A confocal study of mechanism of 5-hydroxytryptamino induced intracellular calcium dynamics in cultured rat stomach fundus smooth muscle cells with a new Ca2+ indicator STDln-AM

A new fluorescent Ca²⁺ indicator STDln-AM for detecting [Ca²⁺], transients in cultured smooth muscle cells is presented. By making a comparison, the difference between STDln and fluo-3 is discussed in detail. Using the new Ca²⁺ indicator, the mechanism of 5-hydroxytryptamino (5-HT) induced intracellular calcium dynamics in stomach fundus smooth muscle cells (SFSMC) of rats is investigated. It is shown that in contrast with fluo-3, STDln is uniformly distributed in the cytosolic compartment but excluded from the nucleus, when it is transfected into cells. This feature makes it a real cytosol Ca²⁺ indicator and can reflect changes of cytosol [Ca²⁺] more accurately than that of fluo-3. In addition, STDln responds to the [Ca²⁺], transients more sensitive and faster than fluo-3. The results also show that, the L-type Ca²⁺ channel inhibitor Mn9202 and the PLC inhibitor Compound 48/80 can significantly inhibit the [Ca²⁺], elevation induced by 5-HT, while the PKC inhibitor D-Sphingosine can enhance the effect of 5-HT. The results suggest that 5-HT acts by the way of 5-HT₂ receptors on SFSMC, then through 5-HT₂ receptors coupled IP₃/Ca²⁺ and GC/PKC double signal transduction pathways to make Ca²⁺ release from intracellular Ca²⁺ stores and Ca²⁺ influx possibly through L-type calcium channels.     

A confocal study of mechanism of 5-hydroxytryptamino induced intracellular calcium dynamics in cultured rat stomach fundus smooth muscle cells with a new Ca~(2+) indicator STDIn-AM

A new fluorescent Ca2+ indicator STDIn-AM for detecting [Ca2+]i transients in cultured smooth muscle cells is presented. By making a comparison, the difference between STDIn and fluo-3 is discussed in detail. Using the new Ca2+ indicator, the mechanism of 5-hydroxytryptamino (5-HT) induced intracellular calcium dynamics in stomach fundus smooth muscle cells (SFSMC) of rats is investigated. It is shown that in contrast with fluo-3, STDIn is uniformly distributed in the cytosolic compartment but excluded from the nucleus, when it is transfected into cells. This feature makes it a real cytosol Ca2+ indicator and can reflect changes of cytosol [Ca2+] more accurately than that of fluo-3. In addition, STDIn responds to the [Ca2+]i transients more sensitive and faster than fluo-3. The results also show that, the L-type Ca2+channel inhibitor Mn9202 and the PLC inhibitor Compound 48/80 can significantly inhibit the [Ca2+]i elevation induced by 5-HT, while the PKC inhibitor D-Sphingosine can enhance the effect of     

希土化合物对离体巨噬细胞的影响

本文应用细胞培养法和单细胞阳离子测定系统研究了希土化合物以地巨噬细胞的影响，结果表明，在培养介质中ＳｍＣｌ３和Ｙｃｌ３的浓度大于１ｍｍｏｌ．ｄｍ＾－３时，有明显的细胞毒性，Ｙｃｌ３的细胞毒性大于ＳｍＣｌ３，ＳｍＣｌ３和ＹＣｌ３的细胞毒性明显大于Ｓｍ（Ａｌａ）３Ｃｌ３和Ｙ（Ａｌａ）２Ｃｌ３。希土化合物的作用使细胞（Ｃａ＾２＋ｉ）升高；毒性越大，Ｃａ＾２＋ｉ升高越甚。低浓度Ｓｍ＾３＋和Ｙ６３＋对细胞膜     

Comparison of Ca2+ and Mg2+ binding to calmodulin. An enthalpy,entropy, and gibbs free energy analysis

