scholarly journals Econazole inhibits thapsigargin-induced platelet calcium influx by mechanisms other than cytochrome P-450 inhibition

1993 ◽  
Vol 295 (2) ◽  
pp. 525-529 ◽  
Author(s):  
J G Vostal ◽  
J C Fratantoni
Keyword(s):  
Plasma Membrane ◽  
Antifungal Agents ◽  
Calcium Influx ◽  
Calcium Stores ◽  
Alternative Mechanism ◽  
Resting Cells ◽  
Calcium Efflux ◽  
Plasma Membrane Permeability ◽  
Cytochrome P 450 ◽  
Internal Calcium

Cytochrome P-450 has been suggested as a mediator of the signal between depleted platelet calcium stores and an increase in plasma membrane permeability to calcium which follows depletion of the stores. This hypothesis is based on the observations that inhibitors of cytochrome P-450, such as the imidazole antifungal agents, also inhibit influx of a calcium surrogate (manganese) into calcium-depleted platelets. We tested the effects of econazole and of a cytochrome P-450 inhibitor, carbon monoxide (CO), on thapsigargin (TG)-induced platelet 45Ca2+ influx. TG specifically depletes internal calcium stores and activates store-regulated calcium influx. Econazole blocked 45Ca2+ influx when it was added before TG (IC50 11 microM). Econazole at a concentration (20 microM) that inhibited 83% of TG-induced calcium influx was not inhibitory to TG-induced calcium efflux from 45Ca(2+)-loaded platelets, and did not affect calcium fluxes in resting platelets. This econazole concentration was also inhibitory to calcium influx even when it was added after the stores had been calcium-depleted by EGTA and TG for 15 min and the signal to increase calcium influx had already been generated. Inhibition of cytochrome P-450 with CO bubbled through platelet suspensions did not change calcium influx in resting cells and potentiated TG-induced calcium influx (160% of control calcium accumulation at 20 min). This effect appeared to be concentration-dependent, such that a 5 min exposure to CO produced a greater influx potentiation than a 3 min exposure. These observations indicate that (1) cytochrome P-450 does not mediate store-regulated calcium influx, and (2) econazole probably inhibits store-regulated calcium influx by an alternative mechanism, such as interaction with plasma membrane calcium channels.

scholarly journals Agonist-induced Ca2+ influx in human neutrophils is secondary to the emptying of intracellular calcium stores

1991 ◽  
Vol 277 (1) ◽  
pp. 73-79 ◽  
Author(s):  
M Montero ◽  
J Alvarez ◽  
J Garcia-Sancho
Keyword(s):  
Plasma Membrane ◽  
Intracellular Calcium ◽  
Time Course ◽  
Human Neutrophils ◽  
Calcium Stores ◽  
Free Medium ◽  
Prolonged Incubation ◽  
Intracellular Calcium Stores ◽  
Cytochrome P 450 ◽  
In Cells

Emptying of the intracellular calcium stores of human neutrophils, by prolonged incubation in Ca(2+)-free medium, by treatment with low concentrations of the Ca2+ inophore ionomycin, or by activation with cell agonists, increased the plasma-membrane permeability to Ca2+ and Mn2+. The chemotactic peptide formylmethionyl-leucyl-phenylalanine and the natural agonists platelet-activating factor and leukotriene B4 released different amounts of calcium from the stores and induced Ca2+ (Mn2+) uptake, the rate of which correlated inversely with the amount of calcium left in the stores. The increased Mn2+ uptake induced by these agonists was persistent in cells incubated in Ca(2+)-free medium, but returned to basal levels in cells incubated in Ca(2+)-containing medium, with the same time course as the refilling of the calcium stores. The calcium-stores-regulated Mn2+ influx, including that induced by agonists, was prevented by cytochrome P-450 inhibitors. We propose that agonist-induced Ca2+ (Mn2+) influx in human neutrophils is secondary to the emptying of the intracellular stores which, in turn, activates plasma-membrane Ca2+ channels by a mechanism involving microsomal cytochrome P-450, similar to that described previously in thymocytes [Alvarez, Montero & Garcia-Sancho (1991) Biochem. J. 274, 193-197].

