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1.
An aqueous solution of alizarin red S containing chloral hydrate both clears intact chlorophyllous gemma cells of Vittaria graminifolia and stains for protoplasmic calcium. Verification that the stain was protoplasmic rather than in the cell wall was shown by a positive reaction in extruded protoplasm. Similar staining was found in extruded protoplasm of Onoclea sensibilis spores. Differentiating gemma cells show localized protoplasmic accumulations of Ca2+ at sites where asymmetric cell divisions initiate the formation of rhizoids, antheridia or vegetative cells. The staining properties of the dye depend on careful control of pH and the addition of appropriate amounts of KCl to the mixture. Treatment of Onoclea spores and Vittaria gemmae with 100 mM EGTA for 30 min nearly abolishes staining of their extruded protoplasts and also of intact cells of gemmae. The use of alizarin red S with and without chloral hydrate demonstrates different pools of protoplasmic Ca2+. When Onoclea spores are ruptured to extrude the protoplasm, both dye mixtures stain a peripheral, granular protoplasmic component. However, the chloral hydrate-containing dye also reveals Ca2+ associated with small particulate protoplasmic components. Extruded protoplasm of gemma cells stains intensely with alizarin-chloral hydrate, but does not stain with alizarin lacking chloral hydrate.  相似文献   

2.
Light-dependent Ca2+ efflux via the Ca2+/H+ antiport in the photosynthetic purple sulfur bacterium Chromatium vinosum was inhibited by three phenothiazines: chlorpromazine; trifluoperazine and phenothiazine. The inhibitors had no effect on Ca2+ uptake by C. vinosum in the dark nor any effect on the light-dependent efflux of either Na+ or Tl+ catalyzed, respectively, by the C. vinosum Na+/H+ or K+/H+ antiports. Ruthenium red and LaCl3, neither of which inhibited light-dependent Ca2+ efflux in C. vinosum, markedly inhibited Ca2+ uptake in the dark by C. vinosum cells. Ruthenium red had no effect on the uptake of either Na+or the K+ analog T1+ by C. vinosum cells in the dark. These results have been interpreted in terms of two separate Ca2+ transport systems in C. vinosum: (i) a phenothiazine-sensitive and ruthenium red, La3+-insensitive Ca2+/H+ antiport responsible for Ca2+ efflux in the light; and (ii) a ruthenium red and La3+-sensitive but phenothiazine-insensitive Ca2+ uptake system.  相似文献   

3.
Calcium activation of oxygen evolution from French-press preparations of Phormidium luridum is largely reversible upon removal of added Ca2+. Activation occurs via a first-order binding with a dissociation constant of 2.8 mM. An 8-fold increase in oxygen evolution rate observed upon Ca2+ addition is accounted for by a 4-fold increase in the number of active photosynthetic units, and a doubling of turnover rate. While both Ca2+ and Mg2+ stimulate turnover, unit activation is Ca2+ specific. Under optimal conditions, 30% of the units functioning in the intact cell can be recovered in the Ca2+-activated preparation.

The Ca2+ requirement of P. luridum preparations is not relieved by proton-carrying uncouplers, or by rate-saturating concentrations of the Hill acceptor, ferricyanide. Taken together with the reported stimulation by Ca2+ of oxygen evolution in the presence of DCMU (Piccioni, R.G. and Mauzerall, D.C. (1976) Biochim. Biophys. Acta 423, 605–609) these observations strongly suggest a site of Ca2+ action within Photosystem II.

