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1.
Schneider EM Menzl I Weber O Hug H 《Biochemical and biophysical research communications》2003,301(1):159-166
We constructed a CD95 overexpressing HeLa cell line which was extremely sensitive towards CD95 mediated apoptosis. In these CD95 overexpressing cells, CD95 blocks the nuclear calcium signal induced by perforin positive and CD95 ligand positive killer cells. This phenomenon is highly relevant in states of inflammatory syndromes such as systemic inflammatory response syndrome (SIRS) and sepsis which are associated with a high probability to reactivate latent viruses due to a functional deficiency of cytotoxic effectors. 相似文献
2.
Differential regulation of intracellular calcium oscillations by mitochondria and gap junctions 总被引:1,自引:0,他引:1
Fluctuations of intracellular Ca2+ ([Ca2+]i) regulate a variety of cellular functions. The classical Ca2+ transport pathways in the endoplasmic reticulum (ER) and plasma membrane are essential to [Ca2+]i oscillations. Although mitochondria have recently been shown to absorb and release Ca2+ during G protein-coupled receptor (GPCR) activation, the role of mitochondria in [Ca2+]i oscillations remains to be elucidated. Using fluo-3-loaded human teratocarcinoma NT2 cells, we investigated the regulation
of [Ca2+]i oscillations by mitochondria. Both the muscarinic GPCR agonist carbachol and the ER Ca2+-adenosine triphosphate inhibitor thapsigargin (Tg) induced [Ca2+]i oscillations in NT2 cells. The [Ca2+]i oscillations induced by carbachol were unsynchronized among individual NT2 cells; in contrast, Tg-induced oscillations were
synchronized. Inhibition of mitochondrial functions with either mitochondrial blockers or depletion of mitochondrial DNA eliminated
carbachol—but not Tg-induced [Ca2+]i oscillations. Furthermore, carbachol-induced [Ca2+]i oscillations were partially restored to mitochondrial DNA-depleted NT2 cells by introduction of exogenous mitochondria. Treatment
of NT2 cells with gap junction blockers prevented Tg-induced but not carbachol-induced [Ca2+]i oscillations. These data suggest that the distinct patterns of [Ca2+]i oscillations induced by GPCR and Tg are differentially modulated by mitochondria and gap junctions. 相似文献
3.
To investigate Ca2+ uptake by Ca2+-depleted bovine chromaffin cells we depleted these cells of Ca2+ by incubating them in Ca2+-free buffer, then measured changes in cytoplasmic Ca2+ concentration ([Ca2+
1)45Ca2+ uptake, and Mn2+ uptake in response to added Ca2+ or MN2+. In depleted cells, the increase in [Ca2+]i after Ca2+ addition, and the Mn2+ and45Ca2+ uptakes were higher than in control cells, and were inhibited by verapamil. The size of the intracellular Ca2+ pools in depleted cells increased after Ca2+ addition. The times for [Ca2+]i rise and Mn2+ entry to reach plateau levels were much shorter than the time for refilling of intracellular Ca2+ stores. In Ca2+-depleted cells and cells which had been loaded with BAPTA,45Ca2+ uptake was much higher than in control cells. These results suggest that extracellular Ca2+ enters the cytoplasm first before refilling the intracellular stores. The rate of Mn2+ influx depended on the level of filling of the Ca2+ stores, suggesting that some signalling takes place between the intracellular stores and Ca2+ entry pathways through the plasma membrane.Abbreviations used BAPTA
1,2-bis(2-aminophenoxy)ethane-N,N,N', N'-tetraacetic acid
- BAPTA/AM
acetoxymethyl ester of BAPTA
- [Ca2+]i
cytosolic Ca2+ concentration
- IP3
inositol 1,4,5-trisphosphate
- tBHQ
2,5-di-(t-butyl)-1,4-benzohydroquinone
This work was included in a thesis submitted by A.-L. Sui to the Department of Biochemistry, National Yang-Ming Medical College, in partial fulfillment of the requirements for the degree of Doctor of Philosophy 相似文献
4.
