SD乳鼠心肌细胞的培养及其低温下钙波-搏动耦联的初步研究

分离培养3 d以内SD乳鼠心肌细胞,Fluo-4负载后分别于37 ℃,24 ℃环境中在激光扫描共焦显微镜 (Laser Scanning Confocal Microscopy, LSCM)下进行细胞搏动及胞内Ca2+运动的观测.结果显示,心肌细胞在体外呈集落生长.当37 ℃时,集落中无钙波产生,但集落细胞同步搏动;当24 ℃时,集落中个别细胞先发生钙波,随后集落细胞才同步搏动.证明心肌细胞搏动与胞内Ca2+浓度变化相关,在24 ℃低温下个别细胞产生的钙波可能对其所在集落细胞的同步搏动有诱导并维持其搏     

2型糖尿病鼠胰腺β细胞中SERCA泵的变化对钙振荡波形影响的建模研究

在原内质网Ca2+-ATP酶(SERCA)模型的基础上建立了2型糖尿病鼠胰腺β细胞的SERCA模型,并将两模型分别应用于钙振荡模型里来模拟SERCA泵的变化对胞质钙浓度([Ca2+])的影响.分析结果发现:(1)病鼠胰腺β细胞中的[Ca2+]升高,并且钙振荡周期和幅值增大;(2)细胞中ATP浓度([ATP])降低,ATP振荡周期增大,但幅值变化不明显;(3)内质网(ER)中钙振荡波形的周期增大, 幅值减小.结论:在2型糖尿病鼠胰腺β细胞中,其[Ca2+]相对于正常大鼠明显升高的现象与细胞中SERCA泵转运钙的能力下降有关.     

基因表达的钙离子调控机制研究

基于基因调节系统的动力学模型,利用数值模拟方法研究了Ca2+振荡对基因表达活性的影响.结果表明:细胞内Ca2+振荡能降低活化转录因子的有效钙阈值;转录因子的活性随Ca2+浓度平均水平增高而增大,但随着细胞内Ca2+振荡周期的增大而减小.     

细胞内钙离子振荡的调制作用

分别考虑了简单周期性和复杂性钙离子振荡对一些特殊规律的细胞功能的影响,例如,肝细胞中磷酸化-去磷酸化循环的肝糖磷酸化激活酶的控制和基因表达等.结果表明简单周期的钙离子振荡表现出许多共同的表型且有很多数学模型.在简单钙离子振荡情况下细胞内钙离子振荡的周期随IP3浓度的增大而减少,Ca2+浓度的平均值在振荡发生区域内随IP3浓度的增大而增大.爆发性、混沌和准周期的复杂钙离子振荡会调制肝糖磷酸化激活酶在同样的区域内出现振荡.     

离子种类和质量浓度对不同温度稠油油水界面张力影响的实验研究

为了研究在不同温度下离子种类和质量浓度对稠油油水界面张力的影响,分别配制了含不同质量浓度Na+、Ca2+、Mg2+的盐水溶液,应用旋转油滴法测量了不同温度时稠油与盐水的界面张力,分析了温度、离子种类与质量浓度对界面张力的影响规律.实验结果表明:①盐水成分为Na+和Mg2+时,随离子质量浓度的增大,界面张力先减小后增大,存在极小值;成分为Ca2+时,界面张力先增大后稳定再增大.②随温度升高,在Na+和Ca2+作用下界面张力明显减小,在Mg2+作用下先减小后稳定.③温度和离子质量浓度均影响界面张力;离子质量浓度低时界面张力主要受温度影响;离子质量浓度高时在Na+作用下界面张力受温度影响较大,在Ca2+作用下界面张力受温度的影响减弱;在Mg2+作用下界面张力受离子质量浓度的影响较大.     

圆柱超声换能器的负载特性及其电声效率

对圆柱超声换能器在液体中的负载特性进行了研究,推导得出了换能器在液体介质中径向声辐射阻抗,并得出了有载圆柱压电超声换能器的频率方程;探讨了有负载的圆柱换能器输入阻抗随液面半径变化的关系.理论计算表明:随着液面半径的增加,换能器径向辐射阻抗在-∞～∞周期性变化;换能器在液体中会产生谐波共振,谐波数随液面半径的增大而增加.通过实验测量了圆柱换能器在水介质中的谐波共振频率,理论和实验测量结果吻合较好.此外,实验测定了换能器在水介质中的电声效率.     

