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1.
Abstract: Modes of Ca2+ activation by bradykinin, serotonin, and ATP and the possible receptor cross-talk were investigated in mouse neuroblastoma × rat glioma hybrid cells (108CC15) by monitoring fura-2 fluorescence in single cells. A transient rise of cytosolic Ca2+ activity was induced by short pulses of the hormones. Brief exposure of cells to ionomycin, which depletes intracellular Ca2+ stores, reduced the size of subsequent responses to bradykinin or ATP, but not to serotonin. Superfusion of the cells with Ca2+-free medium abolished the Ca2+ response to serotonin, whereas the responses to bradykinin and to ATP were only slightly reduced. This indicates that ATP, like bradykinin, Induces the release of Ca2+ from intracellular stores. Serotonin, in contrast, activates Ca2+ entry from the extracellular space. To investigate whether ATP releases Ca2+ from the same stores as bradykinin, we examined the interaction of the hormones by applying them consecutively. When ATP was applied after bradykinin, the nucleotide did not evoke any response, irrespective of the presence or absence of extracellular Ca2+. The application of ATP before that of bradykinin reduced the size of a following bradykinin-induced Ca2+ response in Ca2+-free medium, but not in Ca2+-containing medium. This suggests that bradykinin may interact with the ATP-activated mechanism by cross-desensitization. Possibly, bradykinin receptors are coupled to additional Ca2+ stores not accessible to ATP that are refilled by extracellular Ca2+. Cyclic AMP and cyclic GMP apparently do not affect the Ca2+ responses to bradykinin and serotonin, as shown by the lack of influence of preincubation of the cells with forskolin or sodium nitroprusside. 相似文献
2.
3.
Sandie I. Briand Sylvie G. Bernier Gatan Guillemette 《Journal of cellular biochemistry》1996,63(3):292-301
We provided evidence that calcium-calmodulin plays a major role in bradykinin-induced arachidonic acid release by bovine aortic endothelial cells. In cells labeled for 16 hr with 3H-arachidonic acid, ionomycin and Ca2+-mobilizing hormones such as bradykinin, thrombin and platelet activating factor induced arachidonic acid release. However, arachidonic acid release was not induced by agents known to increase cyclic AMP (forskolin, isoproterenol) or cyclic GMP (sodium nitroprusside). Bradykinin induced the release of arachidonic acid in a dose-dependent manner (EC50 = 1.6 ± 0.7 nM). This increase was rapid, reaching a maximal value of fourfold above basal level in 15 min. In a Ca2+-free medium, bradykinin was still able to release arachidonic acid but with a lower efficiency. Quinacrine (300 μM), a blocker of PLA2, completely inhibited bradykinin-induced arachidonic acid release. The B2 bradykinin receptor antagonist HOE-140 completely inhibited bradykinin-induced arachidonic acid release. The B1-selective agonist DesArg9-bradykinin was inactive and the B1-selective antagonist [Leu8]DesArg9-bradykinin had no significant effect on bradykinin-induced arachidonic acid release. The phospholipase C inhibitor U-73122 (100 μM) decreased bradykinin-induced arachidonic acid release. The calmodulin inhibitor W-7 (50 μM) drastically reduced the bradykinin- and ionomycin-induced arachidonic acid release. Also, forskolin decreased bradykinin-induced arachidonic acid release. These results suggest that the activation of PLA2 by bradykinin in BAEC is a direct consequence of phospholipase C activation. Ca2+-calmodulin appears to be the prominent activator of PLA2 in this system. © 1996 Wiley-Liss, Inc. 相似文献
4.
Preincubation of synaptosomes in standard physiological medium stimulates 2-fold Ca2+ uptake as compared to non-preincubated synaptosomes. When the sodium concentration in the preincubation medium has been halved, Ca2+ uptake was reduced by approximately 50 percent. The addition of ouabain to the preincubation medium decreases depolarization-stimulated Ca2+ uptake by about 40 percent. A steady-state level of Ca2+ uptake is achieved by synaptosomes preincubated for 0,5 or 10 min. These findings suggest that Ca2+ uptake might depend on the Na-gradient formed during the preincubation of synaptosomes under control conditions. 相似文献
5.