Emeran A. Mayer Anatoly Kodner Xiao Ping Sun Jonathan Wilkes David Scott George Sachs 《The Journal of membrane biology》1992,125(2):107-118
Summary Intracellular calcium [Ca2+]
i
measurements in cell suspension of gastrointestinal myocytes have suggested a single [Ca2+]
i
transient followed by a steady-state increase as the characteristic [Ca2+]
i
response of these cells. In the present study, we used digital video imaging techniques in freshly dispersed myocytes from the rabbit colon, to characterize the spatiotemporal pattern of the [Ca2+]
i
signal in single cells. The distribution of [Ca2+]
i
in resting and stimulated cells was nonhomogeneous, with gradients of high [Ca2+]
i
present in the subplasmalemmal space and in one cell pole. [Ca2+]
i
gradients within these regions were not constant but showed temporal changes in the form of [Ca2+]
i
oscillations and spatial changes in the form of [Ca2+]
i
waves. [Ca2+]
i
oscillations in unstimulated cells (n = 60) were independent of extracellular [Ca2+] and had a mean frequency of 12.6 +1.1 oscillations per min. The baseline [Ca2+], was 171 ± 13 nm and the mean oscillation amplitude was 194 ± 12 nm. Generation of [Ca2+]
i
waves was also independent of influx of extracellular Ca2+. [Ca2+]
i
waves originated in one cell pole and were visualized as propagation mostly along the subplasmalemmal space or occasionally throughout the cytoplasm. The mean velocity was 23 +3 m per sec (n = 6). Increases of [Ca2+]
i
induced by different agonists were encoded into changes of baseline [Ca2+]
i
and the amplitude of oscillations, but not into their frequency. The observed spatiotemporal pattern of [Ca2+]
i
regulation may be the underlying mechanism for slow wave generation and propagation in this tissue. These findings are consistent with a [Ca2+]
i
regulation whereby cell regulators modulate the spatiotemporal pattern of intracellularly generated [Ca2+]
i
oscillations.The authors thank Debbie Anderson for excellent technical assistance with the electron microscopy and Dr. M. Regoli for providing the NK-1 agonist [Sar9,Met(O2)11]-SP. This work was supported by National Institutes of Health Grants DK 40919 and DK 40675 and Veterans Administration Grant SMI. 相似文献
5.
The characteristics and properties of the increase in cytosolic [Ca2+] that occurs in bovine adrenal medullary chromaffin cells on exposure to angiotensin II have been investigated. In fura-2 loaded cells exposure to a maximally effective concentration of angiotensin II (100 nM) caused a rapid, but transient increase in cytosolic [Ca2+] followed by a lower plateau that was sustained as long as external Ca2+ was present. In the absence of external Ca2+ only the initial brief transient was observed. In cells previously treated with thapsigargin in Ca2+-free medium to deplete the internal Ca2+ stores, angiotensin II caused no increase in cytosolic [Ca2+] when external Ca2+ was absent. Reintroduction of external Ca2+ to thapsigargin-treated, store-depleted cells caused a sustained increase in cytosolic [Ca2+] that was not further increased upon exposure to angiotensin II. Analysis of the data suggests that in bovine chromaffin cells angiotensin II causes Ca2+ entry via a pathway(s) activated as a consequence of internal store mobilization, and entry through this pathway(s) forms the majority of the sustained Ca2+ influx evoked by angiotensin II. 相似文献
6.
《生物化学与生物物理学报:生物膜》2004,1661(2):204-211
The basal 45Ca2+ influx in human red blood cells (RBC) into intact RBC was measured. 45Ca2+ was equilibrated with cells with t1/2=15-20 s and the influx reached the steady state value in about 90-100 s and the steady state level was 1.5±0.2 μmol/lpacked cells (n=6) at 37 °C. The average value of the Ca2+ influx rate was 43.2±8.9 μmol/lpacked cells hour. The rate of the basal influx was pH-dependent with a pH optimum at pH 7.0 and on the temperature with the temperature optimum at 25 °C. The basal Ca2+ influx was saturable with Ca2+ up to 5 mmol/l but at higher extracellular Ca2+ concentrations caused further increase of basal Ca2+ influx. The 45Ca2+ influx was stimulated by addition of submicromolar concentrations of phorbol esters (phorbol 12-myristate-13-acetate (PMA) and phorbol-12,13-dibutyrate (PDBu)) and forskolin. Uncoupler (3,3′,4′,5-tetrachloro-salicylanilide (TCS) 10−6-10−5 mol/l) inhibited in part the Ca2+ influx. The results show that the basal Ca2+ influx is mediated by a carrier and is under control of intracellular regulatory circuits. The effect of uncoupler shows that the Ca2+ influx is in part driven by the proton-motive force and indicates that the influx and efflux of Ca2+ are coupled via the RBC H+ homeostasis. 相似文献
7.