模拟酸雨胁迫下Ca2 对小白菜的调控作用的研究

通过大棚试验,研究了模拟酸雨胁迫下Ca2+对小白菜体内6种不同形态钙以及生理和营养品质指标的影响。结果表明,施加适宜浓度的Ca2+可以增加小白菜体内各形态钙的浓度,施加过高浓度钙的小白菜体内各形态钙含量相对较低,说明过高浓度的钙在一定程度上抑制了小白菜对钙的吸收。模拟酸雨胁迫下,小白菜体内的果胶酸钙和磷酸三钙浓度随施加Ca2+浓度的增大而增大,达到浓度峰值后下降,且在较强酸雨胁迫下增加程度更明显;小白菜体内总钙以及残渣态钙浓度增加,无机钙和水溶性钙浓度下降.随着Ca2+浓度增大,叶绿素和可溶性糖含量逐渐增大,说明外源Ca2+在一定程度上起到了抵抗酸雨的作用;相对于低浓度Ca2+,高浓度Ca2+调控作用下的叶绿素和可溶性糖含量在强酸性酸雨与弱酸性酸雨胁迫下的差异更小,说明适当地提高的外源Ca2+浓度有助于缓解强酸性酸雨对小白菜造成的损害。果胶酸钙能够提高细胞壁刚性,维持细胞膜的稳定性是抵御酸雨的基本原理之一。     

对称性对二维三角晶格光子晶体简并态的影响

本文从介质圆柱在空气中排列的二维三角晶格光子晶体出发，引入了一种复式格子二维光子晶体结构，使原三角晶格C6v的对称性降低至C2v，从而使能带简并分立，产生了一些新的带隙。采用超元胞的方法来计算能带结构，结果表明，随着两种圆柱半径比的增大，带隙结构越来越丰富。我们将由原二维三角晶格光子晶体第一条能带折叠形成的四条能带定义为超元胞第一组能带，以此类推。发现随着两种圆柱半径比的增大，第一组能带之间出现了3条带隙，总体上逐渐展宽，而第一组能带和第二组能带之间的带隙逐渐减小。最后我们还计算了绝对禁带随着两种圆柱半径比增大的变化情况，发现随着半径比的增大，绝对禁带从无到有，为我们提供了一种新的获得绝对禁带的方法。     

Glucose-induced Ca2+ signals in rat pancreatic β cells

Using microfluorometry to assay intracellular Ca2+ , the influences of varied factors on glucose induced Ca22+ signals, such as glucose-induced initial decline phase (GIDP), Ca2+ oscillation, and Ca2+ release from internal stores, were investigated in single rat pancreatic β cells. Glucose was able to evoke GIDP even at non-stimulus concentration (5 mol/L), which is insufficient to induce Ca2+ spikes. GIDP was dependent on neither membrane depo larization nor extraeellular Ca2+ . However, GIDP was inhibited by thapsigargin, indicating a dependence on Ca2+ up take by Ca22+ stores. The glucose-induced calcium oscillation was inhibited when external Ca2+ was removed. However, thapsigargin could not block the Ca2+ oscillation. These results suggest that maintenance of Ca22+ oscillation requires ex tracellular Ca2+ but not Ca2+ stores. Glucose was able to evoke Ca2+ signals even in the absence of external Ca2+ . The glucose-induced Ca2+ release from intracellular Ca2+ stores was blocked by TTX. However, TTX had no effect on high K--induced Ca2+ store release, suggesting that membrane depolarization can directly release Ca2+ from some internal Ca2+ stores in β cells.     

矩形管道内低雷诺数圆柱绕流尾迹演化特性

为了揭示有限管道壁面对圆柱绕流尾迹演化特性的影响,采用基于有限容积法的数值模拟方法,对矩形管道内雷诺数为100的三维圆柱绕流尾迹流场进行计算分析,探讨阻流比和长径比对圆柱表面和尾迹流场中压力分布的影响。研究结果表明:圆柱前、后驻点的压力系数受展向位置影响较大,后驻点压力系数与旋涡脱落状态有关;时均压力系数受管道端壁影响显著,沿前驻点至后驻点圆柱表面时均压力系数先减小后增大;在远离端壁一定区域内,尾迹流态为涡街;而在靠近端壁的一定区域内,尾迹呈现出近似双子涡形态;圆柱表面相同位置处的时均压力系数基本上随阻流比的增大而增大,随长径比的增大而减小。     

A calcium sensor in the sodium channel modulates cardiac excitability.

Sodium channels are principal molecular determinants responsible for myocardial conduction and maintenance of the cardiac rhythm. Calcium ions (Ca2+) have a fundamental role in the coupling of cardiac myocyte excitation and contraction, yet mechanisms whereby intracellular Ca2+ may directly modulate Na channel function have yet to be identified. Here we show that calmodulin (CaM), a ubiquitous Ca2+-sensing protein, binds to the carboxy-terminal 'IQ' domain of the human cardiac Na channel (hH1) in a Ca2+-dependent manner. This binding interaction significantly enhances slow inactivation-a channel-gating process linked to life-threatening idiopathic ventricular arrhythmias. Mutations targeted to the IQ domain disrupted CaM binding and eliminated Ca2+/CaM-dependent slow inactivation, whereas the gating effects of Ca2+/CaM were restored by intracellular application of a peptide modelled after the IQ domain. A naturally occurring mutation (A1924T) in the IQ domain altered hH1 function in a manner characteristic of the Brugada arrhythmia syndrome, but at the same time inhibited slow inactivation induced by Ca2+/CaM, yielding a clinically benign (arrhythmia free) phenotype.     

