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N Chakravarty

Publications and source records attributed to N Chakravarty.

At least 19 recordsLinked to original sources
PubMed1993

Inhibition of histamine release from mast cells by nigellone.

Nigellone is the carbonyl polymer of thymoquinone, isolated from Nigella Sativa L. seeds. The polymer is far less toxic but retains much of the pharmacologic properties of thymoquinone, which is the active principle. Our investigations, carried out on rat peritoneal mast cells in vitro, show that nigellone in relatively low concentrations is very effective in inhibiting histamine release induced by the secretagogues: antigen in sensitized cells, compound 48/80, and the calcium ionophore A23187. The mechanism of action seems to be through decreasing intracellular calcium by inhibiting its uptake and stimulating the efflux, and by an inhibition on protein kinase C. There is also indication for a mild inhibition of oxidative energy metabolism contributing to some inhibition of the release.

Animals↗
PubMed1992

The roles of calmodulin and protein kinase C in histamine secretion from mast cells.

The effect of a new calmodulin-antagonist, 5-iodo-1-C8, with a high selectivity for calmodulin in comparison to protein kinase C, has been investigated on histamine secretion from mast cells. It has been found to be much more sensitive for the inhibition of histamine secretion than the earlier calmodulin-antagonists, trifluoperazine and W7. The effect of four inhibitors of protein kinase C, viz. staurosporine, K252a, tamoxifen and sphingosine, has also been studied on histamine secretion from mast cells. All of them caused dose-dependent inhibition of histamine secretion induced by the three secretagogues used: antigen, compound 48/80 and the calcium ionophore A23187. K252a was tested against histamine release, induced by the stimulation of protein kinase C alone with the phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA) or the synthetic diacylglycerol, 1-oleoyl-2-acetyl-rac-glycerol (OAG). In both the cases K252a caused dose-dependent inhibition of histamine release. Staurosporine was also tested against TPA and was found to inhibit the release induced by it. Potentiation and inhibition (modulation) of secretagogue-induced histamine release by simultaneous protein kinase C stimulation with TPA or OAG have been demonstrated before. The potentiation and inhibition are shown to be antagonized by staurosporine. The observations point to the involvement of both calmodulin and protein kinase C in the histamine secretion process from mast cells.

Alkaloids↗
PubMed1990

The involvement of protein kinase C in exocytosis in mast cells.

Diacylglycerol generated from inositolphospholipid hydrolysis and tumor-promoting phorbol esters stimulate protein kinase C. The synthetic diacylglycerol 1-oleoyl-2-acetyl-rac-glycerol and the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) have been used in pure rat peritoneal mast cells. Both caused histamine release associated with exocytosis. The release by the stimulation of protein kinase C alone in the absence of secretagogues was slow although up to 50% of the histamine content was released by TPA in 120 min. Remarkable potentiation of histamine release was observed when the mast cells were preincubated with TPA before exposure to the calcium ionophore A23187. The potentiation of histamine release corresponded with an intensification of exocytosis. The potentiation is consistent with a participation of protein kinase C in the secretory process. An inhibitory effect due to protein kinase C activity was also demonstrated using TPA and mast cells from sensitized rats. When sensitized mast cells preincubated with 50 nM TPA for 5 min were exposed to the antigen, the histamine release was substantially reduced compared to the sum of the release by the antigen and TPA or by the antigen alone. There was a corresponding decrease in exocytosis. The inhibition of exocytosis and histamine release seems to reflect a regulatory function of protein kinase C for the termination of the response, as demonstrated in other types of cells apparently acting through an inhibition of inositolphospholipid hydrolysis.

Animals↗
PubMed1990

The role of protein kinase C in histamine secretion from mast cells.

Receptor activation on the cell surface is coupled through a guanine nucleotide regulatory protein to polyphosphoinositide phosphodiesterase. The activation of this enzyme catalyses the hydrolysis of phosphatidylinositol biphosphate. One of the products of this hydrolysis is diacylglycerol, which activates protein kinase C. It can also be activated by tumour-promoting phorbol esters. The synthetic diacylglycerol, 1-oleoyl-2-acetyl-rac-glycerol (OAG) and the phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA) have been used to stimulate protein kinase C in a pure population of rat peritoneal mast cells. Both of them caused histamine release, but the rate of release with TPA or OAG alone was slow. The release was inhibited by blocking the oxidative energy metabolism with antimycin A, and was associated with progressive exocytosis, showing that it is a secretory process. Studies on the interaction between the stimulation of protein kinase C by OAG/TPA and the secretagogues showed a dual effect, both potentiation and inhibition. Antigen (in sensitized cells) and compound 48/80 showed this pattern of response. With the calcium ionophore, A23187, potentiation was the dominant effect, although some inhibition could be shown with TPA. This is possibly related to the large calcium influx which causes translocation of protein kinase C to the membranes and enhances its activity. The potentiation suggests that protein kinase C is involved in the secretion process by the secretagogues, while the inhibition reflects a regulatory function, which is apparently exerted through an inhibition of phosphatidylinositol breakdown. Calcium uptake was enhanced by both TPA and OAG. Protein kinase C may thus contribute to the replenishment of the intracellular calcium stores after the secretory response.

