Multiple lines of evidence indicate that serotonin (5-hydroxytryptamine [5-HT]) and voltage-gated K ⁺ (K _V ) channels play a central role in the pathogenesis of pulmonary hypertension (PH). We hypothesized that 5-HT might modulate the activity of K _V channels, therefore establishing a link between these pathogenetic factors in PH. Here, we studied the effects of 5-HT on K _V channels present in rat pulmonary artery smooth muscle cells (PASMC) and on hK _V 1.5 channels stably expressed in Ltk ⁻ cells. 5-HT reduced native K _V and hK _V 1.5 currents, depolarized cell membrane, and caused a contraction of isolated pulmonary arteries. The effects of 5-HT on K _V currents and contraction were markedly prevented by the 5-HT _2A receptor antagonist ketanserin. Incubation with inhibitors of phospholipase C (U73122), classic protein kinase Cs (Gö6976), or tyrosine kinases (genistein and tyrphostin 23), the cholesterol depletion agent β-cyclodextrin or concanavalin A, an inhibitor of endocytotic processes, also prevented the effects of 5-HT. In homogenates from pulmonary arteries, 5-HT _2A receptors and caveolin-1 coimmunoprecipitated with K _V 1.5 channels, and this was increased on stimulation with 5-HT. Moreover, K _V 1.5 channels were internalized when cells were stimulated with 5-HT, and this was prevented by concanavalin A. These findings indicate that activation of 5-HT _2A receptors inhibits native K _V and hK _V 1.5 currents via phospholipase C, protein kinase C, tyrosine kinase, and a caveolae pathway. K _V channel inhibition accounts, at least partly, for 5-HT-induced pulmonary vasoconstriction and might play a role in PH.

The flavonoid quercetin is metabolized into isorhamnetin, tamarixetin, and kaempferol, the vascular effects of which are unknown. In the present study, the effects of quercetin and its metabolites were analyzed on isometric tension in isolated rat thoracic and abdominal aorta, in isolated intact and β-escin-permeabilized iliac arteries, and on perfusion pressure in the isolated mesenteric resistance vascular bed. In noradrenaline-precontracted vessels, the four flavonoids produced a vasodilator effect, which was inversely correlated with the diameter of the vessel studied; i.e., quercetin, isorhamnetin, tamarixetin, and kaempferol were 5-, 25-, 4-, and 6-fold, respectively, more potent in the resistance mesenteric bed (−log IC₅₀ = 5.35 ± 0.15, 5.89 ± 0.11, 5.34 ± 0.10, and 5.66 ± 0.06, respectively) than in the thoracic aorta (−log IC₅₀ = 4.68 ± 0.08, 4.61 ± 0.08, 4.73 ± 0.11, and 4.81 ± 0.13, respectively;n = 4–6). The vasodilator responses of quercetin and isorhamnetin were not significantly modified after removal of the endothelium in the thoracic aorta or in the mesenteric bed. Furthermore, the guanylate cyclase inhibitor ODQ (1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one; 10⁻⁶ M), the adenylate cyclase inhibitor SQ22536 [9-(tetrahydro-2-furanyl)-9H-purin-6-amine; 10⁻⁶ M], KCl (40 mM), or ouabain (10⁻³ M) had no effect on isorhamnetin-induced vasodilation in the mesenteric bed. In permeabilized iliac arteries stimulated with Ca²⁺(pCa of 5.9), isorhamnetin was also significantly more potent (−log IC₅₀ = 5.27 ± 0.15) than quercetin (−log IC₅₀ = 4.56 ± 0.15). In conclusion, quercetin and its metabolites showed vasodilator effects with selectivity toward the resistance vessels. These effects are not due to or modulated by endothelial factors and are unrelated to changes in cytosolic Ca²⁺.

Voltage-gated K ⁺ channels (K _V ) and thromboxane A ₂ (TXA ₂ ) play critical roles in controlling pulmonary arterial tone under physiological and pathological conditions. We hypothesized that TXA ₂ might inhibit K _V channels, thereby establishing a link between these two major pathogenic pathways in pulmonary hypertension. The TXA ₂ analogue U46619 inhibited I _K(V) (E _max =56.1±3.9%, EC ₅₀ =0.054±0.019 μmol/L) and depolarized pulmonary artery smooth muscle cells via activation of TP receptors. In isolated pulmonary arteries, U46619 simultaneously increased intracellular Ca ²⁺ concentration and contractile force, and these effects were inhibited by nifedipine or KCl (60 mmol/L). U46619-induced contractions were not altered by the inhibitors of tyrosine kinase genistein or Rho kinase Y-27632 but were prevented by the nonselective protein kinase C (PKC) inhibitors staurosporine and calphostin C. Furthermore, these responses were sensitive to Gö-6983 but insensitive to bisindolylmaleimide I and Gö-6976. Based on the specificity of these drugs, we suggested a role for an atypical PKC in U46619-induced effects. Thus, treatment with a PKCζ pseudosubstrate inhibitor markedly prevented the vasoconstriction, the inhibition of I _K(V) , and the depolarization induced by U46619. Western blots showed a transient translocation of PKCζ from the cytosolic to the particulate fraction on stimulation with U46619. These results indicate that TXA ₂ inhibits I _K(V) , leading to depolarization, activation of L-type Ca ²⁺ channels, and vasoconstriction of rat pulmonary arteries. We propose PKCζ as a link between TP receptor activation and K _V channel inhibition.