Location of high affinity Ca2+-binding sites within the predicted transmembrane domain of the sarcoplasmic reticulum Ca2+-ATPase

Cation pumps bind and translocate ions with the intermediate formation of a phosphoenzyme. In spite of extensive knowledge of the primary and even secondary structures of several of these cation transport enzymes, however, no high affinity cation binding sites have yet been determined. Here we report the use of oligonucleotide-directed, site-specific mutagenesis to identify the amino acids involved in Ca2+ binding in one of these transport enzymes, the Ca2+-ATPase of sarcoplasmic reticulum. Alteration of Glu 309, Glu 771, Asn 796, Thr 799, Asp 800 or Glu 908, each of which is predicted to lie near the centre of the transmembrane domain in putative transmembrane sequences M4, M5, M6 and M8 resulted in complete loss of Ca2+ transport function and of Ca2+-dependent phosphorylation of the enzyme by ATP. Phosphorylation of each of the mutant enzymes with inorganic phosphate was observed, however, even in the presence of Ca2+, which inhibits phosphorylation in the wild-type enzyme possessing an intact high affinity Ca2+-binding site. These results suggest that at least six polar, oxygen-containing residues lying near the centre of the transmembrane domain provide ligands for one or both of the two high affinity Ca2+ binding sites in the Ca2+-ATPase.     

Interaction of SynaptotagminⅠ with Phospholipid Membrane: A Monolayer Study

IntroductionSynaptotagmin is a family of vesicletransmembrane proteins present in synapticvesicles and large secretary granules of neuronsand endocrine cells[1 ] .It is a major constituent ofsynaptic vesicle membranes,comprising7% 8%of the total vesicle protein,characterized by ashort intravesiclar N-terminus,a singletransmembrane region,and a long plasmicdomain.　The best-charaterized form of synaptotagmin,syt ,is found abundantly in rostrol brain.Syt was first described in 1 981 [2 ] ,and i…     

Structure of the calcium regulatory muscle protein troponin-C at 2.8 A resolution

Crystals of turkey skeletal muscle troponin-C reveal a molecule of two domains with an unusual structure. Two Ca2+ ions are bound to the C-terminal domain. The two cation-binding sites of the regulatory (N-terminal) domain are Ca2+ free; this domain adopts a markedly different conformation from the C-terminal domain. The two domains are connected by a long nine-turn alpha-helix; three of these turns are exposed fully to solvent.     

Different domains of synaptotagmin control the choice between kiss-and-run and full fusion

Exocytosis-the release of the contents of a vesicle--proceeds by two mechanisms. Full fusion occurs when the vesicle and plasma membranes merge. Alternatively, in what is termed kiss-and-run, vesicles can release transmitter during transient contacts with the plasma membrane. Little is known at the molecular level about how the choice between these two pathways is regulated. Here we report amperometric recordings of catecholamine efflux through individual fusion pores. Transfection with synaptotagmin (Syt) IV increased the frequency and duration of kiss-and-run events, but left their amplitude unchanged. Endogenous Syt IV, induced by forskolin treatment, had a similar effect. Full fusion was inhibited by mutation of a Ca2+ ligand in the C2A domain of Syt I; kiss-and-run was inhibited by mutation of a homologous Ca2+ ligand in the C2B domain of Syt IV. The Ca2+ sensitivity for full fusion was 5-fold higher with Syt I than Syt IV, but for kiss-and-run the Ca2+ sensitivities differed by a factor of only two. Syt thus regulates the choice between full fusion and kiss-and-run, with Ca2+ binding to the C2A and C2B domains playing an important role in this choice.     

Alpha-neurexins couple Ca2+ channels to synaptic vesicle exocytosis

Synapses are specialized intercellular junctions in which cell adhesion molecules connect the presynaptic machinery for neurotransmitter release to the postsynaptic machinery for receptor signalling. Neurotransmitter release requires the presynaptic co-assembly of Ca2+ channels with the secretory apparatus, but little is known about how synaptic components are organized. Alpha-neurexins, a family of >1,000 presynaptic cell-surface proteins encoded by three genes, link the pre- and postsynaptic compartments of synapses by binding extracellularly to postsynaptic cell adhesion molecules and intracellularly to presynaptic PDZ domain proteins. Using triple-knockout mice, we show that alpha-neurexins are not required for synapse formation, but are essential for Ca2+-triggered neurotransmitter release. Neurotransmitter release is impaired because synaptic Ca2+ channel function is markedly reduced, although the number of cell-surface Ca2+ channels appears normal. These data suggest that alpha-neurexins organize presynaptic terminals by functionally coupling Ca2+ channels to the presynaptic machinery.     

