Functional characterization of adenosine receptors and coupling to ATP-sensitive K+ channels in guinea pig urinary bladder smooth muscle

Although multiple adenosine receptors have been identified, the subtype and underlying mechanisms involved in the relaxation response to adenosine in the urinary bladder remain unclear. The present study investigates changes in the membrane potential, as assessed by fluorescence-based techniques, of bladder smooth muscle cells by adenosine receptor agonists acting via ATP-sensitive potassium (K-ATP) channels. Membrane hyperpolarization evoked by adenosine and various adenosine receptor subtype-selective agonists was attenuated or reversed by the K-ATP channel blocker glyburide. Comparison of adenosine receptor agonist potencies eliciting membrane potential effects showed a rank order of potency 5'-N-ethylcarboxamido adenosine (NECA; -log EC50 = 7.97) similar to 2-p-(2-carboxethyl)phenethyl-amino-5'-N-ethylcarboxamidoadenosine hydrochloride (CGS-21680; 7.65) > 2-chloro adenosine (5.90) similar to 2-chloro-N-6-cyclopentyladenosine (CCPA; 5.51) similar to N-6-cyclopentyladenosine similar to N-6-(R)-phenylisopropyladenosine > 2 chloro-N-6-(3-iodobenzyl)-adenosine-5'-N-methyl-carboxamide (2Cl-IB- MECA; 4.78). Membrane potential responses were mimicked by forskolin, a known activator of adenylate cyclase, and papaverine, a phosphodiesterase inhibitor. The A(2A)-selective antagonist 4-(2-[7-amino-2-(2-furyl)[1,2,4]triazolo[2,3-a][1,3,5]triazin-5-yl-amino] ethyl)phenol (ZM-241385), and the adenylate cyclase inhibitor N-(cis-2-phenyl-cyclopentyl) azacyclotridecan-2-imine-hydrochloride (MDL-12330A) inhibited the observed change in membrane potential evoked by adenosine and adenosine-receptor agonists. The rank order potency for relaxation of K+-stimulated guinea pig bladder strips, NECA (-log EC50 = 6.41) similar to CGS-21680 (6.38) > 2-chloro adenosine (5.90) >> CCPA similar to 2Cl-IB-MECA (>4.0) was comparable to that obtained from membrane potential measurements. Collectively, these studies demonstrate that adenosine-evoked membrane hyperpolarization and relaxation of bladder smooth muscle is mediated by A(2A) receptor-mediated activation of K-ATP channels via adenylate cyclase and elevation of CAMP.