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Objective:To explore the changes of dendritic spine morphology and structure in dentate gyrus(DG) and CA1 areas of hippocampus of young rats, so as to provide a direct morphological basis for studying the molecular mechanism of radiation cognitive impairment.Methods:21-day-old Sprague-Dawley (SD) rats were given a single dose of 10 Gy whole brain irradiation. The changes of cognitive function, dendritic spine density and morphological changes in DG and CA1 areas of hippocampus were observed 1 and 3 months after irradiation, and the expression of postsynaptic density protein (PSD95) was detected by Western blot.Results:The cognitive impairment was observed in young rats 3 months after irradiation. The density of dendritic spines in DG area of hippocampus was decreased significantly by 39.06% and 29.27% at 1 and 3 months after irradiation ( t=14.96, 12.35, P<0.05), respectively. The density of dendritic spines in the basal dendrites of hippocampal CA1 area was decreased by 33.40% ( t=10.39, P<0.05) 1 month after irradiation, but had no significant change at 3 months after irradiation. While the density of dendritic spines in the apical dendrites of CA1 region did not change significantly at 1 and 3 months after irradiation. In addition, the morphology of dendritic spines in DG and CA1 regions of hippocampus was dynamically changed after irradiation. The expression of PSD95 protein was decreased by 24.6% and 50.5% ( t=2.97, 9.27, P<0.05) at 1 and 3 months after irradiation, respectively. Conclusions:This study reported the density and morphological changes of dendritic spines in different brain regions of hippocampus of young rats after ionizing radiation, suggesting that PSD95 may participate in the occurrence of radiation-induced cognitive impairment by affecting the structure and morphology of dendritic spines and reducing synaptic plasticity.
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Objective:To investigate the effect of postsynaptic density-95(PSD-95)on long-term learning and memory impairment in neonatal rats induced by sevoflurane anesthesia.Methods:A total of 54 SD rats aged 7 days of SPF grade were randomly divided into 3 groups: control group (exposed to air), model group (exposed to 2.1% sevoflurane, 4 h/d, consecutive 3 days) and PSD-95 inhibitor group (inhaled sevoflurane+ intraperitoneal injection NA-1, consecutive 5 days), with 18 rats in each group.Morris water maze test and new object recognition test were used to detect the ability of visuospatial learning and memory and recognition memory of rats in each group.RT-qPCR was used to detect the mRNA levels of kalirin, Rac1 and PSD-95 in rat hippocampus.The expressions of kalirin, Rac1, PSD-95 and apoptosis related proteins Caspase-3, Bcl-2 and Bax in rat hippocampus were detected by Western blot.The expression levels of kalirin and Rac1 in hippocampus were detected by immunohistochemistry.SPSS 23.0 software was used for statistical analysis.Repeated measurement ANOVA and one-way ANOVA was used for comparing among groups.Results:Repeated measurement ANOVA showed that in the water maze test, the interaction between time and group of platform seeking latency and swimming distance of the three groups were significant ( Ftime×group=36.539, 41.548, both P<0.01). Simple effect analysis showed that the platform latency and swimming distance in the model group from day 2 to 6 were longer than those in the control group (platform latency from day 2 to 6: t=14.039, 17.147, 13.155, 13.831, 27.247, all P<0.01; swimming distance from day 2 to 6: t=10.122, 20.987, 7.267, 10.011, 8.121, all P<0.01). Compared with the model group, from day 2 to 6, the platform latencies of PSD-95 inhibitor group were prolonged( t=7.948, 14.768, 11.582, 12.832, 24.346, all P<0.01) and the swimming distances were increased( t=8.235, 24.325, 11.234, 12.031, 7.036, all