Inhibition by endothelin-1 of cholinergic nerve-mediated acetylcholine release and contraction in sheep isolated trachea

1 The relative roles of ET(A) and ET(B) receptor activation on cholinergic nerve-mediated contraction and acetylcholine (ACh) release were examined in sheep isolated tracheal smooth muscle. 2 Electrical field stimulation (EFS; 90 V, 0.5 ms duration, 1 Hz, 10 s train) applied to sheep isolated tracheal smooth muscle strips induced monophasic contractile responses that were abolished by either 1 mu M tetrodotoxin or 0.1 mu M atropine, but were insensitive to 10 mu M hexamethonium and 100 mu M L-NAME. Thus, EFS-induced contractions resulted from the spasmogenic actions of ACh released from parasympathetic, postganglionic nerves. 3 As expected, sheep isolated tracheal smooth muscle preparations did not contract in response to the ET(B) receptor-selective agonist, sarafotoxin S6c (0.1-100 nM). However, sarafotoxin S6c caused a concentration-dependent and transient inhibition of EFS-induced contractions. The inhibitory effect induced by a maximally effective concentration of sarafotoxin S6c (10 nM; 72.1 +/- 5.7%, n=6) was abolished in the presence of the ET(B) receptor-selective antagonist BQ-788 (1 mu M). Contractile responses to exogenously administered ACh (10 nM-0.3 mM) were not inhibited by sarafotoxin S6c (1 or 10 nM; n = 7). 4 In contrast to sarafotoxin S6c, endothelin-1 induced marked contractions in sheep isolated tracheal smooth muscle. These contractions were inhibited by BQ-123, consistent with an ET(A) receptor-mediated response. In the presence of BQ-123 (3 mu M), endothelin-1 produced a concentration-dependent inhibition of EFS-induced contractions (30 nM endothelin-1, 68.9 +/- 10.2% inhibition, n = 5). These responses were inhibited by 1 mu M BQ-788, indicative of an ET(B) receptor-mediated process. Endothelin-1 was about 3 fold less potent than sarafotoxin S6c. 5 EFS (90 V, 0.5 ms duration, 1 Hz, 15 min train) induced the release of endogenous ACh (1.94 +/- 0.28 pmol. mg(-1) tissue, n = 12), as assayed by h.p.l.c. with electrochemical detection. EFS-induced release of ACh was inhibited to a similar extent by 100 nM endothelin-1 (47 +/- 4%, n = 9) and 10 nM sarafotoxin S6c (46 +/- 9%, n = 3). These effects of endothelin-1 on ACh release were inhibited by 1 mu M BQ-788 alone (n = 4), by BQ-788 in the presence of 3 mu M BQ-123 (n = 4), but not by 3 mu M BQ-123 alone (n = 5). 6 In summary, sheep isolated tracheal smooth muscle contains two anatomically and functionally distinct endothelin receptor populations. ET(A) receptors located on airway smooth muscle mediate contraction, whereas ET(B) receptors appear to exist on cholinergic nerves that innervate tracheal smooth muscle cells and mediate inhibition of ACh release. The inhibitory effect of ET(B) receptor stimulation on cholinergic neurotransmission is in stark contrast to the enhancing effects hitherto described in the airways.