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BACKGROUND AND PURPOSEAdenosine A3 receptors mediate beneficial effects in experimental colitis, but their involvement in enteric neuromuscular functions during bowel inflammation is undetermined. This study investigated the regulatory role of A3 receptors on colonic motility in the presence of experimental colitis. EXPERIMENTAL APPROACHColitis was induced in rats by 2,4-dinitrobenzenesulfonic acid. A3 receptors and adenosine deaminase (ADA, adenosine catabolic enzyme) mRNA were examined by RT-PCR. Tissue distribution of A3 receptors was detected by confocal immunofluorescence. The effects of 2,3-ethyl-4,5-dipropyl-6-phenylpyridine-3-thiocarboxylate-5-carboxylate (MRS1523) (MRS, A3 receptor antagonist), 2-chloro-N6-(3-iodobenzyl)-adenosine-5′-N-methyluronamide (2Cl-IB-MECA) (CIB, A3 receptor agonist), dipyridamole (DIP, adenosine transport inhibitor) and ADA were assayed on contractile responses evoked by electrical stimulation (ES) or carbachol in colonic longitudinal muscle preparations (LMP). KEY RESULTSRT-PCR showed A3 receptors and ADA mRNA in normal colon and their increased level in inflamed tissues. Immunofluorescence showed a predominant distribution of A3 receptors in normal myenteric ganglia and an increased density during colitis. MRS enhanced ES-induced cholinergic contractions in normal LMP, but was less effective in inflamed tissues. After pretreatment with dipyridamole plus ADA, to reduce extracellular adenosine, CIB decreased cholinergic motor responses of normal LMP to ES, with enhanced efficacy in inflamed LMP. A3 receptor ligands did not affect carbachol-induced contractions in LMP from normal or inflamed colon. CONCLUSIONS AND IMPLICATIONSNormally, adenosine modulated colonic cholinergic motility via activation of A3 receptors in the myenteric plexus. A3 receptor-mediated tonic inhibitory control by adenosine was impaired in inflamed bowel, despite increased density of functioning and pharmacologically recruitable A3 receptors. Abbreviations

Adenosine plays an important role in regulating intestinal motility and inflammatory processes. Previous studies in rodent models have demonstrated that adenosine metabolism and signalling are altered during chronic intestinal inflammatory diseases. However, the involvement of the adenosinergic system in the pathophysiology of gut dysmotility associated to a primary neurodysfunction is still unclear. Recently, we showed that the neurotropic Herpes simplex virus type-1 (HSV-1), orally inoculated to rodents, infects the rat enteric nervous system (ENS) and affects gut motor function without signs of systemic infection. In this study we examined whether changes in purinergic metabolism and signaling occur during permanent HSV-1 infection of rat ENS. Using isolated organ bath assays, we found that contraction mediated by adenosine engagement of A1 or A2A receptors was impaired at 1 and 6 weeks post-viral administration. Immunofluorescence studies revealed that viral infection of ENS led to a marked redistribution of adenosine receptors: A1 and A2B receptors were confined to the muscle layers whereas A2A and A3 receptors were expressed mainly in the myenteric plexus. Viral-induced ENS neurodysfunction influenced adenosine metabolism by increasing adenosine deaminase and CD73 levels in longitudinal muscle-myenteric plexus with no sign of frank inflammation. This study provides the first evidence for involvement of the adenosinergic system during HSV-1 infection of the ENS. As such, this may represent a valid therapeutic target for modulating gut contractility associated to a primary neurodysfunction.

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