Role of Rho-kinase and BK channels in β3-AR mediated in vitro relaxation of rat and human bladder smooth muscle strips.

H Cernecka1, K Kersten1, H Maarsingh1,5, IJ de Jong3, C Korstanje4, MC Michel2, M Schmidt1. 1University of Groningen, Groningen, The Netherlands, 2Johannes Gutenberg University, Mainz, Germany, 3University Medical Center, Groningen, The Netherlands, 4Astellas Pharma Europe BV, Leiderdorp, The Netherlands, 5Palm Beach Atlantic University, West Palm Beach, USA

Previous studies suggest that Rho-kinase may play a major role in the control of bladder tone and that the large-conductance voltage and Ca2+-activated K+ (BK) channel is a major regulator of detrusor smooth muscle (DSM) contractility (1,2). Other studies have shown that β3-adrenoreceptor (β3-AR) stimulation of the urinary bladder can cause DSM relaxation (3). This study evaluated the role of BK channels and Rho-kinase in rat and human DSM relaxation induced by the β3-AR agonist, mirabegron (MIRA) with the focus on potential qualitative differences in both species. Human detrusor tissue was obtained with ethical committee approval from patients undergoing cystectomy for bladder cancer (n=4-6 per group) and rat detrusor tissue was obtained from adult male Wistar rats (n=8 per group). Rat DSM strips were pre-contracted with carbachol (1 µM) or KCl (80 mM) and the relaxant effect of MIRA (10-9-10-4 M) or isoproterenol (ISO; 10-9-10-4 M) evaluated in the presence or absence of the Rho-kinase inhibitor, Y27,632 (1 µM). Similar studies evaluating the effect of Rho-kinase inhibition or BK channel inhibition (iberiotoxin; IBX 100 nM) on MIRA and ISO induced relaxation of human DSM strips pre-contracted with carbachol (1 µM) were also performed. Y27,632 did not affect ISO- or MIRA-induced relaxation in KCl pre-contracted rat bladder strips but augmented relaxation in carbachol pre-contracted strips (Table 1). In carbachol pre-contracted human bladder strips Y27,632 also enhanced MIRA-induced relaxation but had no effect on ISO-mediated relaxation. IBX increased MIRA- and ISO-mediated relaxation in human bladder strips pre-contracted by carbachol (Table 1). As the concentration-response curves for β-agonists were shallow, the curves in the presence or absence of inhibitors were compared by 2-way ANOVA testing the effect of treatment.

Table 1 Effects of Y27,632 and IBX on maximum ISO- and MIRA-mediated relaxation of rat and human DSM

Data are means ± S.E.M. of 4 to 8 experiments and expressed as a percentage relaxation. *<0.05 vs. corresponding vehicle, in a 2-way ANOVA.

These data showed that ISO and MIRA induced relaxation of rat and human DSM in vitro. The relative role of Rho-kinase and BK channels in DSM relaxation depends on the pre-contracting stimulus, species and β3-adrenoceptor agonist.

(1) Peters SL et al, Trends Pharmacol Sci 9:492-7, 2006.

(2) Afeli SA & Petkov GV, Eur J Pharmacol 711:50-6, 2013.

(3) Takasu T et al, J Pharmacol Exp Ther 321:642-7, 2007.