The peripheral Ca pump activity of macrophages and neutrophils

Exposure of either alveolar macrophages or blood neutrophils to 0.2 – 1 μM ionophore A23187 in the presence of 0.1 – 1 mM CaCl₂ causes a rapid extracellular release of Ca²⁺, which can be measured by a Ca²⁺-selective electrode. The initial rate at which the cation is extruded from the cells is about 0.1 – 0.2 μg-ions/min/ml of cell water. ATP depletion, but not replacement of extracellular Na⁺ with choline, produces a marked inhibition of Ca²⁺ release from macrophages. When the movements of Ca²⁺ between neutrophils and the incubation medium are followed by an isotopic technique, a transient increase in cell-associated ⁴⁵Ca²⁺ is detected a few seconds after the addition of the ionophore. We suggest that the ionophore A23187 mobilises Ca²⁺ from intracellular stores, with a subsequent cell extrusion of the bivalent cation catalysed by a pump localised at the cell surface. These and other data are consistent with the conclusion that the peripheral Ca²⁺ pump system of macrophages and neutrophils is very similar to the well know Ca²⁺ pump of the red cells with regard to mechanism and capacity.     

Effects of ionophore A23187 on calcium flux and amylase release in isolated mouse pancreatic acini

The divalent cation ionophore A23187 has been used extensively to demonstrate the importance of Ca²⁺ in the control of pancreatic enzyme secretion. The relative importance, however, of the ability of the ionophore to facilitate Ca²⁺ movement across plasma and intracellular membranes in the stimulation of amylase release is not clear. We therefore studied these relationships in isolated pancreatic acini, a preparation in which it is possible to precisely measure both ⁴⁵Ca²⁺ fluxes, Ca²⁺ content and amylase release. A23187 increased the initial rates of both ⁴⁵Ca²⁺ uptake and washout. In addition, the content of both exchangeable ⁴⁵Ca²⁺ and total Ca²⁺ were reduced. These results indicated, therefore, that A23187 increases Ca²⁺ fluxes across both plasma and intracellular membranes. Consistent with this observation, the initial stimulation of amylase release by A23187 was independent of extracellular Ca²⁺. In the absence of extracellular Ca²⁺, however, A23187 caused a rapid fall in acinar Ca²⁺ and subsequent amylase release was abolished. Depletion of intracellular Ca²⁺ by the ionophore also blocked the subsequent stimulation by cholecystokinin (CCK). The results indicate certain similarities in the actions of A23187 and CCK on pancreatic acini; both the agonists have striking effects on intracellular Ca²⁺ which in turn mediates their actions.     

Secretion of granule enzymes from alveolar macrophages : Regulation by intracellular Ca-buffering capacity

Rabbit alveolar macrophages exposed to the ionophore A23187 in the presence of extracellular Ca²⁺ take up about 12 nmoles of Ca²⁺/1 × 10⁶ cells. This uptake induces a slight, but significant, extracellular release of granule enzymes, β-glucuronidase and lysozyme, but not of the cytoplasmic marker, lactate dehydrogenase. If either EGTA is added to the medium or Mg²⁺ replaces Ca²⁺ no stimulation of secretion is observed. If the energy supply is decreased by treating the macrophages with mitochondrial inhibitors such as oligomycin, cyanide, or rotenone and antimycin, Ca²⁺-dependent secretion is potentiated several fold. Selective release of granule enzymes from macrophages exposed to A23187 and Ca²⁺ is also stimulated by cytochalasin B (CB).     

