β−Adrenergic receptor subtype expression in myocyte and non-myocyte cells in human female bladder

Abstract

β3-Adrenergic receptor agonists are currently under clinical development for the treatment of overactive bladder, a condition that is prevalent in postmenopausal women. These agents purportedly relax bladder smooth muscle through a direct action at the myocyte β3-receptor. The aim of this study was to examine the expression of the individual beta-adrenergic receptors in full thickness sections from ageing human female bladder. We obtained a series of rabbit polyclonal antibodies generated against each of the three β-adrenergic receptors, and validated their receptor specificity in CHOK1 cells expressing each of the individual receptors. Immunostaining for β1, β2, and β3 were each more prominent in the urothelium than in the detrusor, with all receptors expressed in the same cell types, indicating co-expression of all three receptors throughout the urothelium in addition to the detrusor. Staining of all receptors was also observed in suburothelial myofibroblast-like cells, intramural ganglion cells, and in Schwann cells of intramural nerves. The β3-receptor in the human urothelium appears to be functional, as two different selective β3-receptor agonists, TAK677 and BRL37344, stimulate cAMP formation in UROtsa cells. Densitometry analysis indicates a persistent expression of all receptors throughout the bladder with increasing age, with the exception of the β2-receptor in the urothelium of the trigone, which appears to decrease slightly in older women. These data indicate that β3-receptor expression is maintained with age, but may function in concert with other β-receptors. Activation of the myocyte receptor may be influenced by action on non-myocyte structures including the intramural ganglion cells and myofibroblasts.

Rat β3-adrenoceptor protein expression: antibody validation and distribution in rat gastrointestinal and urogenital tissues

“…For human β 3 -adrenoceptors, recent efforts have identified two antibodies which, at least to some degree, appear suitable for selective labelling (Limberg et al 2010;Cernecka et al 2012). For rodent β 3 -adrenoceptors, the antibody AB15688 was proposed to be selective based on immunocytochemical experiments in transiently transfected Chinese hamster ovary (CHO) cells (Kullmann et al 2009).…”

Urothelial Signaling

“…Thus the ␣ 1A -AR mRNA appeared to be transcribed, but not translated. (183,224), rat (165,166), and pig (201). In the human bladder wall, immunostainings for all three ␤-AR subtypes were found not only in the urothelium and detrusor, but also in suburothelial myofibroblast-like cells, intramural ganglion cells, and Schwann cells of intramural nerves.…”

Mirabegron: a review of recent data and its prospects in the management of overactive bladder

“…However, β3-ARs account for more than 95% of all β-AR mRNA in the human bladder and are thought to be the main β-ARs mediating human detrusor relaxation [Takeda et al 1999[Takeda et al , 2000Igawa et al 2010]. β3-ARs have been found to be highly and preferentially expressed on urinary bladder tissues, including the urothelium, interstitial cells, and detrusor smooth muscle [Andersson et al 2009;Andersson and Arner, 2004;Otsuka et al 2008;Limberg et al 2010;Fujimura et al 1999].…”