P<0.01). The new object recognition test found that the new object exploration time in the model group was significantly longer than that in the control group ((21.30±2.27)s, (19.21±1.42)s, t=1.843, P<0.01), and the new object exploration time in the PSD-95 inhibitor group was significantly longer than that in the model group ((26.83±2.13)s, t=4.844, P<0.01). The difference index of novel objects in the model group was significantly lower than that in the control group ((0.41±0.12), (0.59±0.10), t=3.416, P<0.01), and the difference index of novel objects in the PSD-95 inhibitor group was significantly lower than that in the model group ((0.37±0.08), t=0.696, P<0.05). The qRT-PCR results showed that the expressions of Rac1, kalirin and PSD-95 mRNA in the model group were significantly lower than those in the control group ( t=9.969, 3.954, 6.561, P<0.05), and the expressions of Rac1, kalirin and PSD-95 mRNA in the PSD-95 inhibitor group were significantly lower than those of the model group ( t=2.132, 2.251, 3.502, all P<0.05). Immunohistochemistry results showed that the kalirin in the hippocampus CA1 area of the model group was significantly lower than that in the control group((8.18±1.94) vs (15.47±3.35), t=11.47, P<0.01), and kalirin in the PSD-95 inhibitor group was significantly lower than that in the model group((4.98±1.53), t=10.28, P<0.01); Rac1 in the model group was significantly lower than that in the control group ((3.72±1.53), (8.17±2.91), t=6.76, P<0.01), and the Rac1 in the PSD-95 inhibitor group(2.73±0.37) was significantly lower than the model group ( t=4.72, P<0.05). Western blot results showed that Caspase-3((1.37±0.16) vs (0.54±0.01), t=5.71, P<0.01) and Bax((1.87±0.31) vs (1.23±0.25), t=12.01, P<0.01) protein levels in the model group were significantly higher than those in the control group.Caspase-3 and Bax protein levels in the PSD-95 inhibitor group were significantly higher than those in model group (Caspase-3: (1.79±0.17), t=9.87, P<0.01; Bax: (2.19±0.21), t=16.19, P<0.01). The Bcl-2 protein level in the model group was significantly lower than that of the control group ((1.22±0.21) vs (1.96±0.38), t=11.92, P<0.01). And the Bcl-2 protein level in the PSD-95 inhibitor group (1.01±0.19) was significantly lower than that in the model group ( t=10.73, P<0.01). Conclusion:Sevoflurane anesthesia can damage the long-term learning and memory function and reduce the expression of PSD95 protein in neonatal rats.Inhibiting the expression of PSD95 can aggravate this damage, which may be related to the synaptic plasticity and apoptosis of neurons involved in PSD95.
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Aim To investigate the distribution of postsynaptic density protein 95 (PSD95) and AMPA receptor GluA2 subunit during the maturation of rat hippocampal neurons in vitro. Methods Cultured rat hippocampal neurons at 4(days in vitro, DIV), 7DIV, 14DIV and 20DIV were used in an immunofluorescence assay to test co-localization of PSD95 and GluA2 by laser confocal microscope at a magnification of 60 ×. Results PSD95 puncta were gradually distributed densely in dendrites and dendritic spines during neuron maturation progresses. GluA2 subunits were uniformly distributed in the cell body, axons, dendrites and dendritic spines. The Pearson' s correlation of PSD95 and GluA2 was 0. 830 ±0. 033 and 0. 734 ±0. 019 respectively at the dendrites of 4DIV and 7DIV neurons, which showed strong co-localization. While at the dendrites of 14DIV and 20DIV neurons, a moderate correlation was demonstrated by the Pearson correlation 0. 547 ± 0. 021 and 0. 574 ± 0. 024. Conclusions GluA2 has no specific intracellular distribution during neuron maturation, while the functional localization of PSD95 depends on the appearance of dendritic spines. The co-localization of GluA2 with PSD95 in dendritic spines suggests that there are stable synapses containing AMPA receptors.