Animals↗
PubMed1988

Effect of TMB-8 on calcium transport in mast cells in relation to histamine secretion.

We have previously reported an inhibition of histamine release by TMB-8 both in the presence and absence of calcium and with glucose in the medium. In the present investigation we have studied the effect of TMB-8 on calcium transport. The observations show that TMB-8 inhibits calcium uptake and enhances calcium efflux in mast cells. As antigen-induced histamine release from sensitized mast cells is primarily dependent on extracellular calcium, the inhibition of anaphylactic histamine release by TMB-8 is probably mainly due to an inhibition of calcium influx into the mast cells. We have shown an increased calcium efflux during histamine release from mast cells induced by compound 48/80 in the absence of calcium in the medium, suggesting the release of intracellular calcium stores. The increased calcium efflux was not inhibited by TMB-8. On the contrary, the enhanced calcium efflux caused by compound 48/80, was added to that by TMB-8. TMB-8 thus had no effect on the calcium release from intracellular stores by compound 48/80 but the enhanced calcium efflux by TMB-8 would tend to inhibit histamine release.

Animals↗
PubMed1987

Calcium uptake in mast cells, energy metabolism and histamine secretion.

The inhibition of energy metabolism of mast cells causes an inhibition of histamine secretion. As the secretion is generally initiated by the influx of calcium into the cell, we have made correlative studies of the effect of blocking the energy metabolism on calcium uptake and histamine secretion. When the influx of calcium is increased by exposing the cells to low concentrations of saponin or ionophore A23187, histamine release occurs, having the character of a secretory response. Brief incubation of the cells with antimycin A, 10(-9) M-10(-7) M, prior to exposure to saponin or the calcium ionophore gave similar dose-response curves for the inhibitory effect of antimycin A on calcium uptake and histamine release. The inhibition of calcium uptake in untreated mast cells by antimycin A, 10(-9) M-10(-7) M, showed good correlation to the inhibition of anaphylactic histamine release and the release induced by compound 48/80. The antigen-induced histamine release is dependent on extracellular calcium and an inhibition of its uptake by antimycin A could by itself inhibit the release. Compound 48/80 on the other hand induces histamine release both in the presence and absence of calcium, and both are similarly inhibited by 10(-9) M-10(-7) M antimycin A. This indicates that antimycin A has other sites of action apart from the inhibition of the influx of extracellular calcium. The inhibitory effect of antimycin A on compound 48/80-induced histamine secretion in the absence of extracellular calcium may be due to an inhibition of energy requiring steps in the final phase of the secretory process.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1987

Role of a Ca2+-Mg2+ ATPase on the mast cell surface in calcium transport and histamine secretion.

We have previously reported the presence of an ATPase, stimulated by calcium and magnesium, on the outer surface of the rat peritoneal mast cell. Experiments in which the enzyme activity was enhanced or inhibited showed a relationship to histamine secretion. Enhanced enzyme activity with increasing concentrations of the substrate (ATP) was associated with a potentiation of histamine release, and a pronounced inhibition of the enzyme caused an inhibition of the release. In the present work we have studied the influx and efflux of calcium in mast cells in relation to the activity of the Ca2+-Mg2+ ATPase on the mast cell membrane. The enzyme activity is shown to be related to calcium influx and has no effect on calcium efflux. Stimulation of the enzyme with ATP is associated with increased calcium influx into the mast cell, and inhibition of the enzyme with AMP causes inhibition of the calcium uptake. In both cases calcium efflux is unaffected. The function of the enzyme is thus different from the calcium efflux enzyme on the cytoplasmic surface, described in other cells. In addition, the Ca2+-Mg2+ ATPase on the mast cell surface is neither stimulated by calmodulin nor inhibited by the calmodulin antagonists, trifluoperazine and W-7. In mast cells the low cytosolic calcium concentration seems to be maintained by Na+-Ca2+ countertransport. Phosphorylation of the Ca2+-Mg2+ ATPase on the mast cell is likely to be associated with Ca2+ release at the cytoplasmic surface of the plasma membrane. It is thus possible that ATP hydrolysis in the membrane stimulates the contraction of microfilaments in the membrane and the cytoskeleton, and promotes the migration of the granules to the plasma membrane.

