Hepatic uptake and biliary excretion of organic cations--I. Characterization of three new model compounds

Three quaternary ammonium compounds (QACs) with different lipophilicity, triethylmethyl ammonium iodide (TEMA), tripropylmethyl ammonium iodide (TPMA) and tri-n-butylmethyl ammonium iodide (TBuMA) were given as a bolus injection of 10 mumole and 1 mumole in an isolated perfused liver. TPMA and TBuMA exhibited saturation kinetics at a dose of 10 mumole, but not when 1 mumole of the agents was given. Biliary clearance of TEMA was equal to the bile flow (0.010 ml/min), whereas for TPMA and TBuMA much higher values of 0.8 ml/min and 2.2 ml/min were found respectively. Partition coefficients of TEMA, TPMA and TBuMA between n-octanol and Krebsbicarbonate solution were 0.0013, 0.013 and 0.14 respectively. Liver-to-plasma concentration ratios were 4, 16 and 30 in the post-distribution phase, whereas bile-to-liver ratios were calculated to be 0.1, 1.3 and 14 respectively. The latter parameter varied roughly proportionally to the lipophilicity of the compounds. The liver/plasma concentration ratios corrected for intracellular binding exceeded a value of 12 indicating that accumulation in the liver of these agents cannot soley be explained by passive equilibration according to the membrane potential. Transport from liver into the bile of TPMA and TBuMA presumably also occurred against an electrochemical gradient. It was inferred that the small molecular weight compounds such as TEMA, can be transported from plasma into bile paracellularly by a passive process. Rapid uptake into the liver of such compounds may not lead to an appreciable biliary output and can even reduce the rate of biliary excretion. QACs with intermediate or high lipophilicity are transported by carrier mediated processes both at the level of hepatocyte uptake and bile canalicular transport. The influence of choleresis on hepato-biliary transport of the three QACs was investigated by giving sodium taurocholate (Tc) by constant infusion of 60 mumole/hr, increasing bile flow from 9 to 16 microliter/min. The biliary output of TEMA appeared to be basically unaffected, whereas the biliary excretion of TPMA and TBuMA was clearly elevated when the bile flow was increased. The stimulatory influence of taurocholate on the biliary output of the latter organic cations is explained by an increased net uptake of these agents into the liver and an increased net canalicular transport. This effect is proposed to be due to a reduced reabsorption from the biliary tree as a consequence of the higher bile flow and/or biliary micelle binding. Taurocholate increased liver-to-plasma ratios.(ABSTRACT TRUNCATED AT 400 WORDS)     

Pharmacokinetics of steroidal muscle relaxants in isolated perfused rat liver.

Both in humans and animals hepatic elimination is an important factor determining the duration of action of non-depolarizing neuromuscular blocking drugs. To elucidate the hepato-biliary disposition of muscle relaxants the pharmacokinetics of several structurally related but physicochemically distinct steroidal neuromuscular blocking drugs were studied in isolated perfused rat livers. Pharmacokinetics analysis with the DIFFIT computer program enabled the simultaneous fitting of independently measured perfusate disappearance and biliary excretion rate curves using a numerical approach. The hepatic disposition of the steroidal muscle relaxants could be adequately described by a three compartment model with elimination from the peripheral compartment V2 (biliary excretion) and storage in a deep compartment (V3) connected to V2. In addition, for vecuronium only slow ester hydrolysis occurring in V2 and V3 was included in the model. The lipophilicity rather than the relative mobility of the muscle relaxants showed a positive relationship with biliary clearance (Cl20) and the initial hepatic uptake (Cl12), indicating that hepato-biliary transport of these organic cations is highly dependent on the hydrophobic character of the compounds. In addition, net hepatic uptake of the steroidal cations was influenced markedly by transport from the liver to perfusate (hepatic efflux). This hepatic efflux (k21) decreased with increasing lipophilicity. In contrast, the extent of intracellular sequestration into deep compartments, indicated by high k23/k32 ratios, seemed to be inversely related to the lipophilicity of the muscle relaxants and might explain the observed prolonged hepatic storage of some of these compounds. In combination with data from subfractionation studies the results indicate that the pharmacokinetic analysis of the hepatic disposition of steroidal muscle relaxants may be used to evaluate actual transport phenomena participating in the hepatic disposition of these drugs.     

