We aimed to assess intrinsic smooth muscle mechanisms contributing to greater nitric oxide (NO) responsiveness in pulmonary vascular vs. airway smooth muscle. Porcine pulmonary artery smooth muscle (PASM) and tracheal smooth muscle (TSM) strips were used in concentration-response studies to the NO donor (Z)-1-[N-2-aminoethyl-N-(2-ammonioethyl)amino]diazen-1-ium-1,2-diolate (DETA-NO). PASM consistently exhibited greater relaxation at a given DETA-NO concentration (NO responsiveness) than TSM NO responsiveness, with DETA-NO log EC50 being -6.55 ± 0.11 and -5.37 ± 0.13 for PASM and TSM, respectively (P < 0.01). We determined relationships between tissue cGMP concentration ([cGMP]i) and relaxation using the particulate guanylyl cyclase agonist atrial natriuretic peptide. Atrial natriuretic peptide resulted in nearly complete relaxation, with no detectable increase in [cGMP]i in PASM and only 20% relaxation (10-fold increase in [cGMP]i) in TSM, indicating that TSM is less cGMP responsive than PASM. Total cGMP-dependent protein kinase I (cGKI) mRNA expression was greater in PASM than in TSM (2.23 ± 0.36 vs. 0.93 ± 0.31 amol mRNA/μg total RNA, respectively; P < 0.01), but total cGKI protein expression was not significantly different (0.56 ± 0.07 and 0.49 ± 0.04 ng cGKI/μg protein, respectively). The phosphotransferase assay for the soluble fraction of tissue homogenates demonstrated no difference in the cGMP EC 50 between PASM and TSM. The maximal phosphotransferase activityindexed to the amount of total cGKI in the homogenate differed significantly between PASM and TSM (1.61 ± 0.15 and 1.04 ± pmol·min-1·ng cGKI-1, respectively; P < 0.05), suggesting that cGKI may be regulated differently in the two tissues. A novel intrinsic smooth muscle mechanism accounting for greater NO responsiveness in PASM vs. TSM is thus greater cGMP responsiveness from increased cGKI-specific activity in PASM. Copyright © 2006 the American Physiological Society.