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IQSEC2 is an X-linked gene highly expressed at the excitatory synapses where it plays a crucial role in a-amino-3-hydroxy-5- methyl-4-isoxazolepropionic acid receptor trafficking and synaptic plasticity. To date, several males and females with severe to profound intellectual disability have been reported harbouring frameshift and nonsense variants in this gene, whereas a milder phenotype has been recognized in females carrying missense pathogenic variants. Here, we report two novel IQSEC2 variants in four females with psychiatric features and otherwise variable cognitive impairment. A female (case 1) with severe verbal language learning disorder and a psychotic episode (precipitated by exposure to anti-contraceptive pill) harboured a de novo pathogenic frameshift variant (c.1170dupG,p.Gln391Alafs*5), whereas the female proband of family 2, displaying severe psychomotor regression and complex psychiatric features carried a missense variant of uncertain significance (c.770G[A,p.Ser257Asn) that was maternally inherited. Skewed X-inactivation was noted in the carrier mother. The maternal aunt, affected by schizophrenia, was found to bear the same IQSEC2 variant. We discuss the variable clinical presentation of IQSEC2 spectrum disorders and the challenging genotype–phenotype correlation, including the possible role of environmental factors as triggers for decompensation. Our report highlights how psychiatric features may be the main clinical presentation in subtle IQSEC2 phenotype, suggesting that the prevalence of IQSEC2 mutations in patients with psychiatric disorders may be underestimated.
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OBJECTIVE: From the perspective of β-amyloid (Aβ) toxicity and synaptic plasticity, the mechanism of electroacupuncture to improve learning and memory ability in the early pathological stages of Alzheimer's disease was explored. METHODS: Twelve male amyloid-protein precursor (APP)/γ-secretase (PS1) double transgenic AD mice were randomly and equally divided into electroacupuncture (EA) group and model group, and other 6 male C57BL/6 mice were used as the normal group. EA (1 Hz/50 Hz, 0.5 mA) was applied to "Baihui" (GV20) and bilateral "Yongquan"(KI1) for 15 min, once every other day for 6 weeks. Immunofluorescence was used to observe the positive expression of Aβ in the left hippocampus. Immunohistochemistry was used to observe the positive expression of postsynaptic density-95 (PSD-95) in the left hippocampus. Western blot was used to detect the expression of PSD-95 and synaptophysin (SYN)in the right hippocampus. RESULTS: Immunofluorescence results showed that extracellular Aβ was seen in the model group and electroacupuncture group, but no senile plaques were seen. Compared with the normal group, the expression level of Aβ in the hippocampus of the model group increased significantly (P<0.01). Compared with the model group, the expression of Aβ in the hippocampus of the EA group decreased (P<0.05). Immunohistochemical results showed that compared with the normal group, the PSD-95 positive expression in the model group was decreased(P<0.05). Compared with the model group, the expression of PSD-95 in the EA group was increased (P<0.05). Western blot results showed that compared with the normal group, the expression levels of PSD-95 and SYN in the hippocampus of the model group were decreased (P<0.05, P<0.01). Compared with the model group, the expression levels of PSD-95 and SYN in the EA group were increased (P<0.05，P <0.01). CONCLUSION: EA can reduce the expression of Aβ in the hippocampus of APP/PS1 mice and increase the expression of PSD-95 and SYN, which may contribute to its effect in improving the synaptic plasticity.
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Objective To observe the effects of Wutou decoction(WTD)on Anxiety and Depression induced by the L5 spinal nerve ligation (SNL) mice. Methods The ICR male mice were randomly divided into sham,SNL,high-dose,medium-dose and low-dose group of Wutou decoction,and Pregabalin group,with 24 in each group. Except the sham group, SNL models were prepared in the remaining groups of mice. After successful modeling, the high, medium, and low dosages were administered with 12.60, 6.30, 3.15 g/kg of Wutou decoction,and pregabalin group was administered with 0.25 g/kg of pregabalin,and the sham group and the model group was given an equal volume of normal saline. All the administration was once per day, continuously last for 21 days. On the 7/14/21thdays after the operation, the depressive and eanxiety behavior were evaluated