PRIMARY ACTIVE TRANSPORT OF PRAVASTATIN ACROSS THE LIVER CANALICULAR MEMBRANE IN NORMAL AND MUTANT EISAI HYPERBILIRUBINEMIC RATS

We have previously demonstrated that the HMG-CoA reductase inhibitor pravastatin is efficiently taken up by the liver via the ‘multispecific anion transporter’ in an active manner.³ To further examine the fate of pravastatin within the liver, its biliary excretion was studied in a single-pass liver perfusion system and isolated liver canalicular membrane vesicles (CMVs) using normal (Sprague–Dawley rats; SDRs) and mutant Eisai hyperbilirubinemic rats (EHBRs). In the liver perfusion experiments, the outflowing drug concentration reached a steady state at 30 min and the extraction ratio was approximately 0·7 in both rat strains. Both the steady-state biliary excretion rate and bile flow rate of the EHBR group were 40% those of SDRs. At steady state, the fraction of unchanged drug in bile was 25–34% in both groups. The concentration ratios of unbound drug in cytosol versus that in sinusoid and in bile versus cytosol were, respectively, 11 and 87 in SDRs, and 13 and 94 in EHBRs. After correction for membrane potential (−40 mV in cytosol), the ratios became 49 and 19 in SDRs and 58 and 21 in EHBRs, respectively. The finding that all of these values were much larger than unity suggested that active transport occurred from liver to bile, as well as from plasma to liver, in both rat strains. Furthermore, ATP-dependent uptake of pravastatin was clearly observed in CMVs prepared from EHBRs as well as SDRs, whereas the stimulation by ATP of DNP-SG transport in CMV was observed only in SDRs. It was concluded that pravastatin is excreted into bile in high concentrations and a primary active transport mechanism which is maintained in EHBRs contributes to the biliary excretion of this drug.     

PRIMARY ACTIVE TRANSPORT OF PROVASTATIN ACROSS THE LIVER CANALICULAR MEMBRANE IN NORMAL AND MUTANT EISAI HYPERBILIRUBINAEMIC RATS

We have previously demonstrated that the HMG-CoA reductase inhibitor pravastatin is efficiently taken up by the liver via the ‘multispecific anion transporter’ in an active manner.³ To further examine the fate of pravastatin within the liver, its biliary excretion was studied in a single-pass liver perfusion system and isolated liver canalicular membrane vesicles (CMVs) using normal (Sprague–Dawley rats; SDRs) and mutant Eisai hyperbilirubinaemic rats (EHBRs). In the liver perfusion experiments, the outflowing drug concentration reached a steady state at 30 min and the extraction ratio was approximately 0·7 in both rat strains. Both the steady state biliary excretion rate and bile flow rate of the EHBR group were 40% of those of SDRS. At steady state, the fraction of unchanged drug in bile was 25–34% in both groups. The concentration ratios of unbound drug in cytosol versus that in sinusoid and of that in bile versus that in cytosol were, respectively, 11 and 87 in SDRs, and 13 and 94 in EHBRs. After correction for the membrane potential (−40 mV in cytosol), the ratios became 49 and 19 in SDR and 58 and 21 in EHBRs, respectively. The finding that all of these values were much larger than unity suggested that active transport occurred from liver to bile, as well as from plasma to liver, in both rat strains. Furthermore, ATP-dependent uptake of pravastatin was clearly observed in CMVs prepared from EHBRs as well as SDRs, whereas the stimulation by ATP of DNP-SG transport in CMVs was observed only in SDRs. It was concluded that pravastatin is excreted into bile in high concentrations and a primary active transport mechanism which is maintained in EHBRs contributes to the biliary excretion of this drug.     

On the pharmacology of Org 6338 (2beta, 16beta-dipiperidino - 5alpha - androstan - 3alpha - ol acetate dimethobromide), a new steroidal neuromuscular blocking agent.