Wang GJ Lin LC Chen CF Cheng JS Lo YK Chou KJ Lee KC Liu CP Wu YY Su W Chen WC Jan CR 《Life sciences》2002,71(9):1081-1090
The effects of timosaponin A-III (TA-III), from Rhizoma Anemarrhenae, on Ca(2+) mobilization in vascular endothelial cells and smooth muscle cells and on vascular tension have been explored. TA-III increased intracellular Ca(2+) concentrations ([Ca(2+)](i)) in endothelials cells at a concentration larger than 5 microM with an EC(50) of 15 microM, and increased [Ca(2+)](i) in smooth muscle cells at a concentration larger than 1 microM with an EC(50) of 8 microM. Within 5 min, the [Ca(2+)](i) signal was composed of a gradual rise, and the speed of rising depended on the concentration of TA-III. The [Ca(2+)](i) signal was abolished by removing extracellular Ca(2+) and was recovered after reintroduction of Ca(2+). The TA-III-induced [Ca(2+)](i) increases in smooth muscle cells were partly inhibited by 10 microM nifedipine or 50 microM La(3+), but was insensitive to 10 microM verapamil and diltiazem. TA-III (10-100 microM) inhibited 0.3 microM phenylephrine-induced vascular contraction, which was abolished by pretreatment with 100 microM N(omega)-nitro-L-arginine (L-NNA) or by denuding the aorta. TA-III also increased [Ca(2+)](i) in renal tubular cells with an EC(50) of 8 microM. Collectively, the results show for the first time that TA-III causes [Ca(2+)](i) increases in the vascular system. TA-III acted by causing Ca(2+) influx without releasing intracellular Ca(2+). TA-III induced relaxation of phenylephrine-induced vascular contraction via inducing release of nitric oxide from endothelial cells. 相似文献
8.
Deval E Levitsky DO Constantin B Raymond G Cognard C 《Experimental cell research》2000,255(2):291-302
Previous investigations have demonstrated molecular and functional expression, at early phases of development of skeletal muscle cells in primary culture, of cardiac isoforms of proteins involved in calcium transport and regulation, like the L-type calcium channel. Here the expression of the cardiac isoform of the Na(+)/Ca(2+) exchanger (NCX1) was studied in skeletal muscle cells developing in vitro, by using biochemical, immunological, and electrophysiological techniques. Northern and Western blot experiments revealed the presence of this cardiac exchanger and its increasing expression during the early phases of development. Confocal imaging of myotubes showed an NCX1 distribution that was predominantly sarcolemmal. The whole-cell patch-clamp technique allowed us to record ionic currents, the direction and the amplitude of which depended on extracellular sodium and calcium concentrations. The developmental changes of this functional expression could be correlated with the molecular NCX1 expression changes. Taken together these data demonstrate the presence of the NCX1 isoform of the Na(+)/Ca(2+) exchanger during in vitro myogenesis and reinforce the theory that significant levels of cardiac-type proteins are transiently expressed during the early phases of the skeletal muscle cell development. 相似文献
9.