Animals↗
PubMed1986

Histamine secretion from permeabilized mast cells by calcium.

A transient increase in the permeability of the mast cell membrane was caused by the exposure of the cells to low concentrations of saponin, 5 or 10 micrograms/ml. These concentrations had very little effect in the absence of calcium but caused 35 to 50% histamine release, having the character of a secretory response, when 0.25 mM or more calcium was added to the medium. The dose-response curve was steep between 25 microM and 250 microM calcium and tended to flatten with higher concentrations. The release was associated with a pronounced increase in calcium uptake, which was faster than the histamine release. The membrane changes were slight as indicated by only 7 to 12% leakage of lactate dehydrogenase and by the absence of any detectable change in the electron micrographs. The transient nature of the membrane change is shown by the following experiment. When the cells were first exposed to saponin in the absence of calcium, the amount of histamine released by the subsequent incubation with calcium varied inversely with the time interval that elapsed before calcium was added. If calcium was added after 15 minutes no histamine release occurred. When calcium uptake was studied in the same manner, the stimulation of calcium uptake in saponin-treated cells also declined progressively with increasing intervals after the exposure to saponin when calcium was added. Stimulation of both histamine release and calcium uptake was inhibited by antimycin A, the inhibition curves with 10(-9)M to 10(-7)M antimycin A being similar. The effect on the calcium uptake by itself could explain the inhibition of histamine release. But the release was also inhibited by the calmodulin antagonists, W-7 and mepacrine, suggesting that the influx of calcium in the permeabilized cells acts primarily through calmodulin-mediated enzyme activation.

Animals↗
PubMed1986

Regulatory role of calcium on histamine secretion.

Calcium seems to have two opposing effects on histamine secretion from mast cells. A rise in the cytosol calcium concentration initiates the chain of reactions leading to histamine secretion. On the other hand, calcium appears to have a regulatory role, limiting the secretion. Removal of cell surface calcium enhances histamine secretion. The present work demonstrates an inhibitory effect of calcium in the medium, using low concentrations of compound 48/80 as the secretagogue. Histamine secretion in response to compound 48/80 primarily utilizes intracellular calcium. When low concentrations of compound 48/80 were used (usually 20-50 ng/ml), calcium (1 mM) inhibited the secretion, the inhibition being more pronounced as the pH was increased from 6.5 to 8.5. The higher pH conceivably promotes the binding of calcium to the phospholipids in the cell membrane. Calcium at this site seems to depress the efflux of calcium from the intracellular stores to the cytosol. The possibility that the removal of calcium from the cell surface causes increased sodium permeability was considered. However, the sodium channel blocker tetrodotoxin (10(-5) M) was equally ineffective in influencing histamine release in the presence and absence of calcium, indicating that a change of sodium permeability was not involved. Antigen-induced (anaphylactic) histamine secretion depends mainly on extracellular calcium, although some secretion occurs in a calcium-free medium. Addition of calcium alone to the medium caused only slight increase in the secretion, but when both phosphatidylserine and calcium were added histamine secretion was remarkably stimulated, apparently through the effect of phosphatidylserine on calcium transport across the plasma membrane.

Anaphylaxis↗
PubMed1986

Histamine release from saponin-permeabilized rat mast cells by calcium.

The first effect of receptor activation on the mast cell surface, initiating histamine secretion, is an increase in the cytosol Ca2+ concentration. It should then be possible to induce histamine secretion by calcium alone, if the calcium permeability of the cell membrane could be increased without any significant interference with the physiological cell functions. This was achieved in the present study by adding low concentrations of saponin (0.0005% and 0.001% w/v) to the medium. When calcium was added to the saponin-permeabilized cells, around 40% histamine release occurred with 0.25 mM extracellular calcium (free Ca2+ 0.15 mM). The release was inhibited by antimycin A (1 microM). Transmission electron microscopy showed formation of vacuoles containing granules stripped of their membranes, which characterize a secretory response. The observations are consistent with a limited increase in the calcium permeability of the cell membrane for a brief period. There was apparently an increase in the cytoplasmic calcium concentration, which acted through calmodulin, since the histamine release induced by calcium from the permeabilized mast cells could be inhibited by a calmodulin-antagonist, mepacrine (10-30 microM).

Animals↗
PubMed1985

Calmodulin in mast cells and its role in histamine secretion.

