Role of plasma albumin in renal elimination of a mercapturic acid. Analyses in normal and mutant analbuminemic rats

Biosynthesis of N-acetylcysteine S-conjugates of xenobiotics (mercapturic acids) occurs via interorgan metabolism and the renal transtubular transport system plays an important role in elimination of the final metabolites from the organism. To assess the behavior of a mercapturic acid in the circulation, plasma clearance of radioactive S-benzyl-N-acetylcysteine and its interaction with plasma proteins were studied in normal and mutant analbuminemic rats (NAR). Intravenously injected S-benzyl-N-acetylcysteine rapidly disappeared from the circulation both in NAR and normal animals. However, its plasma clearance was significantly higher in NAR (45.7 ml kg-1 min-1) than in normal rats (25.2 ml kg-1 min-1). Ultrafiltration analysis revealed that 18.4% and 80.1% of the mercapturate bound to plasma protein(s) from NAR and normal rats, respectively, at 50 microM ligand concentration. The mercapturic acid bound to plasma albumin with an association constant of 2.24 X 10(5) M-1 and the number of binding sites was 1.18/mol albumin. The binding was competitively inhibited by probenecid and L-tryptophan. Concomitant administration of this mercapturic acid with equimolar amounts of albumin resulted in a marked decrease in the plasma clearance (26.2 ml kg-1 min-1) and an increase in the urinary secretion of this ligand in NAR. 30 min after injection of the mercapturic acid (10 mumol/kg body weight), 27.3% and 60.4% of the injected dose was recovered from urine and kidneys of NAR and normal rats respectively. About 41% of the dose was recovered in NAR urine when the ligand was injected bound to an equimolar amount of albumin. These results suggested that albumin is important for the renal accumulation and urinary elimination of the circulating mercapturic acid.     

The role of albumin in the hepatic transport of bilirubin: studies in mutant analbuminemic rats

Bilirubin and other cholephilic organic anions are bound to albumin in the circulation; their hepatic uptake involves a carrier-mediated process. To investigate the possible role of serum albumin in the transhepatic transport of a cholephilic ligand, plasma clearance of radioactive bilirubin and its biliary excretion as well as its interaction with plasma proteins were compared between normal and mutant analbuminemic rats (NAR). With a tracer amount of 3H-labeled bilirubin, its plasma clearance and biliary excretion were comparable in both animal groups. However, the plasma clearance of a loading dose of the ligand was significantly increased and its biliary recovery was low in NAR as compared with normal animals. In accord with these findings in vivo, gel permeation chromatographic analysis revealed that the bilirubin binding capacity of serum proteins was significantly lower in NAR than in control animals. When bilirubin was administered to NAR as a mixture with equimolar albumin, its plasma disappearance was considerably decreased and its biliary recovery was increased. Similar effects were observed when albumin was replaced by an equimolar amount of glutathione S-transferases (ligandins). These observations indicate that, although ligand-protein interaction in the circulation is important for directing bilirubin to the plasma membranes of the hepatocyte, this mechanism is not specific for albumin.     

Hepato-renal transport of phenolsulfonphthalein in analbuminemic rats: albumin is essential for hepatic compensatory elimination of a nephrophilic ligand in animals with renal dysfunction

To investigate a possible function of plasma albumin in partitioning organic anions into bile and urine, phenolsulfonphthalein (PSP) was administered intravenously and its in vivo fate was studied in normal and analbuminemic mutant rats (NAR). No significant change in the rate of PSP disappearance was observed in bilaterally nephrectomized normal rats. However, biliary excretion of the injected dye increased remarkably in nephrectomized normal rats. Intravenously injected PSP disappeared very rapidly from the circulation of NAR. Thus, the plasma clearance and distribution volume of PSP were significantly larger in NAR than in normal rats. Bilateral nephrectomy also failed to decrease the plasma clearance and distribution volume of the dye in NAR. In striking contrast to the experiments in normal rats, bilateral nephrectomy did not increase the biliary secretion of PSP in NAR. When PSP bound to equimolar albumin was injected into bilaterally nephrectomized NAR, the biliary excretion of PSP increased significantly with concomitant decrease in both plasma clearance and distribution volume of the dye. These results indicate that, in cases of renal transport dysfunction, albumin plays a critical role in hepatic compensatory excretion of PSP, a nephrophilic organic anion, whose molecular weight (MW 354) is close to the threshold value for partitioning a ligand to the eliminatory routes in liver and kidney of a rodent.     

