N-Methyl-D-aspartate receptors（NMDARs） play vital roles in the central nervous system,as they are primary mediators of Ca2＋influx during synaptic activity.The subunits that compose NMDARs share similar topological ...N-Methyl-D-aspartate receptors（NMDARs） play vital roles in the central nervous system,as they are primary mediators of Ca2＋influx during synaptic activity.The subunits that compose NMDARs share similar topological structures but are distinct in distribution and pharmacological properties,as well as physiological and pathological functions,which make the NMDAR one of the most complex and elusive ionotropic glutamate receptors.In this review,we focus on GluN2A and GluN2B,the primary NMDAR subunits in the cortex and hippocampus,and discuss their differences in developmental expression,brain distribution,trafficking,and functional properties during neuronal activity.展开更多
Fear memories are critical for survival.Nevertheless,over-generalization of these memories,depicted by a failure to distinguish threats from safe stimuli,is typical in stress-related disorders.Previous studies have su...Fear memories are critical for survival.Nevertheless,over-generalization of these memories,depicted by a failure to distinguish threats from safe stimuli,is typical in stress-related disorders.Previous studies have supported a protective role of ketamine against stress-induced depressive behavior.However,the effect of ketamine on fear generalization remains unclear.In this study,we investigated the effects of ketamine on fear generalization in a fear-generalized mouse model.The mice were given a single sub-anesthetic dose of ketamine(30 mg/kg,i.p.)1 h before,1 week before,immediately after,or 22 h after fear conditioning.The behavioral measure of fear(indicated by freezing level)and synaptic protein expression in the basolateral amygdala(BLA)and inferior-limbic pre-frontal cortex(IL-PFC)of mice were examined.We found that only ketamine administered 22 h after fear conditioning significantly decreased the fear generalization,and the effect was dose-dependent and lasted for at least 2 weeks.The fear-generalized mice showed a lower level of brainderived neurotrophic factor(BDNF)and a higher level of GluN2B protein in the BLA and IL-PFC,and this was reversed by a single administration of ketamine.Moreover,the GluN2B antagonist ifenprodil decreased the fear generalization when infused into the IL-PFC,but had no effect when infused into the BLA.Infusion of ANA-12(an antagonist of the BDNF receptor TrkB)into the BLA or ILPFC blocked the effect of ketamine on fear generalization.These findings support the conclusion that a single dose of ketamine administered 22 h after fear conditioning alleviates the fear memory generalization in mice and the GluN2B-related BDNF signaling pathway plays an important role in the alleviation of fear generalization.展开更多
Activation of N-methyl-D-aspartate receptors（NMDARs）mediates changes in the phosphorylation status of the glutamate receptors themselves.Previous studies have indicated that during synaptic activity,tyrosine kinases...Activation of N-methyl-D-aspartate receptors（NMDARs）mediates changes in the phosphorylation status of the glutamate receptors themselves.Previous studies have indicated that during synaptic activity,tyrosine kinases（Src and Fyn）or phosphatases（PTPαand STEP）are involved in regulating the phosphorylation of NMDARs.In this study,we used immunoblotting to investigate the role of an NMDAR subpopulation on the phosphorylation level of the GluN2B subunit at the Y1336 and Y1472sites in rat brain slices after NMDA treatment.We found that NMDA stimulation dramatically decreased the phosphorylation level of GluN2B at Y1472 in a dose-and time-dependent manner,but not at Y1336.Extrasynaptic NMDAR activation did not reduce the phosphorylation of GluN2B at Y1472.In addition,ifenprodil,a selective antagonist of GluN2Bcontaining NMDARs,did not abolish the decreased phosphorylation of GluN2B at Y1472 triggered by NMDA.These results suggest that the activation of synaptic GluN2A-containing NMDARs is required for the decreased phosphorylation of GluN2B at Y1472that is induced by NMDA treatment in rat brain slices.展开更多
