LAUSR

BACKGROUND AND PURPOSE The way β3 receptor agonists (e.g. mirabegron) control bladder overactivity may involve adenosine release from human and rat detrusor smooth muscle. Retrograde activation of adenosine A1 receptors reduces acetylcholine (ACh) release from cholinergic bladder nerves. β3 -Adrenoceptors usually couple to adenylyl cyclase. Here, we investigated the cyclic AMP target, either protein kinase A (PKA) or the exchange protein directly activated by cAMP (EPAC), most likely participating in β3 -induced cholinergic inhibition of the urinary bladder. EXPERIMENTAL APPROACH [3 H]ACh and adenosine release from urothelium-denuded detrusor samples of cadaveric human organ donors and Wistar rats were measured by liquid scintillation spectrometry and HPLC, respectively. In vivo cystometry recordings were performed in urethane-anaesthetized rats. KEY RESULTS The EPAC inhibitor, ESI-09, prevented mirabegron- and isoprenaline-induced adenosine release from human and rat detrusor strips, respectively. ESI-09, but not the PKA inhibitor, H-89, attenuated inhibition of [3 H]ACh release from stimulated (10Hz) detrusor strips caused by drugs activating β3 -adrenoceptors, adenylyl cyclase (forskolin), and EPAC1 (8-CTP-2Me-cAMP). Isoprenaline-induced inhibition of [3 H]ACh release was also prevented by inhibitors of protein kinase C (PKC; chelerythrine and Go6976) and of the equilibrative nucleoside transporter 1 (ENT1) (dipyridamole and NBTI), but not by phospholipase C inhibition with U73122. Pretreatment with ESI-09, but not with H-89, prevented the reduction of the voiding frequency caused by isoprenaline and forskolin in vivo. CONCLUSION AND IMPLICATIONS Data suggest that β3 -adrenoceptor-induced inhibition of cholinergic neurotransmission in human and rat urinary bladders involves activation of an EPAC1/PKC pathway downstream cyclic AMP production resulting in adenosine outflow via ENT1.