Intracellular uptake and α-amylase and lactate dehydrogenase releasing actions of the divalent cation ionophore A23187 in dissociated pancreatic acinar cells

Summary Intracellular uptake of A23187 and the increased release of amylase and lactate dehydrogenase (LDH) accompanying ionophore uptake was studied using dissociated acinar cells prepared from mouse pancreas. Easily detected changes in the fluorescence excitation spectrum of A23187 upon transfer of the ionophore from a Tris-buffered Ringer's to cell membranes were used to monitor A23187 uptake. Uptake was rapid in the absence of extracellular Ca²⁺ and Mg²⁺ (t1/2=1 min) and much slower in the presence of Ca²⁺ or Mg²⁺ (t1/2=20 min). Cell-associated ionophore was largely intracellular as indicated by fluorescence microscopy, lack of spectral sensitivity to changes in extracellular Ca²⁺ and Mg²⁺, and by equivalent interaction of ionophore with membranes of whole and sonicated cells.A23187 (10 m) increased amylase release 200% in the presence of extracellular Ca²⁺ and Mg²⁺. In the absence of Ca²⁺ (but in the presence of Mg²⁺) A23187 did not increase amylase release. A23187 (10 m) also produced Ca²⁺-dependent cell damage, as judged by increased LDH release, increased permeability to trypan blue, and by disruption of cell morphology. The cell damaging and amylase releasing properties of A23187 were distinguished by their time course and dose-response relationship. A23187 (1 m) increased amylase release 140% without increasing LDH release or permeability to trypan blue.     

Requirements for different Ca2+ pools in the activation of rabbit platelets: I. Release reaction and protein phosphorylation

We examined the role of Ca²⁺, both extracellular and intracellular in origin, in the release reaction and protein phosphorylation in rabbit platelets stimulated with platelet activating factor (acetylglyceryl ether phosphorylcholine), thrombin, or ionophore A23187. In the presence of extracellular Ca²⁺, 8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate (TMB-8), a putative antagonist of intracellular Ca²⁺ transport, blocked platelet activating factor-initiated serotonin release at a half-maximal inhibitor concentration of 40 μM, compared to 350 μM for thrombin-induced release and greater than 500 μM, for A23187-induced release. Platelet activating factor-induced phosphorylation of two platelet proteins of M_r=41 000 (P7P) and 20 000 (P9P) was inhibited by TMB-8, an effect which was additive to that caused by removing extracellular Ca²⁺. TMB-8 demonstrated only minor to non-existant inhibitory effects on phosphorylation in thrombin- or A23187-stimulated platelets. In contrast to P9P phosphorylation, phosphorylation of P7P caused by platelet activating factor was more dependent on a TMB-8 sensitive step than on the availability of extracellular Ca²⁺. Experiments with buffers containing fixed concentrations of free Ca²⁺ revealed that both processes (release and phosphorylation), when stimulated by platelet activating factor and thrombin, had the same threshold requirement (1–3 μM) for extracellular free Ca²⁺. These studies provide evidence that stimulation of rabbit platelets by platelet activating factor is more dependent on a TMB-8-sensitive intracellular Ca²⁺ source than is stimulation caused by thrombin. Furthermore, our data indicate that activation of different intracellular processes involved in platelet secretion (such as P7P and P9P phosphorylation) may require Ca²⁺ from different pools.     

Calcium and cyclic nucleotide involvement in exocrine pancreatic enzyme secretion studied with the ionophore A-23187.

The ionophore, A-23187, was an effective pancreatic secretagogue. The response to A-23187 was Ca²⁺-dependent; Mg²⁺ reduced the secretory response to the ionophore. A-23187-stimulated enzyme release was potentiated by dibutyryl cyclic AMP; in the presence of carbachol, output of pancreatic protein paralleled the response to A-23187 alone. The time-course for secretion with A-23187 was similar to that observed with carbachol. The ionophore did not affect basal cyclic AMP levels but did stimulate a rapid Ca²⁺-dependent production of pancreatic cyclic GMP which preceded the onset of the secretory response. A-23187 did not significantly alter basal or carbachol-stimulated ⁴⁵Ca efflux from isotope preloaded glands; yet in Ca²⁺-lowered media, it inhibited (reversed) the secretory response to carbachol, an effect which may have been due to an outward transport by the ionophore of cholinergic-mobilized intracellular Ca²⁺. Like carbachol, A-23187, inhibits the incorporation of amino acid into new protein, the effect being partially dependent on extracellular Ca²⁺. The data suggest that the pancreatic cholinergic receptor acts as a Ca²⁺-ionophore and that extracellular Ca²⁺ is utilized in the synthesis of cyclic GMP.     