Org 6368 is a homologue of pancuronium bromide. Its interactions with other agents in the cat sciatic nerve-gastrocnemius muscle preparation revealed that paralysis was of the non-depolarizing type. This was confirmed in experiments using avian muscle. Org 6368 is a potent muscle relaxant being 2-4 times as potent as (+)-tubocurarine in the cat. Paralysis in the cat is rapid in onset and of appreciably shorter duration than that of pancuronium and (+)-tubocurarine. Repeated injections of the same dose of Org 6368 show no cumulative effect. Muscle relaxant doses generally cause a slight increase in both blood pressure and heart rate. Although its histamine-releasing capacity is greater than that of pancuronium it is less than that of (+)-tubocurarine. Org 6368 shares with pancuronium a very weak effect on both the muscarinic receptor and ganglionic transmission. Differences in the muscle relaxant profiles of Org 6368 and pancuronium are discussed.     

Pharmacokinetics of the hepatic transport of organic anions: Influence of extra- and intracelluar binding on hepatic storage of dibromosulfophthalein and interactions with indocyanine green

The influence of intracellular and extracellular protein binding on the hepatic storage and biliary elimination of dibromosulfophthalein (DBSP) was studied in isolated perfused rat liver. Under first order kinetic conditions the amount of DBSP in the liver at a given plasma concentration (hepatic storage) was determined by extracellular binding to albumin and intracellular binding to the cytosolic Y and Z proteins as well as concentrative membrane transport from plasma into the liver. At higher doses, extensive binding of DBSP to intracellular organelles also occurred while liver cytosol/plasma concentration gradients of unbound DBSP were much lower. Hepatic storage increased with decreasing albumin concentration in the perfusate of isolated perfused rat livers. However, it was shown that this parameter is dose-dependent, and errors can be introduced in its calculation if nonlinearity of sinusoidal and canalicular transport processes as well as nonlinear protein binding are not taken into account. The influence of another organic anion, indocyanine green (ICG) on the hepatic storage, subcellular distribution, and elimination of DBSP was subsequently studied. At equimolar amounts the presence of ICG resulted in a 50% decrease in hepatic clearance and hepatic distribution volume of DBSP. It was inferred that these changes are due to an inhibition of carrier-mediated transport across the sinusoidal and canalicular membrane and preferential displacement from intracellular binding sites. In contrast DBSP in equimolar amount enhanced the initial disappearance rate and biliary excretion of ICG, probably due to increasing its free fraction in plasma. It is concluded that the level and mechanism of interaction of two drugs within the eliminating organ can be characterized by combining clearance studies with data on subcellular and extracellular binding.     

A new method for studying the relationship between hepatic uptake of drugs and their pharmacodynamic effects in anaesthetized cats.

1 A new in vivo experimental method is described whereby the liver can be temporarily excluded from the general circulation by means of a portocaval shunt operation. The influence of this manoeuvre upon the effects of pancuronium and Org 6368 was investigated using the tibialis muscle preparation of anaesthetized cats. 2 The procedure also allowed intraportal injections of the drugs to be made so that the effect of first-passage uptake by the liver could be compared with hapatic exclusion in the same animal. 3 Hepatic exclusion greatly increased the duration of action of both drugs. Whereas intraportal injection did not significantly alter the effect of pancuronium on the tibialis muscle, the effect of Org 6368 was greatly diminished when given by this route. 4 The liver appears to tolerate short periods of hepatic exclusion and it is concluded that this technique may become a useful tool for studying the handling of drugs by this organ.     

