Relaxation of Endothelin-1-Induced Pulmonary Arterial Constriction by Niflumic Acid and NPPB: Mechanism(s) Independent of Chloride Channel Block1

Abstract

We investigated the effects of the Cl⁻ channel blockers niflumic acid, 5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB) and 4,4′-diisothiocyanatostilbene-2,2′-disulphonic acid (DIDS) on endothelin-1 (ET-1)-induced constriction of rat small pulmonary arteries (diameter 100–400 μm) in vitro, following endothelium removal. ET-1 (30 nM) induced a sustained constriction of rat pulmonary arteries in physiological salt solution. Arteries preconstricted with ET-1 were relaxed by niflumic acid (IC₅₀: 35.8 μM) and NPPB (IC₅₀: 21.1 μM) in a reversible and concentration-dependent manner. However, at concentrations known to block Ca⁺⁺-activated Cl⁻ channels, DIDS (≤500 μM) had no effect on the ET-1-induced constriction. Similar results were obtained when pulmonary arteries were preincubated with these Cl⁻ channel blockers. When l-type Ca⁺⁺ channels were blocked by nifedipine (10 μM), the ET-1-induced (30 nM) constriction was inhibited by only 5.8%. However, niflumic acid (30 μM) and NPPB (30 μM) inhibited the ET-1-induced constriction by ∼53% and ∼60%, respectively, both in the continued presence of nifedipine and in Ca⁺⁺-free physiological salt solution. The Ca⁺⁺ ionophore A23187 (10 μM) also evoked a sustained constriction of pulmonary arteries. Surprisingly, the A23187-induced constriction was also inhibited in a reversible and concentration-dependent manner by niflumic acid (IC₅₀: 18.0 μM) and NPPB (IC₅₀: 8.8 μM), but not by DIDS (≤500 μM). Our data suggest that the primary mechanism by which niflumic acid and NPPB inhibit pulmonary artery constriction is independent of Cl⁻ channel blockade. One possibility is that these compounds may block the Ca⁺⁺-dependent contractile processes.