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.

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 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 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 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 Oxidation-reduction properties of the cytochrome b found in the plasma-membrane fraction of human neutrophils. A possible oxidase in the respiratory burst

1981 ◽  
Vol 194 (2) ◽  
pp. 599-606 ◽  
Author(s):  
A R Cross ◽  
O T Jones ◽  
A M Harper ◽  
A W Segal
Keyword(s):  
Plasma Membrane ◽  
Cytochrome B ◽  
Plasma Membranes ◽  
Mixing Time ◽  
Human Neutrophils ◽  
Half Time ◽  
Sulphur Compounds ◽  
Midpoint Potential ◽  
Oxidation Reduction ◽  
Membrane Bound

The oxidation-reduction midpoint potential of the cytochrome b found in the plasma membrane of human neutrophils has been determined at pH 7.0 (Em,7.0) from measurements of absorption spectra at fixed potentials. In both unstimulated and phorbol myristate acetate-stimulated cells Em,7.0 was -245 mV. Changes in pH affected the Em of the cytochrome b, with a slope of approx. 25 mV/pH unit change. The Em,7.0 of the haem group(s) of the membrane-bound myeloperoxidase of human neutrophils was found to be +34 mV. The plasma membranes contained no detectable ubiquinone, and no iron-sulphur compounds were detected by e.p.r. spectroscopy at 5-20 K. No flavins were detected by e.p.r. spectroscopy. The cytochrome b-245 was not reduced by added NADH or NADPH. Dithionite-reduced cytochrome b-245 formed a complex with CO, supplied as a saturated solution, which was dissociated with 26 microseconds illumination from a xenon flash lamp, and the recombination with CO had a half-time of approx. 6 ms. Partly (80%) reduced cytochrome b-245 was oxidized by added air-saturated buffer with a half-time faster than 1 s at 20 degrees C, a resolution limited by mixing time. These results are compatible with cytochrome b-245 acting as an oxidase.

scholarly journals Pharmacology of a Ca2+-influx pathway activated by emptying the intracellular Ca2+ stores in HL-60 cells: evidence that a cytochrome P-450 is not involved

1994 ◽  
Vol 302 (1) ◽  
pp. 187-190 ◽  
Author(s):  
B D Koch ◽  
G F Faurot ◽  
M V Kopanitsa ◽  
D C Swinney
Keyword(s):  
Plasma Membrane ◽  
Selective Inhibitors ◽  
High Affinity ◽  
Voltage Dependent ◽  
Influx Pathways ◽  
Subsequent Activation ◽  
Cytochrome P 450 ◽  
Haem Iron ◽  
Dependent Pathway ◽  
Ca2 Channels

In HL-60 cells, inhibition of the endoplasmic-reticular Ca2+ pump by thapsigargin leads to the emptying of this intracellular Ca2+ store and a subsequent activation of plasma-membrane Ca2+ influx through a non-voltage-dependent pathway. The elevated intracellular free Ca2+ concentration ([Ca2+]i) produced and maintained by this Ca2+ inflow was used to examine the potency of various compounds to inhibit this influx mechanism. As expected, specific blockers of known Ca2+ channels, such as nifedipine, omega-conotoxin GVIA and ryanodine were without effect. The less selective inhibitors La3+, SKF-96365 and L-651,582, which are thought to inhibit both voltage-dependent and voltage-independent Ca2+ channels, decreased [Ca2+]i back to resting levels, with pIC50 values of 5.2, 5.9 and 6.2 respectively. It has been proposed that a cytochrome P-450 is involved in activating Ca(2+)-influx pathways in thymocytes, neutrophils and platelets. Consistent with this idea, the imidazole cytochrome P-450 inhibitors miconazole, econazole, clotrimazole and ketoconazole inhibited the thapsigargin-elevated [Ca2+]i with pIC50 values of 7.1, 7.1, 7.1 and 5.8 respectively. The high affinity of imidazoles for cytochromes P-450 is due to co-ordinate binding to the haem. This interaction is greatly decreased in 2-substituted imidazoles. We examined whether the inhibition of Ca2+ influx was due to an interaction of the inhibitor imidazole nitrogen with the haem iron of the putative cytochrome P-450 by comparing the activity of two compounds, identical except that one was methylated at the imidazole 2-position. They were found to block thapsigargin-activated Ca2+ influx with equal potency. These results strongly suggest that a cytochrome P-450 is not involved in the activation of the Ca2+ influx produced by emptying the intracellular Ca2+ stores.

scholarly journals A Ca2+ influx associated with exocytosis is specifically abolished in a Paramecium exocytotic mutant.

