Detection of Nitric Oxide Release Induced by Bradykinin in Guinea Pig Trachea and Main Bronchi Using a Porphyrinic Microsensor

Abstract

Indirect evidence using nitric oxide (NO) synthase (NOS) inhibitors suggests that in guinea-pig airways bradykinin releases bronchoprotective NO. In this study, using a recently developed electrochemical method of NO measurement based on a porphyrinic microsensor, we investigated whether bradykinin releases NO from guinea-pig airways and whether the epithelium is the main source of NO. Further, the Ca(2+)-dependence of bradykinin-induced NO release was assessed stimulating airway preparations with bradykinin in Ca(2+)-free conditions. We also studied the immunohistochemical distribution of the Ca(2+)- dependent constitutive isoforms of NOS (constitutive NOS [cNOS]: neuronal and endothelial [ecNOS]) in our preparations. The porphyrinic microsensor was placed in the bathing fluid onto the mucosal surface of tracheal or main bronchial segments. Addition of bradykinin vehicle (0.9% saline) did not cause any detectable change of the baseline signal. Addition of bradykinin caused an upward shift of the baseline that reached a maximum within 1 to 2 s. The amplitude of the response to bradykinin was concentration-dependent between the range 1 nM to 10 microM, with a maximum effect at 10 microM. Bradykinin-induced NO release was higher in tracheal than in main bronchial segments. The selective bradykinin B(2) receptor antagonist D-Arg(0)-[Hyp(3), Thi(5), D-Tic(7), Oic(8)]bradykinin (1 microM) inhibited NO release induced by a submaximum concentration of bradykinin (1 microM). The ability of bradykinin to release NO was markedly reduced in epithelium-denuded segments, and abolished in Ca(2+)-free conditions and after pretreatment with N(G)-monomethyl-L-arginine (100 microM), but not with N(G)-monomethyl-D-arginine. Both cNOS isoforms were present in trachea and main bronchi, ecNOS being the predominant isoform in the epithelium. The study shows that bradykinin via B(2) receptor activation caused a rapid and Ca(2+)-dependent release of NO, mainly, but not exclusively, derived from the epithelium. It also shows that both cNOS isoforms may be involved in bradykinin-evoked NO release.

The effect of overexpression of endothelial nitric oxide synthase on eosinophilic lung inflammation in a murine model

“…Neurally mediated bronchoconstriction by bradykinin is limited by several epithelium-derived factors, including peptidases [36], prostaglandin E2 [37], and an especially important factor, NO [38,39]. In addition, eNOSdeficient mice are markedly hyperresponsive to inhaled methacholine [7].…”

Expression of vascular remodelling markers in relation to bradykinin receptors in asthma and COPD

“…The B2R is constitutively expressed on most cell types and its activation leads to various intracellular events, including nitric oxide release 15. By contrast, B1Rs can be induced during inflammatory insults 14.…”

Acid-induced modulation of airway basal tone and contractility: Role of acid-sensing ion channels (ASICs) and TRPV1 receptor

“…In addition, mRNA of ASIC1a has been evidenced in human isolated skeletal muscle cells (Gitterman et al, 2005). In the absence of epithelium, a residual NO production (40%) persists in the guinea pig airways probably via sensory nerves, airway smooth muscle or vascular endothelium (Ricciardolo et al, 2000). Our results plead for the involvement of airway smooth muscle in the acid-stimulated NO release but the contribution of endothelium in this mechanism is thinkable.…”

“…In rodents, inhalation or instillation of relatively strong acids (pKa b 5) stimulate sensory nerves and the release of tachykinins, mediating bronchoconstriction (Ricciardolo et al, 2000;Lai and Huang, 2002;Morris et al, 2003). We used in our experiments a lesser acid stimulation (pH 6.8) in order to approach the physiologic conditions observed in asthma or in critically ill patients (Hunt et al, 2000;Wemmie et al, 2006;Diochot et al, 2007).…”