Phosphoinositide hydrolysis was studied in primary cultures of rat cerebellar astrocytes pre-labeled with [3H]myo-inositol. Among the agonists examined, the rank order of efficacies in causing phosphoinositide hydrolysis was bradykinin > endothelin-1 > ATP > norepinephrine. The bradykinin response was robust (24-fold increase) with EC50 value of 30 nM and saturating concentration of 1 μM. Preincubation of cells with pertussis toxin did not affect the activation of phosphoinositide turnover by bradykinin. Although short-term (within 90 min) treatment of cells with phorbol dibutyrate attenuated bradykinin-induced phosphoinositide breakdown, the inhibitory effect was lost after 3–6 h of phorbol dibutyrate treatment. Extended (24 h) preincubation resulted in a potentiation of bradykinin response. Homologous desensitization of bradykinin response was observed in cells prestimulated with bradykinin for up to 6 h. However, similar to the effect of phorbol dibutyrate. 24-h pretreatment with bradykinin selectively sensitized the response to bradykinin. Up-regulation of the bradykinin response was also observed in cells prestimulated with endothelin-1 or norepinephrine for 24 h, although these treatments resulted in only homologous desensitization to their own response. Our results suggest that cultured cerebellar astrocytes express bradykinin receptors coupled to phospholipase C and in these cells protein kinase C plays a more prominent role in the negative-feedback regulation of bradykinin-evoked phosphoinositide response. 相似文献
6.
Vasopressin and bradykinin bind to receptors coupled to GTP-binding proteins and rapidly induce polyphosphoinositide breakdown leading to Ca2+ mobilization and activation of protein kinase C. Both peptides are known to induce mitogenesis in the presence of growth factors that act through receptors with intrinsic tyrosine kinase activity. Surprisingly, addition of a combination of vaso-pressin and bradykinin to Swiss 3T3 cells synergistically stimulates DNA synthesis in the absence of any other growth factors. This effect is induced at nanomolar concentrations of the peptides and could be inhibited by addition of specific receptor antagonists or broad spectrum neuropeptide antagonists. Bradykinin, which stimulates transient activation of protein kinase C, induces DNA synthesis in synergy with substances that cause long-term activation of protein kinase C, like vasopression or phorbol 12, 13-dibutyrate. Down-regulation of protein kinase C inhibited the induction of mitogenesis by the combination of vasopressin and bradykinin, thus demonstrating the importance of long-term activation of this enzyme for DNA synthesis. Analysis of tyrosine phosphorylated proteins of Mr = 110,000–130,000 and Mr = 70,000–80,000 revealed a biphasic response after stimulation with bradykinin, whereas the response induced by vasopressin declined after the initial maximum. The combination of bradykinin with vasopressin caused an enhanced and prolonged increase in tyrosine phosphorylation of these proteins as compared with the individual peptides. Inhibition of tyrosine phosphorylation by tyrphostin was paralleled by inhibition of DNA synthesis. Together, these results demonstrate synergistic stimulation of DNA synthesis by bradykinin and vasopressin via prolonged stimulation of multiple signaling pathways and imply that the interactive effects of Ca2+ -mobilizing peptides on mitogenesis may be more general than previously thought. © 1994 Wiley-Liss, Inc. 相似文献
7.
The effects of cytokine and fatty acid treatment on signal transduction in dermal fibroblasts from type 1 diabetics and matched controls were compared. Chronic exposure to TNF, accentuated Ca2+ mobilization in response to bradykinin (BK) in cells from both controls and diabetics; responses were three-fold greater in cells from diabetics than in controls. Similarly, with chronic exposure to IL-1β, BK-induced Ca2+ mobilization was accentuated in cells from type 1 diabetics compared to the controls. Pretreatment with the protein synthesis inhibitor cycloheximide or the protein kinase C inhibitor calphostin C prior to the addition of TNF completely abrogated the TNF-induced increment in peak bradykinin response. Ca2+ transients induced by depleting endoplasmic reticulum (ER) Ca2+ with thapsigargin were also greater in TNF treated fibroblasts than in untreated cells, with greater increases in cells from diabetics. Exposing fibroblasts for 48 hours to 2 mM oleate also increased both the peak bradykinin response and the TNF-induced increment in peak response, which were significantly greater in diabetics than controls. These data indicate that cells from diabetic patients acquire elevated ER Ca2+ stores in response to both cytokines and free fatty acids,and thus exhibit greater sensitivity to environmental inflammatory stimuli and elevated lipids. 相似文献
8.
