Pulmonary, Gastrointestinal and Urogenital Pharmacology Lack of evidence that nebivolol is a β 3 -adrenoceptor agonist Elfaridah P. Frazier a , Martina B. Michel-Reher a , Pieter van Loenen a , Carsten Sand b , Tim Schneider c , Stephan L.M. Peters a , Martin C. Michel a, ⁎ a Department of Pharmacology and Pharmacotherapy, University of Amsterdam, The Netherlands b Department of Pharmacology, University of Duisburg-Essen, Germany c Department of Urology, University of Duisburg-Essen, Germany abstract article info Article history: Received 7 October 2010 Received in revised form 9 November 2010 Accepted 12 November 2010 Available online 21 December 2010 Keywords: Nebivolol Urinary bladder Chinese hamster ovary cell β 3 -Adrenoceptor Nebivolol is a selective β 1 -adrenoceptor antagonist which, in addition, displays endothelium-dependent vasodilating properties in humans and other species. β 3 -Adrenoceptors have been proposed to be a molecular target of nebivolol-induced vasodilatation. Therefore, we have investigated possible β 3 -adrenoceptor agonism by nebivolol for relaxation of the human and rat urinary bladder (prototypical β 3 -adrenoceptor-mediated responses) as well as for cAMP accumulation in Chinese hamster ovary cells stably transfected with the human β-adrenoceptor subtypes. Nebivolol concentration-dependently relaxed both human and rat isolated urinary bladder strips but with low potency, similar to that reported for vasodilatation. However, nebivolol- induced bladder relaxation in either species was not inhibited by the β 3 -adrenoceptor antagonist SR 59,230A (10 μM), although this compound inhibited the isoprenaline-induced relaxation with the expected potency. In radioligand binding studies nebivolol had lower affinity for human β 3 -adrenoceptors than the other two β-adrenoceptor subtypes, but this low affinity was in line with its potency to relax the bladder or isolated blood vessels. In functional studies nebivolol even in high concentrations did not stimulate cAMP formation via any of the three cloned human β-adrenoceptors or in rat bladder smooth muscle cells. Taken together these data demonstrate that nebivolol can relax not only vascular but also urinary bladder smooth muscle. However, they do not support the hypothesis that nebivolol is an agonist at cloned human β 3 -adrenoceptors or in rat or human urinary bladder. © 2010 Elsevier B.V. All rights reserved. 1. Introduction Nebivolol is a β-adrenoceptor antagonist which is selective for β 1 - as compared to β 2 -adrenoceptors (Pauwels et al., 1988; Bundkirchen et al., 2003; Baker, 2010). In contrast to many other β-adrenoceptor antagonists, nebivolol has direct dilating effects in both animal and human isolated blood vessels, which typically are endothelium- dependent and in most cases involve the generation of NO (Ignarro, 2008; Tran Quang et al., 2009; Kamp et al., 2010). As the nebivolol concentrations required for vasodilatation are much higher than those needed to occupy β 1 -adrenoceptors, it is generally assumed that this vasodilatation occurs via a molecular target distinct from the β 1 -adrenoceptor. Further support for this conclusion may come from the clinical finding that the β 1 -adrenoceptor antagonist effects primarily reside in the D-enantiomer of nebivolol, whereas the vasodilating effects of nebivolol are mainly associated with its L-enantiomer in some studies (van Nueten and De Cree, 1998); however, others found nebivolol- induced vasodilatation in vitro not to be stereo-selective (Chlopicki et al., 2002; Tran Quang et al., 2009). There is considerable controversy what the molecular target for nebivolol-induced endothelium-dependent vasodilatation might be. Proposed nebivolol targets include effects on NO bioavailability (Evangelista et al., 2007), oestrogen receptors (Garban et al., 2004), purinergic receptors (Kalinowski et al., 2003), α 1 -adrenoceptors (Rozec et al., 2006), β 2 -adrenoceptors (Broeders et al., 2000; Tran Quang et al., 2009), and β 3 -adrenoceptors (de Groot et al., 2003; Tran Quang et al., 2009; Rozec et al., 2009). Evidence for the latter possibility comes from several sources including the use of antagonists such as cyanopindolol (de Groot et al., 2003), SR 59,230A (Evangelista et al., 2007) and L-748,337 (Rozec et al., 2006, 2009; Tran Quang et al., 2009). However, many of the antagonists which have previously been used have ancillary properties. For example, cyanopindolol is also a ligand at serotonin receptors (Schlicker et al., 1985), and SR 59,230A can act on muscarinic receptors, does not discriminate human β-adrenoceptor subtypes and can be a β 3 -adrenoceptor partial agonist in some cases (Vrydag and Michel, 2007). While L-748,337 has one of the highest β 3 -selectivities among generally available antagonists (Candelore et al., 1999; Baker, 2010), the concentrations used in the above mentioned studies may also block β 1 - and β 2 -adrenoceptors. Therefore, these antagonist data can only be seen as circumstantial evidence for an European Journal of Pharmacology 654 (2011) 86–91 ⁎ Corresponding author. Academic Medical Center, Department Pharmacology and Pharmacotherapy, Meibergdreef 15, 1105 AZ Amsterdam, The Netherlands. Tel.: + 31 20 566 6762; fax: +31 20 696 5976. E-mail address: m.c.michel@amc.nl (M.C. Michel). 0014-2999/$ – see front matter © 2010 Elsevier B.V. All rights reserved. doi:10.1016/j.ejphar.2010.11.036 Contents lists available at ScienceDirect European Journal of Pharmacology journal homepage: www.elsevier.com/locate/ejphar