Transport of organic anions into liver cells and bile

The distribution of [¹⁴C]para-acetylaminohippuric acid ([¹⁴C]PAAH) and [³⁵S]sulfobromophthalein ([³⁵S]BSP) among plasma, liver cells and bile of rats was measured under steady-state conditions. The electrical potential difference (PD) was measured across the sinusoidal membrane of the liver cell and between plasma and bile in the common duct. A PD of 40.3 ± 1.0 mV (mean ± S.E.M.) was recorded across the sinusoidal membrane (cell negative with respect to plasma). The PD between plasma and bile was 3.9 ± 0.5 mV (bile negative with respect to plasma). The electrical potentials, considered together with the concentrations of PAAH and BSP in plasma, liver cells and bile, indicate that these organic anions are actively transported into liver cells across the sinusoidal membrane. The concentrations of PAAH and BSP in liver cells and bile suggest that they are transferred across the canalicular membrane by active transport processes. When the plasma concentration of PAAH was increased, the bile/liver cell ratio of PAAH reached an asymptote, suggesting that the transport of PAAH from liver cells into bile is a carrier-mediated system which can be saturated. An infusion of sodium taurocholate, which stimulated bile flow and increased the excretion of BSP in bile, did not influence the biliary excretion of PAAH. The transport of PAAH from liver cells into bile appears to act as a ‘sink’, thereby maintaining a low liver cell PAAH concentration. The membrane transport system for the uptake and excretion of PAAH may be relevant to the biliary excretion of other exogenous and endogenous organic anions. The results of this study also demonstrate the importance of considering electrical potentials when studying the distribution of a charged compound in the hepato-biliary system.     

Pharmacokinetics of the hepatic transport of organic anions: influence of extra- and intracellular binding on hepatic storage of dibromosulfophthalein and interactions with indocyanine green

The influence of intracellular and extracellular protein binding on the hepatic storage and biliary elimination of dibromosulfophthalein (DBSP) was studied in isolated perfused rat liver. Under first order kinetic conditions the amount of DBSP in the liver at a given plasma concentration (hepatic storage) was determined by extracellular binding to albumin and intracellular binding to the cytosolic Y and Z proteins as well as concentrative membrane transport from plasma into the liver. At higher doses, extensive binding of DBSP to intracellular organelles also occurred while liver cytosol/plasma concentration gradients of unbound DBSP were much lower. Hepatic storage increased with decreasing albumin concentration in the perfusate of isolated perfused rat livers. However, it was shown that this parameter is dose-dependent, and errors can be introduced in its calculation if nonlinearity of sinusoidal and canalicular transport processes as well as nonlinear protein binding are not taken into account. The influence of another organic anion, indocyanine green (ICG) on the hepatic storage, subcellular distribution, and elimination of DBSP was subsequently studied. At equimolar amounts the presence of ICG resulted in a 50% decrease in hepatic clearance and hepatic distribution volume of DBSP. It was inferred that these changes are due to an inhibition of carrier-mediated transport across the sinusoidal and canalicular membrane and preferential displacement from intracellular binding sites. In contrast DBSP in equimolar amount enhanced the initial disappearance rate and biliary excretion of ICG, probably due to increasing its free fraction in plasma. It is concluded that the level and mechanism of interaction of two drugs within the eliminating organ can be characterized by combining clearance studies with data on subcellular and extracellular binding.     

ATP-Dependent Transport of a Novel Thromboxane A2 Receptor Antagonist, [2-(4-Chlorophenylsulfonylaminomethyl)-Indan-5-yl]Acetate (Z-335) and Its Xenobiotic Taurine Conjugate (Z-335-Tau) by Rat Bile Canalicular Membrane Vesicles

PURPOSE: The characteristics of bile canalicular transport processes for xenobiotic taurine conjugates have not yet been clarified. To elucidate the biliary excretion characteristics of xenobiotic taurine conjugates, we investigated the transport of a novel thromboxane A2 receptor antagonist, Z-335, and its taurine conjugate (Z-335-Tau) across the bile canalicular membrane. METHODS: We examined the uptake of Z-335 and Z-335-Tau by isolated bile canalicular membrane vesicles (CMVs) from Sprague Dawley and Eisai-hyperbilirubinemic rats (EHBRs) which EHBRs have a hereditary defect of canalicular multidrug resistance-associated protein 2 (Mrp2) function. Also, the in vitro and in vivo kinetics of Z-335-Tau uptake and excretion were compared. RESULTS: Z-335 uptake by CMVs from normal rats exhibited marked ATP-dependence, whereas ATP-dependent uptake of Z-335 into CMVs from EHBRs was not observed. In contrast, Z-335-Tau uptake into CMVs from both normal rats and EHBRs was ATP dependent. The initial uptake velocity was concentration-dependent, with an in vitro Michaelis constant for initial uptake of 189 microM, which was similar to the in vivo value. CONCLUSIONS: The biliary excretion of Z-335 involves Mrp2, whereas that of Z-335-Tau involves active transport systems that remain intact in EHBRs and show marked ATP dependence, which ATP-dependent transport is involved in the biliary excretion of Z-335-Tau in vivo.     

