Endothelin-1 (ET-1) contracted the rabbit tracheal smooth muscle (RTSM), yielding a bell-shaped tension-concentration curve. Moreover, ET-1 induced concentration- and time-dependent increases in cAMP concentrations in RTSM (EC50, 58 nM; t(1/2), 2.4 min). Pretreatment with the AC inhibitors, SQ-22536, or 2'-5'-dideoxyadenosine, enhanced contraction to ET-1 and converted its bell-shaped tension curve into a sigmoidal one, but left contraction to carbachol and KCI unaltered. The potent ETB-receptor agonists, ET-3 or sarafotoxin-c, mimicked ET-1's effects on cAMP levels (EC50 values 55 and 50 nM). Further, cAMP formation by ETs was inhibited by BQ-788 (selective ETB receptor blocker; IC50, 8 nM), but not by BQ-610 (selective ETA receptor blocker). Removal of the epithelium did not prevent ET-induced increases in cAMP levels. Unlike isoproterenol, ETs failed to activate AC in membrane fractions from RTSM. In intact RTSM, the c-PLA(2) inhibitor, AACOCF3, and the cyclooxygenase inhibitor, indomethacin, blocked ET-induced increases in cAMP levels. These findings reveal a novel, nonepithelial, c-PLA(2)-mediated, regulatory mechanism downstream from ETB receptors.