Calcium requirements in the action of cholecystokinin-pancreozymin on pancreatic acinar cell metabolism.

The present study utilized ionophore A23187 to determine the role of Ca²⁺ in pancreatic acinar cell metabolism. The ionophore A23187 in the presence of EGTA increased efflux of Ca²⁺ from the rat pancreatic fragments. Ionophore and CCK-PZ were equally effective in the presence of extracellular Ca²⁺ in stimulating ¹⁴C-labeled protein secretion. The ionophore decreased synthesis of new protein more effectively than CCK-PZ in the presence of extracellular Ca²⁺. The effect of ionophore and CCK-PZ in combination was greater than either agent alone. Phospholipid labeling was not stimulated by A23187 in the presence of extracellular Ca²⁺ in contrast to CCK-PZ. With CCK-PZ, the effect was dependent on the concentration of extracellular Ca²⁺. Protein phosphorylation was stimulated ～ 109% by CCK-PZ and ～ 39% by ionophore. CCK-PZ stimulated protein phosphorylation in the 100,000 g supernatant whereas A23187 was ineffective. Ionophore A23187 inhibited glucose oxidation whereas CCK-PZ stimulated glucose oxidation. These data suggest that more than one kinase system might be involved in metabolic responses to hormonal stimulation of the pancreas viz. a phosphorylase kinase may be directly activated by Ca²⁺ causing protein discharge whereas other kinase system may require binding of the hormone to receptor leading to other events besides protein discharge.     

Participation of the microtubular-microfilamentous system on intracellular Ca transport and acid secretion in dispersed parietal cells

Biphasic responses of amino[¹⁴C]pyrine accumulation and oxygen consumption were registered by gastrin stimulation in dispersed parietal cells from guinea pig gastric mucosa, and this was mimicked with the calcium ionophore A23187. The characteristics of these phases (first phase and second phase) were distinguished by the differences in the requirements of extracellular Ca²⁺. The first phase evoked by gastrin or ionophore A23187 was independent of extracellular Ca²⁺, whereas the second phase was not. In the first phase, fluorescence of a cytosolic Ca²⁺ indicator (quin2-AM) increased with the stimulation of ionophore A23187 and carbamylcholine chloride in the presence of extracellular Ca²⁺. In addition, an increase in cytosolic Ca²⁺ induced by ionophore A23187, but not by carbamylcholine chloride was also observed in the absence of extracellular Ca²⁺, suggesting that Ca²⁺ pool(s) in parietal cells might be present in the intracellular organelle. Cytochalasin B and colchicine, but not oligomycin, could eliminate this cytosolic Ca²⁺ increase induced by A23187 in a Ca²⁺-free medium. On the other hand, in a Ca²⁺-free medium, addition of ATP after pretreatment with digitonin could diminish the cytosolic Ca²⁺ increase brought about by A23187. This was also observed with oligomycin-treated cells, but not with cytochalasin B-treated cells. Similarly, subcellular fractionation of a parietal cell which had been pretreated with cytochalasin B or colchicine in an intact cell system reduced the rate of ATP-dependent Ca²⁺ uptake. These observations indicate that intracellular Ca²⁺ transport in dispersed parietal cells may be regulated by the microtubular-microfilamentous system. In conclusion, this study demonstrates the possibility of the existence of intracellular Ca²⁺ transport mediated by gastrin or ionophore A23187 and regulated by the microtubular-microfilamentous system in parietal cells.     

Ionophore monensin induces Na+-dependent secretion from rabbit neutrophils. Requirement for intracellular Ca2+ stores

Rabbit (and human) neutrophils release the secretory enzyme β-glucuronidase when treated with the ionophore monensin in the presence of Na⁺. Release of β-glucuronidase occurs without loss of the cytosol enzyme lactate dehydrogenase and a number of other features of the release process lead us to conclude that a normal exocytotic mechanism is involved. These include sensitivity to metabolic inhibition, enhancement of release induced by cytochalasin B and a requirement for internal sources of Ca²⁺ when the cells are stimulated with monensin in the absence of extracellular Ca²⁺. The release process due to monensin differs from that due to receptor directed agonists such as fMet-Leu-Phe and the Ca²⁺ ionophores A23187 and ionomycin in respect of a prolonged time-course which extends over 20 min; nor do monensin-stimulated neutrophils generate the superoxide anion. The results are discussed in the light of reports which indicate a rôle for Na⁺ in the activation of neutrophils by other ligands.     