Yorimasa Ogata Naomi Niisato Tetsuo Negishi Takeshi Sakurai Shunsuke Furuyama Hiroshi Sugiya 《Cell biology international》1995,19(8):637-646
In fura-2-loaded human periodontal ligament (HPDL) cells, bradykinin induced a rapidly transient increase and subsequently sustained increase in cytosolic Ca2+ ([Ca2+]i). When external Ca2+ was chelated by EGTA, the transient peak of [Ca2+]i was reduced and the sustained level was abolished, implying the Ca2+ mobilization consists of intracellular Ca2+ release and Ca2+ influx. Thapsigargin, a specific Ca2+-ATPase inhibitor for inositol 1,4,5-trisphosphate (1,4,5-1P3)-sensitive Ca2+ pool, induced an increase in [Ca2+]i in the absence of external Ca2+. After depletion of the intracellular Ca2+ pool by thapsigargin, the increase in [Ca2+]i induced by bradykinin was obviously reduced. Bradykinin also stimulated formation of inositol polyphosphates including 1,4,5-IP3. These results suggest that bradykinin stimulates intracellular Ca2+ release from the 1,4,5-1P3-sensitive Ca2+ pool. Bradykinin stimulated prostaglandin E2 (PGE2) release in the presence of external Ca2+, but not in the absence of external Ca2+. Ca2+ ionophore A23187 and thapsigargin evoked the release of PGE2 in the presence of external Ca2+ despite no activation of bradykinin receptors. These results indicate that bradykinin induces Ca2+ mobilization via activation of phospholipase C and PGE2 release caused by the Ca2+ influx in HPDL cells. 相似文献
9.
William D. Ehringer Michael J. Edwards Frederick N. Miller 《Journal of cellular physiology》1996,167(3):562-569
α-Thrombin, bradykinin, and histamine are endogenous mediators that increase endothelial permeability. We examined the mechanism by which these three vasoactive mediators could alter permeability to albumin of human umbilical vein endothelial cells (HUVEC). HUVEC were grown to confluence on Transwell membranes and we monitored the flux of fluorescein isothiocyanate-labeled human serum albumin across the membrane from the upper to lower chamber of the Transwell. Addition of α-thrombin, bradykinin, or histamine increased the permeability coefficient of the HUVEC monolayer. At 30 min the permeability coefficient for α-thrombin was 4.92 × 10−6 cm/sec while histamine was 4.47 × 10−6 cm/sec. Maximum changes in the permeability coefficient were about three-fold control baseline values (1.59 × 10−6 cm/sec). There was also a temporal difference in the magnitude of the permeability coefficient. α-Thrombin and bradykinin induced HUVEC permeability was increased for the first 90 min after which it returned to control levels. In contrast, histamine increased the permeability of the HUVEC monolayer throughout the 2 h experiment. To determine a possible intracellular mechanism of the altered permeability coefficients, HUVEC were labeled with FURA-2 and intracellular calcium was monitored by digital fluorescence ratio imaging. Maximum intracellular calcium in HUVEC was increased by α-thrombin (245 ± 20 nM) and histamine (210 ± 22 nM), but not by bradykinin (70 ± 7 nM) as compared to control (69 ± 10). Fluorescent photomicrographs of HUVEC stimulated with the three agonists indicated that α-thrombin and histamine substantially altered HUVEC f-actin arrangement, while bradykinin had no effect on HUVEC f-actin distribution. These data support previous in vitro and in vivo studies demonstrating increased permeability by all three agonists. These data also show, for the first time, that histamine and α-thrombin increased permeability by calcium-dependent intracellular pathways, but bradykinin operates through a calcium-independent mechanism. © 1996 Wiley-Liss, Inc. 相似文献
10.