Elevation of serum albumin concentration in analbuminemic rats by administration of 3'-methyl-4-dimethylaminoazobenzene

Albumin-like protein was detected in the serum of analbuminemic rats which have a mutation affecting albumin mRNA processing and genetically lack serum albumin. This protein was identified as rat serum albumin on the basis of molecular weight, immunological property and digestion patterns with proteases. The serum albumin level in analbuminemic rats was 5 μg/ml 4 weeks after birth and increased slightly during aging. By continuous administration of a hepatocarcinogen, 3′-methyl-4-dimethylaminoazobenzene, the level of serum albumin in the mutant rats increased over that of untreated rats and reached a maximum of 32 μg/ml at the 15th week of the carcinogen administration, whereas that of untreated rats was 16 μg/ml at the same time.     

Reduced activity of androgen biosynthesis in the testes of rats with analbuminemia

Steroid metabolism in Nagase Analbuminemia Rats (NAR), a mutant strain established from Sprague-Dawley rats, was studied. NAR are characterized by lack of serum albumin and hyperlipidemia. Total testosterone concentration in the serum of NAR was lower than that of normal rats, while the serum free testosterone, LH and FSH concentrations were similar. The half lives of 14C-labeled testosterone administered intravenously in NAR and normal Sprague-Dawley (SD) rats were 4.4 and 4.1 min, respectively. The plasma clearance rates of testosterone in NAR and normal rats were 34.7 and 39.1 ml/min per kg body weight. On Sephadex G-100 chromatography, a mixture of [3H]testosterone and normal rat serum gave two protein peaks eluted in the void volume and the albumin fraction, and the radioactivity was eluted all in the albumin fraction. In contrast, a mixture of [3H]testosterone and NAR serum gave a single protein peak eluted in the void volume and the radioactivity was mainly eluted with this protein peak. The association constants of testosterone to NAR and normal rat sera were 1.25 and 2.24 X 10(4) M-1. Enzyme activities related to the synthesis of testosterone by the testicular microsomal fractions of NAR and normal rats were examined. The activities of 3 beta-hydroxysteroid dehydrogenase, 5-ene-4-ene isomerase, 17 alpha-hydroxylase, C-17-C-20 lyase and 17 beta-hydroxysteroid dehydrogenase were lower in NAR than in normal rats. The activity for synthesis of testosterone from pregnenolone by the testicular microsomal fraction of NAR was about 40% of that of normal rats. These findings indicate that the low serum concentration of testosterone in NAR is mainly attributable to decreased biosynthesis of testosterone in the testes.     

Contribution of serum albumin to the transport of orally administered L-tryptophan into liver of rats with L-tryptophan depletion

Summary The role of serum albumin in the transport of orally administered L-tryptophan (Trp) into rat tissues was examined using analbuminemic and Sprague-Dawley (SD) rats with and without a-methyl-DL-tryptophan (AMT)-induced Trp depletion. Trp was orally administered to rats 16h after AMT or 0.85% NaCl administration, when liver tryptophan 2,3-dioxygenase and protein synthetic activities in AMT-treated rats were similar to those of 0.85% NaCl-treated rats. After oral Trp administration, regardless of the presence or absence of Trp depletion, free serum Trp concentrations were similar in the analbuminemic and SD rats, while total serum Trp concentrations were lower in analbuminemic rats than in SD rats. Although liver, brain, and muscle Trp concentrations after oral Trp administration under Trp depletion were lower in analbuminemic rats than in SD rats, the ratio of the liver Trp concentration in analbuminemic rats to that in SD rats was smaller than that of the brain or muscle Trp concentration. These results suggest that variations in serum albumin levels could affect the transport of orally administered Trp into the liver of rats with Trp depletion.     

