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Increasing evidence has recently demonstrated that soluble heparan sulfate (HS), a degradation product of extracellular matrix produced by elastase, plays a key role in the aggravation of acute pancreatitis (AP) and associated lung injury. However little is known about the detailed mechanism underlying HS-induced inflammatory cascade. Our previous work has provided a valuable clue that a large-conductance K(+) channel (MaxiK) was involved in the HS-stimulated activation of murine macrophages. Here we attempted to ask whether pharmacological inhibition of the MaxiK channel will exert beneficial effects on the treatment of AP and secondary lung injury. The protective effects of paxilline, a specific blocker of MaxiK, on rats against sodium taurocholate induced AP were evaluated. Our data showed that paxilline substantially attenuated AP and resultant lung injury, mainly by limiting the burst of inflammatory responses, as proven by decreased plasma concentrations of tumor necrosis factor-α and macrophage inflammatory protein-2, together with unimpaired pancreatic enzyme activities in rats suffering from AP. Compared with the therapeutic administration, pre-treatment of paxilline showed superior potential to slow down the progress of AP. Furthermore, AP rats received paxilline exhibited improved histopathologic alterations both in the pancreas and the lungs, and even lower lung MPO activity. Taken together, our study provides evidence that MaxiK is involved in the spread of inflammatory responses and the following lung injury during the attack of AP, indicating that this ion channel is a promising candidate as a therapeutic target for AP., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Lactation is associated with increased expression of bile acid transporters and an increased size and hydrophobicity of the bile acid pool in rats. ATP-binding cassette (ABC) transporters multidrug resistance protein 2 (Mdr2), Abcb11 [bile salt export pump (Bsep)], and Abcg5/Abcg8 heterodimers are essential for the biliary secretion of phospholipids, bile acids, and cholesterol, respectively. We investigated the expression of these transporters and secretion of their substrates in female control and lactating Sprague Dawley rats and C57BL/6 mice. Expression of Abcg5/Abcg8 mRNA was decreased by 97 and 60% by midlactation in rats and mice, respectively; protein levels of Abcg8 were below detection limits in lactating rats. Mdr2 mRNA expression was decreased in lactating rats and mice by 47 and 59%, respectively. Despite these changes in transporter expression, basal concentrations of cholesterol and phospholipid in bile were unchanged in rats and mice, whereas increased Bsep mRNA expression in early lactation coincided with an increased basal biliary bile acid concentration in lactating mice. Following taurocholate infusion, coupling of phospholipid and taurocholate secretion in bile of lactating mice was significantly impaired relative to control mice, with no significant changes in maximal secretion of cholesterol or bile acids. In rats, taurocholate infusion revealed a significantly impaired coupling of cholesterol to taurocholate secretion in bile in lactating vs. control animals. These data reveal marked utilization of an Abcg5/Abcg8-independent mechanism for basal biliary cholesterol secretion in rats during lactation, but a dependence on Abcg5/g8 for maximal biliary cholesterol secretion.

Background: Cholangiopathies impair the balance between proliferation and apoptosis of cholangiocytes leading to the disappearance of bile ducts and liver failure. Taurocholic acid (TC) is essential for cholangiocyte proliferative and functional response to cholestasis. Bile acids and neurotransmitters co-operatively regulate the biological response of the biliary epithelium to cholestasis. Adrenergic denervation of the liver during cholestasis results in the damage of bile ducts., Aim: To verify whether TC feeding prevents the damage of the biliary tree induced by adrenergic denervation in the course of cholestasis., Methods: Rats subjected to bile duct ligation (BDL) and to adrenergic denervation were fed a TC-enriched diet, in the absence or presence of daily administration of the phosphatidyl-inositol-3-kinase (PI3K) inhibitor wortmannin for 1 week., Results: TC prevented the induction of cholangiocyte apoptosis induced by adrenergic denervation. TC also restored cholangiocyte proliferation and functional activity, reduced after adrenergic denervation. TC prevented AKT dephosphorylation induced by adrenergic denervation. The cytoprotective effects of TC were abolished by the simultaneous administration of wortmannin., Summary/conclusions: TC administration prevents the damage of the biliary tree induced by the adrenergic denervation of the liver. These novel findings open novel perspectives in the understanding of the potential of bile acids especially in post-transplant liver disease.