Soldati L Lombardi C Adamo D Terranegra A Bianchin C Bianchi G Vezzoli G 《Biochemical and biophysical research communications》2002,293(3):974-978
Recently, we have measured in erythrocytes a voltage-modulated and dihydropyridine-inhibited calcium influx. Since arachidonic acid and other polyunsaturated fatty acids influence the activities of most ion channels, we studied their effects on the erythrocyte Ca(2+) influx. It was measured on fresh erythrocytes, isolated from healthy donors, using the fluorescent dye Fura 2 as indicator of [Ca(2+)](i). AA (5-50 microM) and EPA (20-30 microM) stimulated a concentration-dependent increase in [Ca(2+)](i), deriving from extracellular calcium (1 mM), without affecting the intra- and extracellular pH and membrane voltage. The Ca(2+) influx rate varied from 0.5 to 3 nM Ca(2+)/s in the presence of AA and from 0.9 to 1.7 nM Ca(2+)/s with EPA. The Ca(2+) influx elicited by AA and EPA was not inhibited by dihydropyridines, while cyclooxygenase inhibitors were effective and PGE1 or PGE2 did not produce any effect. We conclude that AA could activate an erythrocyte voltage-independent Ca(2+) transport via an intermediate product of cyclooxygenase pathway; however, a direct interaction with the membrane lipid-protein cannot be excluded. 相似文献
10.
Summary Regulation of passive potassium ion transport by the external calcium concentration and temperature was studied on cell cultures of 3T3 mouse cells and their DNA-virus transformed derivatives. Upon lowering of external calcium concentration, passive potassium efflux generally exhibits a sharp increase at about 0.1mm. The fraction of calcium-regulated potassium efflux is largely independent of temperature in the cases of the transformed cells, but shows a sharp increase for 3T3 cells upon increasing temperature above 32°C. In the same range of temperature, the 3T3 cells exhibit the phenomenon of high-temperature inactivation of the residual potassium efflux at 1mm external calcium. At comparable cellular growth densities, the transformed cell lines do not show high-temperature inactivation of residual potassium efflux. These results are consistent with the notion of a decisive role of the internal K+ concentration in the cell-density dependent regulation of cell proliferation. In particular, the growth-inhibiting effect of lowering the external Ca2+ concentrations is considered as largely due to a rise of passive K+ efflux and a subsequent decrease of internal K+ concentration. The experimental data on the Ca2+ dependence of passive K+ flux are quantitatively described by a theoretical model based on the constant field relations including negative surface charges on the external face of the membrane, which cooperatively bind Ca2+ ions and may concomitantly undergo a lateral redistribution. The present evidence is consistent with acidic phospholipids as representing these negative surface charges.This work is dedicated to the memory of Max Delbrück (deceased March 10, 1981), in whose laboratory in 1966 the earlier version of the present theoretical model was developed by one of the authors. 相似文献
11.
Persistent tumour necrosis factor alpha (TNF-alpha) exposure uncouples proximal T-cell receptor (TCR)-signalling events. Here, we demonstrate that chronic TNF-alpha exposure also attenuates signalling distal to the TCR, by specifically inhibiting Ca2+ influx evoked by thapsigargin in CD4+ T-cells. Mitogen-induced Ca2+ responses were impaired in a dose dependent manner, and TCR-induced Ca2+ responses were also significantly reduced. The impairment of Ca2+ influx strongly correlated with poor function as proliferative responses to both mitogen and anti-CD3/CD28 stimulation were suppressed. Our findings show that persistent TNF-alpha exposure of T-cells specifically inhibits store operated Ca2+ influx. This may affect gene activation and contribute to the poor T-cell function in chronic inflammatory disease. 相似文献
12.
The localization of Ca2+ in cells of the periblem and dermatogen in the root meristem and the columella and peripheral cells of the root cap of maize
was examined by the precipitation method of potassium pyroantimonate and EGTA-treatment. In periblem and dermatogen cells,
Ca2+ was found to be localized in the nucleoplasm and granular zone of the nucleolus. Ca2+ was also found in most cell organelles: in the matrix in mitochondria, on the thylakoid membrane in proplastids, in the vacuoles
and on the plasma membranes. Ca2+ was also distributed throughout the cytoplasmic ground matrix. Much Ca2+ was present in the cell wall soon after its formation during the cell division. Ca2+ was also conspicuous in the vesicles of Golgi in the dermatogen cells. In columella and peripheral cells, there was less
Ca2+ in the organelles and cytoplasmic ground matrix, but Ca2+ was present in Golgi vesicles in the peripheral cells. Electron microscopic and X-ray microanalysis showed that Ca2+ was also present in the mucilaginous layer, the outermost cell wall of the peripheral cells. 相似文献
13.