Subcellular distribution of bile acids, bile salts, and taurocholate binding sites in rat liver

We have quantitated bile acids and their conjugates in rat liver using high-pressure liquid chromatography. Over 95% of the hepatic bile acid pool in rat liver homogenates is present as taurocholate and tauromuricholate. Although over 60% of the bile acid pool is recovered in the supernatant, evidence is presented suggesting that taurocholate redistributes among the subcellular fractions during their isolation. Taurocholate (TC) binding to purified subcellular fractions from rat liver was determined by using equilibrium dialysis in a TC concentration range from 0.1 to 100 microM. This is well below the critical micellar concentration of taurocholate (3 mM). All of the fractions investigated exhibited low-affinity binding with dissociation constants from 80 to 240 microM as did membrane lipid vesicles. Therefore, low-affinity binding appears referable to taurocholate nonspecifically partitioning into the lipid bilayer. High-affinity binding is present in plasma membranes, Golgi, and cell supernatant. The high-affinity binding sites in Golgi have a mean dissociation constant (A1) of 1.0 microM and bind 0.15 nmol of TC/mg of protein. Similarly, the high-affinity binding sites of plasma membrane have an A1 of 1.3 microM and bind 0.15 nmol of TC/mg of protein. For cell supernatant, the A1 was 4.8 microM, and 0.35 nmol of TC was bound per mg of protein. Mitochondria, smooth and rough microsomes, and Golgi liposomes showed no detectable amounts of high-affinity binding. These results are compatible with a role for the Golgi complex, cytoplasmic component(s), and plasma membranes in transhepatic bile acid transport.     

Biliary protein output by isolated perfused rat livers. Effects of bile salts.

The output of proteins into bile was studied by using isolated perfused rat livers. Replacement of rat blood with defined perfusion media deprived the liver of rat serum proteins (albumin, immunoglobulin A) and resulted in a rapid decline in the amounts of these proteins in bile. When bovine serum albumin was incorporated into the perfusion medium it appeared in bile within 20 min and the amount in the bile was determined by the concentration of the protein in the perfusion medium. The use of a defined perfusion medium also deprived the livers of bile salts and the amounts of these, and of plasma-membrane enzymes [5'-nucleotidase (EC 3.1.3.5) and phosphodiesterase I], in bile declined rapidly. Introduction of micelle-forming bile salts (taurocholate or glycodeoxycholate) to the perfusion medium 80 min after liver isolation markedly increased the output of plasma-membrane enzymes but had no effect on the other proteins. The magnitude of this response was dependent on the bile salt used and its concentration in bile; there was little effect on plasma-membrane enzyme output until the critical micellar concentration of the bile salt had been exceeded in the bile. A bile salt analogue, taurodehydrocholate, which does not form micelles, did not produce the enhanced output of plasma-membrane enzymes. This work supports the view that the output of plasma-membrane enzymes in bile is a consequence of bile salt output and also provides evidence for mechanisms by which serum proteins enter the bile.     

Characterization of the bile acid transport system in normal and transformed hepatocytes. Photoaffinity labeling of the taurocholate carrier protein

