The presence and the possible mechanism of action of the inhibitory nonadrenergic, noncholinergic nerve system (i-NANC) were investigated in guinea pig pulmonary artery (PA) precontracted with U44069 (a thromboxane analog). In the presence of alpha adrenergic blockage, electrical field stimulation induced a frequency-dependent, tetrodotoxin-sensitive relaxation. This relaxation was reduced by 9.1 +/- 1.9 and 19.4 +/- 2.8% by atropine (1 microM) and combined atropine and propranolol (both 1 microM), indicating that the main component is mediated by i-NANC neural mechanisms. In the branch PA rings, this i-NANC relaxation was unaffected by pretreatment with a cyclooxygenase inhibitor (indomethacin, 10 microM), 5-lipoxygenase inhibitor (A63162, 1 microM) or substance P desensitization, but was inhibited markedly by the P2y-purinoceptor antagonist reactive blue 2 (30 microM) and slightly potentiated by the peptidase alpha-chymotrypsin (2 U/ml). L-NG-monomethyl-arginine(L-NMMA), a nitric oxide synthesis inhibitor, caused a concentration-dependent inhibition of the i-NANC relaxation (53.9 +/- 4.1% at 100 microM), but had no effect on equivalent nitroprusside-induced relaxation. The inhibitory effect of L-NMMA was reversed completely by L-arginine (300 microM), but not by D-arginine (300 microM). Removal of vascular endothelium greatly reduced the i-NANC relaxation in the branch PA rings, but had no effect on i-NANC relaxation in main PA rings. Both in vivo capsaicinization and in vitro desensitization with capsaicin (1 microM) caused a significant reduction of the i-NANC relaxation in main PA, but had no significant effect in the branch PA.(ABSTRACT TRUNCATED AT 250 WORDS)