J W Wang W Su Y P Law C H Lu Y C Chen J L Wang H J Chang W C Chen C R Jan 《Hormone research》2001,55(6):265-270
BACKGROUND: The effect of bradykinin on intracellular free Ca(2+) levels ([Ca(2+)](i)) in MG63 human osteosarcoma cells was explored using fura-2 as a Ca(2+) dye. METHODS/RESULTS: Bradykinin (0.1 nM-1 microM) increased [Ca(2+)](i) in a concentration-dependent manner with an EC(50) value of 0.5 nM. The [Ca(2+)](i) signal comprised an initial peak and a fast decay which returned to baseline in 2 min. Extracellular Ca(2+) removal inhibited the peak [Ca(2+)](i )signals by 35 +/- 3%. Bradykinin (1 nM) failed to increase [Ca(2+)](i) in the absence of extracellular Ca(2+ )after cells were pretreated with thapsigargin (an endoplasmic reticulum Ca(2+) pump inhibitor; 1 microM). Bradykinin (1 nM)-induced intracellular Ca(2+) release was nearly abolished by inhibiting phospholipase C with 2 microM 1-(6-((17 beta-3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione (U73122). The [Ca(2+)](i )increase induced by 1 nM bradykinin in Ca(2+)- free medium was abolished by 1 nM HOE 140 (a B2 bradykinin receptor antagonist) but was not altered by 100 nM Des-Arg-HOE 140 (a B1 bradykinin receptor antagonist). Pretreatment with 1 pM pertussis toxin for 5 h in Ca(2+) medium inhibited 30 +/- 3% of 1 nM bradykinin-induced peak [Ca(2+)](i) increase. CONCLUSIONS: Together, this study shows that bradykinin induced [Ca(2+)](i) increases in a concentration-dependent manner, by stimulating B2 bradykinin receptors leading to mobilization of Ca(2+) from the thapsigargin-sensitive stores in a manner dependent on inositol-1,4,5-trisphosphate, and also by inducing extracellular Ca(2+) influx. The bradykinin response was partly coupled to a pertussis toxin-sensitive G protein pathway. 相似文献
11.
Mark E. Rowin Ralph E. Whatley Ted Yednock John F. Bohnsack 《Journal of cellular physiology》1998,175(2):193-202
Human polymorphonuclear leukocytes (PMNs) express β1 integrins that mediate adhesion to extracellular matrix proteins following stimulation with agonists that induce an increase in intracellular calcium. The purpose of these studies was to determine the contribution made by alterations in intracellular calcium ([Ca++]i) to inside-out activation of β1 integrins using dimethyl sulfoxide (DMSO)-differentiated granulocytic HL60 cells as a model of human PMNs. Activation of β1 integrins was determined by measuring the expression of an activation-dependent epitope on the β1 subunit that is recognized by monoclonal antibody (mAb) 15/7. Exposure of granulocytic HL60 cells to calcium ionophore ionomycin (800 nM) alone did not increase the binding of mAb 15/7 to the cell surface, nor did it increase β1 integrin-mediated adhesion of the cells to fibronectin. Similarly, exposure of the cells to the direct protein kinase C (PKC) activator, dioctanoylglycerol (di-C8) at 100 μM, neither increased binding of mAb 15/7 to these cells nor adhesion to fibronectin. Simultaneous addition of di-C8 and ionomycin, however, caused a significant increase in the expression of the 15/7 epitope and cell adhesion, suggesting synergy between elevating [Ca++]i and stimulating PKC in β1 integrin activation. Chelation of [Ca++]i with Quin-2 and EGTA reduced both basal (unstimulated) expression of the 15/7 epitope and basal adhesion of granulocytic HL60 cells to fibronectin. In addition, chelation of [Ca++]i caused a significant decrease in 15/7 binding and adhesion stimulated by low (1 ng/ml) concentrations of phorbol myristate acetate (PMA). The inhibitory effect of [Ca++]i chelation on β1 integrin activation was reversed by repleting [Ca++]i with ionomycin in a Ca++-containing buffer, or by the addition of higher concentrations of PMA (10 ng/ml). These data suggest a role for [Ca++]i in inside-out activation of β1 integrins, probably through a synergistic effect with PKC activation. J. Cell. Physiol. 175:193–202, 1998. © 1998 Wiley-Liss, Inc. 相似文献
12.