Cholecystokinin induces a decrease in Ca2+ current in snail neurons that appears to be mediated by protein kinase C

Three distinct classes of protein kinases have been shown to regulate Ca2+ current in excitable tissues. Cyclic AMP-dependent protein kinase mediates the action of noradrenaline on the Ca2+ current of cardiac muscle cells. Cyclic GMP-dependent protein kinase mediates the serotonin-induced modulation of the Ca2+ current in identified snail neurons. The Ca2+/diacylglycerol-dependent protein kinase (protein kinase C) has also been found to regulate Ca2+ currents of neurons. However, no neurotransmitter has yet been shown to regulate Ca2+ current through the activation of protein kinase C. We now report that cholecystokinin, a widely occurring neuropeptide which is present in molluscan neuron, modulates the Ca2+ current in identified neurons of the snail Helix aspersa, and that this effect appears to be mediated by protein kinase C. Specifically, sulphated cholecystokinin octapeptide 26-33 (CCK8), activators of protein kinase C, and intracellular injection of protein kinase C, all shorten the Ca2+-dependent action potential and decrease the amplitude of the Ca2+ current in these cells. All these effects are not reversible within the duration of the experiments. Moreover, intracellular injections of low concentrations of protein kinase C, which are ineffective by themselves, enhance the effectiveness of low concentrations of CCK8 on the Ca2+ current.     

Amino-acid sequence of a Ca2+ + Mg2+-dependent ATPase from rabbit muscle sarcoplasmic reticulum, deduced from its complementary DNA sequence

We have cloned and sequenced complementary DNA encoding a Ca2+-ATPase of rabbit muscle sarcoplasmic reticulum. We propose a model of the protein which has 3 cytoplasmic domains joined to a set of 10 transmembrane helices by a narrow, penta-helical stalk. In this model, ATP bound to one cytoplasmic domain would phosphorylate an aspartate in an adjoining cytoplasmic domain, inducing translocation of Ca2+ from binding sites on the stalk.     

The structural basis of calcium transport by the calcium pump

The sarcoplasmic reticulum Ca2+-ATPase, a P-type ATPase, has a critical role in muscle function and metabolism. Here we present functional studies and three new crystal structures of the rabbit skeletal muscle Ca2+-ATPase, representing the phosphoenzyme intermediates associated with Ca2+ binding, Ca2+ translocation and dephosphorylation, that are based on complexes with a functional ATP analogue, beryllium fluoride and aluminium fluoride, respectively. The structures complete the cycle of nucleotide binding and cation transport of Ca2+-ATPase. Phosphorylation of the enzyme triggers the onset of a conformational change that leads to the opening of a luminal exit pathway defined by the transmembrane segments M1 through M6, which represent the canonical membrane domain of P-type pumps. Ca2+ release is promoted by translocation of the M4 helix, exposing Glu 309, Glu 771 and Asn 796 to the lumen. The mechanism explains how P-type ATPases are able to form the steep electrochemical gradients required for key functions in eukaryotic cells.     

Calmodulin bifurcates the local Ca2+ signal that modulates P/Q-type Ca2+ channels

Acute modulation of P/Q-type (alpha1A) calcium channels by neuronal activity-dependent changes in intracellular Ca2+ concentration may contribute to short-term synaptic plasticity, potentially enriching the neurocomputational capabilities of the brain. An unconventional mechanism for such channel modulation has been proposed in which calmodulin (CaM) may exert two opposing effects on individual channels, initially promoting ('facilitation') and then inhibiting ('inactivation') channel opening. Here we report that such dual regulation arises from surprising Ca2+-transduction capabilities of CaM. First, although facilitation and inactivation are two competing processes, both require Ca2+-CaM binding to a single 'IQ-like' domain on the carboxy tail of alpha1A; a previously identified 'CBD' CaM-binding site has no detectable role. Second, expression of a CaM mutant with impairment of all four of its Ca2+-binding sites (CaM1234) eliminates both forms of modulation. This result confirms that CaM is the Ca2+ sensor for channel regulation, and indicates that CaM may associate with the channel even before local Ca2+ concentration rises. Finally, the bifunctional capability of CaM arises from bifurcation of Ca2+ signalling by the lobes of CaM: Ca2+ binding to the amino-terminal lobe selectively initiates channel inactivation, whereas Ca2+ sensing by the carboxy-terminal lobe induces facilitation. Such lobe-specific detection provides a compact means to decode local Ca2+ signals in two ways, and to separately initiate distinct actions on a single molecular complex.