Major depressive disorder(MDD)is a common neuropsychiatric disorder characterized by diverse symptoms.There are big limitations of clinic medicine which highlighted an urgent and clear need for more efficacious and fa...Major depressive disorder(MDD)is a common neuropsychiatric disorder characterized by diverse symptoms.There are big limitations of clinic medicine which highlighted an urgent and clear need for more efficacious and faster-acting therapeutic agents to treat patients with MDD,especially those who are refractory to the traditional antidepressants.In the present study,we assessed a novel compound,YY-21,from timosaponin B-Ⅲ derived from sarsasapogenin of Anemarrhenae Rhizoma.We found that YY-21 obviously increased presynaptic glutamate release and enhanced long-term synaptic activity within 10 min as determined by excitatory postsynaptic current(EPSC) and field excitatory postsynaptic potential(fEPSP) in medial prefrontal cortex(mPFC) slices.YY-21 demonstrated anxiolytic-like effects following acute administration in animals and reversed the depressivelike and anxiety phenotypes induced by chronic unpredictable mild stress(CMS) with a relatively fast therapeutic onset.Our mechanism research reveals that NMDA receptors and two-pore domain potassium(K2P)(TREK1) channels emerged as new drug targets for faster acting antidepressants.K2 P channels generate leak currents that are responsible the maintenance of resting membrane potential.They are potential targets for the treatment of multiple diseases.Here we identify TKDC,an inhibitor of the TREK subfamily,including TREK1,TREK2 and TRAAK channels.Using TKDC as a chemical probe,a combined study of computations,mutagenesis,and electrophysiology reveal an allosteric ligand-binding site in the extracellular cap of the channels.The molecular dynamics simulations suggest that ligand-induced allosteric conformational transitions cause a blockage of the ion conductive pathway.The identification of the extracellular ligand-binding site is confirmed by the discovery of new inhibitors targeting this site using virtual screening.These results suggest that the extracellular cap of a K2P channel can act as a new allosteric site and may serve as a direct drug target.展开更多
目的:观察弱内向整流相关及花生四烯酸激活的弱内向整流相关双孔钾离子通道(tandem of pore domains in a weak inward rectifying related-tandem of pore domains in a weak inward rectifying related arachidonic acid activated tw...目的:观察弱内向整流相关及花生四烯酸激活的弱内向整流相关双孔钾离子通道(tandem of pore domains in a weak inward rectifying related-tandem of pore domains in a weak inward rectifying related arachidonic acid activated two pore-domain potassium channels, TREK-TRAAK K2P)激活对氧化损伤的人视网膜色素上皮(retinal pigment epithelial, RPE)细胞吞噬功能的影响。方法:免疫荧光法检测TREK-1、TREK-2及TRAAK通道蛋白在人RPE细胞的表达,以不同时间和浓度梯度的叔丁基过氧化氢(tertbutyl hydroperoxide, t-BHP)诱导人RPE细胞氧化损伤。分对照组、t-BHP组、利鲁唑加t-BHP组,利鲁唑组,向各组加入2×107/mL的荧光微球或FITC标记猪感光细胞外节膜盘孵育6 h,固定染色后荧光显微镜拍照,Image-Pro Plus 6.0软件分析计算RPE细胞吞噬率和吞噬指数。结果:TREK-1、TREK-2、TREEK通道蛋白亚型在人RPE细胞质中均高表达。人RPE细胞在t-BHP干预后存活率呈浓度和时间依赖性,200μmol/L t-BHP干预6 h与对照组存活率比较差异无统计学意义(P>0.05),400μmol/L t-BHP干预6 h致半数死亡。特异性吞噬指数:t-BHP组低于其他各组(P<0.001),而利鲁唑组高于对照组,差异无统计学意义(P>0.05)。特异性吞噬率:t-BHP组低于其他各组,差异无统计学意义(P>0.05)。非特异性吞噬指数:t-BHP组低于其他各组(P<0.001);非特异性吞噬率:t-BHP组低于利鲁唑加t-BHP组,差异无统计学意义(P>0.05),两两比较,仅利鲁唑组高于t-BHP组(P<0.05)。结论:激活TREK-TRAAK K2P通道可保护氧化损伤人RPE细胞的吞噬功能。展开更多
Two-pore domain potassium (K2P) channels gate primarily within the selectivity filter, termed ‘C-type’ gating. Due to the lack of structural insights into the nonconductive (closed) state, ‘C-type’ gating mechanis...Two-pore domain potassium (K2P) channels gate primarily within the selectivity filter, termed ‘C-type’ gating. Due to the lack of structural insights into the nonconductive (closed) state, ‘C-type’ gating mechanisms remain elusive. Here, molecular dynamics (MD) simulations on TREK-1, a K2P channel, revealed that M4 helix movements induce filter closing in a novel ‘deeper-down’ structure that represents a ‘C-type’ closed state. The ‘down’ structure does not represent the closed state as previously proposed and instead acts as an intermediate state in gating. The study identified the allosteric ‘seesaw’ mechanism of M4 helix movements in modulating filter closing. Finally, guided by this recognition of K2P gating mechanisms, MD simulations revealed that gain-of-function mutations and small-molecule activators activate TREK-1 by perturbing state transitions from open to closed states. Together, we reveal a ‘C-type’ closed state and provide mechanical insights into gating procedures and allosteric regulations for K2P channels.展开更多