Indu S. Ambudkar Timothy Lockwich Yukiharu Hiramatsu Bruce J. Baum 《Molecular and cellular biochemistry》1992,114(1-2):73-77
Conclusions While it is generally accepted that Ca2+ plays an important regulatory role in the physiology of a number of non-excitable cells, the mechanisms which regulate intracellular [Ca2+ are far from well established. Ca2+ transporting mechanisms which distribute Ca2+ intracellularly as well as those which allow influx of extracellular Ca2+ are involved in mediating intracellular Ca2+ homestasis. In this paper we have described recent studies on the regulation of the Ca2+ influx system in the data, it appears that the process of Ca2+ entry is extremely complex and may involve several levels of regulation. Understanding the molecular basis of these regulatory mechanisms presents a challeging problem for future studies. 相似文献
14.
《Electromagnetic biology and medicine》2013,32(1):77-84
AbstractCalcium transients play an essential role in cardiomyocytes and electromagnetic fields (EMF) and affect intracellular calcium levels in many types of cells. Effects of EMF on intracellular calcium transients in cardiomyocytes are not well studied. The aim of this study was to assess whether extremely low frequency electromagnetic fields (ELF-EMF) could affect intracellular calcium transients in cardiomyocytes. Cardiomyocytes isolated from neonatal Sprague-Dawley rats were exposed to rectangular-wave pulsed ELF-EMF at four different frequencies (15?Hz, 50?Hz, 75?Hz and 100?Hz) and at a flux density of 2?mT. Intracellular calcium concentration ([Ca2+]i) was measured using Fura-2/AM and spectrofluorometry. Perfusion of cardiomyocytes with a high concentration of caffeine (10?mM) was carried out to verify the function of the cardiac Na+/Ca2+ exchanger (NCX) and the activity of sarco(endo)-plasmic reticulum Ca2+-ATPase (SERCA2a). The results showed that ELF-EMF enhanced the activities of NCX and SERCA2a, increased [Ca2+]i baseline level and frequency of calcium transients in cardiomyocytes and decreased the amplitude of calcium transients and calcium level in sarcoplasmic reticulum. These results indicated that ELF-EMF can regulate calcium-associated activities in cardiomyocytes. 相似文献
15.
Studies of Ca2+ transport pathways in exocrine gland cells have been useful, chiefly because of the polarized nature of the secretory epithelial cells. In pancreatic acinar cells, for example, Ca2+ reloading of empty intracellular stores can occur solely via Ca2+ entry through the basal part of the plasma membrane. On the other hand, the principal site for intracellular Ca2+ release—with the highest concentration of inositol 1,4,5-trisphosphate (IP3) receptors—is in the apical secretory pole close to the apical plasma membrane. This apical part of the plasma membrane contains the highest density of Ca2+ pumps and is therefore the principal site for Ca2+ extrusion. On the basis of the known properties of Ca2+ entry and exit pathways in exocrine gland cells, the mechanisms controlling Ca2+ exit and entry are discussed in relation to recent direct information about Ca2+ transport into and out of the endoplasmic reticulum (ER) and the mitochondria in these cells. 相似文献
16.