Canalicular membrane transport is primarily responsible for the difference in hepatobiliary excretion of triethylmethylammonium and tributylmethylammonium in rats.

Two structurally similar quaternary ammonium compounds, triethylmethylammonium (TEMA, M(r) 116) and tributylmethylammonium (TBuMA, M(r) 200) were used as model compounds to identify the unit process of hepatobiliary excretion that is responsible for markedly different biliary excretion of organic cations (OCs). Cumulative biliary excretion (in percentage of dose; i.v., 12 micromol/kg) was 0.17 for TEMA and 34.5 for TBuMA. In vivo uptake clearance into the liver was 0.686 +/- 0.020 ml/min for TEMA and 0.421 +/- 0.028 ml/min for TBuMA. When the uptake clearance was examined in an isolated hepatocyte system, comparable clearance between TEMA and TBuMA was obtained, consistent with the in vivo result. These observations suggest that uptake into the liver is not the major determinant for the difference in biliary excretion of the OCs. Coadministration of colchicine, an inhibitor of microtubule formation, had no effect on biliary excretion of the model compounds, and the primary site of subcellular distribution of the OCs appears to be the cytosol, suggesting that intracellular movement does not play a major role in the markedly different biliary excretion of the OCs. In contrast, in vivo excretion clearance across the canalicular membrane for TBuMA was 180-fold greater than that for TEMA, and in vitro efflux clearance of TBuMA was smaller than that of TEMA (p <.01), indicative of involvement of these processes in the markedly different biliary excretion of the OCs. Therefore, these data indicate that canalicular transport is primarily responsible for the markedly different biliary excretion of TEMA and TBuMA.     

Glutathione S-Transferases as a Cefpiramide Binding Protein in Rat Liver

Abstract: To clarify the intrahepatical transport mechanism of cefpiramide, we investigated effects of various agents mainly excreted into the bile by several different mechanisms on the biliary excretion of cefpiramide in rats. Sulfobromophthalein, indocyanine green, bilirubin and probenecid, known to be bound to glutathione S-transferases (GST) (EC 2.5.1.18) in liver cytosol, reduced the biliary excretion of cefpiramide, while neither secretory IgA, which is transported via vesicles in the liver, nor colchicine, which inhibits movements of vesicles, had any effect on the excretion of cefpiramide. Propranolol and metoprolol, metabolized by mixed function oxidases, had no effect on the biliary excretion of cefpiramide. In the chromatography of liver cytosol, the amount of sulfobromophthalein or benzylpenicillin bound to the GST fraction decreased in the presence of cefpiramide or probenecid. The study showed that cefpiramide was transported in the liver without relation to mixed function oxidases or vesichle-mediated transporting system, but in relation to GST which binds cefpiramide, sulfobromophthalein, benzylpenicillin and probenecid, indicating an important role of GST in the cefpirami de excretion into the bile.     