by the tests of the sugar water consumption, the open field and forced swimming detection. In addition, the number of the positive cells expressing the extracellular signal-regulated kinase (p-ERK) in hippocampus were analyzed on Day 7/14/21 in each group by Immunohistochemistry and the expression of both p-ERK and postsynaptic density protein 95 (PSD95) in the hippocampus were analyzed on Day 21 by Western blot. Results At 7, 14, and 21thday after administration, compared with the model group, the paw withdrawal mechanical threshold of mice(0.76 ± 0.21 vs.0.05 ± 0.03),(0.93 ± 0.33 vs.0.05 ± 0.01),(1.12 ± 0.20 vs.0.04 ± 0.01)in the high dose group of Wutou decoction respectively increased(P<0.01),and sugar consumption(1.86 ± 0.44 g vs. 0.84± 0.23 g), (1.84 ± 0.10 g vs. 1.02 ± 0.18 g), (1.63 ± 0.31 g vs. 0.75 ± 0.23 g) significantly increased(P<0.01),and immobility time 4 min after swimming(128.90 ± 35.27 s vs.180.30 ± 21.81 s),(125.50 ± 23.07 s vs.195.30 ± 27.70 s),(169.50 ± 23.07 s vs.207.50 ± 7.46 s)significantly decreased(P<0.01),and the retention time in the central area of the open field(35.35 ± 7.61 s vs.13.90 ± 2.27 s),(26.26 ± 3.61 s vs.13.08 ± 1.98 s),(24.04 ± 6.57 s vs.10.62 ± 3.38 s)significantly increased(P<0.01).After 21 days,the number of p-ERK positive cells in the high-dose group of Wutou decoction(11.00 ± 4.89 vs.33.67 ± 7.35)was siginificantly lower than that of the model group,and the hippocampus p-ERK protein(0.15 ± 0.09 vs.0.54 ± 0.04)siginificantly decreased,and the expression of PSD95 protein(0.32 ± 0.04 vs.0.57 ± 0.09)siginificantly decreased(P<0.01). Conclusions The Wutou decoction achieved significant anti-depressant and anti-anxiety effects,and regulated p-ERK expression levels of hippocampus in the late stage of NP.
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Liquiritigenin (LQ) is a flavonoid that can be isolated from Glycyrrhiza radix. It is frequently used as a tranditional oriental medicine herbal treatment for swelling and injury and for detoxification. However, the effects of LQ on cognitive function have not been fully explored. In this study, we evaluated the memory-enhancing effects of LQ and the underlying mechanisms with a focus on the N-methyl-D-aspartic acid receptor (NMDAR) in mice. Learning and memory ability were evaluated with the Y-maze and passive avoidance tests following administration of LQ. In addition, the expression of NMDAR subunits 1, 2A, and 2B; postsynaptic density-95 (PSD-95); phosphorylation of Ca2+/calmodulin-dependent protein kinase II (CaMKII); phosphorylation of extracellular signal-regulated kinase 1/2 (ERK 1/2); and phosphorylation of cAMP response element binding (CREB) proteins were examined by Western blot. In vivo, we found that treatment with LQ significantly improved memory performance in both behavioral tests. In vitro, LQ significantly increased NMDARs in the hippocampus. Furthermore, LQ significantly increased PSD-95 expression as well as CaMKII, ERK, and CREB phosphorylation in the hippocampus. Taken together, our results suggest that LQ has cognition enhancing activities and that these effects are mediated, in part, by activation of the NMDAR and CREB signaling pathways.
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Animals , Mice , Behavior Rating Scale , Blotting, Western , Calcium-Calmodulin-Dependent Protein Kinase Type 2 , Cognition , Glycyrrhiza , Hippocampus , In Vitro Techniques , Learning , Medicine, East Asian Traditional , Memory , N-Methylaspartate , Phosphorylation , Phosphotransferases , Protein Kinases , Receptors, N-Methyl-D-Aspartate , Response ElementsABSTRACT
Objective To study the changes of hippocampal caspase-3 and PSD-95 expression levels in the mice exposed to ketamine 30 mg/(kg·d)for three months. Methods Forty C57BL/6 mice were randomly divided into two groups,and the chronic ketamine addiction model was established by giving mice a three month course of daily intraperitoneal injections of ketamine. Immunohistochemical study and Western blot-ting were applied to observe the expression of caspase-3 and PSD-95 protein. Results There were more expression of caspase-3 and less of PSD-95 in ketamine group as detected by immuohistochemistry. Western blotting results showed caspase-3 active fragment level significantly increased com-pared to saline group,but PSD-95 protein level was decreased. Conclusion The increased level of caspase-3 protein and reduced expression of PSD-95 are observed after long-term ketamine administration. These findings may provide an evidence for the neurotoxicity in mouse hippocampus of chronic ketamine addition as a recreational drug.
