title: Vasorelaxation induced by dodoneine is mediated by calcium channels blockade and carbonic anhydrase inhibition on vascular smooth muscle cells.
abstract: BACKGROUND
  Dodoneine (Ddn) is one of the active compounds identified from Agelanthus dodoneifolius (DC.) Polhill and Wiens, a medicinal plant used in traditional medicine for the treatment of hypertension. This dihydropyranone exerts hypotensive and vasorelaxant effects on rats, and two molecular targets have been characterized: the carbonic anhydrase and the L-type calcium channel in cardiomyocytes with biochemical and electrophysiological techniques, respectively. To further evaluate the involvement of these two molecular targets in vasorelaxation, the effect of Ddn on rat vascular smooth muscle was investigated.
  
  
  METHODS
  The effects of Ddn on L-type calcium current and on resting membrane potential were characterized in A7r5 cell line using the whole-cell patch-clamp configuration. The molecular identities of carbonic anhydrase isozymes in smooth muscle cells were examined with RT-PCR. Vascular response was measured on rat aortic rings in an organ bath apparatus and the effect of Ddn on intracellular pH was determined by flow cytometry using the pH-sensitive fluorescent probe BCECF-AM [2,7-Bis-(2-Carboxyethyl)-5-(and-6)-Carboxyfluorescein, Acetoxymethyl Ester].
  
  
  RESULTS
  100µM Ddn reduced calcium current density of about 30%. In addition, carbonic anhydrase II, III, XIII and XIV were shown to be expressed in rat aorta and inhibited in smooth muscle cells by Ddn. This inhibition resulted in a rise in pHi of about 0.31, leading to KCa channel activation, thereby inducing membrane hyperpolarization and vasorelaxation. The results of vascular reactivity experiments obtained with pharmacological tools acting on the L-type calcium current and carbonic anhydrase suggest that Ddn produces its vasorelaxant effect via the inhibition of these two molecular targets.
  
  
  CONCLUSIONS
  This study demonstrates that Ddn induced vasorelaxation by targeting two proteins involved in the modulation of excitation-contraction coupling: L-type calcium channels and carbonic anhydrase.
authors.affiliation: Signalisation et Transports Ioniques Membranaires, Université de Poitiers, CNRS ERL 7368; 1 rue Georges Bonnet F-86073 Poitiers Cedex 09, France.
authors.forename: Grégoire
authors.initials: G
authors.lastname: Carre
authors.affiliation: Laboratoire de Physiologie Animale, Université de Ouagadougou, 03 BP 7021, Ouagadougou 01, Burkina Faso.
authors.forename: Maurice
authors.initials: M
authors.lastname: Ouedraogo
authors.affiliation: Signalisation et Transports Ioniques Membranaires, Université de Poitiers, CNRS ERL 7368; 1 rue Georges Bonnet F-86073 Poitiers Cedex 09, France.
authors.forename: Christophe
authors.initials: C
authors.lastname: Magaud
authors.affiliation: Superacid group in "Organic Synthesis" team, Université de Poitiers, CNRS UMR 7285 IC2MP, 4 avenue Michel Brunet, Poitiers 86022 Cedex, France.
authors.forename: Hélène
authors.initials: H
authors.lastname: Carreyre
authors.affiliation: Signalisation et Transports Ioniques Membranaires, Université de Poitiers, CNRS ERL 7368; 1 rue Georges Bonnet F-86073 Poitiers Cedex 09, France.
authors.forename: Frédéric
authors.initials: F
authors.lastname: Becq
authors.affiliation: Signalisation et Transports Ioniques Membranaires, Université de Poitiers, CNRS ERL 7368; 1 rue Georges Bonnet F-86073 Poitiers Cedex 09, France.
authors.forename: Patrick
authors.initials: P
authors.lastname: Bois
authors.affiliation: Università degli Studi di Firenze, Laboratorio di Chimica Bioinorganica, Rm 188, Via della Lastruccia 3, I-50019 Sesto Fiorentino (Firenze), Italy.
authors.forename: Claudiu T
authors.initials: CT
authors.lastname: Supuran
authors.affiliation: Superacid group in "Organic Synthesis" team, Université de Poitiers, CNRS UMR 7285 IC2MP, 4 avenue Michel Brunet, Poitiers 86022 Cedex, France.
authors.forename: Sébastien
authors.initials: S
authors.lastname: Thibaudeau
authors.affiliation: Signalisation et Transports Ioniques Membranaires, Université de Poitiers, CNRS ERL 7368; 1 rue Georges Bonnet F-86073 Poitiers Cedex 09, France.
authors.forename: Clarisse
authors.initials: C
authors.lastname: Vandebrouck
authors.affiliation: Signalisation et Transports Ioniques Membranaires, Université de Poitiers, CNRS ERL 7368; 1 rue Georges Bonnet F-86073 Poitiers Cedex 09, France. Electronic address: Jocelyn.bescond@univ-poitiers.fr.
authors.forename: Jocelyn
authors.initials: J
authors.lastname: Bescond
mesh_terms: D000818:Animals; D001011:Aorta / Q000187:drug effects / Q000502:physiology; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D020746:Calcium Channels, L-Type / Q000502:physiology; D002257:Carbonic Anhydrase Inhibitors / Q000494:pharmacology*; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000502:physiology*; D009924:Organ Culture Techniques; D010636:Phenols / Q000494:pharmacology*; D011753:Pyrones / Q000494:pharmacology*; D051381:Rats; D017208:Rats, Wistar; D014664:Vasodilation / Q000187:drug effects / Q000502:physiology*; D014665:Vasodilator Agents / Q000494:pharmacology*
chemical_list: D002121:Calcium Channel Blockers; D020746:Calcium Channels, L-Type; D002257:Carbonic Anhydrase Inhibitors; D010636:Phenols; D011753:Pyrones; D014665:Vasodilator Agents; C000593123:dodoneine
keywords: Carbonic anhydrase; Dodoneine; Hypertension; L-type calcium current; Natural substance; Vasorelaxation