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.

Endocytosed proteins in hepatocytes are transported to lysosomes for degradation. Metabolites accumulating in these organelles are released into bile by exocytosis, a process that seems to be regulated by the bile salt taurocholate (TC). In this study we examined if TC is also involved in the control of the lysosomal degradation of endocytosed proteins. We used [(14)C]sucrose-labeled horseradish peroxidase ([(14)C]S-HRP), a probe suitable to evaluate lysosomal proteolysis. TC-infused rats as well as isolated rat hepatocytes exposed to TC showed a significant inhibition in the lysosomal degradation of [(14)C]S-HRP (approximately 30%), with no change in either the uptake or the amount of protein reaching lysosomes. Under these conditions, the in vitro assay of lysosomal cathepsins B, L, H, and D revealed no change in their activities, suggesting that a reversible inhibition (lysosomal alkalinization?) was taking place in hepatocytes. Nevertheless, lysosomal pH measured using fluorescein isothiocyanate-dextran was shown not to be altered by TC. In addition, TC was unable to inhibit proteolysis in [(14)C]S-HRP loaded lysosomes or interfere in cathepsin assays. The results suggest that TC inhibits the lysosomal degradation of endocytosed proteins in hepatocytes and that the mechanism does not involve an effect of the bile salt per se or a rise in lysosomal pH.

Objective: To compare the effect of short term feeding of ursocholic acid, a hydrophilic bile acid, as the unconjugated acid and the taurine conjugate, on clinical and biochemical features and bile acid metabolism with that of ursodeoxycholic acid in four patients with primary biliary cirrhosis., Methods: Four patients with stage II primary biliary cirrhosis were studied. Two were fed ursocholic acid (900 mg/day), and two were given tauroursocholate (900 mg/day) in three divided doses. After 1 month, all patients were given 900 mg/day of ursodeoxycholic acid. Fasting serum, bile, and 24-hour urine levels were measured before and at the end of ursocholic acid and tauroursocholate feeding and after 1 month of ursodeoxycholic acid feeding. Clinical and biochemical symptoms were measured by routine hospital methods, and bile acids were measured by gas-liquid chromatography., Results: One month of ursocholic acid or tauroursocholate feeding did not improve clinical or biochemical findings in any patient. Approximately 21-25% ursocholic acid was present in the serum and bile, with substantial metabolism to deoxycholic acid. Increased ursocholic acid was excreted in the urine. In comparison, ursodeoxycholic acid improved biochemical parameters and was 45-65% enriched in the serum and bile., Conclusion: Ursocholic acid as the free bile acid or as taurine conjugate, although more hydrophilic, is poorly enriched in serum and bile and is ineffective in patients with primary biliary cirrhosis.

Rationale and Objectives: We studied the intestinal absorption of manganese mesoporphyrin (Mn-mesoporphyrin), a potential oral hepatobiliary contrast agent., Methods: Mn-mesoporphyrin was complexed with monoolein and taurocholate (mixed micelles). Portal venous delivery and biliary excretion were measured after intestinal administration in rats and rabbits, and the mechanism of intestinal transport was studied in a combined lymph-bile fistula model in rats. T1-weighted magnetic resonance (MR) images of the liver were obtained in rats and domestic pigs before and after gastric administration of Mn-mesoporphyrin in mixed micelles., Results: A 2.2-fold increase of portal venous Mn concentration was found 90 min after intestinal administration of the complex. None was found in the lymph collected from the thoracic duct, indicating a transcellular transport mechanism through the intestinal mucosa with portal venous delivery. Mn-mesoporphyrin levels in bile peaked between 240 and 270 min after administration (200-fold increase). The greatest liver enhancement (20-90%) was measured 360 min after administration., Conclusion: The feasibility of intestinal delivery of Mn-mesoporphyrin, a lipophilic hepatobiliary contrast agent was demonstrated.