本研究旨在探讨触液核GluN2B-BDNF通路在神经病理性疼痛中的作用。应用侧脑室注射特异性触液核示踪剂霍乱毒素亚单位B与辣根过氧化物酶复合物(cholera toxin subunit B conjugated with horseradish peroxidase,CB-HRP)的方法标记触液核...本研究旨在探讨触液核GluN2B-BDNF通路在神经病理性疼痛中的作用。应用侧脑室注射特异性触液核示踪剂霍乱毒素亚单位B与辣根过氧化物酶复合物(cholera toxin subunit B conjugated with horseradish peroxidase,CB-HRP)的方法标记触液核;通过免疫荧光双标染色和Western blot观察大鼠触液核GluN2B和BDNF的表达;采用坐骨神经慢性压迫性损伤法(chronic constriction injury of sciatic nerve,CCI)建立大鼠慢性神经病理性疼痛模型;通过侧脑室注射GluN2B拮抗剂和BDNF中和抗体观察CCI大鼠的行为学变化。结果显示,GluN2B和BDNF均在触液核内表达,并且在CCI大鼠表达上调;侧脑室注射GluN2B拮抗剂或BDNF中和抗体能够减轻CCI大鼠的热痛觉过敏和机械性痛觉超敏;而且侧脑室注射GluN2B拮抗剂能够逆转CCI大鼠BDNF的表达上调。以上结果提示,大鼠触液核内有GluN2B和BDNF的表达,并且触液核GluN2B-BDNF通路参与了大鼠神经病理性疼痛的发生。展开更多
Background Synaptic degeneration occurs in the early stage of Alzheimer’s disease(AD)before devastating symptoms,strongly correlated with cognitive decline.Circular RNAs(circRNAs)are abundantly enriched in neural tis...Background Synaptic degeneration occurs in the early stage of Alzheimer’s disease(AD)before devastating symptoms,strongly correlated with cognitive decline.Circular RNAs(circRNAs)are abundantly enriched in neural tissues,and aberrant expression of circRNAs precedes AD symptoms,significantly correlated with clinical dementia severity.However,the direct relationship between circRNA dysregulation and synaptic impairment in the early stage of AD remains poorly understood.Methods Hippocampal whole-transcriptome sequencing was performed to identify dysregulated circRNAs and miRNAs in 4-month-old wild-type and APP/PS1 mice.RNA antisense purification and mass spectrometry were utilized to unveil interactions between circRIMS2 and methyltransferase 3,N6-adenosine-methyltransferase complex catalytic subunit(METTL3).The roles of circRIMS2/miR-3968 in synaptic targeting of UBE2K-mediated ubiquitination of GluN2B subunit of NMDA receptor were evaluated via numerous lentiviruses followed by morphological staining,co-immunoprecipitation and behavioral testing.Further,a membrane-permeable peptide was used to block the ubiquitination of K1082 on GluN2B in AD mice.Results circRIMS2 was significantly upregulated in 4-month-old APP/PS1 mice,which was mediated by METTL3-dependent N6-methyladenosine(m6A)modification.Overexpression of circRIMS2 led to synaptic and memory impairments in 4-month-old C57BL/6 mice.MiR-3968/UBE2K was validated as the downstream of circRIMS2.Elevated UBE2K induced synaptic dysfunction of AD through ubiquitinating K1082 on GluN2B.Silencing METTL3 or blocking the ubiquitination of K1082 on GluN2B with a short membrane-permeable peptide remarkably rescued synaptic dysfunction in AD mice.Conclusions In conclusion,our study demonstrated that m6A-modified circRIMS2 mediates the synaptic and memory impairments in AD by activating the UBE2K-dependent ubiquitination and degradation of GluN2B via sponging miR-3968,providing novel therapeutic strategies for AD.展开更多
To stop the progression of Alzheimer＇s disease in the early stage, it is necessary to identify new therapeutic targets. We examined striatal-enriched phosphatase 61 expression in the brain tissues of 12-month-old APP...To stop the progression of Alzheimer＇s disease in the early stage, it is necessary to identify new therapeutic targets. We examined striatal-enriched phosphatase 61 expression in the brain tissues of 12-month-old APPswe/PSEN1dE9 transgenic mice. Immunohistochemistry showed that striatal-enriched phosphatase 61 protein expression was significantly increased but