference of ?long values of substantia nigra comparing with the other three groups(P>0.05).There was no significant difference in DTI-related parameters between striatum and hippocampus(P>0.05).Immunohistochemical analysis showed that the IOD of substantia nigra TH staining(P<0.01)and NeuN staining(P<0.05)in the high-frequency group were both significantly higher than that of the other three groups.There was no significant difference of the IOD of substantia nigra a-syn staining between the four groups(P>0.05).No significant difference was found between different groups in the IOD values of TH,NeuN and a-syn staining of striatum and hippocampus either(P>0.05).The correlation analysis showed that there was a significant positive correlation between the FA value of the substantia nigra and the IOD of substantia nigra TH staining in the high-frequency group.Conclusion:High-frequency electrical stimulation(130Hz)in subthalamic nucleus and single-stimulation electrode implantation can significantly improve the exercise performance of Parkinson's disease model in rats.In the contrary,low-frequency electrical stimulation(30Hz)can increase motor dysfunction.High-frequency electrical stimulation plays a role in neuroprotection and improvement of exercise performance of Parkinson's disease model in rats by slowing the degeneration of dopaminergic neurons in substantia nigra.At the same time,DTI detection can be used as an effective technique to assess the therapeutic effects and severity of Parkinson's disease.Part II Quantitative Proteomics Study by TMT Markers in Substantia Nigra of Parkinsonian Rats with Deep Brain Stimulation in the Subthalamic NucleusObjective:As a chronic neurodegenerative disease,Parkinson's disease is with a high incidence in people over 60-year-old.The etiology of the Parkinson's disease is related to aging,environment and genetic factors,whose main pathological change is the progressive degeneration of the dopaminergic neurons in substantia nigra pars compacta.Subthalamic nucleus deep brain stimulation is an effective method treating Parkinson's disease.The previous part of our study has showed that high-frequency electrical stimulation can play a significant neuroprotective effect on dopaminergic neurons in the substantia nigra.At present,there are few proteomics research studying the effects of deep brain stimulation in Parkinson's disease models.In this study,quantitative proteomics with TMT marker was used to explore the mechanism of deep brain stimulation from the proteomics aspects.Methods:In our study,rat models of hemiparkinsonism induced by 6-OHDA were made and divided into three groups:a control group,a sham operation group and a high-frequency group.The portable rat brain deep electrical stimulation system was used for implantation and high-frequency(130Hz)deep brain stimulation was given in the subthalamic nucleus.The open field test was used to assess the changes in exercise performance before rat modelling,2 weeks of modeling,4 weeks of modeling,and 1 week after electrodes were implanted or switched on.Quantitative proteomics with TMT marker and bioinformatics analysis were used for rats' substantia nigra tissue analysis,exploring the effects of deep brain stimulation in subthalamic nucleus on proteomics.Results:One week after electrodes in subthalamic nucleus were implanted or switched on,the total movement distance increased in the high-frequency group and the sham operation group,while decreased in the control group.There were significant statistical differences in movement distance changes between the high-frequency group and the control group,and also the sham operation group and the control group(both P<0.05).GO functional annotation and enrichment analysis showed that after high-frequency electrical stimulation,functional proteins related to positive regulatory,cell regeneration,cell processing and metabolism were significantly up-regulated;while proteins related to apoptosis,cell adhesion,and immune were significantly down-regulated.The KEGG pathway enrichment analysis showed that the Calcium signaling pathway and the PI3K-Akt-mTOR signaling pathway were significantly enriched.Conclusion:High-frequency electrical stimulation(130 Hz)in the subthalamic nucleus and single-stimulation electrode implantation can significantly improve the exercise performance of Parkinsonian rats.High-frequency electrical stimulation can promote the expression of proteins related to cell regeneration while inhibit proteins related to apoptosis and immune response,thus playing a role in neuroprotective regulation.At the same time,high-frequency electrical stimulation can regulate the Calcium signaling pathway and the PI3K-Akt-mTOR signaling pathway to exert its protective effect on dopaminergic neurons in substantia nigra. |
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