Kinetic Analysis of Hepatobiliary Transport for Conjugated Metabolites in the Perfused Liver of Mutant Rats (EHBR) with Hereditary Conjugated Hyperbilirubinemia

Purpose. Previously, we found that the biliary excretion of the 6-hydroxy-5,7-dimethyl-2-methylamino-4-(3-pyridylmethyl) benzothiazole (E3040) glucuronide is severely impaired in Eisai hyperbilirubinemic rats (EHBR), while that of sulfate remains normal (Takenaka et al., J. Pharmacol. Exp. Then, 274: 1362–1369, 1995). The purpose of the present study is to clarify the mechanisms for impairment of the biliary excretion of E3040 glucuronide in EHBR. Methods. We kinetically analyzed the disposition of the conjugates in the perfused liver at steady state. The uptake of the conjugates into the isolated canalicular membrane vesicles (CMVs) was also examined. Results. At steady state, the bile/liver unbound concentration ratios of the conjugates were 40-400 in both rat strains, indicating a highly concentrated process. The biliary excretion clearance (CL_u,bile) of the glucuronide, defined for the unbound concentration in the liver, was decreased in EHBR to 1/30 of that in normal rats, whereas the CL_u,bile of the sulfate was comparable between the two rat strains. In vitro, the transport of E3040 glucuronide into CMV prepared from SD rats exhibited the ATP dependency, whereas minimal effect of ATP was observed on the uptake of the glucuronide into CMV from EHBR. In contrast, the uptake of E3040 sulfate was comparable between SD rats and EHBR. Furthermore, ATP did not stimulate the uptake of sulfate into the CMVs. Conclusions. It was suggested (1) that the excretion of E3040 glucuronide across the bile canalicular membrane is mediated by the primary active transporter which is defective in EHBR and (2) that the bile canalicular transport system for E3040 sulfate is different from that for the glucuronide in that the former remains normal in EHBR.     

Mechanisms of metal transport across liver cell plasma membranes

The liver's pivotal role in the homeostasis of essential trace metals and detoxification of exogenous metals is attributed to its ability to efficiently extract metals from plasma, metabolize, store, and redistribute them in various forms either into bile or back into the bloodstream. Bidirectional transport across the sinusoidal plasma membrane allows the liver to control plasma concentrations and therefore availability to other tissues. In contrast, transport across the canalicular membrane is largely, but not exclusively, unidirectional and is a major excretory pathway. Although each metal has relatively distinct hepatic transport characteristics, some generalizations can be made. First, movement of metals from plasma to bile follows primarily a transcellular route. The roles of the paracellular pathway and of ductular secretion appear minimal. Second, intracellular binding proteins and in particular metallothionein play only indirect roles in transmembrane flux. The amounts of metallothionein normally secreted into plasma and bile are quite small and cannot account for total metal efflux. Third, metals traverse liver cell plasma membranes largely by facilitated diffusion, and by fluid-phase, adsorptive, and receptor-mediated endocytosis/exocytosis. There is currently no evidence for primary active transport. Because of the high rate of hepatocellular membrane turnover, metal transport via endocytic vesicles probably makes a larger contribution than previously recognized. Finally, there is significant overlap in substrate specificity on the putative membrane carriers for the essential trace metals. For example, zinc and copper share many transport characteristics and apparently compete for at least one common transport pathway. Similarly, canalicular transport of five of the metals discussed in this overview (Cu, Zn, Cd, Hg, and Pb) is linked to biliary GSH excretion. These metals may be transported as GSH complexes by the canalicular glutathione transport system(s). Unfortunately, none of the putative membrane carrier proteins have been studied at the subcellular or molecular level. Our knowledge of their biochemical properties is rudimentary and rests almost entirely on indirect evidence obtained in vivo or in intact cell systems. The challenge for the future is to isolate and characterize these putative metal carriers, and to determine how they are functionally regulated.     

Effects of pancuronium and vecuronium on creatine phosphokinase in rat isolated heart, liver, kidney and diaphragm.

1. The effects of two muscle relaxants, namely, pancuronium and vecuronium, on creatine phosphokinase (CPK) release from four different types of tissues, namely, heart, liver, kidney and diaphragm, were studied in the rat in vitro. 2. The total, neat and CPK levels (units/ml), released by muscle relaxants were measured using spectrophotometric determination at 340 nm. 3. The results showed that both muscle relaxants, in low concentrations, i.e. 0.34 or 0.32 microM, close to a clinical dose of 0.1 mg/kg, had no significant effect on CPK leakage in all four types of tissues studied. However, in concentrations 12-122 times clinical dose, the two muscle relaxants produced differential adverse effects in the tissues studied. 4. In most concentrations, pancuronium and vecuronium produced significant increases in the CPK release in the kidney and diaphragm. In contrast, pancuronium had no significant effect on CPK release in the liver and the lowest effect in the heart. Similar results were obtained with vecuronium. 5. The clinical relevance and/or implications of the present results are discussed and the results are compared to those previously reported by other workers in other preparations.     

