Oscillations of cytosolic sodium during calcium oscillations in exocrine acinar cells

In acinar cells from rat salivary glands, cholinergic agonists cause oscillations in cytoplasmic free calcium concentration, which then drive oscillations of cell volume that reflect oscillating cell solute content and fluid secretion. By quantitative fluorescence ratio microscopy of an intracellular indicator dye for sodium, it has now been shown that large amplitude oscillations of sodium concentration were associated with the calcium and cell volume oscillations. Both calcium and sodium oscillations were dependent on the continued presence of calcium in the extracellular medium and were abolished by the specific sodium-potassium adenosine triphosphatase inhibitor ouabain. Thus, calcium oscillations in salivary acinar cells, by modulating the activities of ion transport pathways in the plasma membrane, can cause significant oscillations of monovalent ions that may in turn feed back to regulate calcium oscillations and fluid secretion.     

Calcium gradients underlying polarization and chemotaxis of eosinophils

The concentration of intracellular free calcium ([Ca2+]i) in polarized eosinophils was imaged during chemotaxis by monitoring fluorescence of the calcium-sensitive dye Fura-2 with a modified digital imaging microscope. Chemotactic stimuli caused [Ca2+]i to increase in a nonuniform manner that was related to cell activity. In cells moving persistently in one direction, [Ca2+]i was highest at the rear and lowest at the front of the cell. Before cells turned, [Ca2+]i transiently increased. The region of the cell that became the new leading edge had the lowest [Ca2+]i. These changes in [Ca2+]i provide a basis for understanding the organization and local activity of cytoskeletal proteins thought to underlie the directed migration of many cells.     

Calcium rises abruptly and briefly throughout the cell at the onset of anaphase

Continuous measurement and imaging of the intracellular free calcium ion concentration ([Ca2+]i) of mitotic and interphase PtK1 cells was accomplished with the new fluorescent Ca2+ indicator fura-2. No statistically significant difference between basal [Ca2+]i of interphase and mitotic cells was detected. However, mitotic cells showed a rapid elevation of [Ca2+]i from basal levels of 130 nM to 500 to 800 nM at the metaphase-anaphase transition. The [Ca2+]i transient was brief, lasting approximately 20 seconds and the elevated [Ca2+]i appeared uniformly distributed over the entire spindle and central region of the cell. The close temporal association of the [Ca2+]i transient with the onset of anaphase suggests that calcium may have a signaling role in this event.     

Free calcium at rest during "catch" in single smooth muscle cells

Tension and intracellular free calcium concentration [( Ca2+]i) were measured simultaneously in single smooth muscle cells isolated from the anterior byssus retractor muscle (ABRM) of Mytilus edulis that were loaded with the fluorescent Ca2+ indicator fura-2. Electrical stimulation evoked a transient elevation of [Ca2+]i associated with a "catch" contraction. During the catch state, however, [Ca2+]i was effectively at its resting level and was unaffected by 5-hydroxytryptamine, which induced a rapid relaxation from catch. The results indicate that a maintained high [Ca2+]i is not required for the maintenance of catch tension in intact ABRM and that there was no significant change in [Ca2+]i upon abolition of catch.     

Cellular and subcellular heterogeneity of [Ca2+]i in single heart cells revealed by fura-2

Digital imaging of calcium indicator signals (fura-2 fluorescence) from single cardiac cells has revealed different subcellular patterns of cytoplasmic calcium ion concentration ([Ca2+]i) that are associated with different types of cellular appearance and behavior. In any population of enzymatically isolated rat heart cells, there are mechanically quiescent cells in which [Ca2+]i is spatially uniform, constant over time, and relatively low; spontaneously contracting cells, which have an increased [Ca2+]i, but in which the spatial uniformity of [Ca2+]i is interrupted periodically by spontaneous propagating waves of high [Ca2+]i; and cells that are hypercontracted (rounded up) and that have higher levels of [Ca2+]i than the other two types. The observed cellular and subcellular heterogeneity of [Ca2+]i in isolated cells indicates that experiments performed on suspensions of cells should be interpreted with caution. The spontaneous [Ca2+]i fluctuations previously observed without spatial resolution in multicellular preparations may actually be inhomogeneous at the subcellular level.     