With the support of the National Natural Science Foundation of China,a collaborative study by the research groups led by Prof.Yang Huaiyu(阳怀宇)from the East China Normal University and Prof.Li Yang(李扬)from Shangha...With the support of the National Natural Science Foundation of China,a collaborative study by the research groups led by Prof.Yang Huaiyu(阳怀宇)from the East China Normal University and Prof.Li Yang(李扬)from Shanghai Institute of Materia Medica,Chinese Academy of Sciences展开更多
文摘Major depressive disorder(MDD)is a common neuropsychiatric disorder characterized by diverse symptoms.There are big limitations of clinic medicine which highlighted an urgent and clear need for more efficacious and faster-acting therapeutic agents to treat patients with MDD,especially those who are refractory to the traditional antidepressants.In the present study,we assessed a novel compound,YY-21,from timosaponin B-Ⅲ derived from sarsasapogenin of Anemarrhenae Rhizoma.We found that YY-21 obviously increased presynaptic glutamate release and enhanced long-term synaptic activity within 10 min as determined by excitatory postsynaptic current(EPSC) and field excitatory postsynaptic potential(fEPSP) in medial prefrontal cortex(mPFC) slices.YY-21 demonstrated anxiolytic-like effects following acute administration in animals and reversed the depressivelike and anxiety phenotypes induced by chronic unpredictable mild stress(CMS) with a relatively fast therapeutic onset.Our mechanism research reveals that NMDA receptors and two-pore domain potassium(K2P)(TREK1) channels emerged as new drug targets for faster acting antidepressants.K2 P channels generate leak currents that are responsible the maintenance of resting membrane potential.They are potential targets for the treatment of multiple diseases.Here we identify TKDC,an inhibitor of the TREK subfamily,including TREK1,TREK2 and TRAAK channels.Using TKDC as a chemical probe,a combined study of computations,mutagenesis,and electrophysiology reveal an allosteric ligand-binding site in the extracellular cap of the channels.The molecular dynamics simulations suggest that ligand-induced allosteric conformational transitions cause a blockage of the ion conductive pathway.The identification of the extracellular ligand-binding site is confirmed by the discovery of new inhibitors targeting this site using virtual screening.These results suggest that the extracellular cap of a K2P channel can act as a new allosteric site and may serve as a direct drug target.
文摘目的:观察弱内向整流相关及花生四烯酸激活的弱内向整流相关双孔钾离子通道(tandem of pore domains in a weak inward rectifying related-tandem of pore domains in a weak inward rectifying related arachidonic acid activated two pore-domain potassium channels, TREK-TRAAK K2P)激活对氧化损伤的人视网膜色素上皮(retinal pigment epithelial, RPE)细胞吞噬功能的影响。方法:免疫荧光法检测TREK-1、TREK-2及TRAAK通道蛋白在人RPE细胞的表达,以不同时间和浓度梯度的叔丁基过氧化氢(tertbutyl hydroperoxide, t-BHP)诱导人RPE细胞氧化损伤。分对照组、t-BHP组、利鲁唑加t-BHP组,利鲁唑组,向各组加入2×107/mL的荧光微球或FITC标记猪感光细胞外节膜盘孵育6 h,固定染色后荧光显微镜拍照,Image-Pro Plus 6.0软件分析计算RPE细胞吞噬率和吞噬指数。结果:TREK-1、TREK-2、TREEK通道蛋白亚型在人RPE细胞质中均高表达。人RPE细胞在t-BHP干预后存活率呈浓度和时间依赖性,200μmol/L t-BHP干预6 h与对照组存活率比较差异无统计学意义(P>0.05),400μmol/L t-BHP干预6 h致半数死亡。特异性吞噬指数:t-BHP组低于其他各组(P<0.001),而利鲁唑组高于对照组,差异无统计学意义(P>0.05)。特异性吞噬率:t-BHP组低于其他各组,差异无统计学意义(P>0.05)。非特异性吞噬指数:t-BHP组低于其他各组(P<0.001);非特异性吞噬率:t-BHP组低于利鲁唑加t-BHP组,差异无统计学意义(P>0.05),两两比较,仅利鲁唑组高于t-BHP组(P<0.05)。结论:激活TREK-TRAAK K2P通道可保护氧化损伤人RPE细胞的吞噬功能。
基金This work was supported in part by the Ministry of Science and Technology(2018YFA0508100 to Q.Z.and J.G.)the National Natural Science Foundation of China(31800699 to Q.Z.)the Fundamental Research Funds for the Central Universities,and the‘XingFuZhiHua’funding of ECNU(44300-19311-542500/006 to H.Y.).
文摘Two-pore domain potassium (K2P) channels gate primarily within the selectivity filter, termed ‘C-type’ gating. Due to the lack of structural insights into the nonconductive (closed) state, ‘C-type’ gating mechanisms remain elusive. Here, molecular dynamics (MD) simulations on TREK-1, a K2P channel, revealed that M4 helix movements induce filter closing in a novel ‘deeper-down’ structure that represents a ‘C-type’ closed state. The ‘down’ structure does not represent the closed state as previously proposed and instead acts as an intermediate state in gating. The study identified the allosteric ‘seesaw’ mechanism of M4 helix movements in modulating filter closing. Finally, guided by this recognition of K2P gating mechanisms, MD simulations revealed that gain-of-function mutations and small-molecule activators activate TREK-1 by perturbing state transitions from open to closed states. Together, we reveal a ‘C-type’ closed state and provide mechanical insights into gating procedures and allosteric regulations for K2P channels.
文摘With the support of the National Natural Science Foundation of China,a collaborative study by the research groups led by Prof.Yang Huaiyu(阳怀宇)from the East China Normal University and Prof.Li Yang(李扬)from Shanghai Institute of Materia Medica,Chinese Academy of Sciences