phosphorylated N-methyl-D-aspartate receptor 2B levels were significantly decreased in the cortex and hippocampus of APPswe/PSEN1dE9 transgenic mice. Western blotting of a cell model of Alzheimer＇s disease consisting of amyloid-beta peptide （1-42）-treated C57BL/6 mouse cortical neurons in vitro showed that valeric acid （AP5）, an N-methyl-D-aspartate receptor antagonist, significantly inhibited amyloidbeta 1-42-induced increased activity of striatal-enriched phosphatase 61. In addition, the phosphorylation of N-methyl-D-aspartate receptor 2B at Tyr1472 was impaired in amyloid-beta 1-42-treated cortical neurons, but knockdown of striatal-enriched phosphatase 61 enhanced the phosphorylation of N-methyl-D-aspartate receptor 2B. Collectively, these findings indicate that striatal-enriched phosphatase 61 can disturb N-methyl-D-aspartate receptor transport and inhibit the progression of learning and study disturbances induced by Alzheimer＇s disease. Thus, striatal-enriched phosphatase 61 may represent a new target for inhibiting the progression of Alzheimer＇s disease.展开更多
N-methyI-D-aspartate receptors （NMDARs） containing different GluN2 subunits play distinct roles in synaptic plasticity. Such differences may not only be determined by the channel properties, but also by differential...N-methyI-D-aspartate receptors （NMDARs） containing different GluN2 subunits play distinct roles in synaptic plasticity. Such differences may not only be determined by the channel properties, but also by differential surface distribution and synaptic localization. In the present study, using a Cy3-conjugated Fab fragment of the GFP antibody to label surface-located GluN2 subunits tagged with GFP at the N-terminus, we observed the membrane distribution patterns of GluN2A- or GluN2B-containing NMDARs in cultured rat hippocampal neurons. We found that surface NMDARs containing GluN2A, but not those containing GluN2B, were inclined to cluster at DIV7. Swapping the carboxyl termini of the GluN2 subunits completely reversed these distribution patterns. In addition, surface NMDARs containing GluN2A were preferentially associated with PSD-95. Taken together, the results of our study suggest that the clustering distribution of GluN2A- containing NMDARs is determined by the GluN2A C-terminus, and its interaction with PSD-95 plays an important role in this process.展开更多
钙/钙调蛋白依赖的蛋白激酶Ⅱ(Ca2+/calmodulin-dependent protein kinase Ⅱ,CaMKⅡ)在脑内兴奋性突触部位丰富表达。通过催化谷氨酸受体和众多突触蛋白磷酸化,CaMKⅡ调节磷酸化蛋白在基础或细胞兴奋时的转运、分布和功能。谷氨酸NMDA...钙/钙调蛋白依赖的蛋白激酶Ⅱ(Ca2+/calmodulin-dependent protein kinase Ⅱ,CaMKⅡ)在脑内兴奋性突触部位丰富表达。通过催化谷氨酸受体和众多突触蛋白磷酸化,CaMKⅡ调节磷酸化蛋白在基础或细胞兴奋时的转运、分布和功能。谷氨酸NMDA受体是CaMKⅡ的直接底物,有证据表明CaMKⅡ直接与NMDA受体胞内C末端相互结合,催化一特定丝氨酸(S1303)的磷酸化。CaMKⅡ也加强谷氨酸AMPA受体的磷酸化,通过磷酸化AMPA受体C末端特定的丝氨酸(S831),CaMKⅡ增强AMPA受体的功能。此外,CaMKⅡ可与代谢型谷氨酸受体mGluR1亚型的胞内C末端结合,促进一特定苏氨酸(T871)的磷酸化,从而促进受体兴奋后脱敏。CaMKⅡ在正常状态下与mGluR5受体结合以储存于突触内,刺激mGluR5受体时,CaMKⅡ与mGluR5受体分离,转运至NMDA受体,以介导mGluR5信号对NMDA受体的增强作用。总之,CaMKⅡ与谷氨酸受体相互作用,改变受体磷酸化水平,参与受体的数量和功能以及突触传导活动的调节。展开更多
基金supported by grants from the National Basic Research Development Program of China(2010CB912002)the National Natural Science Foundation of China(30730038 and 81171164)
文摘N-Methyl-D-aspartate receptors（NMDARs） play vital roles in the central nervous system,as they are primary mediators of Ca2＋influx during synaptic activity.The subunits that compose NMDARs share similar topological structures but are distinct in distribution and pharmacological properties,as well as physiological and pathological functions,which make the NMDAR one of the most complex and elusive ionotropic glutamate receptors.In this review,we focus on GluN2A and GluN2B,the primary NMDAR subunits in the cortex and hippocampus,and discuss their differences in developmental expression,brain distribution,trafficking,and functional properties during neuronal activity.