Coupling diurnal cytosolic Ca2+ oscillations to the CAS-IP3 pathway in Arabidopsis

Various signaling pathways rely on changes in cytosolic calcium ion concentration ([Ca2+]i). In plants, resting [Ca2+]i oscillates diurnally. We show that in Arabidopsis thaliana, [Ca2+]i oscillations are synchronized to extracellular Ca2+ concentration ([Ca2+]o) oscillations largely through the Ca2+-sensing receptor CAS. CAS regulates concentrations of inositol 1,4,5-trisphosphate (IP3), which in turn directs release of Ca2+ from internal stores. The oscillating amplitudes of [Ca2+]o and [Ca2+]i are controlled by soil Ca2+ concentrations and transpiration rates. The phase and period of oscillations are likely determined by stomatal conductance. Thus, the internal concentration of Ca2+ in plant cells is constantly being actively revised.     

Sodium-calcium exchange in heart: membrane currents and changes in [Ca2+]i

Recordings have been made of changes in intracellular calcium ion concentration ([Ca2+]i) that can be attributed to the operation of an electrogenic, voltage-dependent sodium-calcium (Na-Ca) exchanger in mammalian heart cells. Guinea pig ventricular myocytes under voltage clamp were perfused internally with fura-2, a fluorescent Ca2+-indicator, and changes in [Ca2+]i and membrane current that resulted from Na-Ca exchange were identified through the use of various organic channel blockers and impermeant ions. Depolarization of cells elicited slow increases in [Ca2+]i, with the maximum increase depending on internal [Na+], external [Ca2+], and membrane voltage. Repolarization was associated with net Ca2+ efflux and a decline in the inward current that developed instantaneously upon repolarization. The relation between [Ca2+]i and current was linear, and the slope was made steeper by hyperpolarization.     

姬松茸多糖诱导HepG2 细胞凋亡的线粒体通路研究

为探讨姬松茸多糖诱导HepG2细胞凋亡过程中线粒体膜电位和细胞内Ca2+浓度([Ca2+]i)的变化.以人肝癌细胞系HepG2细胞为对象,使用流式细胞仪检测姬松茸多糖对HepG2细胞凋亡及线粒体膜电位的影响,激光共聚焦显微镜检测HepG2细胞[Ca2+]i的变化.结果表明姬松茸多糖能显著诱导HePG2细胞凋亡,并不同程度降低HepG2细胞线粒体跨膜电位,升高细胞[Ca2+]i,造成钙稳态失衡.姬松茸多糖诱导HepG2细胞凋亡的机制与降低线粒体膜电位,造成细胞内Ca2+超载有关.     

Erratum

In the report "Activation of salivary secretion: Coupling of cell volume and [Ca(2+)](i) in single cells" by J. K. Foskett and J. E. Melvin (30 June, p. 1582), figures 1 and 2 were inadvertently interchanged. The correct figures are printed below. [See Figures in source pdf.].     

Calcium sensitivity of glutamate release in a calyx-type terminal

Synaptic efficacy critically depends on the presynaptic intracellular calcium concentration ([Ca2+]i). We measured the calcium sensitivity of glutamate release in a rat auditory brainstem synapse by laser photolysis of caged calcium. A rise in [Ca2+]i to 1 micromolar readily evoked release. An increase to >30 micromolar depleted the releasable vesicle pool in <0.5 millisecond. A comparison with action potential-evoked release suggested that a brief increase of [Ca2+]i to approximately 10 micromolar would be sufficient to reproduce the physiological release pattern. Thus, the calcium sensitivity of release at this synapse is high, and the distinction between phasic and delayed release is less pronounced than previously thought.     

Generation of calcium oscillations in fibroblasts by positive feedback between calcium and IP3

A wide variety of nonexcitable cells generate repetitive transient increases in cytosolic calcium ion concentration ([Ca2+]i) when stimulated with agonists that engage the phosphoinositide signalling pathway. Current theories regarding the mechanisms of oscillation disagree on whether Ca2+ inhibits or stimulates its own release from internal stores and whether inositol 1,4,5-trisphosphate (IP3) and diacylglycerol (DG) also undergo oscillations linked to the Ca2+ spikes. In this study, Ca2+ was found to stimulate its own release in REF52 fibroblasts primed by mitogens plus depolarization. However, unlike Ca2+ release in muscle and nerve cells, this amplification was insensitive to caffeine or ryanodine and required hormone receptor occupancy and functional IP3 receptors. Oscillations in [Ca2+]i were accompanied by oscillations in IP3 concentration but did not require functional protein kinase C. Therefore, the dominant feedback mechanism in this cell type appears to be Ca2+ stimulation of phospholipase C once this enzyme has been activated by hormone receptors.     

