Perkins WJ, Warner DO, Jones KA. Prolonged treatment of porcine pulmonary artery with nitric oxide decreases cGMP sensitivity and cGMP-dependent protein kinase specific activity. Am J Physiol Lung Cell Mol Physiol 296: L121-L129, 2009. First published October 24, 2008 doi:10.1152/ajplung.90318.2008.-A cultured porcine pulmonary artery (PA) model was used to examine the effects of prolonged nitric oxide (NO) treatment on the response to acutely applied NO, cGMP analog, or atrial natriuretic peptide (ANP). Twenty-fourhour treatment with the NO donor (Z)-1-[N-(2-aminoethyl)-N-(2-ammonioethyl)amino]diazen-1-ium-1,2-diolate (DETA-NO) resulted in Ͼ10-fold decrease in the response to acutely applied DETA-NO. In parallel with this, the relaxant response to acutely applied cGMP analog, -phenyl-1,N 2 -etheno-8-bromoguanosine-3Ј,5Ј-cyclic monophosphorothioate, Sp isomer (Sp-8-Br-PET-cGMPS), and ANP decreased. The reduction in ANP responsiveness in PA was not associated with a reduction in cGMP levels evoked by 10 Ϫ6 M ANP. Twenty-four hours in culture and treatment with DETA-NO decreased total cGMP-dependent protein kinase (cGKI) mRNA level compared with that in freshly prepared PA (1.05 Ϯ 0.12, 0.42 Ϯ 0.08, and 0.11 Ϯ 0.01 amol/g, respectively). Total cGKI protein levels were decreased to a lesser extent by 24 h in culture and further decreased by 24-h DETA-NO treatment compared with that in freshly prepared PA (361 Ϯ 33, 272 Ϯ 20, and 238 Ϯ 25 ng/mg total protein, respectively). Maximal cGMP-stimulated phosphotransferase activity was reduced in 24-h cultured and DETA-NO-treated PA (986 Ϯ 84, 815 Ϯ 81, and 549 Ϯ 78 pmol Pi ⅐ min Ϫ1 ⅐ mg soluble protein Ϫ1 ), but the cGMP concentration resulting in 50% of maximal phosphotransferase activity was not. cGKI specific activity (maximal cGMPactivated phosphotransferase activity/ng cGKI) was significantly reduced in PA treated with DETA-NO for 24 h compared with freshly prepared and 24-h cultured PA (1.95 Ϯ 0.22, 2.64 Ϯ 0.25, and 2.85 Ϯ 0.28 pmol Pi ⅐ min Ϫ1 ⅐ ng cGKI Ϫ1 , respectively). We conclude that prolonged NO treatment induces decreased acute NO responsiveness in PA in part by decreasing cGMP sensitivity. It does so by decreasing both cGKI expression and cGKI specific activity. guanosine 3Ј,5Ј-cyclic monophosphate; guanosine 3Ј,5Ј-cyclic monophosphate-dependent protein kinase THE NITRIC OXIDE (NO) signaling system plays a significant role in the regulation of vascular smooth muscle tone and, thus, of systemic hemodynamics. Multiple factors regulate this system, including NO production and bioavailability, cGMP production by soluble guanylyl cyclase (sGC), cGMP breakdown by phosphodiesterases (PDE), and transduction of the cGMP signal by smooth muscle cGMP-dependent protein kinases (cGKI) and their associated protein targets involved in decreasing myoplasmic calcium (12,19,28) and myofilament calcium sensitivity (44). Acute, short-lived increases in NO transiently activate this system, resulting in short-term vascular smooth muscle relaxation (21). The effects of prolong...