Effects of calcium ions and the bivalent cation ionophore A23187 on the agglutination and fusion of chicken erythrocytes by Sendai virus.
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ABSTRACT:
SUBMITTER: Hart CA
PROVIDER: S-EPMC1163947 | biostudies-other | 1976 Jul
REPOSITORIES: biostudies-other
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Project description:Incubation of hen erythrocytes with Ca2+ and the bivalent-cation ionophore A23187 induced slight cell fusion in 1 h at 37 degrees C, and extensive fusion during a subsequent 15 min at 47 degrees C. Redistributions of intramembranous particles were observed, possibly involving interactions between Ca2+ and phospholipids, which are discussed in relation to molecular mechanimss of cell fusion.
| S-EPMC1164023 | biostudies-other
Project description:Using two cDNA clones which encode hamster genes specifically induced by glucose starvation, we demonstrated that an 8- and 30-fold increase, respectively, in the transcription rates of these genes was coordinately effected by calcium ionophore A23187 treatment, resulting in a similar increase in the steady-state levels of their mRNAs. This response was observed within several hours of ionophore treatment in several mammalian cell types and appeared to be specifically mediated by A23187 but not by other ionophores in general. To define the regulatory sequence which mediates this Ca2+-induced response, we showed by gene transfection techniques that the 5' flanking sequence of a rat glucose-regulated gene contained the region for induction by A23187. The system reported here offers attractive features for the study of specific gene regulation by Ca2+.
| S-EPMC366848 | biostudies-literature
Project description:1. When the ionophore A23187 and Ca2+ were added to normal human erythrocytes, the incorporation of 32P into phosphatidate was enhanced within 1 min, but there was only slight labelling of other phospholipids. 2. Labelling of phosphatidate in these cells did not continue to increase after about 20min at 37 degrees C; by this time, radioactivity in phosphatidate was about ten times higher inionophore A23187-treated cells than in controls. A net synthesis of phosphatidate was measured in response to the increase in intracellular Ca2+ concentration; the content of this phospholipid in the cell was increased by approximately 50%. 3. In the presence of 2.5 mM-Ca2+ a maximum effect was seen with about 0.5 mug of ionophore/ml. 4. The concentration of Ca2+ giving half-maximal labelling of phosphatidate in the presence of 10 mug of ionophore A23187/ml was about 10 muM. 5. A rapid decrease of ATP content in the cell occurred in ionophore-treated cells. 6. Labelling of phosphatidate appeared to be secondary to the production of 1,2-diacylglycerol in the cells; accumulation of 1,2-diacylglycerol was only seen after about 15 min. After 60 min, the 1,2-diacylglycerol content of the cells was five to seven times that of untreated control cells. 7. The change in the shape of erythrocytes treated with Ca2+ and ionophore appeared to be related to accumulation of 1,2-diacylglycerol. 8. The source of 1,2-diacylglycerol has not been definitely identified, but its fatty acid compositon was similar to that of phosphatidylcholine. However, it has an unusually high content of hexadecenoic acid, a fatty acid not common in the major erythrocyte phospholipids. 9. Accumulation of 1,2-diacyglycerol also occurred in energy-starved cells, even in the absence of calcium; in this case it appeared to be produced by phosphatidate breakdown.
| S-EPMC1163740 | biostudies-other
Project description:Previous studies have demonstrated that A23187, an ionophore which selectively transports divalent cations across cell membranes, has profound effects on human erythrocytes: it causes red cells to take up calcium; lose potassium, water, and ATP; convert from biconcave discs to echinocytes and spheroechinocytes; and become more rigid. The present study has explored the influence of calcium uptake induced by the ionophore on the behavior of individual erythrocyte membranes by the micropipette aspiration technique. Exposure of erythrocytes to calcium and A23187 for intervals of up to 30 minutes resulted in marked changes in membrane viscoelastic properties, including the development of increased resistance to aspiration. The most striking manifestation of altered membrane mechanics was apparent after 10 minutes on incubation. Cells pulled into the pipette for a few seconds and the extruded back into the medium retained the deformity imposed by the pipette for several seconds to a few minutes before regaining the form they manifested prior to initial aspiration. The calcium-induced changes in erythrocyte behavior observed in this study strongly support the concept that extrinsic proteins located inside the membrane provide mechanical support to the cell wall, and that increased levels of calcium cause precipitation or cross-linking of the proteins responsible for the increased resistence to deformation and recoil observed after aspiration into micropipettes.
