Structural Insights into the Mechanisms and Pharmacology of K2P Potassium Channels

Leak currents, defined as voltage and time independent flows of ions across cell membranes, are central to cellular electrical excitability control. The K_2P (KCNK) potassium channel class comprises an ion channel family that produces potassium leak currents that oppose excitation and stabilize the resting membrane potential in cells in the brain, cardiovascular system, immune system, and sensory organs. Due to their widespread tissue distribution, K_2Ps contribute to many physiological and pathophysiological processes including anesthesia, pain, arrythmias, ischemia, hypertension, migraine, intraocular pressure regulation, and lung injury responses. Structural studies of six homomeric K_2Ps have established the basic architecture of this channel family, revealed key moving parts involved in K_2P function, uncovered the importance of asymmetric pinching and dilation motions in the K_2P selectivity filter (SF) C-type gate, and defined two K_2P structural classes based on the absence or presence of an intracellular gate. Further, a series of structures characterizing K_2P:modulator interactions have revealed a striking polysite pharmacology housed within a relatively modestly sized (~70 kDa) channel. Binding sites for small molecules or lipids that control channel function are found at every layer of the channel structure, starting from its extracellular side through the portion that interacts with the membrane bilayer inner leaflet. This framework provides the basis for understanding how gating cues sensed by different channel parts control function and how small molecules and lipids modulate K_2P activity. Such knowledge should catalyze development of new K_2P modulators to probe function and treat a wide range of disorders.     

基于网络药理学、分子对接和实验验证探讨朱茯苓用于失眠的作用机制

为探讨朱茯苓治疗失眠的可能作用机制,通过TCMSP、BAT-MAN、TCMID和STITCH数据库以及文献挖掘筛选朱茯苓的活性成分及潜在靶点,利用TTD、OMIM、GeneCards和CTD数据库获取失眠类疾病的相关靶点,采用Cytoscape软件和String数据库构建活性成分-靶点网络和靶点蛋白相互作用网络,通过BioGPS数据库进行靶点的器官定位,基于David数据库进行GO功能和KEGG通路的富集分析,利用Autodock_vina软件进行活性成分和核心靶点的分子对接验证,最后通过免疫印迹法(Western blot)验证朱茯苓对γ-氨基丁酸(GABA)的两种受体蛋白α1亚基基因GABRA1和γ2亚基基因GABRG2的影响。最终筛选得到朱茯苓活性成分33种,潜在靶点267个,与失眠的交集靶点36个,多作用于脑、心脏等器官;分子对接结果显示GABRA1、GABRG2两个靶点能够与活性成分自发结合并借助氢键等分子间作用力形成较为稳定的构象;富集分析共获得189个GO条目和24条KEGG通路,主要涉及GABA信号通路、5-羟色胺(5-HT)信号通路等;Western blot实验证明朱茯苓能够增强小鼠脑内GABRA1和GABRG2蛋白的表达。通过网络药理学、分子对接和Western blot实验验证,发现朱茯苓可能通过多成分、多靶点、多途径的协同作用发挥镇静安神的作用,为深入进行朱茯苓治疗失眠的作用机制研究提供新思路和新方法。     

基于网络药理学及分子对接的浙贝母花-枇杷花药对止咳化痰作用机制研究

采用网络药理学及分子对接阐明浙贝母花—枇杷花药对的止咳化痰作用机制。以口服生物利用度≥30%和类药性≥0.18筛选成分,Swiss Target Prediction数据库进行靶点预测,GenCLiP 3和Drugbank数据库进行疾病靶点分析。以STRING和Cytoscape软件构建成分—靶点互作图,并进行GO、KEGG分析。以AutoDock Vina和Discovery Studio Visualizer对成分与关键靶点蛋白进行分子对接及相互作用分析。结果表明,药对的主要活性成分为生物碱、三萜和黄酮,作用于PTGS2、AKT1、VEGFA、TNF等靶点,通过VEGF、花生四烯酸代谢、白介素-17、肿瘤坏死因子等信号通路,多成分、多靶点、多通路产生止咳化痰的作用。     

基于网络药理学-分子对接研究桑不同入药部位防治糖尿病的作用机制

采用网络药理学-分子对接研究桑不同入药部位防治糖尿病的作用机制,从TCMSP、TCMID等多个中药数据库获得桑不同部位桑叶、桑椹、桑枝中的成分信息,结合OMIM、TTD等疾病数据库获得糖尿病靶点信息,利用Cytoscape 3.7.2软件分别构建不同部位的“活性成分-疾病靶点”的复杂网络及拓扑分析,应用WebGestalt工具进行通路富集分析。使用Autodock vina软件对桑各入药部位主要活性成分与作用靶点进行分子对接。结果得到桑的不同入药部位主要有11个差异活性成分,调控MAPK8、AKT1、VEGFA、IL6、PPARG等32个核心靶点。靶点主要涉及急性炎症反应、有机氮化合物的反应、细胞增殖调控、对胰岛素刺激反应等生物过程。通过介素-17信号通路、肿瘤坏死因子信号通路、PI3K-AKT信号通路和MAPK信号通路等来发挥治疗糖尿病的作用。本研究通过网络药理学分析桑不同入药部位防治糖尿病的活性成分群及作用机制,为桑的不同入药部位防治糖尿病的作用关系给出了参考依据。     