The taurocholate transport system in normal and transformed hepatocytes has been characterized using transport kinetics and photoaffinity labeling procedures. A photoreactive diazirine derivative of taurocholate, (7,7-azo-3 alpha,12 alpha-dihydroxy-5 beta-cholan-24-oyl)-2-amino [ 1,2-3H ]ethanesulfonic acid (7-ADTC), which has been shown to be a substrate for the bile acid carrier system, was photolyzed in the presence of intact hepatocytes, hepatoma tissue culture (HTC) cells, and plasma membranes derived from the hepatocyte sinusoidal surface. Irradiation of membranes in the presence of 7-ADTC resulted in the incorporation of the photoprobe into two proteins with Mr = 68,000 and 54,000. The specificity of labeling was confirmed by the significant inhibition of labeling observed when photolysis was carried out in the presence of taurocholate. The 68,000-Da protein was easily extracted with water and was shown to exhibit electrophoretic properties identical with rat serum albumin. The 54,000-Da protein required Triton X-100 for solubilization, indicating a strong association with the plasma membrane. Labeling of intact hepatocytes also resulted in specific labeling of the 54,000-Da protein. In contrast to hepatocytes, HTC cells derived from Morris hepatoma 7288C as well as H4-II-E cells derived from Reuber hepatoma H-35 exhibited a total loss of mediated bile acid uptake. Photolysis of 7-ADTC in the presence of HTC cells did not result in the labeling of any proteins, a result consistent with the loss of transport activity, and further supporting the specificity of the labeling reaction. The anion transport inhibitor N-(4-azido-2-nitrophenyl)-2-aminoethyl-[ 35S ]sulfonate, which has been shown to be a substrate for the bile acid carrier system also labeled the 54,000-Da plasma membrane protein when photolyzed in the presence of intact hepatocytes. These results suggest that the 54,000-Da protein is a component of the hepatocyte bile acid transport system and that the activity of this system is greatly reduced in several hepatoma cell lines.     

Expression of the hepatocyte Na+/bile acid cotransporter in Xenopus laevis oocytes

The expression of the basolateral Na+/bile acid (taurocholate) cotransport system of rat hepatocytes has been studied in Xenopus laevis oocytes. Injection of rat liver poly(A)+ RNA into the oocytes resulted in the functional expression of Na+ gradient stimulated taurocholate uptake within 3-5 days. This Na(+)-dependent portion of taurocholate uptake exhibited saturation kinetics (apparent Km approximately 91 microM) and could be inhibited by 4,4'-diisothiocyano-2,2'-disulfonic acid stilbene. Furthermore, the expressed taurocholate transport activity demonstrated similar substrate inhibition and stimulation by low concentrations of bovine serum albumin as the basolateral Na+/bile acid cotransport system previously characterized in intact liver, isolated hepatocytes, and isolated plasma membrane vesicles. Finally, a 1.5- to 3.0-kilobase size-class of mRNA could be identified that was sufficient to express the basolateral Na+/taurocholate uptake system in oocytes. These results demonstrate that "expression cloning" represents a promising approach to ultimately clone the gene and to further characterize the molecular properties of this important hepatocellular membrane transport system.     

Albumin catabolism in diabetic rats

The kinetics of albumin catabolism were studied in normal rats and rats with streptozotocin induced diabetes (blood glucose greater than 500 mg%). Whether determined from the clearance of 125I-albumin from plasma or from the whole body, after 10 days of severe, uncontrolled diabetes there was a 30-35% decrease in the catabolic rate for albumin in the diabetic rats compared to normals. There was also about a 35% contraction of the relative extravascular distribution volume for albumin in the diabetic rats, and about a 25% decrease in the total body mass of albumin. However, the concentration of albumin in the circulation was the same in normal and diabetic animals. We conclude that when the rate of albumin synthesis is substantially depressed in diabetes, the rat maintains normal plasma albumin concentration both by decreasing albumin's fractional catabolic rate and by shifting albumin from the extravascular to the vascular compartment.     

Abnormal renal response to twenty-four-hour dehydration and fasting in Nagase analbuminemic rats.

We assessed renal function in fasting adult Nagase analbuminemic rats (NAR). Sodium output in male and female NAR was 68% and 46%, respectively, of the output of age- and sex-matched normal Sprague-Dawley (SD) rats. Potassium excretion was significantly greater in female NAR but there was no difference between male NAR and SD rats. The renal clearances of urea and creatinine were reduced in NAR with corresponding increases in plasma concentrations; however, the urea and creatinine concentrations were not different in plasma samples taken from normally fed and hydrated SD and NAR rats. Exchangeable body sodium and sodium space was significantly larger in normally fed and hydrated NAR than in SD but there were no differences in plasma sodium concentrations or plasma volumes. Although plasma concentrations of albumin in NAR were only about 0.07% of the concentration in SD rats, the renal clearance of albumin in NAR was threefold greater. Kidney weights in NAR were 10 to 16% less than in SD rats but liver weights were 22 to 42% greater. Clearly, renal function was markedly abnormal in Nagase rats during a 24-hour fast.     

