Mirabegron relaxes urethral smooth muscle by a dual mechanism involving β3 -adrenoceptor activation and α1 -adrenoceptor blockade.

BJP DOI:10.1111/bph.13367 www.brjpharmacol.org British Journal of Pharmacology RESEARCH PAPER Correspondence Mirabegron relaxes urethral smooth muscle by a dual mechanism involving β3adrenoceptor activation and α1-adrenoceptor blockade Edson Antunes, Department of Pharmacology, Faculty of Medical Sciences, State University of Campinas (UNICAMP), 13084-971 Campinas, São Paulo, Brazil. E-mail: ; --------------------------------------------------------- Received 19 February 2015 Revised 16 September 2015 Accepted 19 October 2015 E C Alexandre1, L R Kiguti2, F B Calmasini1, F H Silva1, K P da Silva2, R Ferreira3, C A Ribeiro2, F Z Mónica1, A S Pupo2 and E Antunes1 1 2 Department of Pharmacology, University of Campinas (UNICAMP), Campinas, Brazil, Department of Pharmacology, Institute of Biosciences, University of São Paulo State (UNESP), Botucatu, São Paulo, Brazil, and 3Hematology and Hemotherapy Center, Faculty of Medical Sciences, University of Campinas (UNICAMP), Campinas, Brazil LINKED ARTICLE This article is commented on by Michel, M. C., pp. 429-430 of this issue. To view this commentary visit http://dx.doi.org/ 10.1111/bph.13379. BACKGROUND AND PURPOSE Mirabegron is the first β3-adrenoceptor agonist approved for treatment of overactive bladder syndrome. This study aimed to investigate the effects of β3-adrenoceptor agonist mirabegron in mouse urethra. The possibility that mirabegron also exerts α1-adrenoceptor antagonism was also tested in rat smooth muscle preparations presenting α1A- (vas deferens and prostate), α1D- (aorta) and α1B-adrenoceptors (spleen). EXPERIMENTAL APPROACH Functional assays were carried out in mouse and rat isolated tissues. Competition assays for the specific binding of [3H]prazosin to membrane preparations of HEK-293 cells expressing each of the human α1-adrenoceptors, as well as β-adrenoceptor mRNA expression and cyclic AMP measurements in mouse urethra, were performed. KEY RESULTS Mirabegron produced concentration-dependent urethral relaxations that were shifted to the right by the selective β3adrenoceptor antagonist L-748,337 but unaffected by β1- and β2-adrenoceptor antagonists (atenolol and ICI-118,551 respectively). Mirabegron-induced relaxations were enhanced by the PDE4 inhibitor rolipram, and the agonist stimulated cAMP synthesis. Mirabegron also produced rightward shifts in urethral contractions induced by the α1-adrenoceptor agonist phenylephrine. Schild regression analysis revealed that mirabegron behaves as a competitive antagonist of α1-adrenoceptors in urethra, vas deferens and prostate (α1A-adrenoceptor, pA2 ≅ 5.6) and aorta (α1D-adrenoceptor, pA2 ≅ 5.4) but not in spleen (α1B-adrenoceptor). The affinities estimated for mirabegron in functional assays were consistent with those estimated in radioligand binding with human recombinant α1A- and α1D-adrenoceptors (pKi ≅ 6.0). CONCLUSION AND IMPLICATIONS The effects of mirabegron in urethral smooth muscle are the result of β3-adrenoceptor agonism together with α1A and α1D-adrenoceptor antagonism. Abbreviations CR, concentration ratios; CRC, concentration–response curve; KHS, Krebs–Henseleit solution; LUTS, lower urinary tract symptoms; OAB, Overactive bladder syndrome © 2015 The British Pharmacological Society British Journal of Pharmacology (2016) 173 415–428 415 E C Alexandre et al. BJP Tables of Links TARGETS GPCRs a LIGANDS 17β-oestradiol Mirabegron α1A-adrenoceptor Arginine-vasopressin Noradrenaline α1B-adrenoceptor Atenolol ODQ α1D-adrenoceptor Desipramine Propranolol β1-adrenoceptor Endothelin-1 Rolipram β2-adrenoceptor IBMX Yohimbine β3-adrenoceptor ICI-118,551 Enzymes b PDE4 Isoprenaline L-748,337 These Tables list key protein targets and ligands in this article which are hyperlinked to corresponding entries in http://www.guidetopharmacology.org, the common portal for data from the IUPHAR/BPS Guide to PHARMACOLOGY (Pawson et al., 2014) and are permanently archived in the Concise Guide abc to PHARMACOLOGY 2013/14 ( Alexander et al., 2013a, b, c). Introduction Lower urinary tract symptoms (LUTS) represent one of the most common clinical complaints in men and women due to structural or functional abnormalities in one or more parts of the lower urinary tract, which comprises the bladder, bladder neck, prostate, distal sphincter mechanism and urethra (Abrams et al., 2002, 2013). Approximately 1.9 billion individuals worldwide are estimated to experience any LUTS, with numbers of affected individuals projected to 2.3 billion (18.4% increase) in 2018 (Irwin et al., 2011). LUTS can be divided into three groups, namely, storage symptoms (increased daytime urinary frequency, nocturia, urgency and incontinence), voiding symptoms (slow stream, splitting or spraying, intermittent stream, hesitancy, straining and terminal dribble) and post-micturition symptoms (feeling of incomplete emptying and post-micturition dribble). Overactive bladder syndrome (OAB) is a subset of storage LUTS currently defined as urgency, with or without urge incontinence, usually accompanied by frequency and nocturia (Abrams et al., 2002). In ageing men, LUTS has been attributed to bladder outlet obstruction as a result of benign prostatic enlargement resulting from the histological condition of benign prostatic hyperplasia (Abrams et al., 2013). The lower urinary tract stores and releases urine via integrated circuits with brain, spinal cord and peripheral ganglia. During the voiding phase, parasympathetic neurons release ACh to contract the bladder smooth muscle through activation of muscarinic M3 cholinoceptors, resulting in an efficient bladder emptying (Hegde et al., 1997). ATP via purinergic P2X1 purinoceptors may act as a co-transmitter to ACh in parasympathetic nerves, producing bladder contractions in physiological conditions, although this component plays a minor role in human bladder (Burnstock, 2014). The micturition event is followed by the storage phase where activation of sympathetic post-ganglionic fibers causes the release of noradrenaline that acts on β-adrenoceptors to promote bladder relaxation (Igawa et al., 1999, 2001). The β2- and β3-adrenoceptors play important roles to induce bladder relaxations in rodent and humans (Fujimura et al., 1999). 416 British Journal of Pharmacology (2016) 173 415–428 The urethra is composed of an inner longitudinal and a middle circular smooth muscle layer innervated by autonomic nerves, along with an outer striated muscle (rhabdosphincter) innervated by somatic nerves (Pradidarcheep et al., 2011). Besides being a conduit for the urine, the urethra contributes to urinary continence by relaxing during the voiding phase and contracting during the urine storage phase (Michel and Vrydag, 2006). The urethral smooth muscle is richly innervated by sympathetic fibers, the activation of which results in the release of noradrenaline that acts on post-junctional α1-adrenoceptors to produce contractions during the filing/s (...truncated)