The present study has demonstrated that tauroursodeoxycholate (TUDC), but not taurocholate, can reverse chlorpromazine (CPZ)-induced cholestasis in the isolated perfused rat liver. At an infusion rate of 1.5 mumol/min, TUDC led to restoration of bile flow in the perfused rat liver made cholestatic by the addition of 250 microM CPZ. This reversal was accompanied by an increased excretion of CPZ and its metabolites. A higher infusion rate of 5.0 mumols TUDC/min, however, led to only a transient increase in bile flow and to no increase in CPZ excretion. In contrast to the effects of TUDC, infusion of taurocholate led to an exacerbation of CPZ-induced cholestasis. The differences in the efficacy of the two bile salts may be due to their relative detergent (hydrophobic) properties.

The ability of the cholestatic steroid glucuronide, estradiol-17 beta-(beta-D-glucuronide) (E(2)17G), to inhibit the hepatic clearance (ClH) and biliary secretory rate maximum (SRm) of taurocholate was investigated in the recirculating and single pass isolated perfused male rat liver. In the recirculating perfused liver, E(2)17G (0, 2, 4, or 6 mumol) was added as a bolus dose to the reservoir at zero time while taurocholate was infused into the portal vein in increasing amounts (15, 30, 45, or 60 mumol/mL; 1 mL/hr for 15 min each). E(2)17G (4 mumol) caused a significant (P less than 0.05) inhibition of bile flow and bile acid secretion at 10-15 min during infusion of 15 mumol/hr taurocholate but did not inhibit the SRm which occurred at 42 min, indicating that E(2)17G had not caused an irreversible inhibition of taurocholate transport. E(2)17G (6 mumol) caused a profound and irreversible inhibition of bile flow attributable to retention of E(2)17G in the liver. The noncholestatic estradiol-3-(beta-D-glucuronide) (E(2)3G; 6 mumol) had no significant effect on bile flow or the SRm. In the single pass perfused liver (10 mL/min flow rate), E(2)17G (0, 1, 2, 5, or 10 nmol/mL) or E(2)3G (2 nmol/mL) was added to the perfusate resulting in a stable infusion to the liver. [3H]Taurocholate was infused into the portal vein in increasing amounts to give inflow concentrations (Cin) of 25, 50, 75 or 100 nmol/mL. In the absence of E(2)17G, taurocholate ClH decreased from 0.92 to 0.70 mL/min/g liver with increasing taurocholate concentrations. Neither E(2)17G nor E(2)3G altered the ClH of 25 nmol/mL taurocholate. E(2)17G (10 nmol/mL) inhibited bile flow and bile acid secretion first at 20-25 min, followed by inhibition of ClH of 75 and 100 nmol/mL taurocholate (35-60 min). In contrast, E(2)3G stimulated bile acid secretion and increased the SRm by 80%. Thus, at doses that did not block its own elimination, E(2)17G did not cause an irreversible inhibition of taurocholate transport into bile. E(2)17G did not directly inhibit the uptake of taurocholate into the liver but first inhibited the biliary excretion of taurocholate, resulting in its intrahepatic accumulation and decreased clearance from the perfusate.