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AIM: To investigate the change of long-term potentiation ( LTP ) , and the expression of 5-hydroxytryptamine 1A receptor (5-HT1A receptor) and postsynaptic density protein 95 (PSD-95) in the hippocampus of the rats with posttraumatic stress disorder (PTSD), and to explore the mechanism of 5-HT1A receptor in the regulation of spatial memory in the PTSD rats.METHODS:Healthy adult SD rats (n=36) were randomly divided into control group and mod-el group, with 18 rats in each group.The rats in model group were treated with single prolonged stress to construct the mod-el of PTSD.Morris water maze ( MWM) was used to test the learning and memory ability .The LTP induced by high-fre-quency stimulation (HFS) was detected by electrophysiological method .The protein expression of 5-HT1A receptor and PSD-95 in the hippocampus was determined by Western blot and immunofluorescence .RESULTS: The MWM analysis showed that the latency of the rats searching for the underwater platform in model group was significantly longer than that in control group (P<0.01).The results of electrophysiological analysis showed that the amplitude of the evoked potential in both groups were significantly increased after HFS in the hippocampus , but that in model group was significantly lower than that in control group (P<0.01).The results of Western blot and immunofluorescence analysis showed that compared with control group, the protein expression of 5-HT1A receptor was obviously increased (P<0.05), while the expression of PSD-95 was obviously decreased in model group (P<0.05).CONCLUSION:The spatial memory impairment in the PTSD rats may be associated with the increase in the expression of 5-HT1A receptor and the decrease in the expression of PSD-95 in the CA1 region of hippocampus .
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Objective To explore the effect of autophagy on the expression of synaptophysin (Syn) and postsynaptic density material 95 (PSD-95) in CA1 hippocanipal region of vascular dementia rats. Metluxls A total of 96 healthy male SD rals were randomly divided into sham operation group (Sham group)-vascular dementia model group (VD group)-autophagy inhibitor 3-mcthyl adenine preconditioning group (3-MA group) and autophagy agonist rapamycin preconditioning group (Rap group). Each group was further randomized into 4 subgroups 1, 2, 4 and 8 weeks after model establishment. The rat model of vascular dementia was established using a modified Pulsinelli four-vessel occlusion method. The expressions of Syn and PSD 95 in CA1 hippocanipal region of the vascular dementia rals were delected by Western blotting analysis. Results (1) Compared with the Sham group- the ratio of LC3- II LC3 I in CA1 hippocanipal region of the vascular dementia rats was significantly increased and the expressions of Syn and PSD-95 were significantly decreased in VD group at different time points (all P
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Objective To observe the effect of electroacupuncture (EA) at Baihui (GV20), Dazhui (GV14) and Shenshu (BL23) acu-points on cognitive function and the synapse of neurons in hippocampal CA1 in SAMP8 mice,to explore the mechanism of EA in the treat-ment of Alzheimer's disease(AD).Methods A total of 24 seven-month-old SAMP8 mice were randomly divided into model group(n=12) and EA group (n=12), and the same age SAMR1 mice were as control group (n=12).The EA group accepted EA at Baihui, Dazhui and Shenshu for 30 days.They were assessed with Morris maze test.The expression of synaptophysin(SYN)and postsynaptic density protein 95(PSD95)in hippocampal CA1 region were detected with immunohistochemistry.The morphology and density of synapse in hippocampal CA1 region was observed with transmission electron microscopy.Results Compared with the model group,the latency of Morris maze de-creased in EA group(P<0.05),the time staying in the quadrant of the platform increased(P<0.05),as well as the number passing the origi-nal platform(P<0.05),with the more expression of SYN and PSD95 in hippocampal CA1 region(P<0.001),and more and completed syn-apse.Conclusion EA can improve the learning and memory ability of SAMP8 mice by increasing the expression of SYN and PSD95 to pro-tect the ultrastructure of synapses in hippocampal CA1 region.