Receptor-mediated gene delivery in vivo. Partial correction of genetic analbuminemia in Nagase rats

A plasmid (palb3) was constructed containing the structural gene for human serum albumin driven by mouse albumin enhancer-rat albumin promoter elements. Using an asialoglycoprotein-polycation conjugate consisting of asialoorosomucoid coupled to poly-L-lysine, a soluble DNA complex was formed that was capable of targeting specifically to hepatocytes via asialoglycoprotein receptors present on these cells. Groups of Nagase analbuminemic rats were injected with complexed DNA or controls, followed by two-thirds partial hepatectomy to stimulate hepatocyte replication. Using a cDNA probe for the human albumin structural gene, hybridizable sequences were detected in analbuminemic rats treated with complex as determined by Southern blot analysis. Two weeks post-injection, the targeted DNA was found to exist primarily in plasmid form with an average copy number of 1000/diploid cell. Human albumin mRNA was detected by dot-blot hybridization with a specific oligonucleotide cDNA probe and confirmed by RNase protection assay using a vector-specific probe. Circulating human albumin was detected in the serum of palb3-treated Nagase analbuminemic rats by Western blots using an antibody specific for human serum albumin. A time course demonstrated that circulating human albumin was not detectable 24 h after injection, but became measurable at a level of 0.05 micrograms/ml within 48 h and increased in concentration to a maximum of 34 micrograms/ml by 2 weeks post-injection. This level of expression remained stable through 4 weeks after injection and partial hepatectomy.     

The role of transhepatic bile salt flux in the control of hepatic secretion of triacylglycerol-rich lipoproteins in vivo in rodents

Bile salts (BS) have been shown to suppress the secretion of very-low-density lipoprotein-triglyceride (VLDL-TG) in rat and human hepatocytes in vitro. In the present study, we investigated whether the transhepatic BS flux affects VLDL-TG concentration and hepatic VLDL-TG secretion in vivo. In rats, the transhepatic BS flux was quantitatively manipulated by 1-week chronic bile diversion (BD), followed by intraduodenal infusion with taurocholate (TC) or saline for 6 h. In mice, the transhepatic BS flux was manipulated by a 3-week dietary supplementation with TC (0.5 wt.%) or cholestyramine (2 wt.%). In rats, BD followed by saline or TC infusion did not affect plasma triacylglycerol (TG) concentration, hepatic TG production rate or VLDL lipid composition, compared to control rats. In mice supplemented for 3 weeks with TC or cholestyramine, the transhepatic BS flux was increased by 335% and decreased by 48%, respectively, compared to controls. Among the three experimental groups of mice, an inverse relationship between transhepatic BS flux and either plasma TG concentration (R(2)=0.89) or VLDL-TG production rate (R(2)=0.87) was observed, but differences were relatively small. Present data support the concept that BS can reduce VLDL-TG concentration and inhibit hepatic TG secretion in vivo; however, this occurs only at supraphysiological transhepatic BS fluxes in mice.     

Isolation and characterization of denatured serum albumin from rats with endotoxicosis

Due to its rapid breakdown in the body, denatured serum albumin has not been identified in biological samples. In this study we attempted to determine whether denatured albumin could be identified in rats with endotoxicosis. Male Wistar rats were injected with lipopolysaccharide (LPS; 5 mg/kg body weight). Plasma albumin concentration decreased to one-third the normal level at 2 days after the injection. By using the purified IgG against the specific epitope of chemically denatured albumin, two immunoreactive plasma proteins (bands D2 and D3) were identified by native PAGE followed by Western blot analysis. The plasma concentration of these two proteins increased significantly at 1 and 1.5 days after LPS injection. Peptide mass fingerprinting using matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI/TOF-MS) identified these two proteins as serum albumin. In order to characterize their conformational nature, ion-exchange chromatography was used to isolate D2 and D3 albumins from rats injected with LPS. Far- and near-UV circular dichroism (CD), tryptophan and 1-anilino-8-naphthalenesulfonate (ANS) fluorescence, and proteolytic susceptibility showed conformational alterations in the D2 and D3 albumins as compared with native albumin. These data indicate the presence of denatured albumin in circulating rat plasma, and this fact may contribute to a further understanding of the molecular mechanisms of albumin breakdown in physiological and pathophysiological conditions.     