Cyclosporin A (CyA) decreases bile flow and bile salt secretion in the rat. The purpose of this study was to examine the influence of CyA on the hepatic transport of bromosulfophthalein (BSP). Male Sprague-Dawley rats were injected with CyA at the daily dose of 10 mg/kg (treated animals) or solvent (controls) during three weeks. Hepatic uptake of BSP (assessed by the plasma disappearance curve of the dye) and biliary secretion during infusions (95.5 and 178 nmol/min/100 g) were examined in both groups. Administration of CyA resulted in a decrease in both bile flow and BSP biliary secretion at the two infusion rates used. BSP plasma disappearance rate was significantly lower in treated animals than in controls. Conjugation of the dye was unaffected by CyA. There was no modification in ALT activity or in liver histology. These data show that chronic administration of CyA in rats decreases both hepatic uptake and biliary secretion of BSP. Thus, the inhibitory effect of CyA on biliary secretion is not limited to bile salts but also is observed with other cholephilic substances.

A single intravenous injection of [14C]taurocholate was followed up in blood and bile of rats submitted to steady intravenous infusions of taurocholate (TC) at rates of 0.0, 0.5, 1.0, and 1.5 mumol.min-1.100 g body wt-1 for at least 30 min. The transport rate constants and the amounts of TC in different compartments were estimated by weighted least-squares adjustment of a six-compartment model to the experimental data (3 compartments for TC distribution in blood, 2 compartments for liver, and 1 compartment for sinusoidal blood space). The saturation of the TC excretion rate was reached at 0.8 mumol.min-1.100 g body wt-1. It was characterized by a decrease of both the uptake and excretion rate constants, by an increase of the ratio of the amounts of TC in the two intrahepatic compartments (H'/H), and by an intrahepatic TC concentration of approximately 2 mM. When tauroursodeoxycholate (TUDC) was infused at a rate of 0.5 mumol.min-1.100 g body wt-1 together with TC at a rate of 1.5 mumol.min-1.100 g body wt-1, the TC excretion rate increased to 1.2 mumol.min-1.100 g body wt-1, and the excretion rate constant and H'/H decreased toward control values. These results support the hypothesis that the saturation of the transport of TC is due to TC hepatotoxicity and can be reduced by TUDC. Michaelis-Menten parameters, derived from saturation curves for both uptake and excretion steps, closely matched earlier results, thus confirming the good descriptive capacity of the model.

The volume and compositon of hepatic bile was studied in anesthetized dogs before and approximately 18 weeks after cholecystectomy. Dogs had common duct cannulation, cystic duct ligation, and temporary occlusion of the pylorus. An intravenous infusion of sodium taurocholate, 9 muEq per minute, was given throughout the experiment. Bile volume, electrolyte composition, bile salt output, and 14C-mannitol clearance were determined during infusion of sodium taurocholate alone and after addition of secretin, 4 U. per kilogram per hour, to the infusion. Cholecystectomy then was done and the same experiment was repeated 18 weeks later. Cholecystectomy significantly increased bile volume, sodium and chloride output, and 14C-mannitol clearance both in response to taurocholate alone and to taurocholate plus secretin. Bile salt output and bicarbonate output were unchanged. The study shows that cholecystectomy significantly alters the volume and composition of hepatic bile. The data indicate that cholecystectomy does not affect ductular function or hepatic secretion of bile salts but that canalicular function is altered significantly.

An isolated canine liver perfusion technique featuring a second dog as the pump oxygenator was used to compare biliary lipid secretion during randomized, steady-state perfusions at two different rates of cholyl taurine or chenodeoxycholyl taurine infusions. The hepatic extraction of the trihydroxy-conjugated bile acid was considerably greater than that of the dihydroxy conjugate, possibly explained by ultrafiltration experiments which indicated that cholyl taurine was less protein bound than chenodeoxycholyl taurine. Both bile acids induced phospholipid and cholesterol secretion that was linearly proportional to bile acid secretion. However, each mole of secreted chenodeoxycholyl taurine induced a greater relative secretion of phospholipid and cholesterol than did that of cholyl taurine. Thus in the canine liver, the two primary bile acids are extracted at different rates and induce biliary secretion of different relative lipid composition.

