Mechanisms of impaired biliary excretion of acetaminophen glucuronide after acute phenobarbital treatment or phenobarbital pretreatment

Previous studies have demonstrated that phenobarbital (PB) significantly impairs the biliary excretion of acetaminophen glucuronide (AG) in rats. Studies also suggested that Mrp2 mediates AG biliary excretion, and Mrp3 is involved in AG basolateral export. It was hypothesized that inhibition of Mrp2-mediated AG transport by PB or PB metabolites, and PB induction of Mrp3, may contribute to the impaired biliary excretion of AG by PB. In the present study, the hepatobiliary transport of AG in single-pass isolated perfused Wistar and TR(-) rat livers was investigated. The AG biliary clearance was markedly decreased, and the AG basolateral clearance was significantly increased in TR(-) rat livers. Uptake of AG by Mrp2 and Mrp3, and inhibition of Mrp2- and Mrp3-mediated transport by PB and major PB metabolites, were investigated with rat Mrp2- or Mrp3-expressing Sf9 cell plasma membrane vesicles (Sf9-PMVs). AG was transported by Mrp3 (K(m) approximately 0.91 mM). Net ATP-dependent AG uptake into Mrp2-expressing Sf9-PMVs could not be detected directly. However, AG significantly inhibited Mrp2-mediated 5-(and 6)-carboxy-2',7'-dichlorofluorescein (CDF) transport. p-Hydroxyphenobarbital glucuronide (p-OHPBG), but not PB or p-hydroxyphenobarbital, significantly inhibited Mrp2-mediated CDF transport. The IC(50) values for p-OHPBG inhibition of Mrp2-mediated CDF uptake and Mrp3-mediated AG transport were similar (approximately 0.68 and 0.46 mM, respectively). PB treatment (80 mg/kg/day x 4 days) markedly increased hepatic Mrp3 expression in Wistar rats. In conclusion, inhibition of Mrp2-mediated AG transport by p-OHPBG provided one possible explanation for the impaired biliary excretion of AG after acute PB treatment. However, impaired biliary excretion of AG after PB pretreatment may be attributed primarily to the induction of hepatic Mrp3 by PB.