Binding proteins for linear renin-inhibiting peptides in basolateral plasma membranes of rat liver.

A linear hydrophobic peptide, (Code no. EMD 55068), a synthetic renin-antagonist, competitively inhibits the uptake of taurocholate and of another linear peptide (EMD 51921) but not of oleic acid, serine or thiamin hydrochloride into isolated rat liver cells. EMD 55068 was attached to a gel matrix at a position that is not involved in the protein ligand interaction. The gel matrix used did not interact nonspecifically with solubilized proteins from rat liver. The quantity of bound ligand was determined to be 3.6 mg/ml of gel matrix. In the fraction of EDTA extracted hydrophilic membrane-associated proteins, no binding proteins were detected. Affinity chromatography of integral plasma membrane proteins resulted in four protein bands with molecular masses of 46, 49, 53 and 56 kDa in SDS-PAGE. In contrast, solubilized plasma membrane proteins from AS-30D ascites hepatoma cells, which are unable to transport bile acids and linear peptides, did not bind specifically to the affinity matrix.     

Taurine supplementation induces multidrug resistance protein 2 and bile salt export pump expression in rats and prevents endotoxin-induced cholestasis

The effect of oral taurine supplementation on endotoxin-induced cholestasis was investigated in rat liver. At 12h following lipopolysaccharide (LPS) injection (4mg/kg body weight i.p.) bile flow and bromosulfophthalein (BSP) and taurocholate (TC) excretion were determined in the perfused liver and the expression of the canalicular transporters multidrug resistance protein 2 (Mrp2) and bile salt export pump (Bsep) was analyzed. Injection of LPS induced a significant decrease of bile flow ( 2.2+/-0.2 microl/g liver wet weight/min vs 3.3+/-0.1 microl/g liver wet weight in controls), biliary BSP excretion (10.8+/-2.2 nmol/g/min vs 21.0+/-3.8 nmol/g/min), and biliary TC excretion (114+/-23 nmol/g/min vs 228+/-8 nmol/g/min). These effects were due to transporter retrieval from the canalicular membrane and downregulation of Mrp2 and Bsep expression. In taurine-supplemented rats bile flow was 30% higher than that in untreated rats and the expression of Mrp2 and Bsep protein was increased two- to threefold. In taurine-supplemented rats there was no significant reduction of bile flow or of BSP and TC excretion at 12h following LPS injection. This protective effect of taurine was due to higher Mrp2 and Bsep protein levels compared to nonsupplemented LPS-treated rats, whereas relative Mrp2 retrieval from the canalicular membrane induced by LPS was not significantly different. LPS-induced tumor necrosis factor alpha and interleukin-1beta release were lower in taurine-fed rats; however, downregulation of Mrp2 and Bsep expression by LPS was delayed but not prevented. The data show that oral supplementation of taurine induces Mrp2 and Bsep expression and may prevent LPS-induced cholestasis.     