1990 ◽  
Vol 111 (6) ◽  
pp. 2527-2535 ◽  
Author(s):  
D Kerboeuf ◽  
J Cohen
Keyword(s):  
Plasma Membrane ◽  
Membrane Fusion ◽  
Membrane Permeability ◽  
Gene Product ◽  
Transport Mechanism ◽  
External Stimulus ◽  
Plasma Membrane Permeability ◽  
45Ca Uptake ◽  
Per Se ◽  
Ca2 Channels

A Paramecium possesses secretory organelles called trichocysts which are docked beneath the plasma membrane awaiting an external stimulus that triggers their exocytosis. Membrane fusion is the sole event provoked by the stimulation and can therefore be studied per se. Using 3 microM aminoethyl dextran (AED; Plattner, H., H. Matt, H.Kersken, B. Haake, and R. Stürz, 1984. Exp. Cell Res. 151:6-13) as a vital secretagogue, we analyzed the movements of calcium (Ca2+) during the discharge of trichocysts. We showed that (a) external Ca2+, at least at 3 X 10(-7) M, is necessary for AED to induce exocytosis; (b) a dramatic and transient influx of Ca2+ as measured from 45Ca uptake is induced by AED; (c) this influx is independent of the well-characterized voltage-operated Ca2+ channels of the ciliary membranes since it persists in a mutant devoid of these channels; and (d) this influx is specifically abolished in one of the mutants unable to undergo exocytosis, nd12. We propose that the Ca2+ influx induced by AED reflects an increase in membrane permeability through the opening of novel Ca2+ channel or the activation of other Ca2+ transport mechanism in the plasma membrane. The resulting rise in cytosolic Ca2+ concentration would in turn induce membrane fusion. The mutation nd12 would affect a gene product involved in the control of plasma membrane permeability to Ca2+, specifically related to membrane fusion.

Refilling the inositol 1,4,5-trisphosphate-sensitive Ca2+ store in neuroblastoma x glioma hybrid NG108-15 cells

1993 ◽  
Vol 264 (3) ◽  
pp. C641-C653 ◽  
Author(s):  
T. M. Lo ◽  
S. A. Thayer
Keyword(s):  
Plasma Membrane ◽  
Inhibitory Concentration ◽  
Effective Concentration ◽  
Half Time ◽  
Dual Emission ◽  
Spectral Shifts ◽  
Inositol 1,4,5 Trisphosphate ◽  
The Status ◽  
Ca2 Channels

Bradykinin-induced increases in the intracellular free Ca2+ concentration ([Ca2+]i) were recorded in single NG108-15 cells with indo-1-based dual-emission microfluorimetry (50% effective concentration, 16 nM). A 1-min exposure to 30 nM bradykinin completely depleted the inositol 1,4,5-trisphosphate (IP3)-sensitive Ca2+ store; refilling the store required extracellular Ca2+ (half time, 2 min). Refilling the IP3-sensitive store was completely blocked by 1 microM La3+ and 10 microM nitrendipine, but not 10 microM verapamil, 10 microM flunarizine, 1 microM nitrendipine, or 0.1 microM La3+. Thapsigargin irreversibly depleted the Ca2+ store and prevented its refilling (half-maximal inhibitory concentration, 3 nM). Influx of Ca2+ across the plasma membrane did not increase after depletion of the IP3-sensitive store by exposure to bradykinin, although maintained presence of the agonist produced significant Ca2+ influx. Similarly, Mn2+ and Ba2+ influx, as measured by indo-1 quenching and spectral shifts, did not increase following depletion of IP3-sensitive store. In contrast to depletion of the IP3-sensitive Ca2+ store by bradykinin, thapsigargin (10 nM) treatment produced Ca2+ and Ba2+ influx. We conclude that after Ca2+ mobilization, the IP3-sensitive Ca2+ store in NG108-15 cells is refilled with cytoplasmic Ca2+ via a thapsigargin-sensitive Ca(2+)-Mg(2+)-ATPase. Cytoplasmic Ca2+ is replenished by a persistent leak of Ca2+ across the plasma membrane. This leak is not modulated by the status of the intracellular Ca2+ store. In NG108-15 cells, agonist and thapsigargin-evoked Ca2+ entry are mediated by activation of plasmalemmal Ca2+ channels independent of the status of the IP3-sensitive intracellular Ca2+ store.

Synthetic marine guanidines are effective antileishmanial compounds by altering the plasma membrane permeability

Planta Medica ◽  
2014 ◽  
Vol 80 (16) ◽  
Author(s):  
AG Tempone ◽  
LF Martins ◽  
EG Pinto ◽  
JT Mesquita ◽  
EL Bennett ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Membrane Permeability ◽  
Plasma Membrane Permeability
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