Interplay between nitric oxide and vasoactive intestinal polypeptide in inducing fluid secretion in rat jejunum

Abstract

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 microl 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.

Figure 1. Effect of different concentrations of l-NAME (n = 5 in each group) on rat jejunal fluid secretion induced by 20 mml-arginine (l-Arg; n = 10)

All solutions were perfused in the rat jejunum. Results are expressed as medians (horizontal lines) and interquartile ranges (boxes); positive values denote absorption and negative values denote secretion. P < 0.005, Kruskall-Wallis test; * P < 0.01 compared to control; † P < 0.01 compared to l-arginine and to control; ‡ P < 0.01 compared to l-arginine and P > 0.05 compared to control.

Figure 2. Effect of different concentrations of l-NAME (n = 5 in each group) on rat jejunal basal fluid movement (control; n = 11)

Solutions were perfused in the rat jejunum. Results are expressed as medians and interquartile ranges; positive values denote absorption and negative values denote secretion. P < 0.005, Kruskall-Wallis test; * P < 0.01 compared to control.

Figure 3. Effect of neuronal NOS inhibition and VIP antagonism on rat jejunal basal fluid movement and l-arginine-induced secretion

l-Nitroindazole (NI; 50 mg kg⁻¹) was given I.P. and the VIP antagonist [4Cl-D-Phe⁶,Leu¹⁷]VIP (VIPa; 2 μg kg⁻¹ min⁻¹) was given I.V. The intestine was then perfused with a plasma electrolyte solution with or without 20 mml-arginine (l-Arg) (n = 5 in each group). Results are expressed as medians and interquartile ranges; positive values denote absorption and negative values denote secretion. * P < 0.01 compared to l-Arg and P > 0.05 compared to control + l-NI. † P < 0.01 compared to l-Arg and P < 0.05 compared to control + VIPa.

Figure 4. Effect of tetrodotoxin (TTX) and benzylalkonium chloride (BAC) treatment on rat jejunal basal fluid movement and on l-arginine-induced secretion

TTX (0.2 μm) was added to the intestinal perfusate. BAC treatment of the jejunal segment was performed 2–3 weeks prior to the intestinal perfusion. The intestine was perfused with a plasma electrolyte solution with or without 20 mml-arginine (l-Arg; n = 5 in each group). Results are expressed as medians and interquartile ranges; positive values denote absorption and negative values denote secretion. * P < 0.05 compared to TTX control and P < 0.05 compared to l-Arg. † P < 0.05 compared to BAC control and P < 0.01 compared to sham + l-Arg.

Figure 5. Effect of l-NAME on rat jejunal fluid secretion induced by VIP

VIP (0.6 μg kg⁻¹ min⁻¹) was given I.V. The jejunal segment was perfused with either a plasma electrolyte solution alone (n = 7) or after addition of different concentrations of l-NAME (n = 5–6 in each group). Results are expressed as medians and interquartile ranges; positive values denote absorption and negative values denote secretion. P < 0.005, Kruskall-Wallis test. * P < 0.01 compared to control; † P < 0.01 compared to VIP and P < 0.01 compared to control; ‡ P < 0.05 compared to VIP and P < 0.01 compared to control; § P > 0.05 compared to VIP and P < 0.01 compared to control.

Figure 6. Effect of neuronal NOS inhibition on VIP-induced fluid secretion in rat jejunum

l-Nitroindazole (l-NI) (50 mg kg⁻¹) was given I.P. (n = 5) and VIP was administered I.V. (0.6 μg kg⁻¹ min⁻¹). The jejunal segment was perfused with a plasma electrolyte solution. Results are expressed as medians and interquartile ranges; positive values denote absorption and negative values denote secretion. * P < 0.01 compared to control; † P < 0.01 compared to VIP and P < 0.01 compared to control.