Cicletanine reverses vasoconstriction induced by the endogenous sodium pump ligand, marinobufagenin, via a protein kinase C dependent mechanism

Rationale Cicletanine (CIC), an anti-hypertensive compound with direct vasc ular and natriuretic actions, is especially effective in salt-sensitive hyp ertension, in which dysregulation of the sodium pump plays an important pat hogenic role, and digitalis-like cardiotonic steroids contribute to increas ed vascular tone. The purpose of the present study was to investigate wheth er, and by what mechanisms, cicletanine antagonizes the vasoconstrictor eff ects of cardiotonic steroids in isolated human arteries. Methods The effects of cicletanine on vascular tone were studied in isolate d, endothelium-denuded rings of 2nd-3rd-order branches of human mesenteric arteries precontracted with bufodienolide marinobufagenin (MBG), an Na/K-AT Pase inhibitor, or endothelin-1 (ET-1). Na/K-ATPase activity was measured i n sarcolemmal membranes from the mesenteric artery. Activity of rat brain p rotein kinase C (PKC) was measured using the PepTag phosphorylation assay. Results MBG and ET-1 both induced sustained vasoconstriction in human mesen teric artery rings, and cicletanine relaxed rings pre-contracted with eithe r MBG (EC50 = 11 +/- 2 mu mol/l) or ET-1 (EC50 = 6.4 +/- 1.1 mu mol/l). Alt hough 8-Br-cGMP (100 mu mol/l) caused complete vasorelaxation of arterial r ings pre-contracted with ET-1, it did not affect the MBG-induced vasoconstr iction. An activator of PKC, phorbol diacetate (PDA) (50 nmol/l), attenuate d CIC-induced vasorelaxation of mesenteric artery rings pre-contracted with MBG (EC50 >100 mu mol/l), but not rings pre-contracted with ET-1 (EC50 = 6 .5 +/- 1.2 mu mol/l), In mesenteric artery sarcolemma, 100 nmol/l MBG inhib ited the Na/K-ATPase by 68 +/- 5% and cicletanine (100 mu mol/l) attenuated this Na/K-ATPase inhibition by 85 +/- 6%. In the PepTag PKC assay, cicleta nine produced a concentration-dependent inhibition of rat brain PKC activit y (IC50 45 +/- 11 mu mol/l). In the presence of 50 nmol/l PDA, 100 mu mol/l cicletanine did not antagonize the Na/K-ATPase inhibition by MBG, and did not inhibit the PKC from rat brain. Conclusions Cicletanine antagonizes vasoconstriction induced by Na/K-ATPase inhibition via a PKC-dependent mechanism that does not involve inhibition of cyclic GMP phosphodiesterase (cGMP-PDE), This mechanism of action may be relevant to the greater potency of cicletanine in salt-sensitive hypertens ion in which plasma levels of endogenous digitalis-like cardiotonic steroid s are elevated. Our findings also suggest that PKC is an important factor f or cardiotonic steroid - Na/K-ATPase interactions on the vascular tone, and is therefore a potential target for therapeutic intervention in hypertensi on. J Hypertens 2000, 18:209-215 (C) Lippincott Williams & Wilkins.