The pronounced specificity of the Ca2+ requirement appears in preparations of other cyanobacteria (Anabaena flos-aquae and Anacystis nidulans) but not in the eucaryote Chlorella vulgaris. While milder cell-disruption methods bring about some Ca2+ dependence in P. luridum, French-press treatment is required for maximal expression of Ca2+-specific effects. French-press breakage causes a release of endogenous Ca2+ from cells, supporting the view that added Ca2+ restores oxygen evolution by satisfying a physiological requirement for the cation.  相似文献   


4.
Vascular smooth muscle cells respond with an increase in intracellular Ca2+ within seconds after exposure to oxidized low density lipoprotein (oxLDL). This has been suggested to represent a signaling response that may have implications for gene expression. If so, oxLDL may induce increases in nuclear Ca2+ in smooth muscle cells in response to oxLDL. Aortic smooth muscle cells were exposed to 100 μg/ml oxLDL. Large, rapid increases in [Ca2+]i were observed using fluo-3 as an indicator dye to detect intracellular Ca2+ on the stage of a confocal micro-scope. This was also confirmed using ratiometric imaging of indo signals. These elevations appeared to be localized to the nuclear region of the cell. DNA staining of the cells confirmed its localization to the nuclear / perinuclear region of the cell. Our data demonstrate that oxLDL induces a nuclear localized elevation in Ca2+i that may have important implications for nuclear function.  相似文献   

5.
Vertebrate embryos generate striking Ca2+ patterns, which are unique regulators of dynamic developmental events. In the present study, we used zebrafish embryos as a model system to examine the developmental roles of Ca2+ during gastrulation. We found that gastrula stage embryos maintain a distinct pattern of cytosolic Ca2+ along the dorsal–ventral axis, with higher Ca2+ concentrations in the ventral margin and lower Ca2+ concentrations in the dorsal margin and dorsal forerunner cells. Suppression of the endoplasmic reticulum Ca2+ pump with 0.5 μM thapsigargin elevates cytosolic Ca2+ in all embryonic regions and induces a randomization of laterality in the heart and brain. Affected hearts, visualized in living embryos by a subtractive imaging technique, displayed either a reversal or loss of left–right asymmetry. Brain defects include a left–right reversal of pitx2 expression in the dorsal diencephalon and a left–right reversal of the prominent habenular nucleus in the brain. Embryos are sensitive to inhibition of the endoplasmic reticulum Ca2+ pump during early and mid gastrulation and lose their sensitivity during late gastrulation and early segmentation. Suppression of the endoplasmic reticulum Ca2+ pump during gastrulation inhibits expression of no tail (ntl) and left–right dynein related (lrdr) in the dorsal forerunner cells and affects development of Kupffer’s vesicle, a ciliated organ that generates a counter-clockwise flow of fluid. Previous studies have shown that Ca2+ plays a role in Kupffer’s vesicle function, influencing ciliary motility and translating the vesicle’s counter-clockwise flow into asymmetric patterns of gene expression. The present results suggest that Ca2+ plays an additional role in the formation of Kupffer’s vesicle.  相似文献   

6.
The stiffness of the sarcomeres was studied during the diastolic interval of 18 stimulated (0.5 Hz) cardiac trabeculae of rat (pH 7.4; temperature = 25°C). Sarcomere length (SL) and force (F) were measured using, respectively, laser diffraction techniques (resolution: 4 nm) and a silicon strain gauge (resolution: 0.63 μN). Sinusoidal perturbations (frequency = 500 Hz) were imposed to the length of the preparation. The stiffness was evaluated from the corresponding F and SL sinusoids by analysis of both signals together either in the time domain or in the frequency domain. A short burst (duration = 30 ms) of sinusoidal perturbations was repeated at 5 predetermined times during diastole providing 5 measurements of stiffness during the time interval separating two twitches. These measurements revealed that stiffness increases by 30% during diastole, while a simultaneous expansion of the sarcomeres (amplitude = 10-60 nm) was detected. Measurements of the fluorescence of fura-2 under the same conditions revealed a continuous exponential decline of [Ca2+]i from 210 to 90 nM (constant of time 300 ms) during diastole. In order to test the possibility that the increase of sarcomere stiffness and the decline of [Ca2+]i were coupled during diastole of intact trabeculae, we studied the effect of different free Ca2+-concentrations ([Ca2+]) between 1 and 430 nM on sarcomere stiffness in rat cardiac trabeculae skinned by saponin (n = 17). Stiffness was studied using 500 Hz sinusoidal perturbations of muscle length (ML). We found that, below 70 nM, the stiffness was independent of [Ca2+]; between 70 and 200 nM, the stiffness declined with increase of [Ca2+]; above 200 nM, the stiffness increased steeply with [Ca2+]. The data fitted accurately to the sum of two sigmoids (Hill functions): (1) at [Ca2+] < 200 nM the stiffness decreased with [Ca2+] (EC50 = 160 ± 13 nM; n = −2.6±0.7) and (2) at [Ca2+] > 200 nM, stiffness increased with [Ca2+] (EC50 = 3.4±0.3 μM; n = 2.1±0.2) due to attachment of cross-bridges. From these results, it was possible to reproduce accurately the time course of diastolic stiffness observed in intact trabeculae and to predict the effect on stiffness of a spontaneous elevation of the diastolic [Ca2+]. Identical stiffness measurements were performed in 4 skinned preparations exposed to a cloned fragment of titin (Ti I-II) which has been shown to exhibit a strong interaction with F-actin in vitro. It was anticipated that Ti I-II would compete with endogenous titin for the same binding site on actin in the I-band. Below 200 nM, Ti I-II (2 μM) eliminated the Ca2+-dependence of stiffness. These results are consistent with the hypothesis that the Ca2+-sensitivity of the sarcomeres at [Ca2+] < 200 nM, i.e. where the myocytes in intact muscle operate during diastole, involves an association between titin molecules and the thin filament.  相似文献   

