Thromboxane A2-induced inhibition of voltage-gated K+ channels and pulmonary vasoconstriction: role of protein kinase Czeta.

Abstract

Voltage-gated K+ channels (KV) and thromboxane A2 (TXA2) play critical roles in controlling pulmonary arterial tone under physiological and pathological conditions. We hypothesized that TXA2 might inhibit KV channels, thereby establishing a link between these two major pathogenic pathways in pulmonary hypertension. The TXA2 analogue U46619 inhibited IK(V) (Emax=56.1+/-3.9%, EC50=0.054+/-0.019 micromol/L) and depolarized pulmonary artery smooth muscle cells via activation of TP receptors. In isolated pulmonary arteries, U46619 simultaneously increased intracellular Ca2+ 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 PKCzeta pseudosubstrate inhibitor markedly prevented the vasoconstriction, the inhibition of IK(V), and the depolarization induced by U46619. Western blots showed a transient translocation of PKCzeta from the cytosolic to the particulate fraction on stimulation with U46619. These results indicate that TXA2 inhibits IK(V), leading to depolarization, activation of L-type Ca2+ channels, and vasoconstriction of rat pulmonary arteries. We propose PKCzeta as a link between TP receptor activation and KV channel inhibition.