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Objective To study the effects of different types of exercise training on learning and memory, as well as on the expression of synaptophysin (SYP) and on postsynaptic density protein 95 (PSD-95) in rats in which a model of vascular dementia had been created.Methods Forty male Wistar rats were divided randomly into a voluntary exercise group (V-EX) , a forced exercise group (F-EX) , an involuntary exercise group (I-EX) , a vascular dementia group (VD) and a sham-operation group (Sham) , with 8 rats in each group.Two-vessel occlusion (2-VO) of the arteria carotis communis was used to create a model of vascular dementia in all of the rats except those in the sham-operation group.Beginning one week after the surgery, the V-Ex rats were free to run in a running wheel.The F-EX rats were forced to run 270 m a day in an electric wheel.The I-EX rats were stimulated to imitate the gait pattern of their forelimbs running at 9 m/min three times a day for l0 minutes each time.No special training was given to the rats in the other 2 groups.Three weeks after the surgery, their learning and memory were tested using a novel object recognition test.Immediately after the test, their prefrontal cortex was sampled and the expression of SYP and PSD-95 was detected using western blotting.Results The average novel object recognition indices of the rats in the V-EX, F-EX and I-EX groups were all significantly higher than that of the VD group.Average PSD-95 expression was also significandy higher than in the VD group.Conclusion Exercise, whether voluntary, forced or induced by functional electrical stimulation can improve learning and memory in vascular dementia, at least in rats.The mechanism is possibly that the training can increase the expression of PSD-95 in the prefrontal cortex, though not SYP.
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Objective Bone marrow mesenchymal stem cells (BMSCs) were induced to differentiate to the special histological types of neurons in vitro.The morphological change of cells and positive expression of specific antigen on membrane were studied,and the function of connection between the induced BMSCs was also detected.The feasibility of BMSCs differentiate to the special histological types of neurons was investigated.Methods BMSCs were divided into group Ⅰ (induced with bFGF+GDNF),group Ⅱ (induced with bFGF+GDNF+WHI-P131 +Shh),and control group (no revulsive).The morphologic change of cells was observed,and the positive rate of neuron specific surface antigen and the content of dopamine were detected.Formation of mature synaptic structure was detected by immunohistochemical assay of postsynaptic density protein 95 (PSD-95) expression,and synaptic loop was shown by FM1-43 stain synaptic vesicles.Results By immunohistochemical staining,the positive rates of dopamine transporter (DAT) and tyrosine hydroxylase (TH) in group Ⅱ were significantly higher than those in group Ⅰ,and dopamine can been detected in cell culture supematant of group Ⅱ.After BMSCs was induced into dopamine neuron-like cells,number and length of cell protrusions,positive rate of PSD-95 and fluorescence intensity of FM1-43 in group Ⅱ were significantly higher than those of group Ⅰ.Conclusions There were no significant change in positive rate of neuron-specific surface markers,rate of cell survival and differentiation rate after BMSCs differentiated to dopaminergic neuron-like cells.The number and length of cell protrusions,content of dopamine in cell culture supematant,positive rate of dopaminergic neuron-specific surface antigen (DAT and TH),synaptic function index (positive rate of PSD-95 and fluorescence intensity of synaptic loop) of group Ⅱ were all significantly higher than that of group Ⅰ.
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Brain ischemia leads to overstimulation of N-methyl-D-aspartate (NMDA) receptors, referred as excitotoxicity, which mediates neuronal cell death. However, less attention has been paid to changes in synaptic activity and morphology that could have an important impact on cell function and survival following ischemic insult. In this study, we investigated the effects of reperfusion after oxygen/glucose deprivation (OGD) not only upon neuronal cell death, but also on ultrastructural and biochemical characteristics of postsynaptic density (PSD) protein, in the stratum lucidum of the CA3 area in organotypic hippocampal slice cultures. After OGD/reperfusion, neurons were found to be damaged; the organelles such as mitochondria, endoplasmic reticulum, dendrites, and synaptic terminals were swollen; and the PSD became thicker and irregular. Ethanolic phosphotungstic acid staining showed that the density of PSD was significantly decreased, and the thickness and length of the PSD were significantly increased in the OGD/reperfusion group compared to the control. The levels of PSD proteins, including PSD-95, NMDA receptor 1, NMDA receptor 2B, and calcium/calmodulin-dependent protein kinase II, were significantly decreased following OGD/reperfusion. These results suggest that OGD/reperfusion induces significant modifications to PSDs in the CA3 area of organotypic hippocampal slice cultures, both morphologically and biochemically, and this may contribute to neuronal cell death and synaptic dysfunction after OGD/reperfusion.