7.
The role of Ca2+ in glycerol dissimilation under hypoosmotic stress in the halotolerant alga Dunaliella tertiolecta was investigated using a pharmacological approach. A stretch-activated Ca2+ channel blocker, GdCl3, inhibited glycerol dissimilation under hypoosmotic stress. However, addition of voltage-dependent Ca2+ channel blockers and inhibitors of mitochondrial and endoplasmic reticulum Ca2+ channels did not affect the glycerol dissimilation under hypoosmotic stress. The results of the present study suggest that the influx of Ca2+ from the extracellular space via the stretch-activated Ca2+ channels localized in the plasma membrane is required for the transduction of osmotic signal of D. tertiolecta.  相似文献   

8.
Light-dependent Ca2+ influx into intact spinach chloroplasts, measured with the metallochromic indicator arsenazo III, is stimulated by uncouplers (FCCP, CCCP, nigericin) and inhibited by ruthenium red. The data presented demonstrate that light-dependent Ca2+ influx into chloroplasts is electrogenic and mediated by a uniport-type carrier. The characteristics of the carrier system are similar to those of the Ca2+ uniport of mitochondria.  相似文献   

9.
为探讨大黄鱼幼鱼在低氧及酸化胁迫下机体离子调节情况,本研究探讨了低氧(溶解氧量DO 3.5 mg·L-1,pH 8.1)、酸化(DO 7.0 mg·L-1,pH 7.35)以及低氧酸化协同胁迫(DO3.5 mg·L-1,pH 7.35)对大黄鱼幼鱼鳃组织结构以及离子调节相关生理指标的影响.结果 表明:低氧胁迫下,大黄鱼...  相似文献   

10.
To elucidate the relationship between intracellular free Ca2+ concentration ([Ca2+]i) and Ca2+-signalling by the sarcoplasmic reticulum (SR) in Ca2+-overloaded heart muscle cells, the direct effects of “basal” [Ca2+]i on calcium waves were investigated by altering the membrane potential. When basal inter-calcium wave (BCW) [Ca2+]i was maintained at a high level, (i) calcium waves showed more gradual and more rapidly suppressed increase in [Ca2+]-profile (P < 0.005), and (ii) calcium waves occurred at a significantly higher frequency and velocity (259% and 137%), than when low BCW [Ca2+]i was maintained. Similar investigations on inhibition of the Na+-Ca2+ exchanger, however, showed that membrane potential did not elicit direct effects on calcium waves. These results showed that the elevation of BCW [Ca2+]i per se directly influences Ca2+-signalling in heart muscle cells through non-equilibrated release-restoration Ca2+-handling by the SR.  相似文献   

