Characterization of the Na/HCO3(-) cotransport in human neutrophils.

Background: Bicarbonate transport has crucial roles in regulating intracellular pH (pHi) in a variety of cells. The purpose of this study was to evaluate its participation in the regulation of pHi in resting and stimulated human neutrophils.

Methods: Freshly isolated human neutrophils acidified by an ammonium prepulse were used in this study.

Cl⁻/HCO₃⁻ exchange activity in fMLP-stimulated human neutrophils.

It is well known that chemotactic agents active Na(+)/H(+) exchanger, increasing intracellular pH of neutrophils, but their effect on bicarbonate transporters have not been established yet. To study the effect of fMLP on the activity of Cl(-)/HCO(3)(-) exchange, the rate of pH recovery after acute Cl(-) readmission in cell subjected to an alkaline load by CO(2) washout in a Cl-free medium was measured. The activity of the exchanger was reduced to 72% of control when cells were pre-incubated for 5 min with 0.

Effect of glycine on the release of reactive oxygen species in human neutrophils.

The increase of extracellular glycine concentration prevents or mitigates a variety of pathological dysfunctional inflammatory responses. To eliminate the systemic effects of glycine as the reduction in the release of cytokines, this study was performed in isolated human neutrophils. The increase of the intracellular calcium concentration ([Ca2+](i)) and reactive oxygen species (ROS) release in cells incubated with glycine (0.

Inhibition of cytochalasin-primed neutrophils by hyperosmolarity.

Experimental and clinical investigations using hyperosmotic solutions for resuscitation of hemorrhagic shock demonstrated modulation of the inflammatory response. Decreased postinjury hyperinflammation has been attributed to a reduction in neutrophil-mediated tissue damage. This study shows that cytoskeletal disruption with cytochalasinB did not reverse or prevent the inhibitory effect of an osmolarity increase on the neutrophil cytotoxic response to a formyl peptide.

Hydrogen peroxide activates calcium influx in human neutrophils.

The correlation between an increased production of reactive oxygen species (ROS) and an enhanced calcium entry in primed neutrophils stimulated with fMLP suggests that endogenous ROS could serve as an agonist to reinforce calcium signaling by positive feedback. This work shows that exogenous H2O2 produced a rapid influx of Mn2+ and an increase of intracellular calcium. The H2O2 was insufficient to produce significant changes in the absence of extracellular calcium but addition of Ca2+ to H2O2-treated cells suspended in a free Ca2+/EGTA buffer resulted in a great increase in [Ca2+]i reflecting influx of Ca2+ across the cell membrane.

Effect of glycine on the calcium signal of thrombin-stimulated platelets.

In treatment of hemorrhagic shock, small-volume infusion of 7.5% NaCl gives immediate hemodynamic improvement, but in vitro experiments suggest it depresses the hemostatic system. Since previous reports showed that hyperosmotic glycine solutions preserved the platelet function better than hyperosmotic NaCl solutions, we investigated whether glycine changes the intracellular calcium ([Ca]i) signal.

Multiple alterations in Ca2+ handling determine the negative staircase in a cellular heart failure model.

Background: The flat or negative force frequency relationship (FFR) is a hallmark of the failing heart. Either decreases in SERCA2a expression, increases in Na(+)/Ca(2+) exchanger (NCX) expression or elevated Na(+)(i) have been independently proposed as mediators of the negative FFR.

Methods And Results: To determine whether each one of these mechanisms is sufficient to account for the negative FFR of the failing heart or on the contrary, various mechanisms, acting in concert are required.

Selective inhibition of calcium influx by 2-aminoethoxydiphenyl borate.

2-aminoethoxydiphenyl borate (2APB) blocks agonist-induced Ca2+ mobilization from intracellular stores and Ca2+ entry. To compare the sensitivity profiles of these two components of calcium signaling, human platelets were exposed to different concentrations of 2APB. The drug interferes with the Ca2+ mobilization from intracellular stores induced by thrombin with an IC50 of 52+/-2 microM but it has a more potent inhibitory effect on the Ca2+ entry (IC50=16+/-1 microM).

Functional interaction of carbonic anhydrase and chloride/bicarbonate exchange in human platelets.

Recently, our laboratory has reported the presence of one acidifying Cl-/HC exchange mechanism in human platelets. This paper demonstrates that this exchanger decreases its activity after inhibition of carbonic anhydrase. BCECF-loaded platelets, previously equilibrated in a bicarbonate/CO2 buffered solution, were resuspended in a Hepes-buffered, chloride-free (glucuronate) medium to produce a pHi increase.

Thrombin stimulation of Cl-/HCO3- exchange in human platelets.

The presence of one acidifying Cl-/HCO3- exchange mechanism in human platelets has not been previously reported. This paper demonstrates that this mechanism does function and that it increases its activity after stimulation with thrombin. On resuspension of BCECF-loaded platelets in a chloride-free medium (gluconate replaced) that contains bicarbonate, cytosolic pH (pHi) increased and stabilized after 10 min at an alkaline value.

Carbon monoxide inhibits capacitative calcium entry in human platelets.

The cytosolic calcium concentration in human platelets is elevated by several agonists via receptor-operated mechanisms involving both Ca(2+) release from intracellular stores and Ca(2+) entry. In order to get a mechanistic insight in the effect of carbon monoxide (CO)-containing solutions, this work examines the changes in [Ca(2+)](i) induced by 100 microM adenosine 5'diphosphate (ADP), 0.1 IU/ ml thrombin, 0.

Hypertonic saline solutions attenuate agonist-induced changes of intracellular calcium.

The proper fluid for resuscitation of hemorrhagic shock is still controversial. Hypertonic saline solutions would cause an impairment of platelet function, aggravating blood loss in case of uncontrolled hemorrhage. This work examines the in vitro effect of hypertonic NaCl solutions on the changes in [Ca2+]i induced by 100 microM ADP, 0.

Effect of hyperosmolarity on agonist-induced increases of intracellular calcium in human platelets.

Background: Hypertonic solutions are useful for the management of hypovolemic shock but cause impairment in platelet function. This effect would reduce the ischemia/reperfusion damage caused by activated platelets, but it could be the cause of aggravating blood loss in case of uncontrolled hemorrhage. In this paper, it was studied if osmotic shrinkage of platelets affects the changes in intracellular calcium concentration ([Ca(2+)](i)) induced by thrombin or adenosine 5' diphosphate (ADP).

Capacitative calcium influx and intracellular pH cross-talk in human platelets.

This study focuses on the potential interrelationships between changes in pH and capacitative calcium entry in stimulated platelets and on the participation of SOCs in the control of intracellular pH (pH(i)). Extracellular acidification reduces the Mn(2+) entry, measured by the slope of the quenching of FURA 2 fluorescence at the isoemissive wavelength of 360 nm. In thrombin-stimulated platelets Mn(2+) entry is reduced by acidosis (pH(o) = 6.