Effects of bradykinin and angiotensin II on intracellular Ca2+ dynamics in endothelial cells

Abstract

The purpose of this study was to investigate the effects of angiotensin II and bradykinin on intracellular Ca2+ dynamics in cultured endothelial cells. We used the "second-generation" fluorescent Ca2+ indicator fura-2, in conjunction with dual-wavelength fluorescence spectroscopy, in cultured adherent pulmonary arterial endothelial cells. Angiotensin II (up to 2 microM) had no consistent effect on intracellular Ca2+ levels. In contrast, bradykinin (10 nM) elicited a transient increase of cytosolic free Ca2+, from the resting value of 37 +/- 5 to 647 +/- 123 nM, followed by a decline to a steady-state value of 113 +/- 14 nM, which was significantly higher than the resting Ca2+ levels. Bradykinin's Ca-stimulatory effect was dose dependent, having a half-maximally effective concentration of approximately 1 nM and a maximally effective concentration of 10 nM. A B1-receptor agonist, Des-Arg9-bradykinin, was much less effective than bradykinin as modulator of cytosolic Ca2+. Moreover, a B1-receptor antagonist, Des-Arg9, [Leu8]-bradykinin, did not significantly affect the increase of cytosolic Ca2+ elicited by bradykinin. On the other hand, the bradykinin-elicited increase of Ca2+ was almost completely inhibited by a novel B2-receptor antagonist, D-Arg-[Hyp3, Thi5,8, D-Phe7]-bradykinin. Bradykinin increased cytosolic free Ca2+ levels in cells maintained in Ca2+-deficient extracellular medium, suggesting that the peptide mobilized Ca2+ from intracellular stores. However, the absence of extra-cellular Ca2+ resulted in an 80-90% attenuation of the transient Ca2+ response, whereas the posttransient steady-state response was completely absent. These findings are consistent with the notion that the bradykinin-elicited transient Ca2+ response is dependent on both extra- and intracellular Ca2+ and that the posttransient steady-state response is entirely dependent on extracellular Ca2+. Endothelial cells were responsive to a second dose of bradykinin after a 10-min interim period of incubation in the absence of the peptide hormone. The absence of extracellular Ca2+ during the interim period, or the pretreatment of cells with ionomycin in the absence of extracellular Ca2+, prevented the response of the cells to a second dose of bradykinin. Bradykinin- or ionomycin-desensitized cells could be resensitized by a brief incubation period in Ca2+-replete medium. The results are consistent with the notions that cellular resensitization requires the replenishment of intracellular Ca2+ and that bradykinin, but not angiotensin II, modulates intracellular Ca2+ dynamics in endothelial cells by interacting with a B2-type receptor.

Ca(2+)‐dependent non‐selective cation and potassium channels activated by bradykinin in pig coronary artery endothelial cells.

“…Due to the step concentration gradient across the cell membrane, even a low permeability for Cam+ could exert profound effects on cell function. Hoyer et al (1994) (von Tscharner et al 1986; Morgan-Boyd, Stewart, Vavrek & Hassid, 1987;Nilius, 1991). In these cells, the bradykinin-induced rise in cytosolic Ca2+ is reduced by 70-80% in the absence of extracellular Cam+ and markedly shortened (see also Buclhan & Martin, 1991;Sturek et al 1991 Some endothelial cation channels are more selective for Ca2+ than the Ca2P-dependent cation channel we described for the coronary artery.…”

Role of calreticulin in the sensitivity of myocardiac H9c2 cells to oxidative stress caused by hydrogen peroxide

“…[Ca 2ϩ ]i was measured using a dual-excitation wavelength spectrofluorophotometer (RF-5500; Shimadzu, Kyoto, Japan) with fura-2 essentially as described previously (6,36), but with a slight modification. Briefly, cultured cells on glass coverslips were loaded with 5 M fura-2 AM (Dojindo, Kumamoto, Japan) for 20 min in Earle's balanced salt solution (EBSS; in mM: 26 NaHCO 3, pH 7.4, 1 NaH2PO4, 5.4 KCl, 116 NaCl, 5.5 glucose, and 2 CaCl 2) in the presence of 0.01% Pluronic acid F-127.…”

Cyclic AMP selectively enhances bradykinin receptor synthesis and expression in cultured arterial smooth muscle. Inhibition of angiotensin II and vasopressin response.

“…The kinins which are generated can bind to bradykinin recep-tors which have been found on both endothelial cells as well as on cultured vascular smooth muscle cells (4)(5)(6). Kinin receptors in both cell types are coupled to activation ofa phosphoinositide-specific phospholipase C which in endothelial cells leads to increases in intracellular calcium and the release of nitrosovasodilators and prostaglandins (4,6,7). The tissue level ofkinins are further controlled by proteases such as kininase II (angiotensin converting enzyme) and other kininases which are present on endothelial cells and vascular smooth muscle cells (8).…”