用Fura－2双波长荧光法测定神经细胞内游离钙

采用新型Ca²⁺荧光指示剂Fura-2建立双波长荧光法测定分离的大鼠神经细胞内游离钙浓度([Ca²⁺]_i).结果显示,在静息状态下,其[Ca²⁺]_i为109±12nmol/L.30mmol/L KCl可显著增加[Ca²⁺]_i,并且KCl的这种效应呈一定的剂量依赖关系,提示该法灵敏、可靠.     

脉冲电场对粒细胞内Ca2+浓度影响的动态变化

利用激光共聚焦扫描显微镜和装有CCD系统的荧光显微镜 ,研究在单脉冲电场作用下经fluo 3/AM标记的鸡胚小脑粒细胞内自由Ca²⁺浓度 ( [Ca ²⁺]_i)的动态变化过程 .结果表明 :在单个电脉冲作用下 ,细胞内Ca²⁺浓度立刻升高并达到其最大值 .Ca²⁺浓度升高的幅度以及升高的速率具有电场强度的依赖性 .当细胞外Ca²⁺被过量的EGTA络合或细胞膜上的Ca²⁺通道被La ³⁺堵塞后 ,细胞内的Ca²⁺浓度仍然升高 .细胞内不同区域的Ca²⁺浓度同时升高 ;两极内的Ca²⁺浓度早于胞体的Ca²⁺浓度达到最大值并迅速恢复 .     

豚鼠精子质膜Ca2+ -ATPase活性与精子顶体反应关系的研究

用生化测定法首次证实豚鼠精子质膜Ca²⁺-ATPase活性在精子获能和顶体反应过程中显著下降.Ca²⁺-ATPase抑制剂利尿酸(ethacrynic acid)抑制质膜Ca²⁺-ATPase活性,但钙调素(50μg/mL)的拮抗剂三氟拉嗪(TFP,200～500μmol/L)对该酶活性没有影响,说明钙调素不直接参与精子依赖于ATP的Ca²⁺的主动泵出.但钙调素与精子的Ca²⁺内流有关,钙调素拮抗剂TFP显著促进精子顶体反应和精子对Ca²⁺的摄入.Ca²⁺-ATPase抑制剂栎皮酮(quercetin)、原钒酸钠(sodiumorthovandate)、利尿磺胺(furosemide)和利尿酸均显著促进豚鼠精子的顶体反应,但却抑制精子对Ca²⁺的摄入,这无法用它们对质膜Ca²⁺-ATPase活性的抑制作用解释.推测这可能是由于Ca²⁺-ATPase抑制剂在抑制质膜Ca²⁺-ATPase活性的同时也抑制了顶体外膜或线粒体外膜上的该酶的活性,导致Ca²⁺在细胞质内的积累,进而通过负反馈机制抑制Ca²⁺进一步内流所致.另外,Ca²⁺-ATPase抑制剂对糖酵解的抑制作用也可能是Ca²⁺在细胞质中积累和抑制精子Ca²⁺摄入的原因.     

骨骼肌内质网Ca2+泵转运Ca2+的结构基础

Ca²⁺泵(Ca²⁺-ATPase)是调节细胞内Ca²⁺浓度的重要蛋白质之一. Ca²⁺泵在转运Ca²⁺的过程中经历一系列构象变化. 其中,E1状态为外向的Ca²⁺高亲和状态,E2状态则为内向的Ca²⁺低亲和状态. 目前,骨骼肌内质网Ca²⁺泵转运Ca²⁺过程中的几个中间状态,包括E1-2Ca²⁺,E1-ATP,E1-P-ADP,E2-Pi和E2状态的三维晶体结构已经解析. 介绍这几种状态的晶体结构,并分析Ca²⁺泵在执行功能过程中结构与功能的关系.     

共聚焦显微术测定大鼠和黄鼠心肌细胞游离Ca2+浓度与温度的关系

运用共聚焦激光扫描显微成像术对比研究非冬眠动物大鼠和冬眠动物黄鼠心肌细胞胞内Ca²⁺浓度 ( [Ca²⁺]_i)随温度的变化 .首先标定了不同温度下Ca²⁺探针indo 1的解离常数 ,提出并证明按α定态设定标定溶液pH值的必要性 .细胞荧光分析显示 ,大鼠心肌细胞 [Ca²⁺ ]_i 随温度降低显著上升 ,低温下频繁出现自发钙波 ,胞内发生钙超载 ;相比较冬眠动物黄鼠心肌细胞[Ca²⁺ ]_i  在相同条件下保持稳定 ,避免发生钙超载 .认识其中的钙稳态机制可能对有关医学问题有潜在的指导意义 .     