Mitochondrial permeability transition pore is involved in oxidative burst and NETosis of human neutrophils

Neutrophils release neutrophil extracellular traps (NETs) in response to numerous pathogenic microbes as the last suicidal resource (NETosis) in the fight against infection. Apart from the host defense function, NETs play an essential role in the pathogenesis of various autoimmune and inflammatory diseases. Therefore, understanding the molecular mechanisms of NETosis is important for regulating aberrant NET release. The initiation of NETosis after the recognition of pathogens by specific receptors is mediated by an increase in intracellular Ca²⁺ concentration, therefore, the use of Ca²⁺ ionophore A23187 can be considered a semi-physiological model of NETosis. Induction of NETosis by various stimuli depends on reactive oxygen species (ROS) produced by NADPH oxidase, however, NETosis induced by Ca²⁺ ionophores was suggested to be mediated by ROS produced in mitochondria (mtROS).Using the mitochondria-targeted antioxidant SkQ1 and specific inhibitors of NADPH oxidase, we showed that both sources of ROS, mitochondria and NADPH oxidase, are involved in NETosis induced by A23187 in human neutrophils. In support of the critical role of mtROS, SkQ1-sensitive NETosis was demonstrated to be induced by A23187 in neutrophils from patients with chronic granulomatous disease (CGD). We assume that Ca²⁺-triggered mtROS production contributes to NETosis either directly (CGD neutrophils) or by stimulating NADPH oxidase. The opening of the mitochondrial permeability transition pore (mPTP) in neutrophils treated by A23187 was revealed using the electron transmission microscopy as a swelling of the mitochondrial matrix. Using specific inhibitors, we demonstrated that the mPTP is involved in mtROS production, NETosis, and the oxidative burst induced by A23187.     

Toxoplasma gondii: Calcium lonophore A23187-mediated exit of trophozoites from infected murine macrophages

The Ca²⁺ ionophore A23187 consistently induced the exit of Toxoplasma gondii trophozoites from cultured macrophages which they had recently infected. Following exit of toxoplasmas, the host macrophages underwent degeneration. A23187 was active at concentrations higher than 0.25 μM and the activity reached a plateau at the concentration of 1.0 μM. Noninfected macrophages or those engulfing heat-killed toxoplasmas, or some other particles, were not affected by treatment with A23187. The toxoplasmas exiting host cells were capable of infecting and proliferating in normal macrophages. The A23187-mediated exit of toxoplasmas proceeded despite external Ca²⁺ and was enhanced by the addition of ethylene glycol bis(β-aminoethyl ether) N,N,N′,N′-tetraacetic acid (EGTA) in the reaction mixture. On the other hand, the A23187-mediated exit of toxoplasmas was inhibited significantly by exogenous Mg²⁺.     

Physical properties of biological membranes determined by the fluorescence of the calcium ionophore A23187