Jeffrey P. Gardner M. Balasubramanyam George P. Studzinski 《Journal of cellular physiology》1997,172(3):284-295
The physiologically active form of vitamin D, 1α,25-dihydroxyvitamin D3 (1,25D3), induces promyelocytic HL60 cells to differentiate towards monocyte-like cells. During this differentiation increased cytosolic calcium (Cai2+) and expression of surface receptors for chemotactic factors “prime” the cell for the activation of monocyte functions and the triggering of the respiratory burst pathway. We examined whether the Ca2+ influx mediated by store-operated channels (SOC) contributed to the increased Cai2+ following exposure of HL60 cells to 10−7 M 1,25D3. Cells treated with 1,25D3 for 72 hr demonstrated a rapid transient rise in Cai2+ followed by a second, phasic, increase in Cai2+ in response to the purinergic agonist ATP. This second Cai2+ transient was blocked by Ni2+, SKF 96365, or withdrawal of extracellular Ca2++. In cells suspended in Ca2+-free medium, peak changes (Δ) in [Ca2+]i elicited by ATP-induced Ca2+ mobilization occurred with similar EC50 values in differentiated and vehicle (EtOH)-treated cells; however, peak [Ca2+]i was reduced by 55% in 1,25D3-treated cells. Decreased Ca2+ mobilization was associated with a 25–35% reduction in intracellular Ca2+ stores (determined with ionomycin). 1,25D3-treated cells exposed to ATP or thapsigargin (Tg) in Ca2+-free medium for 3 min with subsequent addition of 1 mM Ca2+ exhibited a respective 80% or 120% stimulation in peak [Ca2+]i compared to EtOH-treated cells. Enhanced Ca2+ influx mediated by SOC was also seen in these cells as an increase in the rate of Mn2+ entry after exposure to ATP or Tg. At 96 hr after addition of 1,25D3, when differentiated phenotype was established, basal Ca2+i and Ca2+ entry mediated by SOC returned to control values, but Ca2+ store size remained reduced. Up-regulation of Ca2+ influx via the SOC pathway during 1,25D3-induced differentiation may contribute to the functional properties of the maturing monocyte, or to the resetting of molecular programs responsible for the changing phenotype. J. Cell. Physiol. 172:284–295, 1997. © 1997 Wiley-Liss, Inc. 相似文献
13.
Toshimichi Yoshida Yasuo Takahashi Shiro Takashima 《Journal of cellular physiology》1993,154(2):301-309
The transformation of certain cells reduces the requirement of extracellular Ca2+ for growth. The SV-40 transformed human lung fibroblasts, WI-38 VA13, require less Ca2+ than normal WI-38 cells. Spreading area of normal cells decreases when cultured in 10 μM Ca2+ medium. Intracellular calcium concentration ([Ca2+]i), of the normal and transformed cells cultured in 10μM and 2 mM Ca2+ media was measured by the fluorescence microscope technique using fura-2 as a probe. The [Ca2+], is measured in the resting state and during mobilization by serum or bradykinin stimulation. The lowering of extracellular calcium concentration results in a decrease in the resting state [Ca2+],i of both normal and transformed cells. Although the total decrease in [Ca2+]i is the same for both cell, the rate of decrease is much faster in normal cells than in transformed cells. Low extracellular Ca2+ reduces the number of cells responsive to the serum or bradykinin stimulation and decreases the peak [Ca2+]i value in both cells. In addition, we investigated, using BCECF as a fluorecent probe, the intracellular pH (pHi) of normal and transformed cells maintained at low and normal Ca2+. The low Ca2+ condition makes pHi acidic in normal cells but not in transformed cells. The acidification of the normal cell is accompanied by a decrease in the spreading area of the cells. The decrease of the cell attacment, followed by the reduced spreading area, induced the acidic pHi. These results suggest that the reduced Ca2+ requirement of transformed cells for growth is related to the mechanism of pHi regulation rather than Ca2+ homeostasis and, possibly, to the anchorage-independent growth, which is a unique feature of transformed cells. © 1993 Wiley-Liss, Inc. 相似文献
14.