猪脑突触体质膜(Ca2+-Mg2+)-ATPase的分离纯化与性质

以猪脑为材料,经匀浆、差速离心、蔗糖密度梯度离心分离突触体. 低渗破膜得到突触体膜. Triton X-100增溶后,经钙调蛋白亲和层析可得去脂的质膜Ca²⁺-ATPase. 用大体积亲和柱和大体积低Ca²⁺淋洗液淋洗,可得产率、纯度和活性均较高的质膜Ca²⁺-ATPase. 与大豆磷脂保温后,去脂的Ca²⁺-ATPase的水解活力可恢复达3.32 μmol/(mg·min).SDS-聚丙烯酰胺凝胶电泳银染显示单一蛋白质带,分子质量约为140 ku,纯度在90%以上. 不同Ca²⁺浓度明显影响酶的活力.     

Ca2+在粟酒裂殖酵母细胞周期时相中的作用*

以粟酒裂殖酵母(Schizosaccharomyces pombe)为研究材料,研究了Ca²⁺在细胞周期时相中的作用。当外源Ca²⁺浓度在0.5-20 mmol/L范围内,随Ca²⁺浓度增加,细胞增殖速度加快,延滞期逐渐缩短。但SD-Ca（CaCl₂省略）并不能终止Sch. Pombe的细胞周期。采用缺氮对群体细胞进行同步化,并以EGTA 螯合培养介质中低浓度的Ca²⁺,Sch. Pombe 细胞增殖被完全抑制,细胞流式法测定结果表明：细胞周期被终止在G1期。分析认为Ca²⁺ 对Sch. Pombe 细胞增殖是必不可少的,外源Ca²⁺在G1期向S期转化过程中起着关键性的作用。     

亚硒酸钠对巨噬细胞内游离Ca2+和Mg2+的影响

用单细胞阳离子测定系统研究了SeO^2-₃对巨噬细胞内游离Ca²⁺和Mg²⁺的影响.实验结果表明:SeO^2-₃高于10^-4mol/L时,有显著的细胞毒性.SeO^2-₃对细胞的毒性作用使细胞内游离Ca²⁺和Mg²⁺的浓度升高但Ca²⁺浓度的升高速率比Mg²⁺快.还有,高于10^-4mol/L的SeO^2-₃对红细胞膜上的Ca²⁺-ATP酶活性有明显抑制作用.     

Gonadotropin releasing hormone stimulation of pituitary cell 3′–5′ cyclic AMP in a carp(Cyprinus carpio) is dependent on extracellular calcium

Pituitaries were collected from a common carp,yprinss carpi, belonging to vitellogenic phase and cells were disaggregated by using 0.3% collagenase and 0.05% tsypsin. Enzymatically dispersed cells were incubatedin vitro in Ca²⁺-free medium to observe the effect ofCanna punctatus GnRH (cGnRH) and Ca²⁺ on pituitary cell cAMP accumulation. Addition of cGnRH (20 Big) to pituitary cell incubation (6 × 10⁴ cells/well) containing 4 mM theophylline, a phosphodiesterase inhibitor, caused two-fold increase of cAMP accumulation in comparison to control, Addition of Ca²⁺ (2 mM) to cGnRH further augmented cAMP accumulation, i.e., four-fold as compared to control. Increasing concentrations of cGnRH in the presence of Ca²⁺ resulted in a dose-dependent increase in cAMP accumulation. To examine the specificity of Ca²⁺ augmentory effect on cGnRH-stimulated pituitary cell cAMP accumulation, a specific Ca²⁺-channel blocker, verapamil was used, At 3 μM dose verapamil completely waived Ca²⁺-augmentation of cGnRH stimulatory effect on cAMP. Interestingly, verapamil also significantly inhibited cGnRH stimulation of cAMP in the Ca²⁺-free medium. Extent of Ca²⁺ plus cGnRH stimulatory effect on cAMP was further increased by the addition of 25 pmol of calmodulin, a Ca²⁺-carrier protein, Addition of verapamil to this system strongly inhibited Ca²⁺ and ealmodulin augnientory effect on cGnRH. Reduced level of cAMP in the pituitary cell due to verapamil was even lower than that of cGnRH plus ealmodulin incubation. Data indicates a contamination of Ca²⁺ in an apparently Ca²⁺-free medium, Results suggest that in lower vertebrate, i.e., fish, GnRH stimulation of pituitary cell cAMP is dependent on extracellulnr Ca²⁺ and incubation of pituitary cell in Ca²⁺-free medium is truly not free of Ca²⁺.     