Interactions between the divalent cation ionophore, A23187, and the divalent cations Ca²⁺, Mg²⁺, and Mn²⁺ were studied in sarcoplasmic reticulum and mitochondria. Conductance measurements suggest that A23187 facilitates the movement of divalent cations across bilayer membranes via a primarily electroneutral process, although a cationic form of A23187 does carry some current.On the basis of fluorescence excitation spectra, A23187 can form either a 1:1 or 2:1 complex with Ca²⁺ in organic solvents. However, in biological membranes, only the 1:1 complexes with Ca²⁺, Mg²⁺, or Mn²⁺ are detected. A23187 produces fluorescent transients under conditions of Ca²⁺ uptake in sarcoplasmic reticulum, which appear to represent changes in intramembrane Ca²⁺ content. Changes in A23187 fluorescence due to mitochondrial Ca²⁺ accumulation are much smaller by comparison and fluorescence transients are not detected.Studies of A23187 fluorescence polarization and lifetimes in biological membranes allow a determination of the rotational correlation time (ρh) of the ionophore. In mitochondria at 22 °C, ρh is 11 nsec in the presence of Ca²⁺ and Mg²⁺, and less than 2 nsec in the presence of excess EDTA.The present results are consistent with a model of ionophore-mediated cation transport in which free M²⁺ binds with A23187 at the membrane surface to form the complex M(A23187)⁺. Reaction of this complex with another molecule of A23187 at the membrane surfaces results in the formation of electrically neutral M(A23187)₂, which carries the divalent cation through the membrane.These results are discussed in terms of physical properties of biological membranes in regions in which divalent cation transport occurs.     

Translational suppression by Ca2+ ionophores: Reversibility and roles of Ca2+ mobilization, Ca2+ influx, and nucleotide depletion

The divalent cation selective ionophores A23187 and ionomycin were compared for their effects on the Ca²⁺ contents, nucleotide contents, and protein synthetic rates of several types of cultured cells. Both ionophores reduced amino acid incorporation by approximately 85% at low concentrations (50–300 nmol/L) in cultured mammalian cells without reducing ATP or GTP contents. At these concentrations A23187 and ionomycin each promoted substantial Ca²⁺ efflux, whereas at higher concentrations a large influx of the cation was observed. Ca²⁺ influx occurred at lower ionophore concentrations and to greater extents in C6 glioma and P3X63Ag8 myeloma than in GH₃ pituitary cells. The ATP and GTP contents of the cells and their ability to adhere to growth surfaces declined sharply at ionophore concentrations producing increased Ca²⁺ influx. Prominent reductions of nucleotide contents occurred in EGTA-containing media that were further accentuated by extracellular Ca²⁺. Ionomycin produced more Ca²⁺ influx and nucleotide decline than comparable concentrations of A23187. The inhibition of amino acid incorporation and mobilization of cell-associated Ca²⁺ by ionomycin were readily reversed in GH₃ cells by fatty acid-free bovine serum albumin, whereas the effects of A23187 were only partially reversed. Amino acid incorporation was further suppressed by ionophore concentrations depleting nucleotide contents. Mitochondrial uncouplers potentiated Ca²⁺ accumulation in response to both ionophores. At cytotoxic concentrations Lubrol PX abolished protein synthesis but did not cause Ca²⁺ influx. Nucleotide depletion at high ionophore concentrations is proposed to result from increased plasmalemmal Ca²⁺-ATPase activity and dissipation of mitochondrial proton gradients and to cause intracellular Ca²⁺ accumulation. Increased Ca²⁺ contents in response to Ca²⁺ ionophores are proposed as an indicator of ionophore-induced cytotoxicity.Abbreviations BSA bovine serum albumin - EGTA [ethylenebis(oxyethylenenitrilo)]tetraacetic acid - PKR double-stranded RNA-regulated protein kinase - ER endoplasmic reticulum - eIF eukaryotic initiation factor     

Visualization and quantification of endoplasmic reticulum Ca in renal cells using confocal microscopy and Fluo5F