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Brain Ischemia , Cell Death , Dendrites , Endoplasmic Reticulum , Ethanol , Mitochondria , N-Methylaspartate , Neurons , Organelles , Phosphotungstic Acid , Post-Synaptic Density , Presynaptic Terminals , Protein Kinases , Proteins , Receptors, N-Methyl-D-Aspartate , ReperfusionABSTRACT
Nitric oxide (NO) is a molecular messenger involved in several events of synaptic plasticity in the central nervous system. Ca2+ influx through the N-methyl-D-aspartate receptor (NMDAR) triggers the synthesis of NO by activating the enzyme neuronal nitric oxide synthase (nNOS) in postsynaptic densities. Therefore, NMDAR and nNOS are part of the intricate scenario of postsynaptic densities. In the present study, we hypothesized that the intracellular distribution of nNOS in the neurons of superior colliculus (SC) superficial layers is an NMDAR activity-dependent process. We used osmotic minipumps to promote chronic blockade of the receptors with the pharmacological agent MK-801 in the SC of 7 adult rats. The effective blockade of NMDAR was assessed by changes in the protein level of the immediate early gene NGFI-A, which is a well-known NMDAR activity-dependent expressing transcription factor. Upon chronic infusion of MK-801, a decrease of 47 percent in the number of cells expressing NGFI-A was observed in the SC of treated animals. Additionally, the filled dendritic extent by the histochemical product of nicotinamide adenine di-nucleotide phosphate diaphorase was reduced by 45 percent when compared to the contralateral SC of the same animals and by 64 percent when compared to the SC of control animals. We conclude that the proper intracellular localization of nNOS in the retinorecipient layers of SC depends on NMDAR activation. These results are consistent with the view that the participation of NO in the physiological and plastic events of the central nervous system might be closely related to an NMDAR activity-dependent function.
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Animals , Male , Rats , Dizocilpine Maleate/pharmacology , Excitatory Amino Acid Antagonists/pharmacology , Nitric Oxide Synthase Type I/metabolism , Receptors, N-Methyl-D-Aspartate/antagonists & inhibitors , Superior Colliculi/enzymology , Immunohistochemistry , Superior Colliculi/drug effectsABSTRACT
Objective To investigate the influence of Memantine on changes of N-methyl-D-aspartic acid receptor 2B (NMDAR-2B) level, the expression of postsynaptic density 95 ( PSD-95) and calcium/calmodulin-dependent protein kinase Ⅱ (CaMK Ⅱ)activities in the hippocampus of vascular dementia(VD)rats and their recognition. Methods The VD rat model was established by permanent,bilateral occlusion of the common carotid arteries. One hundred and forty-four Wistar rats were randomly divided into sham-operated group, VD model group and Memantine-treated group. The water maze test was performed to detect the ability of learning and memory of the rats,the changes of NMDAR-2B level were measured by RT-PCR,the changes in the expression of PSD-95 were detected by immunohistochemical staining,and the changes in the CaMK Ⅱ activities were determined by incorporation of ~(32)P into histone. Results Compared with the sham-operated group, the ability of learning and memory of VD rats was decreased significantly(P<0. 01); the level of NMDAR-2B and the expression of PSD-95 increased significantly at the 4th week after operation(F<0. 05 or 0. 01)and decreased at the 8th to 16th week after operation (P<0. 01). Total CaMK Ⅱ activity was increased significantly at the 4th week after operation(P<0. 01) ,then dropped at the 8th week after operation(P>0. 05) ,and decreased significantly at the 12th,16th week after operation(P<0. 01). As for Memantine-treated group,at the 4th week after operation,there was no statistically significant difference in the parameters mentioned above in contrast with the sham-operated group. At the 8th,and 12th week after operation,the ability of learning and memory,the level of NMDAR-2B and the expression of PSD-95 were significantly increased as compared with VD rats(P<0. 01 or 0. 05) ,but total CaMK Ⅱ activity was similar to that of VD rats. Conclusion With the development of VD,NMDAR-2B level,the expression of PSD-95 and CaMK Ⅱ activities were firstly over-activated,and then decreased greatly. Memantine can improve VD rats' recognition by modulating the expression of NMDAR-2B,but its effect on CaMK Ⅱactivity was limited.