11.
为探究H2S信号在苜蓿(Medicago sativa)体内调节气孔运动的作用,及在此过程中H2S与Ca2+的关系,以蒺藜苜蓿(Medicago truncatula)的野生型和钙离子转运体突变体为试验材料,分别从转录水平、细胞水平和生理水平开展研究。采用qRT-PCR比较相关基因的表达量变化、荧光探针显示体内Ca2+含量、电极法测定H2S含量、光学显微镜观察和测量气孔孔径等。结果表明:蒺藜苜蓿突变体NF3011和NF2734体内H2S的含量与野生型相比极显著降低(P<0.01);H2S信号在一定程度上抑制钙离子转运体编码基因MTR_6g027580的表达;外源生理浓度H2S熏蒸可诱导蒺藜苜蓿气孔关闭,与Ca2+通道阻断剂LaCl3联合处理对野生型气孔运动未产生影响,而在突变体中的结果截然相反;利用荧光探针测定保卫细胞内的Ca2+含量,所得结果与气孔孔径的变化规律完全一致。综上所述,H2S信号促进叶片保卫细胞内Ca2+的含量增加,最终表现为植物气孔孔径变小,在此过程中胞内Ca2+含量变化主要通过Ca2+转运体进行,少部分依赖Ca2+离子通道。该研究结果不仅在理论上丰富了H2S信号的作用机制,更具应用于苜蓿生产实践并推广于其他作物的潜力。  相似文献   

12.
The adjustment of Ca2+ entry in cardiac cells is critical to the generation of the force necessary for the myocardium to meet the physiological needs of the body. In this review, we present the concept that Ca2+ can promote its own entry through Ca2+ channels by different mechanisms. We refer to it under the general term of ‘Ca2+-induced Ca2+ entry’ (CICE). We review short-term mechanisms (usually termed facilitation) that involve a stimulating effect of Ca2+ on the L-type Ca2+ current (ICa-L) amplitude (positive staircase) or a lessening of Ca2+-dependent inactivation of ICa-L. This latter effect is related to the amount of Ca2+ released by ryanodine receptors (RyR2) of the sarcoplasmic reticulum (SR). Both effects are involved in the control of action potential (AP) duration. We also describe a long-term mechanism based on Ca2+-dependent down-regulation of the Kv4.2 gene controlling functional expression of the repolarizing transient outward K+ current (Ito) and, thereby, AP duration. This mechanism, which might occur very early during the onset of hypertrophy, enhances Ca2+ entry by maintaining Ca2+ channel activation during prolonged AP. Both Ca2+-dependent facilitation and Ca2+-dependent down-regulation of Ito expression favour AP prolongation and, thereby, promote sustained voltage-gated Ca2+ entry used to enhance excitation–contraction (EC) coupling (with no change in the density of Ca2+ channels per se). These self-maintaining mechanisms of Ca2+ entry have significant functions in remodelling Ca2+ signalling during the cardiac AP. They might support a prominent role of Ca2+ channels in the establishment and progression of abnormal Ca2+ signalling during cardiac hypertrophy and congestive heart failure.  相似文献   

13.
高Ca2+环境对许多植物的生长不利, 因此研究植物对高Ca2+环境的适应机制非常重要。研究发现, 拟南芥(Ara- bidopsis thaliana)镁转运体MGT7功能缺失突变体mgt7-1mgt7-2具有高Ca2+敏感表型: 在高Ca2+培养基上, 相对于野生型Col-0, 突变体叶鲜重显著下降, 但根长无显著差异。高Ca2+MGT7启动子活性和包括MGT7在内的镁转运体基因表达无显著调节作用。Col-0与mgt7突变体之间, 在外加Ca2+诱导细胞质Ca2+瞬时升高和Ca2+含量方面无显著差异; 但是, 在正常和高Ca2+培养基上, mgt7突变体的Mg含量均显著低于Col-0。高Ca2+显著抑制Col-0和mgt7突变体内Mg的积累。因此我们假设, mgt7突变体的高Ca2+敏感表型是由于其体内Mg含量下降导致的。进一步的研究证实, 只有增加培养基中Mg2+的含量, 而不是N、P、K和S, 才可以使突变体的高Ca2+敏感表型得到恢复。  相似文献   