T－2毒素对心肌细胞三型钙通道的阻滞作用

用膜片钳连细胞电压钳法, 在培养的Wistar大鼠单个心肌细胞上记录了T-2毒素对B、L和T三型Ca²⁺通道单通道电活动的影响. 结果表明, T-2毒素浓度为10mg/L时, 心肌细胞B、L和T三型Ca²⁺通道均受到明显的阻滞, 其阻滞作用表现为使Ca²⁺通道的开放概率减小, 开放时间缩短, 关闭时间延长, 而对流过Ca²⁺通道的Ba²⁺流幅值无影响.     

Progress of Lead‐Free Halide Double Perovskites

The field of halide metal perovskite photovoltaics has caught widespread interest in the last decade. This is seen in the rapid rise of power conversion efficiency, which is currently over 23%. It has also stimulated a widespread application of halide metal perovskites in other fields, such as light‐emitting diodes, field‐effect transistors, detectors, and lasers. Despite the fascinating characteristics of the halide metal perovskites, the presence of toxic lead (Pb) in their chemical composition is regarded as one of the major limiting factors preventing their commercialization. Addressing the toxicity issues in these compounds by a careful and strategic replacement of Pb²⁺ with other nontoxic candidate elements represents a promising direction to fabricate lead‐free optoelectronic devices. Such attempts yield a halide double perovskite structure which allows flexibility for various compositional adjustments. Here, the authors present the current progress and setbacks in crystal structures, materials preparation, optoelectronic properties, stability, and photovoltaic applications of lead‐free halide double perovskite compounds. Prospective research directions to improve the optoelectronic properties of existing materials are given that may help in the discovery of new lead‐free halide double perovskites.     

Trinary Layered Double Hydroxides as High‐Performance Bifunctional Materials for Oxygen Electrocatalysis

Layered double hydroxides (LDHs) are a family of high‐profile layer materials with tunable metal species and interlayer spacing, and herein the LDHs are first investigated as bifunctional electrocatalysts. It is found that trinary LDH containing nickel, cobalt, and iron (NiCoFe‐LDH) shows a reasonable bifunctional performance, while exploiting a preoxidation treatment can significantly enhance both oxygen reduction reaction and oxygen evolution reaction activity. This phenomenon is attributed to the partial conversion of Co²⁺ to Co³⁺ state in the preoxidation step, which stimulates the charge transfer to the catalyst surface. The practical application of the optimized material is demonstrated with a small potential hysteresis (800 mV for a reversible current density of 20 mA cm^?2) as well as a high stability, exceeding the performances of noble metal catalysts (commercial Pt/C and Ir/C). The combination of the electrochemical metrics and the facile and cost‐effective synthesis endows the trinary LDH as a promising bifunctional catalyst for a variety of applications, such as next‐generation regenerative fuel cells or metal–air batteries.     

Interaction of Microwaves and a Temporally Incoherent Magnetic Field on Single and Double DNA Strand Breaks in Rat Brain Cells

The effect of a temporally incoherent magnetic field noise on microwave-induced DNA single and double strand breaks in rat brain cells was investigated. Four treatment groups of rats were studied: microwave-exposure (continuous-wave 2450-MHz microwaves, power density 1 mW/cm², average whole-body specific absorption rate of 0.6 W/kg), noise-exposure (45 mG), microwave + noise-exposure, and sham-exposure. Animals were exposed to these conditions for 2h. DNA single- and double-strand breaks in brain cells of these animals were assayed 4h later using a microgel electrophoresis assay. Results show that brain cells of microwave-exposed rats had significantly higher levels of DNA single- and double-strand breaks when compared with sham-exposed animals. Exposure to noise alone did not significantly affect the levels (i.e., they were similar to those of the sham-exposed rats). However, simultaneous noise exposure blocked microwave-induced increases in DNA strand breaks. These data indicate that simultaneous exposure to a temporally incoherent magnetic field could block microwave-induced DNA damage in brain cells of the rat.     

