Biliary excretion of copper,manganese, and horseradish peroxidase in eisai hyperbilirubinemic mutant rats (EHBRs) with defective biliary excretion of glutathione

A mixture of copper (Cu) (0.38 mg/kg), manganese (Mn) (0.038 mg/kg), and horseradish peroxidase (HRP) (5.0 mg/kg) was injected intravenously (iv) into mature Eisai hyperbilirubinemic rats (EHBRs) and Sprague-Dawley rats (SDRs). Bile was collected at 10-min intervals before and after the injection, under anesthesia. The liver, kidneys, and blood were removed 40 min after the injection. The serum-conjugated bilirubin concentration was 0.85 mg/dL in the EHBRs, but was below detection limits in the SDRs. The bile-reduced glutathione (GSH) concentration was much lower in the EHBRs (0.04 mg/mL) than in the SDRs (1.30 mg/mL). However, the hepatic GSH concentration was about 1.6 times higher in EHBRs (2.26 mg/g liver) than in SDRs (1.43 mg/g liver). The low, excretion of biliary GSH was not caused by the activity of GGT in the liver, since there was no significant difference in the activity between the two groups (5.8±3.4 and 4.6±2.4 μmol p-nitroaniline/g protein/30 min in SDR and EHBR groups, respectively). There was a delay of initial biliary excretion of Cu in EHBRs compared to SDRs. The biliary concentration of Mn was slightly lower in EHBRs than in SDRs. Forty min after the injection of metals, however, there was no difference between hepatic concentrations of the two metals in the two groups. Our results suggest that abnormal deposition of the two metals is not observed naturally in EHBRs. Injected HRP was excreted rapidly and notably in the EHBRs compared to SDRs. Furthermore, the biliary concentration of β-N-acetyl-D-glucosaminidase (β-NAG) was significantly higher in EHBRs than in SDRs. Rapid biliary excretion of Cu, but not of Mn, may be related to the hepatobiliary transport of GSH, but the transport and lysosomal function do not originally regulate the biliary excretion of Cu.