Effects of cysteine on the pharmacokinetics and pharmacodynamics of intravenous and oral azosemide in rats with protein-calorie malnutrition

The effects of cysteine on the pharmacokinetics and pharmacodynamics of azosemide were investigated after intravenous (10 mg/kg) and oral (20 mg/kg) administration to male Sprague-Dawley rats fed on 23% protein diet (control rats), and 5% protein diet with (rats with PCMC) or without (rats with PCM) oral cysteine (250 mg/kg, twice daily for the fourth week) for 4 weeks. After intravenous administration to rats with PCMC, some pharmacokinetic parameters restored fully or more than the level of control rats; the time-averaged nonrenal clearance (2.70 versus 2.32 ml/min/kg) and apparent volume of distribution at steady state (160 versus 189 ml/kg) were comparable to those in control rats, however, the terminal half-life (34.7 versus 57.2 min) and mean residence time (73.3 versus 99.3 min) were significantly shorter, area under the plasma concentration-time curve from time zero to time infinity (AUC, 1930 versus 2680 microg min/ml) was significantly smaller, and time-averaged renal (2.24 versus 1.21 ml/min/kg) and total body (CL, 4.98 versus 3.65 ml/min/kg) clearances were significantly faster than those in control rats. This could be mainly due to significantly faster renal clearance and at least partly due to increased cytochrome P450 1A2 activity by cysteine supplementation. After intravenous administration to rats with PCMC, the total amount of 8-hr urinary excretion of unchanged azosemide was significantly greater (457 versus 305 microg/g body weight), however, the 8-hr urine output (15.3 versus 31.1 ml/g kidney) was not significantly different between control rats and rats with PCMC. This could be due to the fact that urine output seemed to reach an upper plateau from 10 mg/kg dose of azosemide in rats.     

Pharmacokinetics of intravenous theophylline in mutant Nagase analbuminemic rats

It was obtained from our laboratories that the expression of hepatic microsomal cytochrome P450 (CYP) 1A2 increased approximately 3.5 times in mutant Nagase analbuminemic rats (NARs, an animal model for human familial analbuminemia), and theophylline was reported to be metabolized to 1,3-dimethyluric acid (1,3-DMU) and 1-methylxanthine (which was further metabolized to 1-methyluric acid, 1-MU, via xanthine oxidase) via CYP1A2 in rats. Hence, the pharmacokinetic parameters of theophylline, 1,3-DMU and 1-MU were compared after intravenous administration of aminophylline, 5 mg/kg as theophylline, to control Sprague-Dawley rats and NARs. In NARs, the total area under the plasma concentration-time curve from time zero to time infinity (AUC) of theophylline was significantly smaller (1,040 versus 1,750 microg min/ml) than that in control rats and this could be due to significantly faster renal clearance (CL(R), 1.39 versus 0.571 ml/min/kg, due to inhibition of renal reabsorption of unchanged theophylline) and nonrenal clearance (CL(NR), 3.36 versus 2.25 ml/min/kg, due to 3.5-fold increase in CYP1A2) than those in control rats. Based on in vitro hepatic microsomal studies, the intrinsic 1,3-DMU formation clearance was significantly faster in NARs than that in control rats (267 versus 180 x 10(-6) ml/min). After intravenous administration of 1,3-DMU, the renal secretion of 1,3-DMU was inhibited in NARs. Inhibition of renal secretion or reabsorption of various compounds in NARs was also discussed.     

Biliary transport maximum of tauroursodeoxycholate is twice as high as that of taurocholate in the rat

Biliary excretory transport maximum (Tm) and choleretic efficiency were compared for tauroursodeoxycholate and taurocholate in Wistar male rats. Under a continuous iv infusion of bile salts with a stepwise increase in infusion rate, the Tm of tauroursodeoxycholate was found to be two times higher (2.25±0.07 μmole/min/100 g body weight, n=8, mean±SD) than that of taurocholate (0.97±0.05 μmole/min/100 g body weight, n=14, p<0.001). On the other hand, the amount of bile water obligated by the excretion of 1μmole of tauroursodeoxycholate was significantly lower than that of taurocholate. (4.71±0.08 μl/μmole for tauroursodeoxycholate, vs. 9.27±0.76 μl/μmole for taurocholate, mean±SD, p<0.001). It was concluded that tauroursodeoxycholate can be excreted into the bile in male rats twice as efficiently as taurocholate. Furthermore, the higher efficiency in the choleretic property of ursodeoxycholate previously reported by Dumont et al. appears to be specific to free ursodeoxycholate and not to its taurine conjugate used in the present experiment.