Nitric oxide (NO) and vasoactive intestinal polypeptide (VIP) interact in the regulation of neuromuscular function in the gut. They are also potent intestinal secretogogues that coexist in the enteric nervous system. The aims of this study were: (1) to investigate the interaction between NO and VIP in inducing fluid secretion in the rat jejunum, and (2) to determine whether the NO effect on intestinal fluid movement is neurally mediated. The single pass perfusion technique was used to study fluid movement in a 25 cm segment of rat jejunum in vivo. A solution containing 20 mM L-arginine, a NO precursor, was perfused into the segment. The effect of the NO synthase inhibitors (L-NAME and L-nitroindazole (L-NI)) and the VIP antagonist ([4Cl-D-Phe⁶,Leu¹⁷]VIP (VIPa)) on L-arginine-induced changes in fluid movement, expressed as µl min^-1 (g dry intestinal weight)^-1, was determined. In addition, the effect of neuronal blockade by tetrodotoxin (TTX) and ablation of the myenteric plexus by benzalkonium chloride (BAC) was studied. In parallel groups of rats, the effect of L-NAME and L-NI on VIP-induced intestinal fluid secretion was also examined. Basal fluid absorption in control rats was (median (interquartile range)) 65 (45-78). L-Arginine induced a significant fluid secretion (-14 (-20 to -5); P < 0.01). This effect was reversed completely by L-NAME (60 (36-65); P < 0.01) and L-NI (46 (39-75); P < 0.01) and partially by VIPa (37 (14-47); P < 0.01). TTX and BAC partially inhibited the effect of L-arginine (22 (15-32) and 15 (10-26), respectively; P < 0.05). The effect of VIP on fluid movement (-23 (-26 to -14)) was partially reversed by L-NAME (24 (8.4-35.5); P < 0.01) and L-NI (29 (4-44); P < 0.01). The inhibition of VIP or NO synthase prevented L-arginine- and VIP-induced intestinal fluid secretion through a neural mechanism. The data suggest that NO enhances the release of VIP from nerve terminals and vice versa. Subsequently, each potentiates the other's effect in inducing intestinal fluid secretion.