A consistent set of G _B , H _B , and S _B parameters have been determined from ion specific electrode, calorimetric, and spectrophotometric studies for the binding of Ca²⁺ and Mg²⁺ to bovine calmodulin at pH=7.0 and an ionic strength I of 0.113M. A non-linear least squares analysis of calcium specific ion electrode data yields, on a molar basis, four calcium dissociation constants: 10^–7 for the first site, 10^–5 for the fourth site, and two constants between these values. Both calorimetric experiments and an indicator method provide evidence that Mg²⁺ binds to calmodulin, probably at the same sites as Ca²⁺, but with affinities about 100 times smaller: 4×10^–5 for the first site and 2×10^–3 for the fourth. Calorimetric titrations on Ca²⁺ binding to calmodulin in three buffers are consistent with 0.46 protons released upon binding at all four sites and yield an average H _B per site of 5.6 and 7.9 kJ-mol^–1 for Ca²⁺ and Mg²⁺, respectively. The entropy of the system increases by 524 and 361 J-K^–1-mol^–1 when Ca²⁺ and Mg²⁺, respectively, bind to four sites on calmodulin, i.e., the selectivity of calmodulin for Ca²⁺ is primarily derived from entropy effects. Further analysis based on elimination of the entropy term for the metal ions demonstrates that calmodulin bound to Ca²⁺ has a larger entropy than the unbound calmodulin; the opposite is true for calmodulin bound to Mg²⁺. These analyses are consistent with the hypothesis that Ca²⁺ forms tight complexes at all sites on calmodulin and that release of waters of hydration upon binding is the source of the increase of entropy in the system.     

八肋游仆虫中心蛋白N-端半分子与Tb3+，Ca2+的结合

用分子生物学方法表达、纯化了游仆虫中心蛋白及N-端半分子，用铽荧光探针法、离子竞争法研究了pH 7.4，0.01 mol· L^-1 Hepes条件下中心蛋白与铽、钙的结合性质。结果表明中心蛋白有4个铽结合部位，其中2个为高亲合结合部位、2个为低亲合结合部位。具有2个低亲合结合部位的中心蛋白半分子与铽结合的条件常数是(2.13±0.10)×10⁵ L·mol^-1，与钙结合的条件常数是(7.52±0.02)×10² L·mol^-1。     

The influence of dihydropyridines calcium antagonists on 5-HT-induced intracellular calcium signal

Confocal laser scanning microscopy (CLSM) was applied to detect the intracellular [Ca2 ] variety of fluorescent intension, with Fluo-3/AM fluorescence loaded in SFSMC. The results show that 10 μmol/L Lacidipine can reduce the frequence which 10 μmol/L 5-HT induced [Ca2 ] spark in SFSMC of calcium over loading to 50%, and amplitude to 50% or so. We can draw a conclusion that dihydropyridines cal-cium antagonists lacidipine can antagonize the release of intracellular [Ca2 ] which 5-HT-induced in dose dependent manner.     

Characteristics of evodiamine-exerted stimulatory effects on rat jejunal contractility

This study was designed to characterise the effects of evodiamine on intestinal contractility and reveal the correlated mechanisms. Evodiamine (2.5–80.0 μM) increased normal jejunal contractility and jejunal hypocontractility established under a variety of experimental conditions. Evodiamine-exerted stimulatory effects were blocked by the L-type Ca²⁺ channel blocker nifedipine or abolished in the Ca²⁺-free assay condition. The stimulatory effects of evodiamine on jejunal contractility were partially blocked in the presence of neurotoxin tetrodotoxin or endogenous acetylcholine synthesis blocker hemicholinium-3 or muscarinic receptor antagonist atropine, respectively. Evodiamine-exerted stimulatory effects were blocked by c-kit receptor tyrosine kinase inhibitor imatinib. Evodiamine increased myosin phosphorylation in jejunal smooth muscle of constipation-prominent rats. These results showed that evodiamine-exerted stimulatory effects on jejunal segments are Ca²⁺-dependent, need the presence of interstitial cell of Cajal, requirement of cholinergic neuron and correlate with increased myosin phosphorylation, implicating the potential value of evodiamine in relieving hypo-motility disorders.     

Effects of BaCl<Subscript>2</Subscript> on slow vacuolar ion channels on radish by patch-clamp

The effects of BaCl₂ on slow vacuolar (SV) currents of radish are studied by using the whole-vacuolar patch-clamp recording mode. The Ca²⁺-dependent SV channel can be activated by cytosolic Ca²⁺. When 1 mmol/L BaCl₂ is added into pipette solution, SV currents are suppressed remarkably. Then adding BaCl₂ of different concentrations into the bath solution, SV currents reflect different effects. The results show that BaCl₂ with a lower concentration (<3 mmol/L) promotes the channel currents and the currents are saturated when BaCl₂ concentrations are between 1 μmol/L and 1 mmol/L, but BaCl₂ with higher concentration (≥ 3 mmol/L) inhibits SV currents.     