Double reduction estimation and equilibrium tests in natural autopolyploid populations

Many bioinformatics pipelines include tests for equilibrium. Tests for diploids are well studied and widely available, but extending these approaches to autopolyploids is hampered by the presence of double reduction, the comigration of sister chromatid segments into the same gamete during meiosis. Though a hindrance for equilibrium tests, double reduction rates are quantities of interest in their own right, as they provide insights about the meiotic behavior of autopolyploid organisms. Here, we develop procedures to (i) test for equilibrium while accounting for double reduction, and (ii) estimate the double reduction rate given equilibrium. To do so, we take two approaches: a likelihood approach, and a novel U-statistic minimization approach that we show generalizes the classical equilibrium χ² test in diploids. For small sample sizes and uncertain genotypes, we further develop a bootstrap procedure based on our U-statistic to test for equilibrium. We validate our methods on both simulated and real data.     

Tuning Electronic Structure of NiFe Layered Double Hydroxides with Vanadium Doping toward High Efficient Electrocatalytic Water Oxidation

Binary NiFe layer double hydroxide (LDH) serves as a benchmark non‐noble metal electrocatalyst for the oxygen evolution reaction, however, it still needs a relatively high overpotential to achieve the threshold current density. Herein the catalyst's electronic structure is tuned by doping vanadium ions into the NiFe LDHs laminate forming ternary NiFeV LDHs to reduce the onset potential, achieving unprecedentedly efficient electrocatalysis for water oxidation. Only 1.42 V (vs reversible hydrogen electrode (RHE), ≈195 mV overpotential) is required to achieve catalytic current density of 20 mA cm^?2 with a small Tafel slope of 42 mV dec^?1 in 1 m KOH solution, which manifests the best of NiFe‐based catalysts reported till now. Electrochemical analysis and density functional theory +U simulation indicate that the high catalytic activity of NiFeV LDHs mainly attributes to the vanadium doping which can modify the electronic structure and narrow the bandgap thereby bring enhanced conductivity, facile electron transfer, and abundant active sites.     

Europium doped Sr2YNbO6 double perovskite phosphor for photoluminescence and thermoluminescence properties

A luminescent double perovskite phosphor Sr₂YNbO₆ doped with Eu³⁺ crystallized to the monoclinic phase and was synthesized successfully via a conventional high-temperature combustion method. The formation of the crystal structure, phase purity, and surface morphology were studied using X-ray diffraction patterns and scanning electron microscopy. The characteristic vibrations between the atoms of the functional groups present in phosphor were studied using Fourier transform infrared spectra analysis. The luminescence properties of the prepared phosphors were investigated in terms of photoluminescence (PL) and thermoluminescence (TL). PL excitation spectra exhibited charge transfer bands and the characteristic 4f⁶ transitions of Eu³⁺. A prominent PL emission was obtained for the phosphor doped with 4 mol% Eu³⁺ under the 396 nm excitation wavelength. PL emission quenching was observed for the higher doping concentrations due to a multipole–multipole interaction. A highly intense PL emission arose due to the hypersensitive ⁵D₀–⁷F₂ electric dipole transition of Eu³⁺ that dominated the emission spectra. The thermal stability of the phosphor was examined through temperature-dependent PL. The TL properties of the Sr₂YNbO₆ double perovskites irradiated with a ⁹⁰Sr beta source at different doses were measured. The double perovskite phosphors under study showed a linear dose–response with increasing beta dose, ranging from 1 Gy to 10 Gy. Trapping parameters of the TL glow curves were determined using Chen's peak shape method and computerized glow curve deconvolution (CGCD). CGCD fitting of the TL glow curves revealed that it was consisted of three major peaks and followed second-order kinetics. The estimated activation energies were determined using different methods and were comparable and significant.     

