Warifteine, a bisbenzylisoquinoline alkaloid, induces relaxation by activating potassium channels in vascular myocytes  Apio CL Assis,* † Islania GA Ara ujo,* † Renata PC Lima,* M^ onica M Almeida,* Alexsandro F Marinho,* Jos e M Barbosa-Filho,* Jader S Cruz, ‡ Darizy F Silva § and Isac A Medeiros* *Laboratory of Pharmaceutical Technology, Federal University of Paraíba, Jo~ ao Pessoa, † Department of Physiology and Pathology, Federal University of Paraíba, Jo~ ao Pessoa, ‡ Institute of Biological Sciences, Federal University of Minas Gerais, Belo Horizonte, and § Department of Bioregulation, Federal University of Bahia, Salvador, Brazil SUMMARY 1. The present study used functional and electrophysiological approaches to investigate the mechanisms by which warifteine, a bisbenzylisoquinoline alkaloid isolated from Cissampelos sym- podialis Eichl., causes vasorelaxation of the rat thoracic aorta. 2. Warifteine (1 pmol/L–10 lmol/L) induced concentra- tion-dependent relaxation (pD 2 = 9.40 ± 0.06; n = 5) of endothelium-intact aortic rings precontracted with nor- adrenaline (10–100 lmol/L). The relaxation effects were not attenuated by removal of the endothelium. Warifteine also induced the relaxation of prostaglandin F 2a (1–10 mmol/L)-precontracted rings (pD 2 = 9.2 ± 0.2; n = 8). In contrast, the relaxant activity of warifteine was nearly abolished in high K + (80 mmol/L)-precontracted aortic rings. In preparations incubated with 20 mmol/L KCl or with the K + channel blockers tetraethylammoni- um (1, 3 and 5 mmol/L), iberiotoxin (20 nmol/L), 4-am- inopyridine (1 mmol/L) or glibenclamide (10 lmol/L), the vasorelaxant activity of warifteine was markedly reduced. However, BaCl 2 (1 mmol/L) had no effect on the relaxant effects of warifteine. 3. In vascular myocytes, warifteine (100 nmol/L) signifi- cantly increased whole-cell K + currents (at 70 mV). Under nominally Ca 2+ -free conditions, warifteine did not reduce extracellular Ca 2+ -induced contractions in rings precontracted with high K + or noradrenaline (100 lmol/L). 4. Together, the results of the present study indicate that warifteine induces potent concentration-dependent relaxation in the rat aorta via an endothelium-independent mechanism that involves the activation of K + channels. Key words: bisbenzylisoquinoline alkaloid, potassium channels, rat aorta, vascular smooth muscle cells, vasodilatation, warifteine. INTRODUCTION Hypertension is a global public health concern that is universally accepted as an important prognostic factor for cardiovascular dis- eases and premature mortality. Therefore, the prevention and treatment of hypertension remain a priority for the medical com- munity. 1 Substances derived from natural products are an important source of new medicines. Alkaloids constitute the largest class of botanical secondary metabolites, exhibiting therapeutic effects, such as antirheumatic, 2 antihyperglycaemic, 3 antiviral 4 and anti- inflammatory actions. 4,5 From this group of naturally occurring organic compounds, the bisbenzylisoquinoline alkaloids (e.g. cycleanine, tetrandine and berbamine) exhibit diverse biological activities, such as suppression of hepatic injury and the produc- tion of tumour necrosis factor in Bacille Calmette–Guerin plus lipopolysaccharide-treated mice. 6,7 In addition to its antihyperten- sive effect on spontaneously hypertensive rats, renal hypertensive rats and deoxycorticosterone acetate-salt hypertensive rats, 8 tetrandine has been shown to decrease systolic and diastolic arte- rial pressure in humans. 9 Dauricine and daurisoline have been shown to have anti-arrhythmic actions, 10 providing further evi- dence of the beneficial cardiovascular effects of bisbenzylisoquin- oline alkaloids. Therefore, further studies into this chemical structure may result in new treatments for cardiovascular prob- lems, such as arrhythmia and hypertension. Cissampelos sympodialis Eichl. (Menispermaceae), a species found in north-eastern and south-eastern Brazil, has been used in folk medicine for the treatment of rheumatism, colds, asthma and other conditions. 11 Phytochemical analyses of the roots of this plant revealed the presence of a group of alkaloids, namely wari- fteine, methylwarifteine, roraimine and milonine. Warifteine (C 36 H 36 N 2 O 6 ; molecular weight 592.7; Fig. 1), a bisbenzyliso- quinoline alkaloid, 12 is the predominant component isolated from the ethanolic extract of C. sympodialis root bark. Several studies have shown promising pharmacological effects of C. sympodialis and warifteine on biological systems, including immunomodula- tion, 13 antidepressant effects 14 and inhibition of eosinophil recruitment. 11 However, few reports have investigated the effects of warifteine on vasomotor tone, despite the known anti- arrhythmic and antihypertensive properties of compounds from the same chemical class as warifteine. Previous studies have demonstrated that warifteine induces concentration-dependent relaxation of the rabbit aorta. 12 This relaxation was shown to be Correspondence: Professor Darizy F Silva, Departamento de Biorregu- lac ß~ ao, Universidade Federal da Bahia (UFBA), Vale do Canela, Salvador, BA 40110–902, Brazil. Email: darizy@gmail.com Received 22 July 2012; revision 17 October 2012; accepted 21 October 2012. © 2012 The Authors Clinical and Experimental Pharmacology and Physiology © 2012 Wiley Publishing Asia Pty Ltd Clinical and Experimental Pharmacology and Physiology (2013) 40, 37–44 doi: 10.1111/1440-1681.12029