14.
Measurements of Ca2+ influx and [Ca2+]i changes in Fura-2/AM-loaded prothoracic glands (PGs) of the silkworm, Bombyx mori, were used to identify Ca2+ as the actual second messenger of the prothoracicotropic hormone (PTTH) of this insect. Dose-dependent increases of [Ca2+]i in PG cells were recorded in the presence of recombinant PTTH (rPTTH) within 5 minutes. The rPTTH-mediated increases of [Ca2+]i levels were dependent on extracellular Ca2+. They were not blocked by the dihydropyridine derivative, nitrendipine, an antagonist of high-voltage-activated (HVA) Ca2+ channels, and by bepridil, an antagonist of low-voltage-activated (LVA) Ca2+ channels. The trivalent cation La3+, a non-specific blocker of plasma membrane Ca2+ channels, eliminated the rPTTH-stimulated increase of [Ca2+]i levels in PG cells and so did amiloride, an inhibitor of T-type Ca2+ channels. Incubation of PG cells with thapsigargin resulted in an increase of [Ca2+]i levels, which was also dependent on extracellular Ca2+ and was quenched by amiloride, suggesting the existence of store-operated plasma membrane Ca2+ channels, which can also be inhibited by amiloride. Thapsigargin and rPTTH did not operate independently in stimulating increases of [Ca2+]i levels and one agent’s mediated increase of [Ca2+]i was eliminated in the presence of the other. TMB-8, an inhibitor of intracellular Ca2+ release from inositol 1,4,5 trisphosphate (IP3)-sensitive Ca2+ stores, blocked the rPTTH-stimulated increases of [Ca2+]i levels, suggesting an involvement of IP3 in the initiation of the rPTTH signaling cascade, whereas ryanodine did not influence the rPTTH-stimulated increases of [Ca2+]i levels. The combined results indicate the presence of a cross-talk mechanism between the [Ca2+]i levels, filling state of IP3-sensitive intracellular Ca2+ stores and the PTTH-receptor’s-mediated Ca2+ influx.  相似文献   

15.
吕俊  于存 《菌物学报》2019,38(6):993-1002
白囊耙齿菌Irpex lacteus是分离自倒木上的一株可以分泌漆酶和锰过氧化物酶的白腐真菌。利用I. lacteus对固体条件下的活性黑、活性红、结晶紫、茜素红和孔雀石绿进行脱色能力的检测,通过单因素和正交试验优化I. lacteus对茜素红的脱色条件,并以3种作物发芽率为指标测定茜素红被I. lacteus脱色前后的毒性变化。结果显示,I. lacteus对5种染料均可脱色,其中对茜素红染料的脱色更为彻底;单因素和正交试验优化I. lacteus对茜素红的脱色条件为:pH 7.0、葡萄糖10.0g/L、硫酸铵0.66g/L、接种量2片(Φ=8.0mm)、100.0mL三角瓶装液20.0mL,优化条件下I. lacteus对茜素红脱色10d时的脱色率为88.26%,与未优化前的脱色率相比提高了60.50%;茜素红染料被I. lacteus脱色前后毒性大小排序为:染料原液>染料脱色后>PDB培养基处理,表明茜素红染料存在一定的毒性,I. lacteus脱色茜素红后可以使其毒性减弱。通过本研究,为I. lacteus在茜素红等染料废水脱色以及降低染料废水毒性方面的应用奠定基础。  相似文献   

