Adenosine receptor-coupled modulation of ATP release from isolated urothelial cells of mouse bladder

Introduction Sensations arising from the urinary bladder, such as those occurring during filling, involve purinergic signalling pathways due to ATP release from the urothelium when it is subject to physical stresses. Excessive urothelial ATP release is associated with sensory and overactive bladder pathologies and there is an unmet need to understand how to regulate such release that may alleviate abnormal sensations. This study tested the hypothesis that adenosine, acting on A1 receptors, has a significant role. Methods Urothelial cellular suspensions were prepared by trypsin/EDTA disruption of the bladder wall mucosa and used immediately for experiments. ATP release from cell suspensions, when subject to mechanical stresses, was measured with a luciferin-luciferase assay. Results ATP release following mechanical stress was attenuated by adenosine which in turn was prevented by a protein kinase-A (PKA) blocker, cAMPs-Rp. A comparable attenuation of release was caused by addition of cinaciguat, a soluble guanylate cyclase (sGC) activator. A greater attenuation resulted from a combination of adenosine and cinaciguat. The A1-selective receptor antagonist DPCPX increased ATP release, that was not overcome by adenosine. Conclusions ATP release from urothelial cells was attenuated by adenosine via A1 receptors, and by the sGC activator cinaciguat. Kinetics of ATP release by the two agents were compatible with them acting independently. The increase of release by DPCPX may be explained by the antagonist exerting inverse agonism at the A1 receptor, the receptor exerting a constitutively active state.