scholarly journals Cytochrome P-450 may link intracellular Ca2+ stores with plasma membrane Ca2+ influx

1991 ◽  
Vol 274 (1) ◽  
pp. 193-197 ◽  
Author(s):  
J Alvarez ◽  
M Montero ◽  
J García-Sancho
Keyword(s):  
Plasma Membrane ◽  
Aryl Hydrocarbon Hydroxylase ◽  
Plasma Membrane Permeability ◽  
Aryl Hydrocarbon ◽  
Oxygen Scavenging ◽  
Aryl Hydrocarbon Hydroxylase Activity ◽  
Cytochrome P 450 ◽  
Degree Of Filling ◽  
Ca2 Channels ◽  
Dependent Mechanism

We have studied the mechanism of the regulation of plasma membrane Ca2+ permeability by the degree of filling of the intracellular Ca2+ stores. Using Mn2+ as a Ca2+ surrogate for plasma membrane Ca2+ channels, we found that Mn2+ uptake by rat thymocytes is inversely related to the degree of filling of the intracellular Ca2+ stores. This store-dependent plasma membrane permeability is inhibited by oxygen scavenging, CO, imidazole antimycotics and other cytochrome P-450 inhibitors. The pattern of inhibition is similar to that reported previously for the inhibition of microsomal cytochrome P-450-mediated aryl hydrocarbon hydroxylase activity of lymphocytes. Several calmodulin antagonists, both phenothiazinic (trifluoperazine, fluphenazine and chlorpromazine) and dibenzodiazepinic (clozapine), accelerate Mn2+ uptake by cells with Ca2(+)-filled stores, and this effect is prevented by imidazole antimycotics. Our results suggest that cytochrome P-450 may be the link between the stores and the plasma membrane Ca2+ pathway. We propose a model in which this cytochrome, sited at the stores, stimulates plasma membrane Ca2+ influx. This stimulatory effect is, in turn, prevented by the presence of Ca2+ inside the stores, possibly via a calmodulin-dependent mechanism.

scholarly journals Control of Ca2+ entry into HL60 and U937 human leukaemia cells by the filling state of the intracellular Ca2+ stores

1993 ◽  
Vol 289 (3) ◽  
pp. 761-766 ◽  
Author(s):  
S R Alonso-Torre ◽  
J Alvarez ◽  
M Montero ◽  
A Sanchez ◽  
J García-Sancho
Keyword(s):  
Plasma Membrane ◽  
Inositol Phosphate ◽  
Platelet Activating Factor ◽  
Membrane Receptors ◽  
Plasma Membrane Permeability ◽  
Prolonged Incubation ◽  
Differentiated Cells ◽  
Cytochrome P 450 ◽  
Filling State ◽  
Ca2 Channels

Differentiation of HL60 cells by treatment with dimethyl sulphoxide induces the expression of membrane receptors for N-formylmethionyl-leucyl-phenylalanine (fMLP) and for platelet-activating factor (PAF). In these cells both agonists produced an increase in the cytosolic Ca2+ concentration ([Ca2+]i) by release of Ca2+ from the intracellular stores, followed shortly by an acceleration of the entry of Ca2+ or Mn2+, used here as a Ca2+ surrogate for Ca2+ channels. Cytochrome P-450 inhibitors blocked the agonist-induced entry of Ca2+ or Mn2+ with no modification of Ca2+ release from the stores. Emptying the intracellular Ca2+ stores either by treatments inducing no inositol phosphate production, such as prolonged incubation in Ca(2+)-free medium or treatment with the Ca2+ ionophore ionomycin, increased the plasma-membrane permeability to Ca2+ and Mn2+. This Ca(2+)-store-regulated Mn2+ entry was inhibited by Ni2+ and by cytochrome P-450 inhibitors. Refilling of the Ca2+ stores by incubation in Ca(2+)-containing medium restored low Mn2+ permeability. The same mechanism is present and functional in non-differentiated cells, before expression of membrane receptors for fMLP and PAF. These results suggest that agonist-induced Ca2+ (Mn2+) entry is secondary to the emptying of the intracellular Ca2+ stores, which in turn activates plasma-membrane channels by a mechanism involving cytochrome P-450.