Ca2+-Mg2+-Zn2+体系在水溶性聚磷酸铵溶液中螯合规律的研究

近年来,水溶性聚磷酸铵在液体肥料和复合肥料的领域受到了广泛的关注,并在发达国家中得到了大面积的推广及应用。在pH值为5.5~8.0、温度为278.15 K~323.15 K的条件下,本文采用滴定法研究Ca_²⁺-Mg_²⁺-Zn_²⁺体系在聚磷酸铵溶液中的螯合规律。实验结果表明：相同质量分数的聚磷酸铵溶液对金属离子的螯合量会随着体系中Ca_²⁺、Mg_²⁺、Zn_²⁺的摩尔浓度的变化而变化;随着温度的升高而逐渐降低;随着pH的增加而逐渐增加;随着聚合度的升高而逐渐增加。采用傅里叶红外光谱对聚磷酸铵和A₁B₃C₃体系的螯合物进行表征。     

A novel red emitting phosphor Ca2SnO4: Eu

A novel red emitting phosphor, Eu³⁺-doped Ca₂SnO₄, was prepared by the solid-state reaction. X-ray powder diffraction (XRD) analysis confirmed the formation of Ca₂SnO₄: Eu³⁺. Field-emission scanning electron-microscopy (FE-SEM) observation indicated a narrow size-distribution of about 500 nm for the particles with spherical shape. Photoluminescence measurements indicated that the phosphor exhibits bright red emission at about 615 nm under UV excitation. The excellent luminescence properties make it possible as a good candidate for plasma display panels (PDP) application. Splitting of the ⁵D₀-⁷F_J transitions of Ca₂SnO₄: Eu³⁺ suggests that the Eu³⁺ ions occupied two nonequivalent sites in the crystallite. The luminescence lifetime measurement showed a bi-exponential decay, providing other evidence for the existence of two different environments for Eu³⁺ ions.     

Novel Ca-selective merocyanine-type chromoionophore derived from calix[4]arene-diamide

The synthesis and calcium-selective chromogenic ion binding properties of benzothiazolium derivative of calix[4]arene are described. The merocyanine-type ionophore derived from calix[4]arene-diamide showed selective chromogenic properties toward Ca²⁺ ions in aqueous methanol. The compound showed significant Ca²⁺ ion dependent changes in UV-vis spectral properties in the presence of other physiologically important metal ions (Na⁺, K⁺, and Mg²⁺). The pH sensitive chromogenic behavior also suggests that the compound can be used as a sensitive pH indicator around pH 6.5.     

A Thermodynamic Study Between 18-Crown-6 with Mg2+, Ca2+, Sr2+and Ba2+ Cations in Water–Methanol and Water–Ethanol Binary Mixtures Using The Conductometric Method

The complexation reactions between Mg²⁺, Ca²⁺, Sr²⁺ and Ba²⁺ cations with the macrocyclic ligand, 18-Crown-6 (l8C6) in water–methanol (MeOH) binary systems as well as the complexation reactions between Ca²⁺ and Sr²⁺ cations with 18C6 in water–ethanol (EtOH) binary mixtures have been studied at different temperatures using conductometric method. The conductance data show that the stoichiometry of all the complexes is 1:1. It was found that the stability of 18C6 complexes with Mg²⁺, Ca²⁺, Sr²⁺ and Ba²⁺ cations is sensitive to solvent composition and in all cases, a non-linear behaviour was observed for the variation of log K _f of the complexes versus the composition of the mixed solvents. In some cases, the stability order is changed with changing the composition of the mixed solvents. The selectivity order of 18C6 for the metal cations in pure methanol is: Ba²⁺ > Sr²⁺ > Ca²⁺ > Mg²⁺. The values of thermodynamic parameters (Δ H _c ° and Δ S _c °) for formation of 18C6–Mg²⁺, 18C6–Ca²⁺, 18C6–Sr²⁺ and 18C6–Ba²⁺complexes were obtained from temperature dependence of the stability constants. The obtained results show that the values of (Δ H _c ° and Δ S _c °) for formation of these complexes are quite sensitive to the nature and composition of the mixed solvent, but they do not vary monotonically with the solvent composition.This revised version was published online in July 2005 with a corrected issue number.     