基金supported by grants from the National Natural Science Foundation of China(81530061 and 81471829)the Pearl River Nova Program of Guangzhou(201610010154)the Natural Science Foundation of Guangdong Province China(2017A030313095).
文摘Fear memories are critical for survival.Nevertheless,over-generalization of these memories,depicted by a failure to distinguish threats from safe stimuli,is typical in stress-related disorders.Previous studies have supported a protective role of ketamine against stress-induced depressive behavior.However,the effect of ketamine on fear generalization remains unclear.In this study,we investigated the effects of ketamine on fear generalization in a fear-generalized mouse model.The mice were given a single sub-anesthetic dose of ketamine(30 mg/kg,i.p.)1 h before,1 week before,immediately after,or 22 h after fear conditioning.The behavioral measure of fear(indicated by freezing level)and synaptic protein expression in the basolateral amygdala(BLA)and inferior-limbic pre-frontal cortex(IL-PFC)of mice were examined.We found that only ketamine administered 22 h after fear conditioning significantly decreased the fear generalization,and the effect was dose-dependent and lasted for at least 2 weeks.The fear-generalized mice showed a lower level of brainderived neurotrophic factor(BDNF)and a higher level of GluN2B protein in the BLA and IL-PFC,and this was reversed by a single administration of ketamine.Moreover,the GluN2B antagonist ifenprodil decreased the fear generalization when infused into the IL-PFC,but had no effect when infused into the BLA.Infusion of ANA-12(an antagonist of the BDNF receptor TrkB)into the BLA or ILPFC blocked the effect of ketamine on fear generalization.These findings support the conclusion that a single dose of ketamine administered 22 h after fear conditioning alleviates the fear memory generalization in mice and the GluN2B-related BDNF signaling pathway plays an important role in the alleviation of fear generalization.
基金supported by grants from the National Natural Science Foundation of China (30900418)the Natural Science Program of Department of Education of Zhejiang Province,China (Y201122469)
文摘Activation of N-methyl-D-aspartate receptors（NMDARs）mediates changes in the phosphorylation status of the glutamate receptors themselves.Previous studies have indicated that during synaptic activity,tyrosine kinases（Src and Fyn）or phosphatases（PTPαand STEP）are involved in regulating the phosphorylation of NMDARs.In this study,we used immunoblotting to investigate the role of an NMDAR subpopulation on the phosphorylation level of the GluN2B subunit at the Y1336 and Y1472sites in rat brain slices after NMDA treatment.We found that NMDA stimulation dramatically decreased the phosphorylation level of GluN2B at Y1472 in a dose-and time-dependent manner,but not at Y1336.Extrasynaptic NMDAR activation did not reduce the phosphorylation of GluN2B at Y1472.In addition,ifenprodil,a selective antagonist of GluN2Bcontaining NMDARs,did not abolish the decreased phosphorylation of GluN2B at Y1472 triggered by NMDA.These results suggest that the activation of synaptic GluN2A-containing NMDARs is required for the decreased phosphorylation of GluN2B at Y1472that is induced by NMDA treatment in rat brain slices.
基金supported by the National Natural Science Foundation of China(No.81571066)the Natural Science Foundation of Jiangsu Province,China(No.BE2015626)。
文摘本研究旨在探讨触液核GluN2B-BDNF通路在神经病理性疼痛中的作用。应用侧脑室注射特异性触液核示踪剂霍乱毒素亚单位B与辣根过氧化物酶复合物(cholera toxin subunit B conjugated with horseradish peroxidase,CB-HRP)的方法标记触液核;通过免疫荧光双标染色和Western blot观察大鼠触液核GluN2B和BDNF的表达;采用坐骨神经慢性压迫性损伤法(chronic constriction injury of sciatic nerve,CCI)建立大鼠慢性神经病理性疼痛模型;通过侧脑室注射GluN2B拮抗剂和BDNF中和抗体观察CCI大鼠的行为学变化。结果显示,GluN2B和BDNF均在触液核内表达,并且在CCI大鼠表达上调;侧脑室注射GluN2B拮抗剂或BDNF中和抗体能够减轻CCI大鼠的热痛觉过敏和机械性痛觉超敏;而且侧脑室注射GluN2B拮抗剂能够逆转CCI大鼠BDNF的表达上调。以上结果提示,大鼠触液核内有GluN2B和BDNF的表达,并且触液核GluN2B-BDNF通路参与了大鼠神经病理性疼痛的发生。
基金supported by the National Natural Science Foundation of China(82372337,81500925 to Xiong Wang,81801062 to Xiaoguang Li)Tongji Hospital(HUST)Foundation for Excellent Young Scientist(2020YQ01-11 to Xiong Wang)the Natural Science Foundation of Hubei Province(2022CFB150 to Huijun Li).