scholarly journals Calcitropic effects of recombinant prolactins in Oreochromis mossambicus

1994 ◽  
Vol 266 (4) ◽  
pp. R1302-R1308 ◽  
Author(s):  
G. Flik ◽  
F. Rentier-Delrue ◽  
S. E. Wendelaar Bonga
Keyword(s):  
Plasma Membrane ◽  
Dose Response ◽  
Calcium Influx ◽  
Oreochromis Mossambicus ◽  
Membrane Preparation ◽  
Calcium Efflux ◽  
Branchial Epithelium ◽  
Plasma Membrane Preparation

Homologous recombinant tilapia prolactin-188 (PRL-I) and tilapia prolactin-177 (PRL-II) were tested for calcitropic activity in tilapia, Oreochromis mossambicus. Injection of PRL-I and PRL-II (4 injections, 12.5 pmol/g, over an 8-day period) induced hypercalcemia that resulted from an enhanced calcium influx via the gills and a decreased calcium efflux. Both PRLs increased the density of the Ca(2+)-transporting Ca(2+)-adenosinetriphosphatase in a plasma membrane preparation of the branchial epithelium. Dose-response studies (doses tested: 0.75-12.5 pmol/g) demonstrated that PRL-I was roughly twofold more potent than PRL-II in inducing hypercalcemia, in enhancing basal levels of cortisol, and in stimulating opercular ionocyte density. PRL-I and PRL-II were equipotent in stimulating the dermal mucocyte frequency. We conclude that in this species PRL-I and PRL-II have calcitropic effects, and that PRL-I is more potent than PRL-II in this respect. We postulate that PRL has corticotrophic activity in this fish.

scholarly journals Theoretical Study of the Function of the IP3 Receptor / BK Channel Complex in a Single Neuron

2021 ◽  
Author(s):  
◽  
M. E. Pérez-Bonilla
Keyword(s):  
Plasma Membrane ◽  
Calcium Influx ◽  
Ryanodine Receptors ◽  
Cytosolic Calcium ◽  
Bk Channels ◽  
Bk Channel ◽  
Ip3 Receptors ◽  
Ip3 Receptor ◽  
Calcium Efflux ◽  
The Central Nervous System

Large conductance calcium-activated potassium (BK) channels carry out many functions in the central nervous system. The opening of BK channels requires a rise in the cytosolic calcium ([Ca2+]cyt) concentration, which can occur in two ways: calcium influx from voltage-gated calcium channels (VGCCs) located on the plasma membrane and calcium efflux through the endoplasmic reticulum (ER) membrane to the cytosol triggered by inositol 1,4,5-trisphosphate (IP3) receptors (IP3-Rs) and ryanodine receptors (RyRs). The BK channel/IP3-R/RyR interaction has been widely reported in smooth muscle but scarce information exist on neurons, where its presence is uncertain. The aim of this study was to develop a computational model of a neuron to replicate the interaction between the release of Ca2+ from the ER (through IP3-Rs and RyRs) and the opening of BK channels on the plasma membrane to regulate the level of [Ca2+]cyt, based on the Hodgkin-Huxley formalism and the Goldbeter model. The mathematical models were implemented on Visual Basic® and differential equations were solved numerically. Various conditions of BK conductance and the efflux of endoplasmic Ca2+ were explored. The results show that an abrupt increase in [Ca2+]cyt (≥ 5 mM) activates the BK channels and either pauses or stops the action potential train.

scholarly journals Agonist-induced Ca2+ influx into human platelets is secondary to the emptying of intracellular Ca2+ stores