Study of Complex Formation between N-Phenylaza-15-Crown-5 with Mg2+, Ca2+, Ag+ and Cd2+ Metal Cations in Some Binary Mixed Aqueous and Non-aqueous Solvents using the Conductometric Method

The complexation reactions between Mg²⁺, Ca²⁺, Ag⁺ and Cd²⁺ metal cations with N-phenylaza-15-crown-5 (Ph-N15C5) were studied in acetonitrile (AN)–methanol (MeOH), methanol (MeOH)–water (H₂O) and propanol (PrOH)–water (H₂O) binary mixtures at different temperatures using the conductometric method. The conductance data show that the stochiometry of all of the complexes with Mg²⁺, Ca²⁺, Ag⁺ and Cd²⁺ cations is 1:1 (L:M). The stability of the complexes is sensitive to the solvent composition and a non-linear behaviour was observed for variation of log K _f of the complexes versus the composition of the binary mixed solvents. The selectivity order of Ph-N15C5 for the metal cations in neat MeOH is Ag⁺>Cd²⁺>Ca²⁺>Mg²⁺, but in the case of neat AN is Ca²⁺>Cd²⁺>Mg²⁺>Ag⁺. The values of thermodynamic parameters (ΔH _c ^o , ΔS _c ^o ) for formation of Ph-N15C5–Mg²⁺, Ph-N15C5–Ca²⁺, Ph-N15C5–Ag⁺ and Ph-N15C5–Cd²⁺ complexes were obtained from temperature dependence of stability constants and the results show that the thermodynamics of complexation reactions is affected by the nature and composition of the mixed solvents.     

New ON-OFF type Ca-selective fluoroionophore having boron-dipyrromethene fluorophores

A new fluorogenic ionophore has been prepared by conjugating calix[4]-crown-5 ether with boron-dipyrromethene fluorophore. The ionophore exhibited a pronounced selective ON-OFF type response toward Ca²⁺ ions over other physiologically important metal ions of Na⁺, K⁺, and Mg²⁺. The interaction with Ca²⁺ ions resulted in very efficient quenching of fluorescence at 507 nm excited by relatively long wavelength irradiation at 480 nm. The selectivity ratios toward Ca²⁺ ions over other tested physiologically important metal ions were larger than 180 in aqueous 95% MeOH solution.     

Study of Complex Formation Between Dicyclohexano-18-Crown-6 (DCH18C6) with Mg 2+, Ca2+, Sr 2+, and Ba2+ Cations in Methanol–Water Binary Mixtures Using Conductometric Method

The complexation reactions between Mg²⁺,Ca²⁺,Sr²⁺ and Ba²⁺ metal cations with macrocyclic ligand, dicyclohexano-18-crown-6 (DCH18C6) were studied in methanol (MeOH)–water (H₂O) binary mixtures at different temperatures using conductometric method . In all cases, DCH18C6 forms 1:1 complexes with these metal cations. The values of stability constants of complexes which were obtained from conductometric data show that the stability of complexes is affected by the nature and composition of the mixed solvents. While the variation of stability constants of DCH18C6-Sr ²⁺ and DCH18C6-Ba²⁺versus the composition of MeOH–H₂O mixed solvents is monotonic, an anomalous behavior was observed for variations of stability constants of DCH18C6-Mg²⁺ and DCH18C6-Ca²⁺ versus the composition of the mixed solvents. The values of thermodynamic parameters (ΔH_c°, ΔS_c°) for complexation reactions were obtained from temperature dependence of formation constants of complexes using the van’t Hoff plots. The results show that in most cases, the complexation reactions are enthalpy stabilized but entropy destabilized and the values of thermodynamic parameters are influenced by the nature and composition of the mixed solvents. The obtained results show that the order of selectivity of DCH18C6 ligand for metal cations in different concentrations of methanol in MeOH–H₂O binary system is: Ba²⁺>Sr²⁺>Ca²⁺> Mg²⁺.     