Cd2+ provokes inositol trisphosphateproduction and releases stored Ca2+, apparently by binding to a zinc site in the external domain of an orphan receptor. One pM Cd2+ evokes an immediate spike in cytosolic free Ca2+, which is similar to that evoked by bradykinin. Platelet-derived growth factor (PDGF) also increases free Ca2+ in human dermalfibroblasts, but there is a distinct lag before free Ca2+ rises in response to PDGF. Genistein, which selectively inhibits tyrosine kinases, markedly inhibited Ca2+ mobilization evoked by PDGF. Calcium mobilization triggered by cadmium or bradykinin was relatively insensitive to genistein. The PDGF receptor is known to be a tyrosine kinase, whichphosphorylates and thereby activatesphospholipase C, whereas a G protein couples the bradykinin receptor to anotherphospholipase C isoform. These findings support the hypothesis that the orphan receptor triggered by cadmium is coupled to phospholipase C via a G protein.Abbreviations BSA
bovine serum albumin
- BK
bradykinin
- [Ca2+]i
cytosolic free calcium
- DME
Dulbecco's modified Eagle's medium
- FBS
fetal bovine serum
- HEPES
4-(2-hydroxyethyl)-1-piperazine-ethanesulfonic acid
- IC50
concentration that produces 50% inhibition
- PDGF
platelet-derived growth factor
- PSS
physiological salts solution
- SE
standard error of the mean 相似文献
15.
Continuous superfusion of rat glioma cells with medium containing bradykinin (from 0.2 nM) induced a transient hyperpolarization followed by regular hyperpolarizing oscillations of the membrane potential. Similar repetitive hyperpolarizing oscillations were caused by extracellularly applied bradykinin or muscarine or by intracellularly injected GTP-gamma-S. The frequency of the oscillations was 1 per minute at bradykinin concentrations ranging from 0.2 nM to 2 microM, but the amplitude and duration increased with rising peptide concentration. The muscarine-induced oscillations were blocked by atropine. In the presence of extracellular Ca2+, the substances thapsigargin, 2,5-di(tert-butyl)-1,4-benzohydroquinone (tBuBHQ), and ionomycin reversibly suppressed the bradykinin-induced oscillations. Thapsigargin and tBuBHA, which are known to block the Ca2+ ATPase of endoplasmic reticulum, caused a transient rise in cytosolic Ca2+ activity, monitored with Fura-2, in suspensions of rat glioma cells or of mouse neuroblastoma-rat glioma hybrid cells. After a transient Ca2+ rise caused by thapsigargin, tBuBHQ, or ionomycin, the Ca2+ response to bradykinin which is known to be due to release of Ca2+ from internal stores was suppressed. This indicates that thapsigargin and tBuBHQ deplete internal Ca2+ stores as already seen previously for ionomycin. Thus, the inhibition of the membrane potential oscillations by thapsigargin, tBuBHQ, and ionomycin indicates that the oscillations are associated with activation of InsP3-sensitive Ca2+ stores. In some cells composite oscillation patterns which consisted of two independent oscillations with different amplitudes that overlapped additively were seen. We discuss that this pattern and the concentration dependency of the oscillations could be due to "quantal" Ca2+ release from stores with different inositol 1,4,5-triphosphate sensitivities. Subsidence of the oscillations after omission of extracellular Ca2+ seems to be due to a lack of replenishment of the intracellular stores with Ca2+, which comes from the extracellular compartment. 相似文献
16.