1991 ◽  
Vol 280 (3) ◽  
pp. 783-789 ◽  
Author(s):  
M T Alonso ◽  
J Alvarez ◽  
M Montero ◽  
A Sanchez ◽  
J García-Sancho
Keyword(s):  
Plasma Membrane ◽  
Platelet Activating Factor ◽  
Human Platelets ◽  
Temperature Dependent ◽  
Plasma Membrane Permeability ◽  
Low Concentrations ◽  
Cytochrome P 450 ◽  
Filling State ◽  
Ca2 Channels ◽  
Stimulation Of

We have studied the relation between the filling state of the intracellular Ca2+ stores and the plasma-membrane permeability to Mn2+, used here as a Ca2+ surrogate for Ca2+ channels. Emptying of the intracellular Ca2+ stores either by incubation in Ca(2+)-free medium or by treatment with low concentrations of the Ca2+ ionophore ionomycin accelerated the influx of Mn2+. Refilling of the Ca2+ stores by incubation in Ca(2+)-containing medium restores low Mn2+ permeability. This Ca(2+)-store-regulated permeability was inhibited by Ni2+ and by cytochrome P-450 inhibitors. Stimulation of platelets with thrombin produced Ca2+ release from the intracellular stores, which was followed, after a temperature-dependent lag (2 s at 37 degrees C; 5 s at 18 degrees C), by an acceleration of Mn2+ influx. Cytochrome P-450 inhibitors prevented the thrombin-induced Mn2+ influx, with little effect on the Ca2+ mobilization from the intracellular stores. Ki values were similar to those estimated for inhibition of the store-regulated permeability in non-stimulated platelets. Similar results were found in platelets stimulated by platelet-activating factor or by ADP. We propose that agonist-induced Ca2+ (Mn2+) influx in platelets is secondary to the emptying of the intracellular Ca2+ stores. The activation of the plasma-membrane Ca2+ (Mn2+) pathway may take place by a mechanism involving microsomal cytochrome P-450, similar to that described previously in thymocytes [Alvarez, Montero & García-Sancho (1991) Biochem. J. 274, 193-197] and neutrophils [Montero, Alvarez & García-Sancho (1991) Biochem. J. 277, 73-79].

scholarly journals Control of plasma-membrane Ca2+ entry by the intracellular Ca2+ stores. Kinetic evidence for a short-lived mediator

1992 ◽  
Vol 288 (2) ◽  
pp. 519-525 ◽  
Author(s):  
M Montero ◽  
J Alvarez ◽  
J García-Sancho
Keyword(s):  
Plasma Membrane ◽  
Time Lag ◽  
Human Neutrophils ◽  
Half Time ◽  
Plasma Membrane Permeability ◽  
Steady State Levels ◽  
Clearance Kinetics ◽  
Cytochrome P 450 ◽  
Degree Of Filling ◽  
Ca2 Channels

We have studied the correlation between the degree of filling of the intracellular Ca2+ stores and the plasma-membrane permeability to Mn2+, a Ca2+ surrogate for plasma-membrane Ca2+ channels, in human neutrophils loaded with fura-2. Refilling of the stores of cells previously depleted of Ca2+ decreased the entry of Mn2+, but the magnitude of this effect depended on the refilling protocol. When refilling was allowed to proceed to steady-state levels by a 3 min incubation with different external Ca2+ concentrations (0.05-1 mM), almost complete inhibition of Mn2+ entry was observed at 40% of maximum refilling. In contrast, when different degrees of store refilling were attained by incubation with 1 mM-Ca2+ for short periods (10-40 s), inhibition of Mn2+ entry was smaller at comparable degrees of refilling. When quick refilling was allowed to proceed up to 40% (about 20 s at 37 degrees C) and then stopped at this level by removal of external Ca2+, the rate of Mn2+ uptake was high just after refilling and then decreased with time within the next few seconds (half-times approximately 7 s at 37 degrees C and approximately 20 s at 25 degrees C). We have proposed previously that the Ca2+ stores, when emptied of Ca2+, may generate a second messenger able to open the plasma-membrane Ca2+ channels by a mechanism involving cytochrome P-450. The results here are consistent with the existence of such a messenger and suggest that it is cleared from the cytoplasm with a half-time of about 7 s at 37 degrees C. In addition, inhibition of Mn2+ entry in cells with empty Ca2+ stores by cytochrome P-450 inhibitors showed a time lag consistent with the clearance kinetics proposed above.