| S-EPMC2032159 | biostudies-other
Project description:1. The bivalent cation ionophore A23187 was used to increase the intracellular concentration of Ca2+ in pigeon erythrocytes to investigate whether the increase in cyclic AMP content caused by adrenaline might be influenced by a change in intracellular Ca2+ in intact cells. 2. Incubation of cells with adrenaline, in the concentration range 0.55--55 muM, resulted in an increase in the concentration of cyclic AMP over a period of 60 min. The effect of adrenaline was inhibited by more than 90% with ionophore A23187 (1.9 muM) in the presence of 1 mM-Ca2+. This inhibition could be decreased by decreasing either the concentration of the ionophore or the concentration of extracellular Ca2+, and was independent of the concentration of adrenaline. 3. The effect of ionophore A23187 depended on the time of incubation. Time-course studies showed that maximum inhibition by ionophore A23187 was only observed when the cells were incubated with the ionophore for at least 15 min before the addition of adrenaline. 4. The inhibition by ionophore A23187 depended on the concentration of extracellular Ca2+. In the absence of Mg2+, ionophore A23187 (1.9 muM) inhibited the effect of adrenaline by approx. 30% without added Ca2+, by approx. 66% with 10 muM-Ca2+ and by more than 90% with concentrations of added Ca2+ greater than 30 muM. However, even in the presence of EGTA [ethanedioxybis(ethylamine)tetra-acetate](0.1--10 mM), ionophore A23187 caused an inhibition of the cyclic AMP response of at least 30%, which may have been due to a decrease in cell Mg2+ concentration. 5. The addition of EGTA after incubation of cells with ionophore A23187 resulted in a partial reversal of the inhibition of the effect of adrenaline. 6. Inclusion of Mg2+ (2 mM) in the incubation medium antagonized the inhibitory action of ionophore A23187. This effect was most marked when the ionophore A23187 was added to medium containing Mg2+ before the addition of the cells. 7. The cellular content of Mg2+ was decreased by approx. 50% after 20 min incubation with ionophore A23187 (1.9 muM) in the presence of Ca2+ (1 mM) but no Mg2+. When Mg2+ (2 mM) was also present in the medium, ionophore A23187 caused an increase of approx. 80% in cell Mg2+ content. Ionophore A23187 had no significant effect on cell K+ content. 8. Ionophore A23187 caused a decrease in cell ATP content under some conditions. Since effects on cyclic AMP content could also be shown when ATP was not significanlty lowered, it appeared that a decrease in ATP in the cells could not explain the effect of ionophore A23187 on cyclic AMP. 9. Ionophore A23187 (1.9 muM), with 1 mM-Ca2+, did not enhance cyclic AMP degradation in intact cells, suggesting that the effect of ionophore A23187 on cyclic AMP content was mediated through an inhibition of adenylate cyclase rather than a stimulation of cyclic AMP phosphodiesterase. 10. It was concluded that in intact pigeon erythrocytes adenylate cyclase may be inhibited by intracellular concentrations of Ca2+ in the range 1-10 muM.
| S-EPMC1163961 | biostudies-other
Project description:The possibility that Ca2+ ions are involved in the control of the increased phosphatidylinositol turnover which is provoked by alpha-adrenergic or muscarinic cholinergic stimulation of rat parotid-gland fragments has been investigated. Both types of stimulation provoked phosphatidylinositol breakdown, which was detected either chemically or radiochemically, and provoked a compensatory synthesis of the lipid, detected as an increased rate of incorporation of 32Pi into phosphatidylinositol. Acetylcholine had little effect on the incorporation of labelled glycerol, whereas adrenaline stimulated it significantly, but to a much lower extent than 32P incorporation: this suggests that the response to acetylcholine was entirely accounted for by renewal of the phosphorylinositol head-group of the lipid, but that some synthesis de novo was involved in the response to adrenaline. The responses to both types of stimulation, whether measured as phosphatidylinositol breakdown or as phosphatidylinositol labelling, occurred equally well in incubation media containing 2.5 mm-Ca2+ or 0.2 mm-EGTA [ethanedioxybis(ethylamine)-tetra-acetic acid]. Incubation with a bivalent cation ionophore (A23187) led to a small and more variable increase in phosphatidylinositol labelling with 32Pi, which occurred whether or not Ca2+ was available in the extracellular medium: this was not accompanied by significant phosphatidylinositol breakdown. Cinchocaine, a local anaesthetic, produced parallel increases in the incorporation of Pi and glycerol into phosphatidylinositol. This is compatible with its known ability to inhibit phosphatidate phosphohydrolase (EC 3.1.3.4) and increase phosphatidylinositol synthesis de novo in other cells. These results indicate that the phosphatidylinositol turnover evoked by alpha-adrenergic or muscarinic cholinergic stimuli in rat parotid gland probably does not depend on an influx of Ca2+ into the cells in response to stimulation. This is in marked contrast with the K+ efflux from this tissue, which is controlled by the same receptors, but is strictly dependent on the presence of extracellular Ca2+. The Ca2+-independence of stimulated phosphatidylinositol metabolism may mean that it is controlled through a mode of receptor function different from that which controls other cell responses. Alternatively, it can be interpreted as indicating that stimulated phosphatidylinositol breakdown is intimately involved in the mechanisms of action of alpha-adrenergic and muscarinic cholinergic receptor systems.
