反复应激引起前额叶皮层物上谷氨酸受体的表达.pptVIP

* * * * * * * * * * * * * * * * * * * * * The ralationship with the time of stress: To understand the complex actions of stress hormones, we exposed animals to various days of restraint stress. To test the different regions: To test the regional speci?city of the effect of repeated stress, we also examined glutamatergic transmission in striatum and hippocampus from young male rats. Biochemical Measurement of Surface and Total Proteins: We performed western blotting and surface biotinylation experiments to detect the total and surface levels of AMPAR and NMDAR subunits in PFC slices from stressed, young (4-week-old) male rats. The amount of the glutamate recptors: Inject the GR antagonist RU486: To test the different regions: No signi?cant changes were found in other brain areas, including striatum and hippocampus. The results in vitro: We next examined whether the effect of repeated stress in vivo may be mimicked by corticosterone (CORT) application in vitro. Inject the GR antagonist and MR antagonist : RU486:The GR antagonist; RU28318:The MR antagonist Measure the cluster density : Ubiquitin/Proteasome-dependent Degradation: Since the total level of NR1 andGluR1 was reduced in repeatedly stressed animals, we examined whether it could be due to the decreased synthesis or increased degradation of glutamate receptors. Ubiquitin/Proteasome-dependent Degradation: The level of ubiquitinated GluR2, NR2A, or NR2B subunits remained unchanged. Repeated stress also failed to alter the ubiquitination of SAP97 (a GluR1 binding protein) and PSD-95. Test other proteasome inhibitor: MG132:proteasome inhibitor Lact: another proteasome inhibitor Chloro: general lysosomal enzyme inhibitor Leupeptin: lysosomal protease inhibitor 11R-CS:membrane-permeable calpain protease inhibitor Test the effect of proteasome inhibitor: ShRNA Lentiviral Knockdown: Given the role of proteasome-dependent degradation of gluta- mate receptors in the detrimental effects of repeated stress, we w