Potentiation by endothelin‐1 of cholinergic nerve‐mediated contractions in mouse trachea via activation of ETB receptors

Abstract

1 We have previously shown that endothelin-1-induced contraction of mouse isolated tracheal smooth muscle was mediated via both ET_A and ET_B receptors. In the current study, we have investigated endothelin-1-induced potentiation of cholinergic nerve-mediated contractions in mouse isolated trachea and have characterized pharmacologically the endothelin receptors mediating this response. 2 Electrical field stimulation (EFS; 70 V, 0.5 ms duration, 10 s train, 0.1–60 Hz) of mouse isolated trachea caused frequency-dependent, monophasic contractions (magnitude of contraction of 60 Hz was 56 ± 4%C_max (n = 6), where C_max is the contractile response to 10 μm carbachol). EFS-induced contractions were abolished by either 0.1 μm atropine or 3 μm tetradotoxin, but were not affected by 1 μm hexamethonium, indicating that they were induced by stimulation of postganglionic cholinergic nerves. In contrast, contractions induced by exogenously applied acetylcholine were inhibited by atropine, but not by either tetrodotoxin or hexamethonium. 3 The ET_B receptor-selective agonist, sarafotoxin S6c, caused marked concentration-dependent potentiation of EFS-induced contractions in mouse isolated tracheal segments. At 0.1 nm, sarafotoxin S6c exerted no direct contractile effect, but significantly increased a standard EFS-induced contraction of 20%C_max by 8 ± 2%C_max (i.e. 1.4 fold, n = 5, P < 0.05). At higher concentrations, 10 nm sarafotoxin S6c induced a large, transient contraction (peak response of 74 ± 2%C_max at 10 min; 3 ± 2%C_max at 45 min) and enhanced the standard EFS-induced contraction by 30 ± 4%C_max (i.e. 2.5 fold, n = 5, P < 0.01). In contrast, 10 nm sarafotoxin S6c did not enhance contractile responses to exogenously applied acetylcholine (n = 6). 4 Endothelin-1 also modulated EFS-induced contractions. At 0.1 nm, endothelin-1 exerted no direct contractile effect, but significantly increased the standard EFS-induced contraction of 20%C_max by 7 ± 2%C_max (i.e. 1.35 fold, n = 5, P < 0.05). At 1 nm, endothelin-1 induced a small, sustained contraction (16 ± 3% C_max) and increased the standard EFS-induced contraction by 19 ± 2%C_max (i.e. 1.95 fold, n = 5, P < 0.01). Finally, 10 nm endothelin-1 induced a large, sustained contraction (98 ± 8%C_max), but the EFS-induced contraction was significantly reduced from 20%C_max to 6 ± 4%C_max (n = 6, P < 0.05). In contrast, in the presence of 3 μm BQ-123 (ET_A receptor-selective antagonist), 10 nm endothelin-1 induced a transient contraction mediated via ET_B receptors (peak response of 59 ± 10%C_max at 10 min; 8 ± 2%C_max at 45 min). Under these conditions, the standard EFS-induced contraction was increased by 26 ± 1%C_max (i.e. 2.3 fold, n = 6, P < 0.01). 5 The potentiation of EFS-induced contractions produced by 1 nm endothelin-1 was not mediated by ET_A receptors, since 3 μm BQ-123 did not diminish this effect (n = 6). Furthermore, 1 nm endothelin-1 did not potentiate EFS-induced contractions in preparations in which the function of the ET_B receptor-effector system had been attenuated by desensitization (n = 6). 6 In summary, endothelin-1 potentiates cholinergic nerve-mediated contractions in mouse isolated trachea, apparently by activating prejunctional ET_B receptors. This neuronal pathway offers an additional mechanism through which endothelin-1 may elevate bronchomotor tone.