The prairie dog was used as a model for human gallstone formation. Stones formed in the gallbladder of all animals on a lithogenic diet. Hepatic bile was nonlithogenic, whereas gallbladder bile promoted cholesterol precipitation. Addition of taurocholate to the diet reduced the number of stones and lithogenicity. Cholecystectomy resulted in an increased bile flow and reduced secretion of cholesterol in the animals on a high cholesterol diet. Reduction of cholesterol and bile acid synthesis by negative feedback was demonstrated in isolated hepatocyte culture. The shift of bile salt production to chenodeoxycholates on a high cholesterol intake was demonstrated both in vivo and in vitro. A theory of gallstone formation is presented which hypothesizes a defect in hepatocyte storage of cholesterol rather than bile acid synthesis as the primary effect, relegating the problems to one of a disease of lipid metabolism.

Two distinct lipoprotein receptors can be expressed in the dog liver. One is the apolipoprotein (apo-) B,E receptor. This receptor binds apo-B-containing low density lipoproteins (LDL), as well as apo-E-containing lipoproteins, such as the cholesterol-induced high density lipoproteins (HDL(c)). The second hepatic lipoprotein receptor is the apo-E receptor. It binds apo-E HDL(c) and chylomicron remnants, but not LDL. The present studies were undertaken to determine whether short-term (acute) regulation of the two receptors can occur in response to perturbations in hepatic cholesterol metabolism. The design used three groups of experimental animals: (a) immature dogs (with both hepatic apo-B,E and apo-E receptors expressed), (b) adult dogs (with predominantly the apo-E receptor expressed and little detectable apo-B,E receptor binding activity), and (c) dogs treated with the bile acid sequestrant cholestyramine or those that have undergone biliary diversion (with apo-E receptors and induced apo-B,E receptors). In the first series of experiments, changes in hepatic lipoprotein receptor expression were studied by delivering cholesterol to the liver via intestinal lymph lipoproteins. Dog lymph (5-11 mg of triglycerides/min per kg of body weight, 0.15-0.3 mg of cholesterol/min per kg) or saline were infused intravenously for 6-8 h into matched pairs of dogs. Serial liver biopsies were obtained at intervals of 1-2 h. A progressive loss of specific (calcium-dependent) binding of LDL was seen in hepatic membranes from both immature and cholestyramine-treated dogs. After 4-6 h of lymph infusion, almost no apo-B,E receptor binding could be detected. The decrease in binding of apo-E HDL(c) to the same membranes was much less pronounced, and could be explained by a loss of binding of HDL(c) to the apo-B,E receptor; there was little or no effect on apo-E receptor binding. In the second series of experiments, the effects of a diminished hepatic demand for cholesterol on lipoprotein receptor expression were studied by suppressing bile acid synthesis. The bile acid taurocholate (2-3 mumol/kg per min) was infused intravenously over a 6-h interval. This resulted in a progressive loss of LDL binding to liver membranes of immature or cholestyramine-treated dogs. The infusion of taurocholate for 6 h did not significantly alter the expression of the apo-E receptor binding activity, whereas apo-B,E receptor activity was rapidly down-regulated. Preparation of a bile fistula in adult dogs markedly induced the expression of the apo-B,E receptor. In this state, the binding activity of the apo-B,E receptor could be almost totally abolished by reinfusion of taurocholate for 6 h, without profoundly affecting apo-E receptor binding. Evidence from the analysis of plasma lipoprotein patterns and tissue culture reactivity suggested that changes in assayed hepatic lipoprotein receptor activity occurred in concert with changes in plasma lipoproteins.The results indicate that the two canine hepatic lipoprotein receptors differ in their metabolic regulation. The apo-B,E receptor responds rapidly to changes in hepatic requirements for cholesterol. The apo-E receptor appears to be more refractory to acute regulation. The rapidity of the changes in the activity of the apo-B,E receptor (within 2-4 h) suggests that the binding activity of this receptor may be regulated by factors independent of protein synthesis.