Ca(2+)-induced Ca2+ release in sea urchin egg homogenates: modulation by cyclic ADP-ribose

Calcium-induced calcium release (CICR) may function widely in calcium-mediated cell signaling, but has been most thoroughly characterized in muscle cells. In a homogenate of sea urchin eggs, which display transients in the intracellular free calcium concentration ([Ca2+]i) during fertilization and anaphase, addition of Ca2+ triggered CICR. Ca2+ release was also induced by the CICR modulators ryanodine and caffeine. Responses to both Ca2+ and CICR modulators (but not Ca2+ release mediated by inositol 1,4,5-trisphosphate) were inhibited by procaine and ruthenium red, inhibitors of CICR. Intact eggs also displayed transients of [Ca2+]i when microinjected with ryanodine. Cyclic ADP-ribose, a metabolite with potent Ca(2+)-releasing properties, appears to act by way of the CICR mechanism and may thus be an endogenous modulator of CICR. A CICR mechanism is present in these nonmuscle cells as is assumed in various models of intracellular Ca2+ wave propagation.     

Inositol 1,3,4,5-tetrakisphosphate induces Ca2+ sequestration in rat liver cells

Inositol 1,4,5-trisphosphate [I(1,4,5)P3] is a second messenger generated along with diacylglycerol upon the binding of various physiological agents with their cell surface receptors. I(1,4,5)P3 mobilizes Ca2+ from intracellular storage sites through a receptor-coupled mechanism, and the subsequent increased intracellular free calcium ion concentration [( Ca2+]i) activates a multitude of cellular responses. Electropermeabilized neoplastic rat liver epithelial (261B) cells were used to study Ca2+ sequestration, a process that reverses the elevated [Ca2+]i to resting levels and replenishes intracellular Ca2+ pools. Although I (1,4,5)P3-mobilized Ca2+ is readily sequestered into storage pools by the action of Ca2+-adenosine triphosphatases, Ca2+ mobilized by addition of the nonmetabolized inositol trisphosphate isomer I(2,4,5)P3 is not sequestered, suggesting that metabolism is necessary to eliminate the stimulus for Ca2+ release. Several inositol phosphate compounds were examined for their ability to lower the buffer [Ca2+] to determine if a specific I(1,4,5)P3 metabolite might be involved in stimulating Ca2+ sequestration; of these, I(1,3,4,5)P4 alone was found to induce Ca2+ sequestration, demonstrating a physiological role for this inositol trisphosphate metabolite.     

TRPM4 regulates calcium oscillations after T cell activation

TRPM4 has recently been described as a calcium-activated nonselective (CAN) cation channel that mediates membrane depolarization. However, the functional importance of TRPM4 in the context of calcium (Ca2+) signaling and its effect on cellular responses are not known. Here, the molecular inhibition of endogenous TRPM4 in T cells was shown to suppress TRPM4 currents, with a profound influence on receptor-mediated Ca2+ mobilization. Agonist-mediated oscillations in intracellular Ca2+ concentration ([Ca2+]i), which are driven by store-operated Ca2+ influx, were transformed into a sustained elevation in [Ca2+]i. This increase in Ca2+ influx enhanced interleukin-2 production. Thus, TRPM4-mediated depolarization modulates Ca2+ oscillations, with downstream effects on cytokine production in T lymphocytes.     

Role of Calcium Ion in Apoptosis of MD Cancer Cells Induced by Arsenic Trioxide

In order to observe the role of calcium ion in apoptosis of MD cancer cells induced by arsenic trioxide, inhibition percentage was detected by MTT assay; morphology changes were examined by fluorescence microscope; apoptosis was examined by DNA Ladder; [Ca^2＋]i was investigated by spectrofluorimeter in vitro on MDCC-MSB 1 cells. The results showed that As2O3 inhibited the proliferation of MDCC-MSB1 cells in concentration dependent manner （P〈0.05 or P〈0.01）; typical apoptosis character was observed by fluorescence microscope; DNA Ladder was observed; the [Ca^2＋]i was elevated significantly after the treatment of As203 （P〈0.05 or P〈0.01） and showed a dose-dependent manner. It is concluded that the calcium may play an important role in apoptosis of MD cancer cells induced by arsenic trioxide.     