184 INHIBITION OF CALCIUM EFFLUX EXTENDS THE DURATION OF CALCIUM SIGNALS IN PIG OOCYTES

2012 ◽  
Vol 24 (1) ◽  
pp. 204
Author(s):  
L. Y. Yan ◽  
C. Wang ◽  
H. L. Luo ◽  
Z. Machaty
Keyword(s):  
Plasma Membrane ◽  
Intracellular Calcium ◽  
Calcium Influx ◽  
Control Group ◽  
Calcium Signal ◽  
Free Calcium ◽  
Cytoplasmic Calcium ◽  
Free Medium ◽  
Calcium Efflux ◽  
Calcium Signals

Calcium signaling involves the transient elevation in the intracellular free-calcium concentration, which is responsible for controlling a great number of biological functions. In many cell types, a signal is generated when calcium stored in the endoplasmic reticulum is released into the cytoplasm, followed by an influx of calcium across the plasma membrane. At the same time, calcium is removed from the cytosol by ATPases, which pump it back into the intracellular store or out of the cell. The size of the calcium signal is thus determined by the amount of calcium moving into and out of the cytoplasm. In the present study, we investigated the effect of inhibiting the movement of calcium across the plasma membrane on a calcium-signal that was artificially induced in pig oocytes. In vitro-matured pig oocytes were loaded with the calcium-indicator dye, fura-2. The release of calcium from the cytoplasmic stores was stimulated by adding ethanol at a final concentration of 7% and changes in the intracellular free-calcium levels were monitored by using InCyt Im2, a dual-wavelength fluorescence imaging system. In the control group, fluorescent measurements were performed in the presence of extracellular calcium. In additional treatment groups, the ethanol treatment was performed in (1) a calcium-free medium (prevents calcium influx); (2) the presence of 1 mM gadolinium (limits calcium fluxes across the plasma membrane in both directions); and (3) the presence of gadolinium in a calcium-free medium. In each group, 15 oocytes were measured; the data were subjected to 1-way ANOVA and differences between treatment means were compared by the Tukey's test. We found that in control oocytes, 7% ethanol induced a rise of 722.1 ± 49.2 nM in the cytoplasmic calcium levels and the average duration of the calcium transient was 309.1 ± 11.3 s. In the calcium-free medium, this increase was significantly smaller (only 117.9 ± 4.8 nM; P < 0.01), probably because the calcium release was not followed by an influx of calcium across the plasma membrane. In the presence of 1 mM gadolinium in the regular calcium-containing medium, the intracellular calcium levels climbed by 278.8 ± 45.4 nM and dropped to baseline levels only after 773.3 ± 79.9 s. Finally, in the calcium-free medium and in the presence of 1 mM gadolinium, 7% ethanol induced only a small rise (64.5 ± 6.3 nM) in the cytoplasmic calcium levels. These latter increases were both significantly lower (P < 0.01) compared to that in the control group. The results indicate that (1) intracellular calcium signals are sustained by a calcium influx from the extracellular medium and (2) preventing calcium efflux extends the duration of the cytoplasmic calcium elevation in the oocyte. This may have relevance in the manipulation of intracellular calcium levels in oocytes for the development of novel parthenogenetic activation methods.

scholarly journals Spatial Relationship and Functional Relevance of Three Lipid Domain Populations at the Erythrocyte Surface

2018 ◽  
Vol 51 (4) ◽  
pp. 1544-1565 ◽  
Author(s):  
Louise Conrard ◽  
Amaury Stommen ◽  
Anne-Sophie Cloos ◽  
Jan Steinkühler ◽  
Rumiana Dimova ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Calcium Influx ◽  
Spatial Relationship ◽  
Calcium Balance ◽  
Lipid Domains ◽  
Calcium Efflux ◽  
Lipid Domain ◽  
Erythrocyte Surface ◽  
Shape Restoration ◽  
Functional Relevance