Sarcoplasmic/endoplasmic reticulum (ER) Ca²⁺ is the most abundant store of intracellular Ca²⁺, and its release is an important trigger of physiological and cell death pathways. Previous work in our laboratory revealed the importance of ER Ca²⁺ in toxicant-induced renal proximal tubular cell (RPTC) death. The purpose of this study was to evaluate the use of confocal microscopy and Fluo5F, a low affinity Ca²⁺ indicator, to directly monitor changes in RPTC ER Ca²⁺. Fluo5F staining reflected ER Ca²⁺, resolved ER structure, and showed no colocalization with tetramethyl rhodamine methyl ester (TMRM), a marker of mitochondrial membrane potential. Thapsigargin, an ER Ca²⁺ pump inhibitor, decreased ER fluorescence by 30% and 55% at 5 and 15 min, respectively, whereas A23187, a Ca²⁺ ionophore caused more rapid ER Ca²⁺ release (55% and 75% decrease in fluorescence at 5 and 15 min).Carbonylcyanide-p-trifluoromethoxyphenylhydrazone (FCCP), a mitochondrial uncoupler, added at the end of the experiment, further decreased ER fluorescence after thapsigargin treatment, revealing that thapsigargin did not release all ER Ca²⁺. In contrast, FCCP did not decrease ER fluorescence after A23187 treatment, suggesting complete ER Ca²⁺ release. ER Ca²⁺ release in response to A23187 or thapsigargin resulted in a modest but significant decrease in mitochondrial membrane potential. These data provide evidence that confocal microscopy and Fluo5F are useful and effective tools for directly monitoring ER Ca²⁺ in live cells.     

Effects of Ca ionophore A23187 and Ca deficiency on pancreatic phospholipids and amylase release in vitro

The Ca²⁺ ionophore A23187 elicits a transient increase in pancreatic amylase release in vitro, and this is accompanied by a transient decrease in phosphatidyl inositol concentration. Effects of ionophore A23187 and carbachol on amylase release and phosphatidylinositol breakdown are dependent on medium Ca²⁺. These results suggest that major secretagogue-induced, pancreatic phospholipid changes follow, rather than precede, changes in Ca²⁺ in the pancreas.     

Effect of ionophore A23187 upon membrane function and ion movement in human and toad erythrocytes

Summary Addition of 0.1–0.3 m A23187, a divalent cation ionophore, to human erythrocytes suspended in a 1.0mm ⁴⁵Ca²⁺-containing buffer results in a small ( two fold) increase in [Ca²⁺] _i , a significant decrease in osmotic fragility, and a decrease in intracellular K⁺ (100 mmoles/liter of cells to 70 mmoles/liter cells) without significant alteration of intracellular [Na⁺]. This decrease in [K⁺] _i is associated with a significant decrease in packed cell volume and correlates directly with the observed alteration is osmotic fragility. Increasing extracellular K⁺ to 125mm prevents the A23187-induced changes in osmotic fragility, K⁺ content and cell volume, but does not prevent the ionophore-induced uptake of⁴⁵Ca²⁺. Addition of 0.1–0.3 m A23187 to toad erythrocytes leads to an increase in⁴⁵Ca²⁺ uptake comparable to that observed in human erythrocytes, but does not alter osmotic fragility, cell volume or K⁺ content. Higher concentrations of ionophore (3.0–10.0 m) cause a 30- to 50-fold increase in⁴⁵Ca²⁺ uptake and concomitant change in K⁺ content, cell volume and osmotic fragility. These changes in cell properties can be prevented by increasing extracellular [K⁺] to 90mm. The difference in sensitivity of the two cell types to A23187 is attributed to the presence of additional intracellular calcium pools within toad erythrocytes that prevent an increase in cytoplasmic Ca²⁺ until Ca²⁺ uptake is increased substantially at the higher concentrations of A23187.     

Hysteretic activation of the Ca pump revealed by calcium transients in human red cells

The enzymatic basis for the Ca²⁺ pump in human red cells is an ATPase with hysteretic properties. The Ca²⁺-ATPase shifts slowly between a ground state deficient in calmodulin and an active state saturated with calmodulin, and rate constants for the reversible shifts of state were recently determined at different Ca²⁺ concentrations (Scharff, O. and Foder, B. (1982) Biochim. Biophys. Acta 691, 133–143). In order to study whether the Ca²⁺ pump in intact red cells also exhibits hysteretic properties we have analysed transient increases of intracellular calcium concentrations (Ca_i), induced by the divalent cation ionophore A23187. The time-dependent changes of Ca_i were measured by use of radioactive calcium (⁴⁵Ca²⁺) and analysed with the aid of a mathematical model, based partly on the Ca²⁺-dependent parameters obtained from Ca²⁺-ATPase experiments, partly on the A23187-induced Ca²⁺ fluxes determined in experiments with intact red cells. According to the model a delay in the activation of the Ca²⁺ pump is a prerequisite for the occurrence of A23187-induced calcium transients in the red cells, and we conclude that the Ca²⁺ pump in human red cells responds hysteretically. It is suggested that Ca²⁺ pumps in other types of cell also have hysteretic properties.     