Male hamsters were fed normal and essential fatty acid (EFA)-deficient diets for at least 12 wk before bile duct cannulation. With [32P]phosphate, hepatic synthesis of lecithin was similar, but biliary excretion of newly synthesized lecithin was significantly reduced in EFA-deficient compared to that in normal hamsters. Hepatic uptake of intravenously infused taurocholate (TC) and taurochenodeoxycholate (TCDC) were similar in both groups of animals. However, biliary excretion of intravenously infused TC was significantly reduced in EFA-deficient hamsters, whereas that of TCDC-was unchanged. The absolute rate of biliary cholesterol excretion was similar in both groups. Canalicular bile flow, as measured by [14C]erythritol clearance after functional nephrectomy, was significantly lower, with both the bile salt-dependent and independent fractions of this flow being diminished in EFA-deficient hamsters infused with TC. It is concluded that EFA deficiency leads to impaired biliary excretion of taurocholate, lecithin, and water, while cholesterol transport is unaffected, and thus results in supersaturation of bile with respect to cholesterol and production of lithogenic bile.

The effect of somatostatin, an inhibitor of release of a number of gastrointestinal and other hormones, on choleresis was investigated in chronic, bile fistula dogs with taurocholate-stabilized bile flow. Somatostatin inhibited both fasting and meal-stimulated choleresis, and bile flows during somatostatin inhibition of both fasting and fed dogs were similar, suggesting a complete suppression of factors causing feeding choleresis. Although a transient decrease in bile salt output was observed, bile salt output was unaffected during most of the period of bile flow inhibition. Hormone suppression by somatostatin, indicated by measurement of serum insulin, occurred over a similar time course as inhibition of choleresis. These observations provide further evidence for physiological humoral regulation of choleresis.

The effect of repeated administration of taurocholate on bile acid pool size, biliary composition and biliary excretory capacity for bile acids and two xenobiotics was determined. The bile acid pool was increased 50 to 60% by oral administration of sodium taurocholate (300--900 mg/kg, 10 ml/kg) every 12 hr for 2 days to male Sprague-Dawley rats. Bile flow, biliary excretion of bile acids, cholesterol and phospholipid and the concentrations of phospholipid and bile acids in bile were increased in rats treated with 750 mg of taurocholate per kg. No effect was observed on Na+,K+ or Cl- levels. The biliary transport maximum for taurocholate was increased by 30% in rats treated with 750 mg/kg. In contrast, the plasma disappearance and biliary excretion of phenol-3,6-dibromphthalein and ouabain were not affected by taurocholate administration.

Sodium taurocholate was administered to the in situ perfused liver of male Sprague Dawley rats at various rates below (57 and 114 nmol/min -g liver) and above (228 and 456 nmol/min -g liver) its biliary transport maximum (Tm) to study its effect on bile formation. As expected, bile flow increased with increasing dose until maximal bile salt excretion was reached. By contrast, during taurocholate infusions exceeding the taurocholate-Tm, bile flow and bile salt excretion decreased. Under those conditions, a given bile salt excretion was associated with a smaller volume of bile. A relationship between these effects and the concentration of taurocholate in the perfusate (160 to 860 nmol/ml) was suggested by the observation that bile formation returned toward normal when the taurocholate concentration was lowered by exchange of the perfusate.