Luminescence and dosimetry approach in terbium(III)-activated tungstate double perovskite

A series of tungstate double perovskite Ca₃WO₆ doped with Tb³⁺ was prepared by a combustion process using urea as a flux. The crystal structure identification of Ca₃WO₆:Tb³⁺ phosphors was done using X-ray diffraction patterns, and a monoclinic structure was discovered. The Fourier transform infrared spectrum of Ca₃WO₆:Tb³⁺ displayed characteristic vibrations of tungstate bonds. Under 278 nm excitation, Ca₃WO₆:Tb³⁺ exhibited intense downconversion green emission, which corresponded to the ⁵D₄–⁷F_J (J = 4,5) transitions of Tb³⁺. The phosphor exhibited the highest photoluminescence (PL) intensity when it was doped with 1 mol% of Tb³⁺; later intensity quenching appeared to be due to the multipolar interaction at higher dopant concentrations. Moreover, high-quality thermoluminescence (TL) was detected when phosphors were irradiated using beta rays. The effects of Tb³⁺ concentration and beta dose on TL intensity were the two major aspects studied in detail. The TL intensity demonstrated excellent linear response to the applied range of beta dose. The trap parameters of the studied phosphors were computed by the peak shape approach and glow curve deconvolution. The fading effect on TL intensity was studied by recording the TL glow curves after 1 month of beta irradiation. Obtained results from the PL and TL characterizations showed that the phosphors under study have the potential to be used in lighting displays and in thermoluminescence dosimetry.     

Processing of multi-dimensional NMR data with the new software PROSA

Summary The new program PROSA is an efficient implementation of the common data-processing steps for multi-dimensional NMR spectra. PROSA performs linear prediction, digital filtering, Fourier transformation, automatic phase correction, and baseline correction. High efficiency is achieved by avoiding disk storage of intermediate data and by the absence of any graphics display, which enables calculation in the batch mode and facilitates porting PROSA on a variety of different computer systems; including supercomputers. Furthermore, all time-consuming routines are completely vectorized. The elimination of a graphics display was made possible by the use of a new, reliable automatic phase-correction routine. CPU times for complete processing of a typical heteronuclear three-dimensional NMR data set of a protein vary between less than 1 min on a NEC SX3 supercomputer and 40 min on a Sun-4 computer system.     

双低两用温敏核不育水稻低温条件下花粉发育的细胞学观察

双低两用温敏核不育水稻96－5－2S在11.31－20.19℃低温条件下,花粉母细胞形成、减数分裂和花粉后期发育均正常,最后形成充满淀粉的成熟花粉,而作对照的两用温敏不育水稻陆18S在相同低温条件下,除花粉母细胞形成早期发育正常外,花粉母细胞晚期、减数分裂期均有异常,败育主要发生在单核小孢子靠边期,没有形成成熟花粉,结果表明,96－5－2S桫胜殖期抗寒性强,低温生理障碍雄性不育临界温度低。     

Double knockout of Akt2 and AMPK predisposes cardiac aging without affecting lifespan: Role of autophagy and mitophagy

Increased age often leads to a gradual deterioration in cardiac geometry and contractile function although the precise mechanism remains elusive. Both Akt and AMPK play an essential role in the maintenance of cardiac homeostasis. This study examined the impact of ablation of Akt2 (the main cardiac isoform of Akt) and AMPKα2 on development of cardiac aging and the potential mechanisms involved with a focus on autophagy. Cardiac geometry, contractile, and intracellular Ca²⁺ properties were evaluated in young (4-month-old) and old (12-month-old) wild-type (WT) and Akt2-AMPK double knockout mice using echocardiography, IonOptix® edge-detection and fura-2 techniques. Levels of autophagy and mitophagy were evaluated using western blot. Our results revealed that increased age (12 months) did not elicit any notable effects on cardiac geometry, contractile function, morphology, ultrastructure, autophagy and mitophagy, although Akt2-AMPK double knockout predisposed aging-related unfavorable changes in geometry (heart weight, LVESD, LVEDD, cross-sectional area and interstitial fibrosis), TEM ultrastructure, and function (fractional shortening, peak shortening, maximal velocity of shortening/relengthening, time-to-90% relengthening, intracellular Ca²⁺ release and clearance rate). Double knockout of Akt2 and AMPK unmasked age-induced cardiac autophagy loss including decreased Atg5, Atg7, Beclin1, LC3BII-to-LC3BI ratio and increased p62. Double knockout of Akt2 and AMPK also unmasked age-related loss in mitophagy markers PTEN-induced putative kinase 1 (Pink1), Parkin, Bnip3, and FundC1, the mitochondrial biogenesis cofactor PGC-1α, and lysosomal biogenesis factor TFEB. In conclusion, our data indicate that Akt2-AMPK double ablation predisposes cardiac aging possibly related to compromised autophagy and mitophagy. This article is part of a Special Issue entitled: Genetic and epigenetic regulation of aging and longevity edited by Jun Ren & Megan Yingmei Zhang.