M. Saotome

The effects of calcitonin gene‐related peptide on tracheal smooth muscle of guinea‐pigs in vitro

1 The effect of calcitonin gene‐related peptide (CGRP) on airway smooth muscle is controversial. The aim of this study was to determine whether the action of CGRP on tracheal strips of guinea‐pigs is modulated by epithelium and whether this peptide‐induced action involves other mediators including nitric oxide (NO) and endothelin (ET)‐1. 2 CGRP produced a weak dose‐dependent increase in guinea‐pig tracheal tension in vitro (‐log EC50 = 8.5 ± 0.1, maximum contraction = 8.3 ± 1.2% of 50 mM KCl‐induced contraction, n = 6). In epithelium‐depleted preparations, CGRP (10−7 m)‐induced contraction was significantly potentiated from 9.0 ± 1.9% to 41.1 ± 6.0% (n = 6). 3 L‐NG‐nitro‐arginine methyl ester (L‐NAME, 10−4 m), which inhibits NO synthesis, enhanced the contractile response to CGRP from 9.0 ± 1.9% to 31.2 ± 1.1% (n = 6). Indomethacin (10−5 m) also enhanced the response to CGRP, although the effect was weak (13.4 ± 3.2%, n = 6). 4 Anti‐ET‐1 serum changed the CGRP‐induced contraction into a relaxation. After incubation of the trachea with ET‐1 (10−7 m) to attenuate ET‐1‐induced responses, the CGRP‐induced contraction also changed into a relaxation. BQ‐123 (an ETA receptor antagonist) and BQ‐788 (an ETB receptor antagonist) caused the same conversion of the CGRP response, from contraction to relaxation, although the relaxing effect elicited by BQ‐788 was more potent than that by BQ‐123. Maximum inhibitory responses were −31.0 ± 3.3% and −13.0 ± 2.3% of 50 mM KCl‐induced contraction, respectively (n = 6). 5 In primary culture, guinea‐pig tracheal epithelial cells released ET‐1, and CGRP (10−5 m) significantly increased the release of ET‐1. 6 These data suggest that the action of CGRP is modulated by airway epithelium and this mechanism involves the release of NO and ET‐1. Especially, the majority of contractile action elicited by CGRP consists of an action of ET‐1 via the predominant ETB receptor.

Endothelin-1-induced relaxation of guinea pig trachealis muscles.

Our accompanying paper demonstrated that endothelin-1 (ET-1 constricts guinea pig airways directly and indirectly through mediators such as histamine and arachidonate metabolites. In order to exclude the role of mast cells in bronchoconstriction, we sensitized guinea pigs and challenged them in vitro with an antigen (ovalbumin). In the postanaphylactic trachea, ET-1 caused a transient relaxation followed by constriction. Such relaxation by ET was also observed in the tracheas constricted with carbamylcholine. The relaxation was completely blocked by nordihydroguaretic acid and AA861, lipooxygenase inhibitors, but not by indomethacin, a cyclooxygenase inhibitor, and FPL 55712, a leukotriene antagonist. Because the relaxation was not affected even in the presence of methylarginine, an inhibitor of NO synthesis, and superoxide dismutase, an enzyme for destroying NO radical, we concluded that ET-1 induces the relaxation of the tracheal muscles by producing lipooxygenase products, probably hydroperoxides of arachidonic acid.

Multiple mechanisms of bronchoconstrictive responses to endothelin-1

We investigated the mechanism of the endothelin-1 (ET-1)-induced bronchoconstriction of guinea pig tracheal smooth muscles. ET-1 contracted the tracheas in a dose-dependent manner. A combination of FPL55712 (leukotriene antagonist), diphenhydramine (histamine antagonist), and indomethacin (cyclooxygenase inhibitor) shifted the dose-response curve of ET-1 to the right and suppressed the maximal constriction. Azelastine, an antiallergic agent, exerted essentially similar results. The present data suggest that ET-1 constricts the airway smooth muscles not only by direct action on the tracheal smooth muscles but also by indirect action mediated through production of various chemical mediators in cells other than muscles.