Melatonin is released from enterochromaffin cells in the intestinal mucosa, but the influence of melatonin on gastrointestinal function is still largely unknown. Recently we found that administration of melatonin decreases basal duodenal mucosal paracellular permeability and increases duodenal bicarbonate secretion (DBS). The aim of the present study was to elucidate the acute and long-term effects of melatonin administration on ethanol-induced alterations of duodenal mucosal paracellular permeability and motility. In this study male Sprague-Dawley (SD) rats (n=44; 250-375 g) were used. For evaluation of long-term effects of melatonin administration rats were given melatonin (0.1 mg/ml), or vehicle, in the drinking water for 14 days, while the other rats were given normal tap water. Rats were anaesthetized with thiobutabarbital sodium (Inactin®) 120 mg/kg body weight intraperitoneally. The surgical and experimental protocol and methodology used have been described previously [1]. A brief summary is given here. A laparatomy was performed and a ~30 mm segment of the proximal duodenum, with an intact blood supply, was perfused in situ with saline for basal recordings followed by 15% ethanol (v/v) for 30 min. Effects on duodenal mucosal permeability [blood-to-lumen clearance of 51Cr-EDTA] and motility were investigated. Values are mean ± SEM, compared by 1-way ANOVA or paired t test when appropriate. Luminal perfusion with 15% ethanol induced significant (p<0.05) increases in mucosal paracellular permeability and duodenal motility from 0.18 ± 0.06 to 1.67 ± 0.29 ml min-1100g-1 (n=9) and from 537 ± 46 to 1209 ± 148 AUC 10 min-1 (n=9) respectively. Pre-treating animals with an intravenous (IV) injection of melatonin 20 mg kg-1 10 min before ethanol exposure reduced the ethanol-induced increase in 51Cr-EDTA clearance to 0.65 ± 0.23 ml min-1 100g-1, p<0.05, n=6). In animals pretreated with 20 mgkg-1 melatonin IV 15% ethanol failed to augment duodenal motility. Hexamethonium (10 mgkg-1 IV followed by continuous IV infusion at a rate of 10 mg kg-1h-1, n=9) abolished the effects of IV melatonin. In animals administered melatonin for 14 days 15% ethanol induced significantly (p<0.05) less increases in mucosal paracellular permeability compared to their corresponding vehicle (from 0.12 ± 0.02 to 2.30 ± 0.46 mlmin-1100g-1 (n=4) compared to 0.18 ± 0.06 to 2.94 ± 0.42 mlmin-1100g-1 (n=6) respectively). No increases in duodenal motility in response to 15% ethanol was observed in melatonin treated animals. Our results show that melatonin reduces ethanol-induced increases in duodenal paracellular permeability partly via enteric neural pathways involving nicotinic receptors. In addition, melatonin inhibits ethanol-induced increases in duodenal motor activity. These results suggest that melatonin may serve important gastrointestinal barrier functions.