文摘Background Synaptic degeneration occurs in the early stage of Alzheimer’s disease(AD)before devastating symptoms,strongly correlated with cognitive decline.Circular RNAs(circRNAs)are abundantly enriched in neural tissues,and aberrant expression of circRNAs precedes AD symptoms,significantly correlated with clinical dementia severity.However,the direct relationship between circRNA dysregulation and synaptic impairment in the early stage of AD remains poorly understood.Methods Hippocampal whole-transcriptome sequencing was performed to identify dysregulated circRNAs and miRNAs in 4-month-old wild-type and APP/PS1 mice.RNA antisense purification and mass spectrometry were utilized to unveil interactions between circRIMS2 and methyltransferase 3,N6-adenosine-methyltransferase complex catalytic subunit(METTL3).The roles of circRIMS2/miR-3968 in synaptic targeting of UBE2K-mediated ubiquitination of GluN2B subunit of NMDA receptor were evaluated via numerous lentiviruses followed by morphological staining,co-immunoprecipitation and behavioral testing.Further,a membrane-permeable peptide was used to block the ubiquitination of K1082 on GluN2B in AD mice.Results circRIMS2 was significantly upregulated in 4-month-old APP/PS1 mice,which was mediated by METTL3-dependent N6-methyladenosine(m6A)modification.Overexpression of circRIMS2 led to synaptic and memory impairments in 4-month-old C57BL/6 mice.MiR-3968/UBE2K was validated as the downstream of circRIMS2.Elevated UBE2K induced synaptic dysfunction of AD through ubiquitinating K1082 on GluN2B.Silencing METTL3 or blocking the ubiquitination of K1082 on GluN2B with a short membrane-permeable peptide remarkably rescued synaptic dysfunction in AD mice.Conclusions In conclusion,our study demonstrated that m6A-modified circRIMS2 mediates the synaptic and memory impairments in AD by activating the UBE2K-dependent ubiquitination and degradation of GluN2B via sponging miR-3968,providing novel therapeutic strategies for AD.
文摘目的观察电针“百会”“神庭”对血管性痴呆(vascular dementia,VD)大鼠学习和记忆功能的影响,并从突触结构及突触相关蛋白表达水平的角度揭示其作用机制。方法将35只雄性SD大鼠随机分为假手术组、模型组、电针穴位组、电针非穴位组和奥拉西坦组,每组7只。采用改良双侧颈动脉结扎模型,电针穴位组大鼠选择“百会”“神庭”两穴治疗,电针非穴位组大鼠选择固定非穴位刺激,每次电针30 min,每日1次,连续干预14 d;奥拉西坦组大鼠选择腹腔注射奥拉西坦,50 mg/kg,每日1次,连续14 d。采用Morris水迷宫检测各组大鼠学习和空间记忆能力;透射电子显微镜观察各组大鼠海马CA1区突触结构;Western blot检测各组大鼠海马突触后致密蛋白95(postsynaptic density protein 95,PSD95)、GluA1、GluN2B和磷酸化GluN2B蛋白表达水平。结果与假手术组比较,模型组大鼠学习期逃避潜伏时间延长,测试期跨越平台次数减少,目标象限停留时间显著缩短,大脑质量显著增加,海马CA1区突触结构数明显减少,海马PSD95、GluA1、GluN2B和磷酸化GluN2B蛋白表达水平均显著降低,差异均有统计学意义(P<0.05);与模型组比较,电针穴位组大鼠的学习期逃避潜伏时间缩短,测试期跨越平台次数增加,目标象限停留时间延长,大脑质量降低,CA1区突触结构数增多,海马PSD95、GluA1、GluN2B和磷酸化GluN2B蛋白表达水平增加,差异均有统计学意义(P<0.05)。结论电针“百会”“神庭”能改善VD大鼠的学习记忆功能,改变海马突触结构,分子机制可能和增加突触蛋白PSD95、GluA1和GluN2B的蛋白表达水平相关。