Differences in the hepatobiliary transport of two quinolone antibiotics, grepafloxacin and lomefloxacin, in the rat

The biliary excretion of grepafloxacin (GPFX) was compared with that of lomefloxacin (LFLX) in rats. The biliary clearances (Cl(plasma)(bile)) of GPFX was 2.9 times greater than LFLX based on the plasma concentration reached during constant intravenous (i.v.) infusion. The liver-plasma unbound concentration ratio, K(pu), of GPFX (1.7) was also higher than that of LFLX (0.7). The hepatic uptake clearance, assessed from an integration plot analysis, of GPFX was comparable with the hepatic blood flow rate, and 1.5 times that of LFLX, indicating that membrane transport in the uptake process is more efficient for GPFX. This was also supported by the difference between the uptake clearance of GPFX and LFLX in isolated rat hepatocytes. The bile-liver unbound concentration ratio of GPFX and LFLX was approximately 6 and 3, respectively, and the biliary clearance based on the unbound liver concentration of GPFX was 1.8 times that of LFLX. These results suggest that the concentrative transport of GPFX also across the canalicular membrane was more efficient than that of LFLX. Thus, the membrane transport activity via both sinusoidal and canalicular membranes determines the net excretion of each compound.     

Effects of butylated hydroxytoluene (BHT) on biliary excretion of xenobiotics and bile flow in rats

A significant enhancement in the biliary excretion of iv injected sulfobromophthalein (BSP), phenol- 3,6 -dibromphthalein disulfonate (DBSP), procaine amide ethobromide (PAEB) and ouabain was observed in rats maintained on diets containing 0.25% BHT for periods of 10 days. The enhanced biliary excretion of these drugs in BHT treated rats appears to be correlated with the increase in bile flow produced by BHT. The increased bile flow was due to an increase in canalicular bile production rather than a change in net ductular secretion or reabsorption of fluid since bile to plasma concentration ratios of erythritol were unchanged and no permeability change in the biliary tree was observed when mannitol was administered by retrograde intrabiliary injection. The increase in bile flow was not due to an enhanced excretion of bile salts into bile, because both the biliary bile acid concentration and total biliary excretion of bile acids were lower in BHT-treated rats than in control rats. It appears that the increase in bile flow produced by BHT is due to the osmotic choleresis related to the secretion of BHT and its metabolites into bile.     

Synergistic role of 3-hydroxy-3-methylglutaryl coenzyme A reductase and cholesterol 7alpha-hydroxylase in the pathogenesis of manganese-bilirubin-induced cholestasis in rats.

Manganese (Mn) and bilirubin (BR) induce intrahepatic cholestasis when injected sequentially. It was suggested that accumulation of newly synthesized cholesterol in the canalicular membrane is an initial step in the development of cholestasis. To clarify the involvement of cholesterol in the pathogenesis of Mn-BR-induced cholestasis, we examined biliary secretion and liver subcellular distribution of lipids in the cholestatic conditions (Mn-BR combination). We also examined hepatic metabolism of cholesterol under cholestatic and noncholestatic (Mn or BR given alone) conditions. The Mn-BR combination reduced bile flow by 50%, and bile acid, phospholipids, and cholesterol output by 42, 75, and 90%, respectively. There was a dramatic impairment of cholate excretion but not chenodeoxycholate excretion. Phosphatidylcholine species secreted into bile were unchanged, and microsomal total phospholipid content was significantly increased. Although there was no changes in liver membrane phospholipid content, the cholesterol/phospholipid ratio was increased by more than 50% in the canalicular fraction. Despite the increased concentration of cholesterol in canalicular membrane the activities of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, key enzyme in cholesterol synthesis, and cholesterol 7alpha-hydroxylase, key enzyme in cholesterol conversion to bile acids, were significantly reduced. However, the injection of Mn alone significantly increased both enzymes, while BR alone inhibited cholesterol 7alpha-hydroxylase by 62%, without affecting HMG-CoA reductase. In conclusion, the critical cholestatic events in Mn-Br-induced cholestasis appear to be mediated through the synergistic effects of Mn and BR acting on synthesis and degradation of cholesterol.     

