title: Cicletanine reverses vasoconstriction induced by the endogenous sodium pump ligand, marinobufagenin, via a protein kinase C dependent mechanism.
abstract: BACKGROUND
  Cicletanine (CIC), an anti-hypertensive compound with direct vascular and natriuretic actions, is especially effective in salt-sensitive hypertension, in which dysregulation of the sodium pump plays an important pathogenic role, and digitalis-like cardiotonic steroids contribute to increased vascular tone. The purpose of the present study was to investigate whether, and by what mechanisms, cicletanine antagonizes the vasoconstrictor effects of cardiotonic steroids in isolated human arteries.
  
  
  METHODS
  The effects of cicletanine on vascular tone were studied in isolated, endothelium-denuded rings of 2nd-3rd-order branches of human mesenteric arteries pre-contracted with bufodienolide marinobufagenin (MBG), an Na/K-ATPase inhibitor, or endothelin-1 (ET-1). Na/K-ATPase activity was measured in sarcolemmal membranes from the mesenteric artery. Activity of rat brain protein kinase C (PKC) was measured using the PepTag phosphorylation assay.
  
  
  RESULTS
  MBG and ET-1 both induced sustained vasoconstriction in human mesenteric artery rings, and cicletanine relaxed rings pre-contracted with either MBG (EC50 = 11 +/- 2 micromol/l) or ET-1 (EC50 = 6.4 +/- 1.1 micromol/l). Although 8-Br-cGMP (100 micromol/l) caused complete vasorelaxation of arterial rings pre-contracted with ET-1, it did not affect the MBG-induced vasoconstriction. An activator of PKC, phorbol diacetate (PDA) (50 nmol/l), attenuated CIC-induced vasorelaxation of mesenteric artery rings pre-contracted with MBG (EC50 > 100 micromol/l), but not rings pre-contracted with ET-1 (EC50 = 6.5 +/- 1.2 micromol/l). In mesenteric artery sarcolemma, 100 nmol/l MBG inhibited the Na/K-ATPase by 68 +/- 5% and cicletanine (100 micromol/l) attenuated this Na/K-ATPase inhibition by 85 +/- 6%. In the PepTag PKC assay, cicletanine produced a concentration-dependent inhibition of rat brain PKC activity (IC50 45 +/- 11 micromol/l). In the presence of 50 nmol/l PDA, 100 micromol/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 hypertension in which plasma levels of endogenous digitalis-like cardiotonic steroids are elevated. Our findings also suggest that PKC is an important factor for cardiotonic steroid-Na/K-ATPase interactions on the vascular tone, and is therefore a potential target for therapeutic intervention in hypertension.
authors.affiliation: Laboratory of Cardiovascular Science, Intramural Research Program, National Institute on Aging, Baltimore, Maryland 21224, USA.
authors.forename: A Y
authors.initials: AY
authors.lastname: Bagrov
authors.forename: R I
authors.initials: RI
authors.lastname: Dmitrieva
authors.forename: N A
authors.initials: NA
authors.lastname: Dorofeeva
authors.forename: O V
authors.initials: OV
authors.lastname: Fedorova
authors.forename: D A
authors.initials: DA
authors.lastname: Lopatin
authors.forename: E G
authors.initials: EG
authors.lastname: Lakatta
authors.forename: M T
authors.initials: MT
authors.lastname: Droy-Lefaix
mesh_terms: D000818:Animals; D000959:Antihypertensive Agents / Q000494:pharmacology*; D001921:Brain / Q000201:enzymology; D002018:Bufanolides / Q000494:pharmacology*; D006152:Cyclic GMP / Q000031:analogs & derivatives / Q000494:pharmacology; D019332:Endothelin-1 / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D066298:In Vitro Techniques; D008297:Male; D008638:Mesenteric Arteries / Q000187:drug effects / Q000502:physiology; D008875:Middle Aged; D011493:Protein Kinase C / Q000378:metabolism*; D011725:Pyridines / Q000494:pharmacology*; D051381:Rats; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors*; D014661:Vasoconstriction / Q000187:drug effects* / Q000502:physiology
chemical_list: D000959:Antihypertensive Agents; D002018:Bufanolides; D019332:Endothelin-1; D004791:Enzyme Inhibitors; D011725:Pyridines; C016276:8-bromocyclic GMP; C093896:marinobufagenin; C038068:cicletanine; D011493:Protein Kinase C; D000254:Sodium-Potassium-Exchanging ATPase; D006152:Cyclic GMP