Stimulatory Effect of Nitric Oxide on Bicarbonate Secretion in Bullfrog Duodenums in vitro

The effect of nitric oxide (NO) on HCO–₃ secretion was examined in vitro using an isolated preparation of bullfrog duodenum. The tissue was bathed in unbuffered Ringer’s solution gassed with 100% O₂ on the mucosal side and HCO–₃ Ringer’s solution gassed with 95% O₂–5% CO₂ on the serosal side. The HCO–₃ secretion was measured by the pH-stat method using 2 mmol/l HCl as the titrant to keep the mucosal pH at 7.4. (±)-(E)-Ethyl-2-[(E)-hydroxyimino]-5-nitro-3-hexenamine (NOR3) was used as a NO donor and added to the serosal solution. To analyze the NOR3 action on HCO–₃ secretion, the effects of dibutyryl adenosine-3′,5′-cyclic monophosphate (dbcAMP), dibutyryl guanosine-3′,5′-cyclic monophosphate (dbcGMP), methylene blue, and indomethacin on the HCO–₃ response were also examined. NOR3 (1×10–⁴ and 3×10–⁴ mol/l) caused an increase in HCO–₃ secretion in a dose-dependent manner, and this effect appeared with an about 30-min time lag, reaching the level of 1.5–2.5 times greater than basal values at 1–2 h later. Both dbcAMP (1×10–³ mol/l) and dbcGMP (1×10–³ mol/l) also caused a significant increase in HCO–₃ secretion in bullfrog duodenums in vitro, although the onset of the HCO–₃ response to dbcGMP was delayed as compared to the former. The stimulatory action of NOR3 on duodenal HCO–₃ secretion was significantly attenuated by methylene blue (5×10–⁵ mol/l) and indomethacin (1×10–⁵ mol/l), the latter also inhibiting the HCO–₃ response to dbcGMP. The release of prostaglandin E₂ in the serosal solution was significantly increased after addition of NOR3 (3×10–⁴ mol/l) and dbcGMP (1×10–³ mol/l) in an indomethacin-sensitive manner. These results suggest that the NO donor increases duodenal HCO–₃ secretion in vitro, and this action of NO donor is cGMP-dependent and mediated by endogenous prostaglandins. Duodenal HCO–₃ secretion may be regulated locally by NO/cGMP in addition to prostaglandin/cAMP.