16.
Protein phosphorylation in vitro was investigated in guard cells from Vicia faba. A number of proteins with apparent molecular masses of 72, 67, 57, 52, 49, 44, 37, and 26 kDa were phosphorylated when guard-cell extract was incubated with [γ-32P]ATP under Ca2+-free conditions. In the presence of Ca2+ at 1 μM, several proteins with apparent molecular masses of 125, 83, 41, 31, and 25 kDa were newly phosphorylated. These Ca2+-dependent protein phosphorylations were suppressed by (8R*,9S*,11S*)-(−)-9-hydroxy-9-methoxycarbonyl-8-methyl-2,3,9,10-tetrahydro-8,11-epoxy-1H,8H,11H-2,7b,11a- triazadibenzo[a,g]cycloocta[cde]trinden-1-one (K-252a), a wide-range inhibitor of protein kinases, suggesting that the protein phosphorylations were mediated by protein kinases. Several proteins were phosphorylated in vitro in mesophyll extract from Vicia. In contrast to guard cells, there was no detectable Ca2+-dependent protein phosphorylation in mesophyll cells. 1-(5-Indonaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine (ML-7), an inhibitor of myosin light chain kinase (MLCK), and an antagonist of calmodulin (CaM), N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7), inhibited Ca2+-dependent phosphorylation of 41- and 25-kDa proteins in guard cells. Fractionation experiments revealed that the Ca2+-dependent phosphorylated proteins with molecular masses of 41 and 25 kDa were present in the mitochondria, and the 125- and 31-kDa proteins in the cytosol. These results suggest that Ca2+-dependent protein phosphorylation occurs markedly in guard cells, and that Ca2+-dependent phosphorylation of 41- and 25-kDa proteins may be catalyzed by MLCK or MLCK-like protein kinase in guard cells.  相似文献   

17.
为了探明褪黑素(MT)和钙离子(Ca2+)在调控植物耐热性中是否存在互作关系,以黄瓜幼苗为试材,分析了内源MT和Ca2+对高温胁迫的响应;并通过叶面喷施100 μmol·L-1 MT、10 mmol·L-1 CaCl2、3 mmol·L-1乙二醇二乙醚二胺四乙酸(EGTA,Ca2+螯合剂)+100 μmol·L-1 MT、0.05 mmol·L-1氯丙嗪(钙调素拮抗剂,CPZ)+100 μmol·L-1 MT、100 μmol·L-1氯苯丙氨酸(p-CPA,MT合成抑制剂)+10 mmol·L-1 CaCl2和去离子水(H2O),研究高温下(42/32 ℃)外源MT和Ca2+对黄瓜幼苗活性氧积累、抗氧化系统及热激转录因子(HSF)和热激蛋白(HSPs)等的影响。结果表明: 黄瓜幼苗内源MT和Ca2+均受高温胁迫诱导;外源MT可上调常温下钙调素蛋白(CaM)、钙依赖蛋白激酶(CDPK5)、钙调磷酸酶B类蛋白(CBL3)、CBL结合蛋白激酶(CIPK2)mRNA表达;CaCl2处理的MT合成关键基因色氨酸脱羧酶(TDC)、5-羟色胺-N-乙酰转移酶(SNAT)和N-乙酰-5-羟色胺甲基转移酶(ASMT)水平也显著升高,MT含量快速增加。MT和CaCl2可显著增强高温下黄瓜的抗氧化能力,减少活性氧(ROS)积累,同时上调HSF7HSP70.1HSP70.11 mRNA表达,从而减轻高温胁迫引起的过氧化伤害,植株热害症状明显减轻,热害指数和电解质渗漏率显著降低。加入EGTA和CPZ后,MT对黄瓜幼苗抗氧化能力和热激蛋白表达的促进效应明显减弱,Ca2+对高温下黄瓜幼苗过氧化伤害的缓解效应也被p-CPA逆转。可见,MT和Ca2+均可诱导黄瓜幼苗的耐热性,二者在热胁迫信号转导过程中存在互作关系。  相似文献   