The release of dopamine from synaptosomes from rat striatum by the ionophores X 537A and A 23187

The antibiotics X 537A and A 23187 are negatively charged divalent cation ionophores. X 537A may, in addition, be an ionophore for amines including catecholamines. The effects of these ionophores were examined on the uptake and release of dopamine by synaptosomes prepared from rat corpus striatum. Both X 537A and A 23187, at concentrations less than 0.5 μM, release both endogenous and [³H]-dopamine from synaptosomes. They had virtually no effect on the uptake of exogenous dopamine. These compounds act by different mechanisms. X 537A causes divalent ion-independent release in which a large fraction of the effluent consists of deaminated products. X 537A, in addition, releases [³H]dopamine from rat adrenal medullary chromaffin granules. The results suggest that X 537A causes release of dopamine from intrasynaptosomal storage vesicles and perhaps is acting as a catecholamine carrier across the vesicular membrane. A 23187, on the other hand, causes a Ca²⁺-dependent release in which only a small fraction of the catechol in the effluent is deaminated. A 23187 has little effect on the release of [³H]dopamine from chromaffin granules. These results suggest that A 23187 carries Ca²⁺ into the synaptosomes and thereby initiates exocytotic release.     

红细胞在钙离子和离子载体A23187作用下的流变特性研究

用新激光衍射法研究了钙离子及离子载体A23187对红细胞流变特性的影响.用不同浓度的钙离子及离子载体A23187分别处理红细胞后,测量其取向指数和小变形指数.结果表明离子载体A23187较细胞外钙离子浓度对红细胞流变特性的影响更大.而且,最大取向指数和最大小变形指数随着钙离子及离子载体A23187浓度的增加而降低.离子载体A23187浓度增加导致红细胞变形能力明显降低.     

Role of membrane associated serine esterase in the activation of phospholipase A2 by calcium ionophore (A23187) in pulmonary arterial smooth muscle cells

Exposure of rabbit pulmonary arterial smooth muscle cells to 10 M of the calcium ionophore A23187 dramatically stimulates cell membrane-associated phospholipase A₂ activity and arachidonic acid release. In addition, A23187 also enhances cell membrane-associated serine esterase activity. Serine esterase inhibitors phenylmethylsulfonylfuoride and diisopropyl fluorophosphate prevent the increase in serine esterase and phospholipase A₂ activities and arachidonic acid release caused by A23187. A23187 still stimulated serine esterase and phospholipase A₂ activities and arachidonic acid release in cells pretreated with nominal Ca²⁺ free buffer. Treatment of the cell membrane with A23187 does not cause any appreciable change in serine esterase and phospholipase A₂ activities. Pretreatment of the cells with actinomycin D or cycloheximide did not prevent the increase in the cell membrane associated serine esterase and phospholipase A₂ activities, and arachidonic acid release caused by A23187. These results suggest that (i) a membrane-associated serine esterase plays an important role in stimulating the smooth muscle cell membrane associated phospholipase A₂ activity (ii) in addition to the presence of extracellular Ca²⁺, release of Ca²⁺ from intracellular storage site(s) by A23187 also appears to play a role in stimulating the cell membrane-associated serine esterase and phospholipase A₂ activities, and (iii) the increase in the cell membrane-associated serine esterase and phospholipase A₂ activities does not appear to require new RNA or protein synthesis.Abbreviations A23187 calcium ionophore - AA arachidonic acid - PMSF phenylmethyl sulfonylfuoride - DFP diisopropyl-fluorophosphate - DMEM Dulbecco's modified Eagles medium - FCS fetal calf serum - PBS phosphate buffered saline - HBPS Hank's buffered physiological saline - PLA₂ phospholipase A₂