Some kinetic parameters of the human red cell Ca2+-ATPase were studied on calmodulin-free membrane fragments following preincubation at 37°C. After 30 min treatment with EGTA(1 mm) plus dithioerythritol (1 mm), a V max of about 0.4 μmol Pi/mg × hr and a K s of 0.3 μm Ca2+ were found. When Mg2+ (10 mm) or Ca2+(10 μm) were also added during preincubation, V maxbut not Kwas altered. Ca2+ was more effective than Mg2+, thus increasing V max to about 1.3 μmol Pi/mg × hr. The presence of both Ca2+ and Mg2+ during pretreatment decreasedKto 0.15 μm, while having no apparent effect on V max. Conversely, addition of ATP (2 mm) with either Ca2+ or Ca2+ plus Mg2+increased Vmax without affecting K. Preincubation with Ca2+ for periods longer than 30 min further increased Vmaxand reduced Kto levels as low as found with calmodulin treatment. The Ca2+ activation was not prevented by adding proteinase inhibitors (iodoacetamide, 10 mm; leupeptin, 200 μm; pepstatinA, 100 μm; phenylmethanesulfonyl fluoride, 100 μm). The electrophoretic pattern of membranes preincubated with or without Mg2+, Ca2+ or Ca2+ plus Mg2+ did not differ significantly from each other. Moreover, immunodetection of Ca2+-ATPase by means of polyclonal antibodiesrevealed no mobility change after the various treatments. The above stimulation was not altered by neomycin (200 μm), washing with EGTA (5 mm) or by both incubating and washing with delipidized serum albumin (1 mg/ml), or omitting dithioerythritol from the preincubation medium. On the other hand, the activation elicited by Ca2+ plus ATP in the presence of Mg2+ was reduced 25–30% by acridine orange (100 μm), compound 48/80 (100 μm) or leupeptin (200 μm) but not by dithio-bis-nitrobenzoic acid (1 mm). The fluorescence depolarization of 1,6-diphenyl-and l-(4-trimethylammonium phenyl)-6-phenyl 1,3,5-hexatriene incorporated into membrane fragments was not affected after preincubating under the different conditions. The results show that proteolysis, fatty acid production, an increased phospholipid metabolism or alteration of membrane fluidity are not involved in the Ca2+ effect. Ca2+ preincubation may stimulate the Ca2+-ATPase activity by stabilizing or promoting the E1 conformation. 相似文献
17.
Njanoor Narayanan 《Biochimica et Biophysica Acta (BBA)/General Subjects》1981,678(3):442-459
Two membrane fractions, one enriched in sarcoplasmic reticulum and the other enriched in sarcolemma, were isolated from the myocardium of young (3–4-months-old) and aged (24–25-months old) rats. ATP-supported Ca2+ binding and accumulating activities as well as (Mg2+ + Ca2+)-ATPase activities of these membrane fractions were studied in an effort to determine the influence of age on the Ca2+ pump function of the two myocardial membrane systems. Sarcoplasmic reticulum from aged hearts showed significantly reduced (approx. 50%) rates of ATP-supported (oxalate-facilitated) Ca2+ accumulation compared to sarcoplasmic reticulum from young hearts; the amount of Ca2+ accumulated by this membrane of aged heart at steady state was also lower. On the other hand, sarcolemma from aged hearts displayed 2-fold higher rates of ATP-supported Ca2+ accumulation compared to sarcolemma from young hearts; at steady state, sarcolemma from aged hearts accumulated significantly higher amounts of Ca2+ than did sarcolemma from young hearts. Similar age-related differences were also observed in the ATP-dependent Ca2+ binding activities of the two membranes, determined in the absence of oxalate. The divergent age-associated changes in Ca2+ binding and accumulating activities of sarcoplasmic reticulum and sarcolemma were seen at varying Ca2+ concentrations (0.24–39.1 μM).With either membrane, kinetic analysis showed 2-fold age-related differences in the V values for ATP-supported Ca2+ accumulation (V (nmol Ca2+/mg protein per min): sarcoplasmic reticulum — young, 119 ± 8; aged, 59 ± 5; sarcolemma — young, 11 ± 2; aged, 21 ± 3); the concentrations of Ca2+ required for half-maximal velocities did not differ significantly with age (K0.5 for Ca2+ (μM): sarcoplasmic reticulum — young, 2.5 ± 0.20; aged, 2.9 ± 0.25; sarcolemma — yount, 2.7 ± 0.25; aged, 3.2 ± 0.30). Kinetic parameters of ATP-dependent Ca2+ binding also indicated that the velocity of Ca2+ binding but not the concentration of Ca2+ required for half-maximal binding was altered due to aging. At identical Ca2+ concentrations, the combined Ca2+ accumulating activity of sarcoplasmic reticulum and sarcolemma from aged hearts was significantly lower (38–47%) than the combined Ca2+ accumulating activity of the two membranes from young hearts. No significant age-related differences were observed in the ATP-independent (passive) Ca2+ binding (or accumulation) by sarcoplasmic reticulum and sarcolemma, the (Mg2+ + Ca2+)-ATPase activities of these membranes, their polypeptide composition or relative purity. These results indicate that differential alterations occur in the ATP-supported Ca2+ pump activities of sarcoplasmic reticulum and sarcolemma in aging myocardium and such alterations may be due to age-associated changes in the efficacy of coupling ATP hydrolysis to Ca2+ transport. Further, the age-related increment in the Ca2+ pump activity of sarcolemma is inadequate to fully compensate for the diminished Ca2+ pump activity of sarcoplasmic reticulum. It is, therefore, suggested that deterioration of the Ca2+ pump function of sarcoplasmic reticulum may contribute to the increased relaxation time observed in aging heart. 相似文献
18.