文摘To stop the progression of Alzheimer＇s disease in the early stage, it is necessary to identify new therapeutic targets. We examined striatal-enriched phosphatase 61 expression in the brain tissues of 12-month-old APPswe/PSEN1dE9 transgenic mice. Immunohistochemistry showed that striatal-enriched phosphatase 61 protein expression was significantly increased but phosphorylated N-methyl-D-aspartate receptor 2B levels were significantly decreased in the cortex and hippocampus of APPswe/PSEN1dE9 transgenic mice. Western blotting of a cell model of Alzheimer＇s disease consisting of amyloid-beta peptide （1-42）-treated C57BL/6 mouse cortical neurons in vitro showed that valeric acid （AP5）, an N-methyl-D-aspartate receptor antagonist, significantly inhibited amyloidbeta 1-42-induced increased activity of striatal-enriched phosphatase 61. In addition, the phosphorylation of N-methyl-D-aspartate receptor 2B at Tyr1472 was impaired in amyloid-beta 1-42-treated cortical neurons, but knockdown of striatal-enriched phosphatase 61 enhanced the phosphorylation of N-methyl-D-aspartate receptor 2B. Collectively, these findings indicate that striatal-enriched phosphatase 61 can disturb N-methyl-D-aspartate receptor transport and inhibit the progression of learning and study disturbances induced by Alzheimer＇s disease. Thus, striatal-enriched phosphatase 61 may represent a new target for inhibiting the progression of Alzheimer＇s disease.
基金supported by grants from the National Basic Research Program of China (2010CB912002)the National Natural Science Foundation of China (81221003 and 81271453)Fundamental Research Funds for the Central Universities of China
文摘N-methyI-D-aspartate receptors （NMDARs） containing different GluN2 subunits play distinct roles in synaptic plasticity. Such differences may not only be determined by the channel properties, but also by differential surface distribution and synaptic localization. In the present study, using a Cy3-conjugated Fab fragment of the GFP antibody to label surface-located GluN2 subunits tagged with GFP at the N-terminus, we observed the membrane distribution patterns of GluN2A- or GluN2B-containing NMDARs in cultured rat hippocampal neurons. We found that surface NMDARs containing GluN2A, but not those containing GluN2B, were inclined to cluster at DIV7. Swapping the carboxyl termini of the GluN2 subunits completely reversed these distribution patterns. In addition, surface NMDARs containing GluN2A were preferentially associated with PSD-95. Taken together, the results of our study suggest that the clustering distribution of GluN2A- containing NMDARs is determined by the GluN2A C-terminus, and its interaction with PSD-95 plays an important role in this process.
文摘钙/钙调蛋白依赖的蛋白激酶Ⅱ(Ca2+/calmodulin-dependent protein kinase Ⅱ,CaMKⅡ)在脑内兴奋性突触部位丰富表达。通过催化谷氨酸受体和众多突触蛋白磷酸化,CaMKⅡ调节磷酸化蛋白在基础或细胞兴奋时的转运、分布和功能。谷氨酸NMDA受体是CaMKⅡ的直接底物,有证据表明CaMKⅡ直接与NMDA受体胞内C末端相互结合,催化一特定丝氨酸(S1303)的磷酸化。CaMKⅡ也加强谷氨酸AMPA受体的磷酸化,通过磷酸化AMPA受体C末端特定的丝氨酸(S831),CaMKⅡ增强AMPA受体的功能。此外,CaMKⅡ可与代谢型谷氨酸受体mGluR1亚型的胞内C末端结合,促进一特定苏氨酸(T871)的磷酸化,从而促进受体兴奋后脱敏。CaMKⅡ在正常状态下与mGluR5受体结合以储存于突触内,刺激mGluR5受体时,CaMKⅡ与mGluR5受体分离,转运至NMDA受体,以介导mGluR5信号对NMDA受体的增强作用。总之,CaMKⅡ与谷氨酸受体相互作用,改变受体磷酸化水平,参与受体的数量和功能以及突触传导活动的调节。
