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 mu M carbachol). EFS-induced contractions were abolished by either 0.1 mu M atropine or 3 mu M tetradotoxin, but were not affected by 1 mu M 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 mu M 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 mu M 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.