Hepatic uptake and biliary excretion of organic cations--II. The influence of ion pair formation

The influence of an anorganic anion iodide (I-) and an organic anion tetraphenylborate (TPB-) on the hepatic uptake and biliary excretion of three organic cations, triethylmethyl ammonium (TEMA), tripropylmethyl ammonium (TPMA) and tri-n-butylmethyl ammonium (TBuMA) was studied. The compounds were injected as a bolus (D = 1 mumole) and studied in isolated perfused livers. In the perfusion medium 25% of the amount of NaCl (3 mmole) was replaced by NaI, whereas in two other experiments TPB- was added to the medium in two concentrations (2 microM and 200 microM). NaI did not affect the biliary output of the three quaternary ammonium compounds (QACs) although an increased net rate of hepatic uptake was found for all compounds, most likely due to a decreased liver to plasma transport. Liver to plasma concentration ratios were increased, while the ratios between bile to liver and bile to plasma were not affected. TPB- in catalytic amounts added to the medium (2 microM) decreased the biliary output of TEMA and TBuMA, whereas the kinetic profile of TPMA was unchanged. The decreased biliary excretion rate of TEMA was explained by a decreased plasma level (due to the increased liver uptake) assuming that the small molecular weight compounds can enter the bile directly from plasma via the junctional complexes between the cells. The bile to plasma (B/P) ratio was not affected. In contrast, the bile to plasma (B/P) ratio and the bile to liver (B/L) ratio of TBuMA were decreased, compared with the control, probably due to an increased reabsorption from the bile, whereas the back transport from the liver into the plasma was also decreased. A large amount of TPB- (200 microM), added to the perfusion medium, dramatically changed the kinetic profile of the three QACs. Ion pair formation between the QACs and TPB- was supposed to be responsible for this effect. Plasma levels dropped more rapidly as a result of an increased rate of liver uptake. The biliary excretion of all compounds was greatly reduced (the excretion rates were 0.022, 0.19 and 0.18 nmole/min, compared with 0.047, 0.71 and 7.5 nmole/min for the controls). It is concluded that ion pair formation may play a role in the hepatobiliary transport. The rate of liver uptake of the QACs is enhanced in the presence of an anion, which is due to an increase in plasma to liver transport (k12) and a reduced liver to plasma transport (k21).(ABSTRACT TRUNCATED AT 400 WORDS)     

Atracurium and vecuronium: two unique neuromuscular blocking agents

Atracurium and vecuronium are two new nondepolarizing skeletal muscle relaxants that were developed to overcome the deficiencies seen with currently available agents (tubocurarine, metocurine, pancuronium, and gallamine). Both compounds have unique metabolic profiles, separating them from other nondepolarizing agents. Neither drug depends on normal renal function for excretion and each can safely be given to patients with renal failure. Atracurium also does not depend on hepatic function for metabolism; however, vecuronium may require dosing adjustments in hepatic disease. Atracurium and vecuronium have similar onset times for muscle relaxation but shorter durations of action than other nondepolarizing muscle relaxants. Both agents produce minimal cumulative effects with repeated doses. A major advantage of both agents is their relative lack of cardiovascular effects when given in clinically effective doses, differing from other nondepolarizing muscle relaxants. Both agents produce a lower degree of histamine release than other agents, although atracurium appears to produce a higher incidence of histamine-like reactions than vecuronium. Vecuronium appears to be the agent of choice in patients with a history of asthma or allergy. The place in therapy for these new agents is discussed.