Hypothesis / aims of study
In patients suffering from overactive bladder (OAB) the detrusor is in a hypersensitive state. The exact nature of this hypersensitive state can vary, especially when comparing OAB patients with an otherwise healthy bladder to those with a concomitant inflammation. Mirabegron, a selective β3-agonist, was recently approved for the treatment of OAB in combination with an antimuscarinic drug. Early reports on mirabegron suggested that part of the relaxatory effect was exerted via release of nitric oxide (NO). However, this remains to be shown in conclusive studies.
Numerous studies utilize an approach in which relaxation of bladder smooth muscle is studied in pre-contracted tissue. However, one needs to be aware that this can be a difficult approach since the pre-contraction tends to fade during the time period that the relaxatory agent is exerting its effect. In the current study, we chose to study the indirect relaxatory influence that activation of β-adrenoceptors exerts on muscarinic receptor-induced contraction.
The aim of the current study was to determine how β-adrenoceptor subtypes (β1-3) contribute to bladder relaxation and if this is influenced by nitric oxide. Further, it was wondered if this is altered in a state of inflammation, similar to treatment with antimuscarinics which have been shown to be less useful in patients with concomitant cystitis.
Study design, materials and methods
A total of 36 adult male Sprague-Dawley rats were used in the current study. Each rat was pre-treated with either cyclophosphamide (CYP; 100 mg/kg i.p.) to induce experimental cystitis or saline (4 mL/kg i.p.), serving as controls. Sixty hours later, the rats were sacrificed and their bladders were excised. From each bladder, two bladder strips (2 x 6 mm) were cut and mounted in an organ bath setup. After an equilibration period, methacholine (1*10-7 – 3*10-4 M) was applied and the contractile force was measured in the absence and presence of increasing concentrations of a selective β-adrenoceptor agonist (5*10-8 M – 5*10-6 M); either dobutamine (β1), isoprenaline (β1/2) or ZD7114 (β3). Further, each series of measurements was performed in the absence or presence of the NO synthase inhibitor L-NNA (1*10-4 M). All data analysis and statistical comparisons were performed using GraphPad Prism 7.03 (GraphPad Software Inc., San Diego, USA). Two-way ANOVA followed by Tukey’s correction for multiple comparisons was used to compare the obtained dose-response curves and one-way ANOVA followed by Tukey’s correction was used to determine statistical significance between logEC50-values.
Interpretation of results
The current data shows that the presence of a β-agonist, regardless of its subtype selectivity (β1, β1/2 or β3), attenuates muscarinic receptor-induced detrusor contraction. Somewhat surprisingly, this attenuation does not seem to be dependent on release of nitric oxide and remains constant in a state of inflammation. It should be noted that the current study does not distinguish between possible sites of action of β-adrenoceptor-induced relaxation, i.e. via β-adrenoceptors located on the detrusor or via β-adrenoceptors in other parts of the bladder, for instance the urothelium.