The Study Of Neurotrophin Receptor P75^NTR Regulates Radiation-induced Cognitive Dysfunction

Part I.Radiation induced cognitive dysfunction: an experimental model in the ratObjective: Radiotherapy is one of the predominant methods for the treatment of primary and secondary brain tumors.Unfortunately,cognitive declines have been observed in patients who survived post-radiotherapy,and radioactive neurotoxicity in normal brain affects the quality of life for cancer survivors.But the pathogenesisremains unknown.The present study aimed to develop a model of radiation-induced cognitive dysfunction in the young rat,in order to further explore the mechanism of radiation-induced cognitive dysfunction.Methods: 100 rats were anesthetized with 3.6% chloral hydrateand placed in a prone position in a linear accelerator.The control rats were treated similarly.The whole brain of each rat received a single dose of 0 Gy or 10 Gy of 4 Mev electron beam.21-day-old Sprague-Dawley male rats were divided randomly into 1month,2months,3months after radiation?n = 50 in each group?.Sequential behavioral tests including open field,novel location and novel object recognition and Morris water maze tests were under taken post-irradiation.Results:?1?In the open field test,there were no radiation-induced alterations in the total distance traveled,or the time spent in the centre zone.?2?In the novel location test,both groups spent more time exploring the object in the novel location,there was no significant difference between groups at any time point.After the novel location trial,rats were tested for novel object recognition,the irradiated rats spent less time exploring the novel object at 3 months post-irradiation.?3?During the place navigationtest,the irradiated group significantly increased latency time compared with control group after irradiation 3months.In the spatial probe test,the spatial working memory of the irradiatied rats was impaired since 3 months after irradiation.Conclusion: Our experiment establishedradiation-induced cognitive dysfunctionmodel of rats successfully.Part II.Effects of ionizing radiation on the hippocampalneurogenesisObjective: With the development of study in pathogenesy of radiation-induced cognitive dysfunction,the traditional vascular injury and demyelinating lesions explanation gradually has gradually replaced.The hippocampal neurogenesis has become the most important content of the field.Therefore,the accurate understanding of ionizing radiation on hippocampus neurogenesisis the basic requirement for exploring the mechanim of radiation-induced cognitive dysfunction.Methods: Our previous study?Part I?had demonstrated 10 Gy whole brain irradiation could induce significant cognitive deficits in 21-day-old SD rats.So,a single dose of 10 Gy 4MV electron beam was given to 100 male SD rats in this study?n=50 per group?.To determine the effects of irradiation on the production of new cells in the DG,1,2 or 3 months following irradiaton,rats received injection of 5-bromo-2'-deoxyuridine?Brd U?.The immature neurons were labeled with antibodies against Doublecortin?Dcx?.To determine the fate of cells produced by surviving precursor cells,either 1 or 3 months after a single dose of 10 Gy,Brd U-positive cells co-expressing Neu N were labeled.The total number of positively labeled cells were counted by confocal microscopy.Results:?1?There was a substantial reduction in the number of proliferating cells in the dentate gyrus?DG?region.Compare to control,Brd U-positive cells in radiated rats reduced 74.2%?82.6% and 73.9%,respectively.?2?Cranial irradiation reduced the average number of Dcx-positive cells at both 1 month?85.4%?and 3 months?73.6%?.?3?One month after irradiation,there was a significant decrease in the percentage of Brd U+/ Neu N+ cells?68.5%?,there are almost no Brd U and Neu N double positive cells were observed 3 months after irradiation.Conclusion: In the present study,we have shown that cranial irradiation could reduce the radiosensitive cells in the DG include proliferating cells and immature neurons,and make persistent damage on the new neurons.Part III.Cranial irradiationalters dendritic spines density and morphology in the hippocampusObjective: Multimodality treatments increase survival rates in brain tumors patients,whereas their quality of life is impaired result from cognitive deficits after cranial irradiation.Changes in dendritic spine number and structure are believed to be associated with hippocampal dependent learning and memory.The present study aimed to gain some insight into the temporal and sub region specific cellular changes in the hippocampusfollowing brain irradiation.methods: To investigate the impact of radiation exposure on spine morphological alteration in brain hippocampus,the experimentalratswerefromprevious study?part II;n=40;20 per group?,1 and 3 months later,changes in spine density and morphology in the DG granule and CA1 pyramidal neurons were assessed using Golgi staining.Moreover,we analyze the synaptic-related proteins PSD-95 and Drebrin within the dendritic spines.Results:?1?Our result revealed significant reductions inspine density at both 1month?40.6%?and 3 months?28.9%?in the DG.?2?Whereas,irradiation decreased spine density only at 1 month?33.3%?in the CA1 basal dendrites and no significant changes in the CA1 apical dendrites at either time after irradiation.?3?Noteworthy among our findings were the significant dynamic changes in spine morphology that persisted 3 months following cranial irradiation.?4?Meanwhile,we found synaptic-related proteins PSD-95 and Drebrin levels depletion coincided in time with alteration in dendritic spines.Conclusion: These data suggest that cranial irradiation decreased the dendritic spine density and altered the morphology by reducing the synaptic-related proteins PSD-95 and Drebrin in young rats,the abnormal dendritic spines may be associated with radiation-induced cognitive dysfunction.Part ?.The p75 neurotrophin receptor regulates cranial irradiation-induced hippocampus-dependent cognitive dysfunctionObjective: Cognitive deficit is the most serious complication of cranial irradiation,which is characterized by progressive hippocampus-dependent learning,memory and spatial processing problems.However,the underlying mechanisms remain obscure.The neurotrophin receptor p75(p75^NTR)is involved in diverse arrays of cellular responses,including neurite outgrowth,neurogenesis,and negatively regulation of spine density,which are associated with various neurological disorders.We hypothesized p75^NTR may involve irradiation-induced cognitive dysfunction.methods: Our previous study?part I,II,III?have successfully establishedradiationinduced cognitive dysfunctionmodel of rats,detected the changes of hippocampal neurogenesis and dendritic spines in rats with radiation-induced cognitive dysfunction.Taken together,21-day-old SD rats were received 10 Gy cranial irradiation and interfered the levels of p75^NTR in the hippocampus by intrahippocampal infusion technique?n=160?in this study.Then rats were tested for learning and memory abilities.After behavioral testing,immunofluorescence and Golgi staining were used to examine hippocampal neurogenesis and dendritic spine changes,respectively.Results:?1?Our experiments showed that increased p75^NTR protein levels were accompanied by increased p75^NTR transcripts.TAp73 could induce the increase in p75^NTR m RNA after irradiation,and the TAp73/p75^NTR axis may underlie the pathology of radiation-induced cognitive dysfunction.?2?Knockdown of p75^NTR by intrahippocampal infusion of AAV8-shp75,ameliorated dendritic spine abnormalities,and synaptic-related proteins,thus prevented memory deficits,likely through normalization the phosphor-AKT activity.?3?Moreover,viral-mediated overexpression of p75^NTR in the normal hippocampus reproduced learning and memory deficits.Conclusion: This report demonstrates that p75^NTR is an important regulator of irradiation-induced cognitive deficits by regulating dendritic development and synapse structure.