Effect of sildenafil on cyclic nucleotide phosphodiesterase activity, vascular tone and calcium signaling in rat pulmonary artery

Abstract

1 Sildenafil (viagra) is a potent PDE5 inhibitor and thus a relaxant drug in corpus carvernosum smooth muscle. In the present work, we evidenced the presence of PDE5 isozyme and investigated the effect of sildenafil on the specific cyclic nucleotide phosphodiesterase (PDE) activity, smooth muscle tone and calcium signaling in the rat main pulmonary artery (MPA). 2 The PDE activity was measured in cytosolic and microsomal fractions. Total cAMP and cGMP-PDE activities were mainly present in the cytosolic fraction. Sildenafil (0.1 mM) reduced by 72% cGMP-PDE activity, whereas zaprinast (10 mM), a relatively selective PDE5 inhibitor, reduced this activity by 63%. Sildenafil (0.1 mM) also inhibited significantly (22%) the cAMP-PDE activity. 3 Western blot analysis revealed the expression of PDE5 mainly in the cytosolic fraction of MPA. Sildenafil concentration-dependently inhibited (IC 50 ¼ 3.4 nM) the activity of MPA PDE5 partially purified by HPLC. 4 Sildenafil (0.1 nM-50 mM) concentration-dependently relaxed MPA rings precontracted with phenylephrine (0.5 mM). The potency of sildenafil (IC 50 ¼ 11 nM) was similar to that of a nitric oxide donor, sodium nitroprusside, but higher than that of zaprinast (IC 50 ¼ 600 nM). The vasorelaxant effect of sildenafil was not altered by endothelium removal or in the presence of KT 5823 (1 mM) and H89 (1 mM), potent inhibitors of PKG and PKA, respectively. 5 In isolated MPA myocytes, which had been loaded with the calcium fluorophore indo-1, sildenafil (10 -100 nM) antagonized ATP-and endothelin-1-induced calcium oscillations but had no effect on the transient caffeine-induced [Ca 2+ ] i response. 6 This study demonstrates the presence of a functional and highly sildenafil-sensitive PDE5 isozyme in rat MPA. Inhibition of this isozyme mainly accounts for the potent pulmonary vasodilator action of sildenafil, which involves alteration in the inositol triphosphate-mediated calcium signaling pathway.

Postnatal Maturation of Phosphodiesterase 5 (PDE5) in Piglet Pulmonary Arteries: Activity, Expression, Effects of PDE5 Inhibitors, and Role of the Nitric Oxide/Cyclic GMP Pathway

“…The PDE5 inhibitors zaprinast (0.1-10 M) and dipyridamole (0.1-10 M), which inhibit PDE1 at higher concentrations, were 100-to 300-fold less potent than sildenafil on vasorelaxation (Table 1), as described on PDE5 activity (31,32), and their effects were similar in both age groups. The effects of zaprinast were also unaffected by endothelium removal but inhibited by ODQ (not shown).…”

Effects of ω-hydroxylase product on distal human pulmonary arteries

“…Although arachidonic acid-induced relaxation in human pulmonary arteries (HPAs) involves cytochrome P-450-dependent metabolites and potassium channels (9), the precise molecular mode of action of 20-HETE remains to be ascertained. Since 20-HETE may be of putative clinical interest for PAH (11,24,25), we have focused the present study on the alternative mechanisms involved in the 20-HETE-induced relaxation in HPAs.The aim of the present work was to assess the physiological effects of 20-HETE on resistance vessels. Complementary approaches used include 1) tension measurements of relaxation on human arterial rings; 2) analyses of the effects of 20-HETE on the Ca 2ϩ sensitivity of the mechanical apparatus; and 3) evaluation of the status and expression of the PKC-potentiated inhibitory protein CPI-17 and p116 Rip proteins, which are involved in the control of the Ca 2ϩ sensitivity in vascular smooth muscle (VSM) cells (17,30).…”

“…In that respect, vasodilators have some beneficial effects on PAH. Among various existing pulmonary arterial vasodilators, such as NO, sildenafil, and cGMP (24,25), 20-hydroxyeicosatetraenoic acid (20-HETE), a cytochrome P-450 4A metabolite of arachidonic acid, consistently dilates pulmonary arteries in several species, including humans (2). 20-HETE induces a relaxation in bovine pulmonary arteries, whereas it induces a contraction in bovine renal arteries (12,15), suggesting a specific mode of action of 20-HETE according to vascular beds.…”

“…Although arachidonic acid-induced relaxation in human pulmonary arteries (HPAs) involves cytochrome P-450-dependent metabolites and potassium channels (9), the precise molecular mode of action of 20-HETE remains to be ascertained. Since 20-HETE may be of putative clinical interest for PAH (11,24,25), we have focused the present study on the alternative mechanisms involved in the 20-HETE-induced relaxation in HPAs.…”

Phosphodiesterase Enzymes and Inhibitors: Review

“…88 PDE5'in, sıçan pulmoner arterlerinde akciğer direncini kontrol ettiği ifade edilmiştir. 89 Son zamanlarda köpek, sıçan ve fareler üzerinde yapılan çalışmalarda, PDE5'in beta (β) reseptörleri aracılığı ile kardiyak yanıtları düzenlediği gösterilmiştir.…”