Thrombin-, bradykinin-, and arachidonic acid-induced Ca2+ signaling in Ehrlich ascites tumor cells 总被引:1,自引:0,他引:1
Jorgensen Nanna Koschmieder; Petersen Stine Falsig; Hoffmann Else Kay 《American journal of physiology. Cell physiology》1999,276(1):C26
Stimulation ofsingle Ehrlich ascites tumor cells with agonists (bradykinin, thrombin)and with arachidonic acid (AA) induces increases in the freeintracellular Ca2+ concentration([Ca2+]i)in the presence and absence of extracellularCa2+, measured using theCa2+-sensitive probe fura 2. Sequential stimulation with two agonists elicits sequential increasesin[Ca2+]i,unlike addition of the same agonist twice. Bradykinin and thrombin haveadditive effects on[Ca2+]iin Ca2+-free medium. Thephosphoinositidase C inhibitor U-73122 inhibits the agonist-inducedincreases in[Ca2+]i,whereas ryanodine has no effect. Pretreatment of cells in Ca2+-free medium with thapsigarginabolishes the bradykinin-induced increase in[Ca2+]ibut not the response to thrombin. The AA-induced response is notinhibited by U-73122 and cannot be mimicked by the inactive structuralanalog trifluoromethylarachidonyl ketone. Pretreatment of the cellswith 50 µM AA (but not with 10 µM AA) abolishes the agonist-inducedincrease in[Ca2+]i.Thus bradykinin, thrombin, and AA induce increases in[Ca2+]iin Ehrlich cells due to Ca2+ entryand release from intracellular stores. Thrombin causes release ofCa2+ from an intracellular storethat is insensitive to bradykinin and is not depleted by thapsigarginbut is depleted by AA. 相似文献
17.
The characteristics and properties of the increase in cytosolic [Ca2+] that occurs in bovine adrenal medullary chromaffin cells on exposure to histamine have been investigated. Specifically, these experiments were conducted to determine how much external Ca2+ enters the cell through a (capacitative) Ca2+ entry pathway activated as a consequence of intracellular Ca2+ store mobilization, relative to that which enters independently of store depletion via other channels activated by histamine. In Fura-2 loaded cells continued exposure to histamine (10 μM) 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 (100 nM) in Ca2+-free medium to deplete the internal Ca2+ stores, histamine caused no increase in cytosolic [Ca2+] when external Ca2+ was absent. Re-introduction of external Ca2+ to thapsigargin-treated store-depleted cells caused a sustained increase in cytosolic [Ca2+] that was further increased (P < 0.0002) upon exposure to histamine. The histamine-evoked increase was prevented by the H1-receptor antagonist, mepyramine (2 μM). A comparison was made between store-dependent Ca2+ entry consequent upon store mobilization with histamine in Ca2+-free medium and plateau phase Ca2+ entry resulting from stimulation with histamine in Ca2+-containing medium. The latter was found to be approximately 3 times greater in magnitude than the former (P ? 0.0001) at the same concentration of histamine (10 μM). It is concluded that histamine causes Ca2+ entry not only via a capacitative entry pathway secondary to internal store mobilization, but also causes substantial Ca2+ entry through other pathways. 相似文献
18.
These experiments were designed to learn the role of bradykinin induced changes in intracellular Ca2+ in the activation of phospholipase D activity in PC12 cells. Ionomycin at a concentration of 0.1M caused an increase in intracellular Ca2+ comparable to bradykinin, but had no effect on phospholipase D activity. Carbachol, ATP, and thapsigargin also increased intracellular Ca2+ but had no effect on phospholipase D activity. Increases in intracellular Ca2+ may be a necessary but not a sufficient factor in the activation of phospholipase D. To investigate this issue, the bradykinin induced increase in intracellular Ca2+ was blocked by preincubating the cells in Ca2+-free media plus EGTA or in media containing the intracellular Ca2+ chelator BAPTA/AM. These preincubations completely blocked the bradykinin induced increase in intracellular Ca2+ but only attenuated the bradykinin mediated activation of phospholipase D. Physiological increases in intracellular Ca2+ apparently do not mediate the effect of bradykinin on phospholipase D. 相似文献
19.