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Objective To investigate the effect of electroacupuncture (EA) combined with repetitive transcranial magnetic stimulation ( rTMS) on postsynaptic density 95 ( PSD- 95) in the hippocampus of rats after cerebral ischemia. Methods Sixty healthy male Wistar rats were divided into normal, model, EA, rTMS and EA + rTMS groups randomly, and the rats in each group were further divided into 7 d, 14 d and 28 d subgroups. A model of middle cerebral artery occlusion ( MCAO) was established, followed by EA, rTMS or EA + rTMS treat-ment. The expression of PSD-95 in the dentate gyros and area CA3 of the hippocampus of the ischemic cerebral hemisphere were investigated. Results The expression of PSD-95 in the dentate gyrus and area CA3 of the ische-mic cerebral hemisphere had decreased 7 d after cerebral ischemia. The expression of PSD-95 in the three treat-ment groups increased after 14 d of treatment and was significantly greater compared with the model group after 28 d of treatment. The EA + rTMS group was especially superior to the EA and rTMS groups in terms of the expression of PSD-95 in area CA3 by the 28th day. Conclusions EA + rTMS can significantly increase the expression of PSD-95 in the hippocampus after cerebral ischemia. This may help explain how EA + rTMS improves learning and mem-ory after cerebral ischemia.
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The 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNP), a protein of unknown function in vivo, is abundantly expressed in myelinating glia in two isoforms, CNP1 and CNP2. In this study, immunoblot analysis showed that CNP1 is the major isoform in adult forebrain, and that both isoforms are included in the postsynaptic density (PSD) fraction and tyrosine-phosphorylated at the basal level. However, subcellular distribution and detergent extraction data showed that CNP is nonspecifically associated with the PSD fraction. Immunocytochemistry revealed that CNP is detected, in a weak but punctate pattern, in dissociated rat hippocampal neurons of 3 days to 2 weeks in vitro. The CNP-positive punctae were distributed throughout soma and dendrites, and distinct from PSD95-positive ones. Immunoblot analysis indicated that CNP is also expressed in neuronal stem cell lines, HiB5 and F11. Interestingly, in addition to the known two isoforms, a new CNP isoform of MW 45 kDa was expressed in these cell lines and was the major type of isoform in F11 cells. Taken together, our data suggest that CNP is expressed in the early stage of in vitro development and nonspecifically included in the adult rat PSD fraction.
Subject(s)
Animals , Rats , 2',3'-Cyclic-Nucleotide Phosphodiesterases/metabolism , Aging/physiology , Cells, Cultured , Hippocampus/cytology , Immunohistochemistry , Nerve Tissue Proteins/metabolism , Neurons/metabolism , Phosphotyrosine/metabolism , Prosencephalon/cytology , Rats, Sprague-Dawley , Substrate SpecificityABSTRACT
Objective To investigate the mechanism of the soluble A? oligomers-induced alteration of synaptic proteins. Methods This study applied immunocytochemistry technique to investigate the changes of the expression of postsynaptic density-95(PSD-95) in primary hippocampal neurons, which was exposed to A?_ 25-35 after NMDAR antagonist or agonist treatment. Results The results showed that A?_ 25-35 downregulated PSD-95 protein in a dose- and time-dependent manner. Treatment of cells with MK801 (a general NMDA receptor antagonist) prevented A?-induced PSD-95 degradation. Moreover, when extrasynaptic NMDA receptors were blocked by ifenprodil (a NR2B subunit specific antagonist), the A?-induced downregulation of PSD-95 was significantly attenuated. Whereas, when synaptic NMDA receptors were blocked by bicuculline (a GABA receptor antagonist) in combination with MK801, the PSD-95 degradation did not change significantly.Conclusion The results suggest that A?-induced downregulation of PSD-95 depends on NMDAR activity, and extrasynaptic NMDA receptors may be involved in A?-induced synaptic protein degradation.