Regulation of phagocytosis and [Ca2+]i flux by distinct regions of an Fc receptor.

The binding of multivalent immunoglobulin G complexes to Fc receptors (Fc gamma Rs) on macrophages activates multiple immune functions. A murine macrophage cell line, but not a fibroblast cell line, that was transfected with human Fc gamma RIIA mediated phagocytosis and an intracellular Ca2+ concentration ([Ca2+]i) flux upon cross-linking of human Fc gamma RIIA. Transfected macrophages that expressed a truncated receptor lacking 17 carboxy-terminal amino acids phagocytosed small antibody complexes. However, only wild-type transfectants phagocytosed labeled erythrocytes and fluxed [Ca2+]i. Thus, the cytoplasmic domain of human Fc gamma RIIA contains distinct functional regions.     

Extracellular and Intracellular Calcium both Involved in the Jasmonic Acid Induced Calcium Mobilization in Arabidopsis thaliana

The objective of this experiment is to evaluate the role of intracellular and extracellular Ca2＋ and calmodulin （CAM） in jasmonic acid （JA） signaling. The laser scanning microscopy was used to detect the changes of [Ca2＋]cyt of Arabidopsis thaliana leaf cells which pretreated with different types of calcium channel blocker. Moreover, the expression of VSP, one of JA response genes, was also investigated after pretreated with the above blocker and antagonist of CaM. The results showed that extracellular and intracellular calcium both involved in the JA-induced Ca2＋ mobilization, and then Ca2＋ exerted its functions through activating the CaM or CaM related proteins. The apoplast calcium influx and the calcium release from the calcium stores are both involved in the JA-induced calcium mobilization, then the JA-induced Ca2＋ transmited the JA signal through CaM or CaM related proteins, and regulated the JA responsive genes.     

Ca（2＋）和茉莉酸对菠菜CBF表达的影响

[目的]为探究钙离子、茉莉酸（Jasmonic acid,JA）对菠菜CBF转录因子表达的影响。[方法]以菠菜（Spinacia oleraceaL.）为材料,对其进行低温（4℃）或JA处理或用钙离子载体A23187预处理菠菜幼苗,利用Northern印迹技术检测各处理时CBF的表达情况。[结果]低温和JA均可诱导菠菜中CBF基因的表达。[结论]推测低温可能首先导致细胞中JA含量升高,JA通过一定的机制诱导细胞质基质中游离钙离子浓度（[Ca2＋]cyt）升高,然后Ca2＋可能通过CaM（Calmodulin）或CaM相关蛋白传递低温信号,从而诱导CBF的表达。     

胞外及胞内Ca~(2+)共同参与拟南芥中茉莉酸诱导的钙动员

【目的】探究茉莉酸(Jasmonic acid,JA)诱导的钙动员的来源及钙调素(CaM)在JA信号通路中的作用。【方法】以拟南芥(Arabidopsis thaliana columbia)为材料,采用不同类型的离子通透性通道抑制剂(或拮抗剂)预处理拟南芥叶肉细胞,利用激光共聚焦显微镜检测其对JA诱导的[Ca2+]cyt升高的影响;同时还利用上述拮抗剂、钙调素(CaM)拮抗剂处理10d的拟南芥幼苗,采用Northern blot方法检测了其对JA诱导的VSP表达的影响。【结果】胞内及胞外钙离子共同参与JA诱导的钙动员,JA诱导的Ca2+通过CaM或CaM相关蛋白进一步传递JA信号。【结论】质外体钙离子的流入和胞内钙库中钙离子的释放均参与JA诱导的钙离子动员,Ca2+可能通过CaM或CaM相关蛋白传递JA信号,进一步诱导JA反应基因的表达。     

IP3 receptor types 2 and 3 mediate exocrine secretion underlying energy metabolism

Type 2 and type 3 inositol 1,4,5-trisphosphate receptors (IP3R2 and IP3R3) are intracellular calcium-release channels whose physiological roles are unknown. We show exocrine dysfunction in IP3R2 and IP3R3 double knock-out mice, which caused difficulties in nutrient digestion. Severely impaired calcium signaling in acinar cells of the salivary glands and the pancreas in the double mutants ascribed the secretion deficits to a lack of intracellular calcium release. Despite a normal caloric intake, the double mutants were hypoglycemic and lean. These results reveal IP3R2 and IP3R3 as key molecules in exocrine physiology underlying energy metabolism and animal growth.