Inhalation by design: dual pharmacology β-2 agonists/M3 antagonists for the treatment of COPD

This paper describes the successful design and development of dual pharmacology β-2 agonists-M3 antagonists, for the treatment of chronic obstructive pulmonary disorder using the principles of ‘inhalation by design’. A key feature of this work is the combination of balanced potency and pharmacodynamic duration with desirable pharmacokinetic and material properties, whilst keeping synthetic complexity to a minimum.     

绿绒蒿属植物化学成分及药理活性研究进展

绿绒蒿属植物主要化学成分有生物碱、黄酮和挥发油等,本文综述了近几十年来绿绒蒿属植物化学成分和药理活性的研究进展,为今后对该属植物进一步的研究提供参考。     

牛磺酸研究进展

牛磺酸具有抗肿瘤、增强免疫、保护心脏、降压、降血脂、降血糖、减轻脂肪肝、降转氨酶、抗衰老等诸多作用。本文查阅国内外相关文献,并将其分析归纳,综述了牛磺酸性质、药理作用的研究进展,以为含有牛磺酸的守宫及含守宫中成药的抗肿瘤等临床作用机理提供研究线索。     

A mutation in CLOCK leads to altered dopamine receptor function

Mice with a mutation in the Clock gene (ClockΔ19) have a number of behavioral phenotypes that suggest alterations in dopaminergic transmission. These include hyperactivity, increased exploratory behavior, and increased reward value for drugs of abuse. However, the complex changes in dopaminergic transmission that underlie the behavioral abnormalities in these mice remain unclear. Here we find that a loss of CLOCK function increases dopamine release and turnover in striatum as indicated by increased levels of metabolites HVA and DOPAC, and enhances sensitivity to dopamine receptor antagonists. Interestingly, this enlarged dopaminergic tone results in downstream changes in dopamine receptor (DR) levels with a surprising augmentation of both D1‐ and D2‐type DR protein, but a significant shift in the ratio of D1 : D2 receptors in favor of D2 receptor signaling. These effects have functional consequences for both behavior and intracellular signaling, with alterations in locomotor responses to both D1‐type and D2‐type specific agonists and a blunted response to cAMP activation in the ClockΔ19 mutants. Taken together, these studies further elucidate the abnormalities in dopaminergic transmission that underlie mood, activity, and addictive behaviors.     

Ex vivo Culturing of Whole,Developing Drosophila Brains

We describe a method for ex vivo culturing of whole Drosophila brains. This can be used as a counterpoint to chronic genetic manipulations for investigating the cell biology and development of central brain structures by allowing acute pharmacological interventions and live imaging of cellular processes. As an example of the technique, prior work from our lab¹ has shown that a previously unrecognized subcellular compartment lies between the axonal and somatodendritic compartments of axons of the Drosophila central brain. The development of this compartment, referred to as the axon initial segment (AIS)², was shown genetically to depend on the neuron-specific cyclin-dependent kinase, Cdk5. We show here that ex vivo treatment of wild-type Drosophila larval brains with the Cdk5-specific pharmacological inhibitors roscovitine and olomoucine³ causes acute changes in actin organization, and in localization of the cell-surface protein Fasciclin 2, that mimic the changes seen in mutants that lack Cdk5 activity genetically.A second example of the ex vivo culture technique is provided for remodeling of the connections of embryonic mushroom body (MB) gamma neurons during metamorphosis from larva to adult. The mushroom body is the center of olfactory learning and memory in the fly⁴, and these gamma neurons prune their axonal and dendritic branches during pupal development and then re-extend branches at a later timepoint to establish the adult innervation pattern⁵. Pruning of these neurons of the MB has been shown to occur via local degeneration of neurite branches⁶, by a mechanism that is triggered by ecdysone, a steroid hormone, acting at the ecdysone receptor B1⁷, and that is dependent on the activity of the ubiquitin-proteasome system⁶. Our method of ex vivo culturing can be used to interrogate further the mechanism of developmental remodeling. We found that in the ex vivo culture setting, gamma neurons of the MB recapitulated the process of developmental pruning with a time course similar to that in vivo. It was essential, however, to wait until 1.5 hours after puparium formation before explanting the tissue in order for the cells to commit irreversibly to metamorphosis; dissection of animals at the onset of pupariation led to little or no metamorphosis in culture. Thus, with appropriate modification, the ex vivo culture approach can be applied to study dynamic as well as steady state aspects of central brain biology.