| S-EPMC1165566 | biostudies-other
Project description:In addition to the microvesicles released during the treatment of human erythrocytes with Ca2+ and ionophore A23187, a new subpopulation of still smaller dense vesicles ('nanovesicles') has now been identified. Nanovesicles are about 60 nm in diameter, have an acetylcholinesterase activity higher than that of microvesicles and appear to be relatively enriched in sphingomyelin and correspondingly depleted of phosphatidylethanolamine. They have a polypeptide composition quite different from those of erythrocyte membranes or microvesicles, consisting largely of components of mol.wts. 60 000 and 26 000 in addition to haemoglobin. These two major polypeptides do not appear to represent contaminating cytoplasmic proteins or proteolytic subfragments of a larger protein.
| S-EPMC1161973 | biostudies-other
Project description:Ionophores are small molecules or peptides that transport metal ions across biological membranes. Their transport capabilities are typically characterized in vitro using vesicles and single ion species. It is difficult to infer from these data which effects ionophores have on living cells in a complex environment (e.g. culture medium), since net transport is influenced e.g. by ion composition, concentration gradients, pH gradient, and active transport. To gain insights into the antibacterial mechanism of the semi-synthetic polyether ionophore 4-Br-A23187, we investigated its effects on the gram-positive model organism Bacillus subtilis. Changes in intracellular ion concentrations were determined using ionomics and the physiological impact was assessed by proteomics. 4-Br-A23187 treatment resulted in an increase in intracellular copper levels, an effect that was dependent on the copper concentration of the medium. The antibiotic effect of 4-Br-A23187 is further aggravated by a decrease in intracellular manganese and magnesium. Effects of copper hyperaccumulation, mirrored by the proteomic response, included oxidative stress, disturbance of proteostasis, metal, and sulfur homeostasis. Especially solvent-exposed iron-sulfur clusters like that of aconitase of the citric acid cycle are readily destabilized by copper. Using calcein fluorescence quenching as a readout, we confirmed in vitro that 4-Br-A23187 acts as copper ionophore.
2022-06-08 | PXD028938 | Pride
Project description:Ca(2+) ionophore A23187 is known to induce the acrosome reaction of mammalian spermatozoa, but it also quickly immobilizes them. Although mouse spermatozoa were immobilized by this ionophore, they initiated vigorous motility (hyperactivation) soon after this reagent was washed away by centrifugation. About half of live spermatozoa were acrosome-reacted at the end of 10 min of ionophore treatment; fertilization of cumulus-intact oocytes began as soon as spermatozoa recovered their motility and before the increase in protein tyrosine phosphorylation, which started 30-45 min after washing out the ionophore. When spermatozoa were treated with A23187, more than 95% of oocytes were fertilized in the constant presence of the protein kinase A inhibitor, H89. Ionophore-treated spermatozoa also fertilized 80% of oocytes, even in the absence of HCO3(-), a component essential for cAMP synthesis under normal in vitro conditions. Under these conditions, fertilized oocytes developed into normal offspring. These data indicate that mouse spermatozoa treated with ionophore are able to fertilize without activation of the cAMP/PKA signaling pathway. Furthermore, they suggest that the cAMP/PKA pathway is upstream of an intracellular Ca(2+) increase required for the acrosome reaction and hyperactivation of spermatozoa under normal in vitro conditions.
| S-EPMC3831971 | biostudies-literature
Project description:Calcium is a crucial factor in regulating the biological behavior of cells. The imbalance of calcium homeostasis in cytoplasm will cause abnormal behavior of cells and the occurrence of diseases. In intracytoplasmic sperm injection (ICSI) cycle, the dysfunction of oocyte activation caused by insufficient release of Ca2+ from endoplasmic reticulum is one of the main reasons for repeated fertilization failure. Calcium ionophore (A23187) is a highly selective calcium ionophore, which can form stable complex with Ca2+ and pass through the cell membrane at will, effectively increasing intracellular Ca2+ levels. It has been reported that calcium ionophore (A23187) can activate oocytes and obtain normal embryos. However, there are few studies on unfertilized oocytes after calcium ionophore (A23187) rescue activation in ICSI cycle. The purpose of this study was to analyze the effects of calcium ionophore (A23187) rescue activation on the activation of unfertilized oocytes, embryonic development potential, embryonic development timing and chromosomal aneuploidy, and to compare and analyze the clinical data of patients with calcium ionophore (A23187) activation in clinical application. The results showed that a certain proportion of high-quality blastocysts with normal karyotype could be obtained after calcium ionophore (A23187) rescue activation of unfertilized oocytes, and it did not have a significant effect on the timing of embryo development. In clinical practice, direct activation with calcium ionophore (A23187) after ICSI was better than rescue activation the next day. In conclusions, the studies on the effectiveness and safety of calcium ionophore (A23187) rescue activation for oocytes with ICSI fertilization failure can enable some patients to obtain usable, high-quality embryos during the first ICSI cycle.
| S-EPMC8320372 | biostudies-literature