Kim JH Jain D Tliba O Yang B Jester WF Panettieri RA Amrani Y Puré E 《American journal of physiology. Lung cellular and molecular physiology》2005,289(4):L511-L520
The molecular mechanisms by which bradykinin induces excessive airway obstruction in asthmatics remain unknown. Transforming growth factor (TGF)-beta has been involved in regulating airway inflammation and remodeling in asthma, although it is unknown whether TGF-beta can modulate bradykinin-associated bronchial hyperresponsiveness. To test whether TGF-beta directly modulates airway smooth muscle (ASM) responsiveness to bradykinin, isolated murine tracheal rings were used to assess whether TGF-beta alters ASM contractile responsiveness to bradykinin. Interestingly, we found TGF-beta-treated murine rings (12.5 ng/ml, 18 h) exhibited increased expression of bradykinin 2 (B(2)) receptors and became hyperreactive to bradykinin, as shown by increases in maximal contractile responses and receptor distribution. We investigated the effect of TGF-beta on bradykinin-evoked calcium signals since calcium is a key molecule regulating ASM excitation-contraction coupling. We reported that TGF-beta, in a dose- (0.5-10 ng/ml) and time- (2-24 h) dependent manner, increased mRNA and protein expression of the B(2) receptor in cultured human ASM cells. Maximal B(2) receptor protein expression that colocalized with CD44, a marker of membrane cell surface, occurred after 18 h of TGF-beta treatment and was further confirmed using fluorescence microscopy. TGF-beta (2.5 ng/ml, 18 h) also increased bradykinin-induced intracellular calcium mobilization in fura-2-loaded ASM cells. TGF-beta-mediated enhancement of calcium mobilization was not attenuated with indomethacin, a cyclooxygenase inhibitor. These data demonstrate for the first time that TGF-beta may play a role in mediating airway hyperresponsiveness to bradykinin seen in asthmatics by enhancing ASM contractile responsiveness to bradykinin, possibly as a result of increased B(2) receptor expression and signaling. 相似文献
20.
Characterization of Bradykinin-Induced Phosphoinositide Turnover in Neurohybrid NCB-20 Cells 总被引:4,自引:2,他引:2
Phosphoinositide hydrolysis was studied in neurohybrid NCB-20 cells prelabeled with myo-[3H]inositol. Among nearly 20 neurotransmitters and neuromodulators examined, only bradykinin, carbachol, and histamine significantly increased the accumulation of [3H]inositol monophosphate (IP1) in the presence of lithium. The EC50 of bradykinin was 20 nM and the saturating concentration was approximately 1 microM. The bradykinin response was robust (10-fold) and was potently and selectively blocked by a bradykinin antagonist, B 4881 [D-Arg-(Hyp3, Thi, D-Phe)-bradykinin], with a Ki of 10 nM. This effect of bradykinin appeared to be additive to that mediated by activation of muscarinic cholinergic and histamine H1 receptors. The accumulation induced by bradykinin or carbachol was dependent on the presence of calcium in the incubation medium; less than twofold stimulation was observed in the absence of exogenous calcium. Bradykinin-induced [3H]IP1 accumulation required high concentration of lithium to elicit its maximal stimulation; the concentration of lithium required for half maximal effect was about 13 mM, similar to the value reported previously for carbachol-induced accumulation in the same cell line. In contrast, using related neurohybrid NG108-15 cells, bradykinin-induced [3H]IP1 accumulation was found to require much less lithium. IN the presence of lithium, bradykinin also evoked a transient increase in the production of [3H]-inositol bis- and trisphosphate. Basal and bradykinin-induced phosphoinositide breakdown was inhibited by 4 beta-phorbol 12,13-dibutyrate, but was unaffected by the biologically inactive 4 beta-phorbol. Pretreatment of cells with pertussis toxin induced only about 30% loss of the bradykinin-induced [3H]IP1 accumulation, without affecting basal activity.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献