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Retrograde activation of adenosine A<jats:sub>1<\/jats:sub> receptors reduces ACh release from cholinergic bladder nerves. \u03b2<jats:sub>3<\/jats:sub>\u2010Adrenoceptors usually couple to adenylyl cyclase. Here we investigated, which of the cAMP targets, protein kinase A or the exchange protein directly activated by cAMP (EPAC) could be involved in this cholinergic inhibition of the bladder.<\/jats:p><\/jats:sec><jats:sec><jats:title>Experimental Approach<\/jats:title><jats:p>[<jats:sup>3<\/jats:sup>H]ACh and adenosine release from urothelium\u2010denuded detrusor strips of cadaveric human organ donors and rats were measured by liquid scintillation spectrometry and HPLC, respectively. In vivo cystometry was also performed in urethane\u2010anaesthetized rats.<\/jats:p><\/jats:sec><jats:sec><jats:title>Key Results<\/jats:title><jats:p>The exchange protein directly activated by cAMP (EPAC) inhibitor, ESI\u201009, prevented mirabegron\u2010 and isoprenaline\u2010induced adenosine release from human and rat detrusor strips respectively. ESI\u201009, but not the PKA inhibitor, H\u201089, attenuated inhibition of [<jats:sup>3<\/jats:sup>H]ACh release from stimulated (10 Hz) detrusor strips caused by activating \u03b2<jats:sub>3<\/jats:sub>\u2010adrenoceptors, AC (forskolin) and EPAC1 (8\u2010CTP\u20102Me\u2010cAMP). Isoprenaline\u2010induced inhibition of [<jats:sup>3<\/jats:sup>H]ACh release was also prevented by inhibitors of PKC (chelerythrine and Go6976) and of the equilibrative nucleoside transporter 1 (ENT1; dipyridamole and NBTI), but not by PLC inhibition with U73122. Pretreatment with ESI\u201009, but not with H\u201089, prevented the reduction of the voiding frequency caused by isoprenaline and forskolin in vivo.<\/jats:p><\/jats:sec><jats:sec><jats:title>Conclusion and Implications<\/jats:title><jats:p>Data suggest that \u03b2<jats:sub>3<\/jats:sub>\u2010adrenoceptor\u2010induced inhibition of cholinergic neurotransmission in human and rat urinary bladders involves activation of an EPAC1\/PKC pathway downstream cAMP production resulting in adenosine outflow via ENT1.<\/jats:p><\/jats:sec>","DOI":"10.1111\/bph.14921","type":"journal-article","created":{"date-parts":[[2019,11,13]],"date-time":"2019-11-13T05:56:44Z","timestamp":1573624604000},"page":"1589-1608","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":12,"title":["\u03b2<sub>3<\/sub> Adrenoceptor\u2010induced cholinergic inhibition in human and rat urinary bladders involves the exchange protein directly activated by cyclic AMP 1 favoring adenosine release"],"prefix":"10.1111","volume":"177","author":[{"given":"Isabel","family":"Silva","sequence":"first","affiliation":[{"name":"Laborat\u00f3rio de Farmacologia e Neurobiologia, Instituto de Ci\u00eancias Biom\u00e9dicas Abel Salazar (ICBAS) Universidade do Porto  Porto Portugal"},{"name":"Center for Drug Discovery and Innovative Medicines (MedInUP), Instituto de Ci\u00eancias Biom\u00e9dicas Abel Salazar (ICBAS) Universidade do Porto  Porto Portugal"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"M. 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