This compound, which corresponds to

the 337.25 m/z band (Fig. 3B), exhibited a modest increment upon UDCA infusion. The instability of GSNO under MS conditions might explain why this band is not predominant in the spectrum. However, FK228 in vitro as shown in Fig. 3B, the relative intensity of a 319.24 m/z band (seemingly corresponding to dehydrated GSNO) was manifestly higher in UDCA-stimulated bile versus basal bile. These data support the concept that UDCA infusion induces an increase of GSNO in bile. We also assessed the involvement of glutathione in the transport of NO to bile by determining biliary NO in rats after depleting their livers of glutathione with BSO. As we previously reported,26 UDCA increased hepatic glutathione levels in normal rats (Fig. 4A). However, in rats that received BSO, liver glutathione was markedly reduced, regardless of UDCA administration (Fig. 4A). An analysis of UDCA in bile from UDCA-infused normal rats and BSO-treated rats showed that biliary UDCA secretion was similar in both situations (Supporting Fig. 2), and this indicates

that the secretion of UDCA to bile is not prevented in the absence of glutathione. In contrast, the secretion of NO species after UDCA infusion does depend on glutathione, as it was virtually abolished in BSO-treated animals (Fig. 4B), even though their hepatic NOS activity was increased IWR-1 to levels similar to those found in UDCA-infused normal rats (data not shown). These findings are consistent with the notion that glutathione has a major role as a carrier for the transport of NO to bile. Glutathione and glutathione conjugates are known to be secreted at the canaliculi through the ABCC/Mrp2 pump. Therefore, we performed UDCA infusion experiments in TR− rats, which exhibit defective canalicular transport of those

compounds because of an ABCC2 mutation.27 In these animals, the levels of biliary glutathione fall 3 logs with respect to normal values, but the compound is still secreted to bile in the micromolar range.28 As shown in Fig. 5A,B, UDCA-infused TR− rats exhibited a significant decrease in both the concentration and biliary output of NO species in comparison with UDCA-infused normal rats. The increment in biliary NO secretion upon UDCA infusion in TR− rats was less than half of that observed in normal animals 上海皓元 (P < 0.05; see the inset in Fig. 5B). In the mutant rats, the levels of both total SNOs and LMw-SNOs increased after UDCA administration, but the values were about one-third of those observed in UDCA-treated normal rats (Supporting Fig. 3). These findings indicate that the glutathione carrier ABCC2/Mrp2 contributes at least partially to biliary NO secretion and provide further support for a role of glutathione as a vehicle for the transport of NO along the biliary tree. To determine whether GSNO could play a role in stimulating ductal secretion in vivo, we performed a retrograde infusion of 150 μL of 250 μM GSNO through the common bile duct in the isPRL model.