Calmodulin content and distribution in rat peritoneal mast cells was determined by radioimmunoassay. Isolated pure mast cells were disrupted by sonication and the total calmodulin content and its distribution were determined. Calmodulin bound to the membranes was released by heating with 0.1% Lubrol PX to 95 degrees C for 5 min. The total calmodulin content of mast cells was found to be 160 +/- 14 ng/10(6) cells (9.4 +/- 0.82 pmoles/10(6) cells). The highest amount (68%) was present in the supernatant representing the cytosol. The next highest amount (26%) was found in the composite fraction consisting of mitochondria, endoplasmic reticulum, Golgi vesicles and plasma membrane (100,000 g pellet). The mast cell granules contained 4% of the total calmodulin. Trifluoperazine (TFP) was used as an antagonist to explore the role of calmodulin in histamine secretion. At 10 microM concentration, TFP caused a negligible spontaneous histamine release by its membrane effect. TFP (10 microM) inhibited histamine release by all the three secretagogues used, but the degree of inhibition varied: 60% with antigen, 40% with compound 48/80 and 20% with ionophore A23187. It is suggested that the TFP effect is due to calmodulin-antagonism and interference with the activation of enzymes, essential to the secretory process.

Animals↗
PubMed1985

Glucose metabolism in rat mast cells. Stimulation of the pentose phosphate pathway by compound 48/80.

The glycogen content of rat peritoneal mast cells (mean: 3 nmoles/10(6) cells) was increased 15% by incubation with glucose (1 mM) and reduced 35% when incubated without glucose at 37 degrees C for 15 min. The storage capacity for glycogen is thus low. Lactate production at 37 degrees C in a substrate-free medium was low (2.5-6.3 nmoles/10(6) cells in 40 min), but was stimulated 5-fold in the aerobic medium and 10-15 fold in the anaerobic medium by glucose. Both aerobic and anaerobic glycolysis in presence of glucose can thus provide energy for histamine secretion. The initial enzymes of the pentose phosphate pathway, glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase, have been demonstrated in mast cells. The enzyme activity in mast cells was, however, low compared to the high activity in the other peritoneal cells. The extent of the pentose cycle activity was determined from the conversion of 14C1- and 14C6-glucose to 14CO2, expressing the specific 14CO2 yields as fractions of the total glucose utilization. The normal pentose cycle activity with 1 mM glucose was 0.4% of the glucose metabolism. This was remarkably simulated by an electron acceptor, phenazine methosulfate. The pentose cycle was enhanced to 0.71% (80% stimulation) after exposure of the mast cells to compound 48/80, causing 68% histamine release. The stimulation of the pentose cycle by compound 48/80 seems to be due to the enhancement of biosynthetic processes during the regenerative phase.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1984

Role of intracellular calcium in histamine release from rat mast cells.

Compound 48/80-induced histamine release may utilize both intracellular and extracellular calcium. The release is inhibited by La3+ and TMB-8 both in the presence and absence of calcium. The observations are consistent with a mobilization of calcium from the inner surface of the plasma membrane supporting the release. TMB-8 probably acts at more than one site. It has been shown to inhibit skeletal muscle contraction and this has been correlated to its inhibitory effect on the release of Ca2+ from the sarcoplasmic reticulum. TMB-8 also inhibits resting cellular influx and efflux of calcium in the ileum. Anaphylactic and dextran-induced histamine release is dependent on extracellular calcium; the inhibition of the release by TMB-8 seems to be primarily due to an interference with calcium transport across the plasma membrane. In the absence of extracellular calcium the inhibition of compound 48/80-induced histamine release by TMB-8 seems to be due to stabilization of Ca2+ binding to cellular stores.

Animals↗
PubMed1984

Effect of arachidonic acid metabolism on the release of histamine and SRS (leukotrienes) from guinea-pig lung.

The effect of arachidonic acid (AA) metabolism on histamine release and SRS (leukotrienes) production has been studied in guinea-pig lung using anaphylactic reaction and Ca2+ ionophore as the triggering agents in vitro. AA and L-cysteine enhanced SRS production without any appreciable effect on histamine release. Two nonsteroid anti-inflammatory agents, indomethacin and ketoprofen, which block prostaglandin production by the cyclooxygenase pathway, stimulated SRS production but had hardly any effect on histamine release, indicating that SRS synthesis is more sensitive to prostaglandin regulation. Enhancement of SRS production was more pronounced for antigen than for Ca2+ ionophore. This might be related to different cellular origin of SRS with the two triggering agents. Using rat peritoneal cells, both mast cells and the other cells were found to produce SRS in response to Ca2+ ionophore, the amount formed by the latter type of cells being higher. Inhibition of lipoxygenase by 5,8,11,14-eicosatetraynoic acid and nordihydroguaiaretic acid depressed SRS production, but had no effect on histamine release. SRS production triggered by Ca2+ ionophore was more sensitive, possibly because of different cellular origin of SRS in response to the two stimuli. The explanation for the discrepancy between the effect on SRS production and histamine release may also have to be sought in their different origins. SRS may mainly stem from cells, which are more sensitive to the inhibitors than the mast cell, which is the source of histamine.

Animals↗