The hepatotoxicity of CPZ was studied in the isolated perfused rat liver in order to more closely define possible mechanisms of phenothiazine-induced cholestasis. Perfusate concentrations of CPZ were increased from 5 x 10(-6) M to 5 x 10(-4) M until bile secretion was significantly inhibited. Measurements were then made of determinants of bile secretory function, including the magnitude of lobar distribution of perfusate flow, BAIF, and liver plasma membrane enzyme activity, Na+,K+-ATPase, Mg++-ATPase and 5'-nucleotidase. BAIF diminished significantly from control values of 1.76 +/- 0.07 microliter min-1gm-1 of liver to 1.34 +/- 0.15 and 0.80 +/- 0.09 following 2.5 and 5 x 10(-4) M CPZ, respectively. Perfusate flow also diminished from 5.64 +/- 0.44 to 1.24 +/- 0.12 ml min-1 gm-1 of liver 20 min following 5 x 10(-4) M CPZ and was associated with reduced flow to peripheral areas of the hepatic lobes as demonstrated by Tc-HAM. By 30 min, perfusate flow had returned to baseline values. CPZ also transiently diminished the excretion of bile acids in livers receiving a constant infusion of 40 mumol hr-1 sodium taurocholate. Defects in hepatic perfusion could not account entirely for the impairment in BAIF, since comparable mechanical restriction of perfusate flow in controls only diminished BAIF to 1.49 +/- 0.08 microliter min-1gm-1 of liver. CPZ signofocamt;u rediced tje secofoc actovotu pf Mg++-ATPase and 5'-nucleotidase but did not affect Na+,K+-ATPase in liver plasma membrane isolated 20 min after 5 x 10(-4) M CPZ. CPZ also resulted in a profound shift in the recovery of protein in isolated liver plasma membrane fractions from the light (density = 1.16) to heavier (density = 1.18) fractions. These findings, together with previous observations demonstrating alterations in hepatic ultrastructure, indicate that CPZ interacts in a complex manner with hepatocyte plasma and cytoplasmic membrane components and suggest that these drug-membrane interactions independently result in diminished hepatic perfusion, impairment of bile acid excretion, and inhibition of bile acid-independent bile secretion.

The purpose of this study was to examine the hepatocellular extraction of taurocholate and to determine the kinetic characteristics of the uptake process. The uptake of taurocholate by the liver of the intact dog was studied by the multiple-indicator dilution method. 51Cr-labeled red blood cells (a vascular indicator), 125I-labeled albumin (an extravascular reference), and [14C]taurocholate were injected into the portal vein. Different doses of unlabeled taurocholate were included in the injection mixture. Hepatic venous dilution curves were obtained. As a consequence of the hepatic uptake, the outflow recovery of [14C]taurocholate was much reduced when compared to that of albumin, but its recovery increased with increasing doses of taurocholate, suggesting a progressive saturation of the uptake process. The analysis of the dilution curves fitted a three-compartment model system well and no return of the extracted taurocholate to the extracellular space could be detected. The initial space of distribution of taurocholate was 1.22 plus or minus 0.12 (SD) times greater than that of albumin. Analysis of the data for uptake was consistent with Michaelis-Menten kinetics. The calculated initial maximal velocity of uptake (Vmax) was 4.53 mumol times s--1 times 100 g of liver--1 and the dose yielding half-maximal velocity (DK) was 7.11 mumol times 100 g of liver--1. These results are consistent with the hypothesis that the uptake of taurocholate is carrier-mediated. The maximal vilocity of uptake was about six times the known maximal capacity of biliary secretion of taurocholate in the dog.

These studies were undertaken to characterize the role of bile salt hydrophobicity in determining the rate of cholesterol recruitment from the liver. Using an in vivo rat model in which the acquisition of hepatic cholesterol from chylomicron remnants, low-density lipoproteins, and de novo synthesis was measured and kept constant, it was found that the amount of sterol recruited from the liver cell increased progressively as the liver was probed with a constant infusion of progressively more hydrophobic bile salts. The absolute secretion rate of both cholesterol and phospholipid increased nearly 50% as the hydrophobic index of the bile salts traversing the liver increased from 1.7 to 4.5, but the ratio of cholesterol to phospholipid secreted in bile remained nearly constant. Thus, when cholesterol entry into the hepatocyte via lipoproteins and de novo cholesterol synthesis is constant, the mass of cellular cholesterol recruited into the bile is directly proportional to the hydrophobic-hydrophilic balance of the secreted bile salts.