18.
以塔里木盆地南缘关键物种疏叶骆驼刺为材料,研究了不同盐渍土壤生境(轻度盐渍土、中度盐渍土、重度盐渍土)下其器官间Na+、K+、Ca2+、Mg2+的分布、吸收及运输特征,以探讨疏叶骆驼刺对自然盐渍生境的适应特性.结果表明: 在轻度和中度盐渍土生境,Na+在各器官中的分布规律为茎≈刺>叶>根,而在重度盐渍土生境,Na+分布规律为叶>茎≈刺>根;Ca2+和Mg2+在疏叶骆驼刺体内的分布规律为叶>刺>茎>根.随着土壤含盐量的增加,疏叶骆驼刺体内各器官Na+含量都增大,而叶片中K+含量呈下降趋势;根和叶器官中K+/Na+值明显降低,各器官中Ca2+/Na+、Mg2+/Na+值都降低.盐渍生境下,疏叶骆驼刺体内Ca2+选择性运输系数和Mg2+选择性运输系数均为茎-叶>茎-刺>根-茎.疏叶骆驼刺为适应盐渍生境,在土壤含盐量较低时,将Na+聚集于茎和刺;而在土壤含盐量较高时,则将Na+聚集于叶片.此外,Ca2+和Mg2+可能是疏叶骆驼适应盐渍生境的无机渗透调节物质.  相似文献   

19.
Ca2+ mobilization elicited by simulation with brief pulses of high K + were monitored with confocal laser scanned microscopy in intact, guinea pig cardiac myocytes loaded with the calcium indicator fluo-3. Single wavelength ratioing of fluorescence images obtained after prolonged integration times revealed non-uniformities of intracellular Ca2+ changes across the cell, suggesting the presence of significant spatial Ca2+ gradients. Treatment with 20 μM ryanodine, an inhibitor of Ca2+ release from the SR, and 10 μM verapamil, a calcium channel blocker, reduced by 42% and 76% respectively the changes in [Ca2+]i elicited by membrane depolarization. The overall spatial distribution of [Ca2+]i changes appeared unchanged. Ca2+ transients recorded in the presence of verapamil and ryanodine (about 20% of the size of control responses), diminished in the presence of 50 μM 2-4 Dichlorbenzamil (DCB) or 5 mM nickel, two relatively specific inhibitors of the exchange mechanism. Conversely, when the reversal potential of the exchange was shifted to negative potentials by lowering [Na+]0 or by increasing [Na+]i by treatment with 20 μM monensin, the amplitude of these Ca2+ transients increased. Ca2+ transients elicited by membrane depolarization and largely mediated by reverse operation of Na+-Ca2+ exchange could be recorded in the presence of ryanodine, verapamil and monensin. These findings suggest that in intact guinea pig cardiac cells, Ca2+ influx through the exchange mechanism activated by a membrane depolarization in the physiological range can be sufficient to play a significant role in excitation-contraction coupling.  相似文献   

20.
The effects of calcium ions (Ca2+) on the stability of artichoke (Cynara scolymus L.) peroxidase (AKPC) have been studied. The thermal stability of AKPC was improved by the addition of Ca2+; the melting temperature increased by 20 °C and the deactivation energy by 26 kJ mol−1. AKPC was stable in a selection of organic solvents but was less active with 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid) than under aqueous conditions. Ca2+-free AKPC retained more activity in the presence of organic solvents due to its better maintenance of the rate of compound I formation with hydrogen peroxide (H2O2) compared to AKPC-Ca2+. AKPC retained at least 75% activity over 24 h in the pH range 3.0–10.5 and about 50% over 1 month at pH 7.0 or 5.5, irrespective of the Ca2+ content. AKPC-Ca2+ was considerably more resistant to inactivation by H2O2 than Ca2+-free AKPC suggesting that the presence of Ca2+ boosts turnover under oxidizing conditions. AKPC has been applied as an alternative to horseradish peroxidase (HRP) in glucose concentration assays; the presence of Ca2+ or of the Ca2+ chelating agent ethylenediaminetetraacetic acid made no difference to the final result. The possibility is discussed that addition and removal of a labile Ca2+ from AKPC could be used to control enzyme activity both in vivo and in vitro.  相似文献   

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