Background/Aims: Red blood cells (RBC) have been shown to exhibit stable submicrometric lipid domains enriched in cholesterol (chol), sphingomyelin (SM), phosphatidylcholine (PC) or ganglioside GM1, which represent the four main lipid classes of their outer plasma membrane leaflet. However, whether those lipid domains co-exist at the RBC surface or are spatially related and whether and how they are subjected to reorganization upon RBC deformation are not known. Methods: Using fluorescence and/or confocal microscopy and well-validated probes, we compared these four lipid-enriched domains for their abundance, curvature association, lipid order, temperature dependence, spatial dissociation and sensitivity to RBC mechanical stimulation. Results: Our data suggest that three populations of lipid domains with decreasing abundance coexist at the RBC surface: (i) chol-enriched ones, associated with RBC high curvature areas; (ii) GM1/PC/chol-enriched ones, present in low curvature areas; and (iii) SM/PC/chol-enriched ones, also found in low curvature areas. Whereas chol-enriched domains gather in increased curvature areas upon RBC deformation, low curvature-associated lipid domains increase in abundance either upon calcium influx during RBC deformation (GM1/PC/chol-enriched domains) or upon secondary calcium efflux during RBC shape restoration (SM/PC/chol-enriched domains). Hence, abrogation of these two domain populations is accompanied by a strong impairment of the intracellular calcium balance. Conclusion: Lipid domains could contribute to calcium influx and efflux by controlling the membrane distribution and/or the activity of the mechano-activated ion channel Piezo1 and the calcium pump PMCA. Whether this results from lipid domain biophysical properties, the strength of their anchorage to the underlying cytoskeleton and/or their correspondence with inner plasma membrane leaflet lipids remains to be demonstrated.

Calcium efflux from an intracellular pool activated by GTP hydrolysis in cultured gastric smooth muscle

1994 ◽  
Vol 266 (3) ◽  
pp. G388-G394 ◽  
Author(s):  
R. J. Gilbert ◽  
M. Khalid
Keyword(s):  
Smooth Muscle ◽  
Calcium Release ◽  
Calcium Influx ◽  
Calcium Stores ◽  
Calcium Efflux ◽  
Gtp Hydrolysis ◽  
Intracellular Pool ◽  
Gastric Smooth Muscle ◽  
Gamma S ◽  
Calcium Pool

In these studies, we have characterized calcium movement due to guanosine triphosphate (GTP) hydrolysis from an ATP-sequestered intracellular calcium pool in cultured gastric smooth muscle. GTP (1-100 microM), when added to an ATP-regenerating medium, resulted in a concentration-dependent and irreversible efflux of calcium from an organellar calcium pool. GTP-induced calcium efflux was not affected by variation of the ATP/ADP ratio (8.5-155.0), indicating that GTP did not act by inhibiting calcium influx via calcium adenosinetriphosphatase. To assess whether the calcium increase was necessarily associated with GTP hydrolysis, experiments were performed with the nonhydrolyzable guanine nucleotide analogues guanosine 5'-[beta-thio]diphosphate (GDP beta S), 5'-guanylyl imidodiphosphate guanosine (GppNHp), and 5'-O-(3-thiotriphosphate) (GTP gamma S). Administration of GDP beta S and GppNHp resulted in no significant calcium efflux. GTP gamma S caused a small steady-state calcium increase (20% of that induced by the hydrolyzable nucleotide) but irreversibly inhibited all subsequent calcium increase due to GTP. The possibility that GTP may either modify the concentration of mobilizable calcium in inositol trisphosphate (IP3)-sensitive calcium stores or the responsivity of IP3-associated calcium channels was assessed by two experiments: 1) prior administration of GTP at concentrations < or = 100 microM had no effect on IP3-induced calcium release, and 2) heparin, which competitively inhibits IP3 binding to its receptor on the endoplasmic reticulum, did not affect GTP-associated calcium increase. These results demonstrate that, in gastric smooth muscle, GTP causes calcium efflux from an intracellular pool that is functionally independent from that pool sensitive to IP3.(ABSTRACT TRUNCATED AT 250 WORDS)

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