Calix[4]arenesulfonylamidines. Synthesis, structure and influence on Mg, ATP-dependent calcium pumps

Calix[4]arenes, functionalized at the wide rim with two or four N²-sulfonylamidine groups were prepared. In the crystalline state, the bowl shaped calix[4]arene-bis-N-sulfonyltrifluoromethylacetamidine 3b is associated through intermolecular hydrogen bonds, NH?OS, while the phenyl rings of the Ph-SO₂-fragments are hosted in the cavities of the nearby molecules of 3b. Calixarene 4b influences Ca²⁺ transport in Mg²⁺, ATP-dependent calcium pumps.     

Eu luminescence—a structural probe in BiCa4(PO4)3O, an apatite related phosphate

Eu³⁺ luminescence is studied in apatite-related phosphate BiCa₄(PO₄)₃O. Compositions of the formula Bi_1−xEu_xCa₄(PO₄)₃O [x=0.05, 0.1, 0.3, 0.5, 0.8 and 1.0] are synthesized and they are isostructural with parent BiCa₄(PO₄)₃O. Room temperature photoluminescence shows the various transitions ⁵D₀→⁷F_J(=0,1,2) of Eu³⁺. The emission results of compositions with different Eu³⁺ content show the difference in site occupancy of Eu³⁺ in Bi_1−xEu_xCa₄(PO₄)₃O. The intense ⁵D₀-⁷F₀ line at 574 nm for higher Eu³⁺ content is attributed to the presence of strongly covalent Eu-O bond that is possible by substituting Bi³⁺ in the Ca(2) site. This shows the preferential occupancy of Bi³⁺ in Ca(2) site and this has been attributed to the 6s² lone pair electrons of Bi³⁺. This is further confirmed by comparing the emission results with La_0.95Eu_0.05Ca₄(PO₄)₃O.     

Palladium catalyzed synthesis of Ca indicators with aryl bithiophene and terthiophene fluorophores

Five new fluorescent indicators for Ca²⁺ were synthesized using the Stille reaction. They all consist of the tricarboxylate chelator APTRA (o-aminophenol-N,N,O-triacetic acid) linked to a (substituted) bithiophene or terthiophene fluorophore. The dissociation constants K_d measured via fluorimetric titrations at 21 °C in 100 mM KCl buffered solution, pH 7.05, for the Ca²⁺ complexes with the new probes are in the range between 10 and 40 μM.     

多吡啶钴(Ⅲ)配合物与Zn2+形成的新型“关-开”荧光探针

近十几年来,对小分子过渡金属配合物与大分子DNA键合与识别机理的研究一直是国际上生物无机化学领域十分活跃的研究课题[1￣3],已发展了一系列具有特定功能的配合物,如DNA结构探针和DNA荧光探针等。与其他类型的金属配合物相比,八面体过渡金属多吡啶配合物具有丰富的光化学和光物理信息,当这些配合物与DNA相互作用时,由于结构匹配或微环境的差异,配合物的光谱特征会出现不同程度的改变,从而达到对DNA的检测。传统的DNA荧光探针有[Ru(bpy)2dppz]2 和[Ru(phen)2dppz]2 (bpy=2,2′-联吡啶,phen=1,10-菲咯啉,dppz=二吡啶[3,2-a∶2′,3′…     

Violet and greenish photoprotein obelin mutants for reporter applications in dual-color assay

Two kinds of Ca²⁺-regulated photoprotein obelin with altered color of bioluminescence were obtained by active-center amino acid substitution. The mutant W92F-H22E emits violet light (λ_max = 390 nm) and the mutant Y139F emits greenish light (λ _max = 498 nm), with small spectral overlap, both display high activity and stability and thus may be used as reporters. For demonstration, the mutants were applied in dual-color simultaneous immunoassay of two gonadotropic hormones—follicle-stimulating hormone and luteinizing hormone. Bioluminescence of the reporters was simultaneously triggered by single injection of Ca²⁺ solution, divided using band-pass optical filters and measured with a two-channel photometer. The sensitivity of simultaneous bioluminescence assay was close to that of a separate radioimmunoassay. Figure Two kinds of Ca²⁺-regulated photoprotein obelin with altered color of bioluminescence were obtained and applied in dual-color simultaneous immunoassay of two gonadotropic hormones.