Xiao-Ying Tien Christopher Katnik Bahaa M. Qasawa Michael D. Sitrin Deborah J. Nelson Thomas A. Brasitus 《The Journal of membrane biology》1993,136(2):159-168
The present studies were conducted to investigate the mechanisms underlying the 1,25-dihydroxycholecalciferol (1,25(OH)2D3)-induced increase in intracellular Ca2+ ([Ca2+]
i
) in individual CaCo-2 cells. In the presence of 2mm Ca2+, 1,25(OH)2D3-induced a rapid transient rise in [Ca2+]
i
in Fura-2-loaded cells in a concentration-dependent manner, which decreased, but did not return to baseline levels. In Ca2+-free buffer, this hormone still induced a transient rise in [Ca2+]
i
, although of lower magnitude, but [Ca2+]
i
then subsequently fell to baseline. In addition, 1,25(OH)2D3 also rapidly induced45Ca uptake by these cells, indicating that the sustained rise in [Ca2+]
i
was due to Ca2+ entry. In Mn2+-containing solutions, 1,25(OH)2D3 increased the rate of Mn2+ influx which was temporally preceded by an increase in [Ca2+]
i
. The sustained rise in [Ca2+]
i
was inhibited in the presence of external La3+ (0.5mm). 1,25(OH)2D3 did not increase Ba2+ entry into the cells. Moreover, neither high external K+ (75mm), nor the addition of Bay K 8644 (1 μm), an L-type, voltage-dependent Ca2+ channel agonist, alone or in combination, were found to increase [Ca2+]
i
, 1,25(OH)2D3 did, however, increase intracellular Na+ in the absence, but not in the presence of 2mm Ca2+, as assessed by the sodium-sensitive dye, sodium-binding benzofuran isophthalate. These data, therefore, indicate that CaCo-2
cells do not express L-type, voltage-dependent Ca2+ channels. 1,25(OH)2D3 does appear to activate a La3+-inhibitable, cation influx pathway in CaCo-2 cells. 相似文献
19.
Yazaki I Abe M Santella L Koyama Y 《Biology of the cell / under the auspices of the European Cell Biology Organization》2004,96(2):153-167
The micromeres, the first cells to be specified in sea urchin embryos, are generated by unequal cleavage at the fourth cell division. The micromeres differentiate autonomously to form spicules and dispatch signals to induce endomesoderm in the neighbouring macromeres cells in the embryo. Using a calcium indicator Fura-2/AM and a mixture of dextran conjugated Oregon green-BAPTA 488 and Rhodamine red, the intracellular calcium ion concentration ([Ca2+]i) was studied in embryos at the 16-cell stage. [Ca2+]i was characteristically elevated in the micromeres during furrowing at the 4th cleavage. Subsequently, Ca2+ oscillated for about 10 min in the micromeres, resulting in episodic high levels of [Ca2+]i. High [Ca2+]i regions were associated with regional localizations of the endoplasmic reticulum (ER), though not with ER accumulated at the vegetal pole of the micromeres during the 4th division. Pharmacological studies, using a blocker of IP3-mediated Ca2+ release (Xestospongin), a store-operated Ca2+ entry inhibitor (2 aminoethoxydiphenyl borate (2-APB)) and an inhibitor of stretch-dependent ion channels (gadolinium), suggest that the high [Ca2+]i and oscillations in the micromeres are triggered by calcium influx caused by the activation of stretch-dependent calcium channels, followed by the release of calcium ions from the endoplasmic reticulum. On the basis of these new findings, a possible mechanism for autonomous formation of the micromeres is discussed. 相似文献