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B Stieger

Publications and source records attributed to B Stieger.

At least 19 recordsLinked to original sources
PubMed1996

Expression of a rat liver phosphatidylcholine translocator in Xenopus laevis oocytes.

A phospholipid translocating protein from rat liver has been expressed in Xenopus laevis oocytes. Injection of oocytes with total rat liver messenger RNA (mRNA) resulted in the function expression of saturable uptake of the water soluble phophatidylcholine derivative L-alpha-dibutyroylglycero-3-phophatidylcholine (diC4PC), Kinetic studies revealed an apparent Km value of approximately 10 mmol/L, which is similar to the value previously obtained in isolated rat liver canalicular plasma membrane vesicles for an adenosine triphosphate (ATP)-independent phosphatidylcholine translocator. Size fractionation of total rat liver mRNA yielded an active mRNA species between 1.8 and 2.6 kb, that stimulated the expressed phophatidylcholine uptake activity approximately fivefold as compared with differently sized mRNA subfractions. This active mRNA size class is too small to code for the mdr2 P-glycoprotein, which has been suggested to function as an ATP-dependent canalicular phosphatidylcholine translocator. Hence, the data indicate that there are at least two separate polypeptides involved in phospholipid translocation from hepatocytes into bile.

ATP Binding Cassette Transporter, Subfamily B↗
PubMed1996

The peptide-based thrombin inhibitor CRC 220 is a new substrate of the basolateral rat liver organic anion-transporting polypeptide.

The peptidomimetic thrombin inhibitor CRC 220, 4-methoxy-2,3,6-trimethylphenylsulfonyl-L-aspartyl-D-4-amidinop henylalanyl- piperidide, is taken up into isolated rat hepatocytes through active, carrier-mediated transport. This uptake is inhibited by bile acids. Functional expression in Xenopus laevis oocytes was performed to identify the transport system responsible for the hepatocellular CRC 220 uptake. Injection of poly(A)+RNA in X. laevis oocytes resulted in a two- to three-times higher uptake of CRC 220, compared with uninjected or water-injected control oocytes. Taurocholate (200 mumol/L) inhibited this uptake completely. No uptake of the peptidomimetic thrombin inhibitor was observed, when X. laevis oocytes were injected with complementary RNA (cRNA) encoding either the cloned rat liver Na(+)-dependent taurocholate transporter Ntcp, the renal oligopeptide carrier rhaPT or the intestinal oligopeptide transporter PepT1. However, after injection of cRNA of the cloned rat liver Na(+)-independent organic anion transporting polypeptide oatp, a specific and saturable CRC 220 uptake was observed (Michaelis-Menten constant 29.5 mumol/L). Cis-inhibition with known oatp-substrates, e.g., 20 mumol/L Bromsulphalein (BSP), 2007 mumol/L taurocholate and 2007 mumol/L cholate, occurred in oatp-expressing X. laevis oocytes, whereas substrates of the two peptide carriers as well as dipeptide- and single-amino acid constituents of the thrombin inhibitor itself lacked any significant inhibitory effects. These data show that the modified dipeptide CRC 220 is a highly selective substrate of the organic anion transporting polypeptide oatp in the basolateral plasma membrane of rat hepatocytes.

Animals↗
PubMed1996

Structure-function correlation of tight junctional impairment after intrahepatic and extrahepatic cholestasis in rat liver.

BACKGROUND & AIMS: Tight junctions, the only barrier between blood and bile, are crucial in bile formation. The aim of this study was to correlate changes in morphology and permeability by comparing structural parameters with marker secretion into normal and cholestatic rat bile. METHODS: Cholestasis was induced by bile duct ligation of 5 and 21 days of ethinylestradiol administration. Quantitated structural parameters induced junctional length, strand number, junctional depth, and spacing of junctional particles. Junctional permeability was probed with horseradish peroxidase and dextrans of increasing sizes. RESULTS: Junctional length was decreased slightly by ethinylestradiol (-16% after 21 days) but increased by ligation (77%). Mean strand number decreased from 4.6 to 3.7 after 21 days of ethinylestradiol and 3.4 after ligation associated with increased junctional depth. The proportions of morphologically horseradish peroxidase-positive junctions increased from 4% to 15% after 21 days of ethinylestradiol and to 56% after ligation. Horseradish peroxidase secretion was increased twofold by ethinylestradiol and 6.5-fold by ligation, paralleled by an increase of dextran size selectivity from 70,000 to 79,000 daltons after ethinylestradiol and to 266,000 daltons after ligation. CONCLUSIONS: Impairment of junctional integrity is paralleled with the degree of cholestasis, whereas correlation of morphological and physiological alterations shows a close structure-function relationship.

Animals↗
PubMed1996

Immunologic distribution of an organic anion transport protein in rat liver and kidney.

A Na(+)-independent organic anion transport protein was recently cloned from rat liver using a Xenopus laevis oocyte expression system [E. Jacquemin, B. Hagenbuch, B. Stieger, A.W. Wolkoff, and P.J. Meier, Proc. Natl. Acad. Sci. USA 91: 133-137, 1994]. Although expression of this protein is sufficient for cells to transport the organic anion bromosulfophthalein, little is known about its cell biology or biochemical characteristics. Northern blot analysis performed under high-stringency conditions revealed hybridization with RNA only from liver and kidney; transcripts appeared the same in these two organs. Within kidney, hybridization was greatest when RNA extracted from the outer medulla was used. Immunoblot analysis revealed that in liver, the transporter was enriched in 0.1 M Na2CO3-extracted membranes and sinusoidal plasma membrane preparations, consistent with its being an integral membrane protein. This 80-kDa protein migrated as a 65-kDa protein after treatment with N-glycanase. Immunomorphological examination of liver revealed basolateral plasma membrane localization. In 0.1 M Na2CO3-extracted membranes of kidney, the transporter migrated as an 83-kDa protein on nonreducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). On reduction, it resolved into peptides of 33 and 37 kDa. SDS-PAGE migration of the liver protein was unaffected by reduction. Immunomorphological examination of kidney revealed apical plasma membrane localization in the S3 segment of the proximal tubule of the outer medulla. Differential processing and trafficking of this transporter in liver and kidney may have important functional and regulatory consequences.

Adenosine Triphosphate↗
PubMed1995

Characterization of L-carnitine transport by rat kidney brush-border-membrane vesicles.

In the presence of a 100 mM Na+ gradient, transport of L-carnitine into rat renal brush-border-membrane vesicles was linear over 30 s and showed an overshoot at 5 min. The uptake of L-carnitine was clearly less active in the presence of other cations such as Li+, K+, Cs+ or choline. In the presence of a Na+ gradient, L-carnitine uptake after 20 s was much higher for chloride as an anion than for SCN-, NO3-, gluconate or SO4(2-). In comparison with conditions with inside positive or no membrane potential, transport was higher in vesicles with an inside negative membrane potential, suggesting an electrogenic mechanism. The kinetic characterization of the Na(+)-dependent portion of L-carnitine transport revealed two transport systems with Km values of 17.4 +/- 3.9 microM and 15.0 +/- 6.0 mM, respectively. The transport could be inhibited in a concentration-dependent fashion by structural analogues such as butyrobetaine, L-acetylcarnitine, trimethyl-lysine and D-carnitine, but not by L-arginine or glycinebetaine.

Animals↗
PubMed1995

Molecular and functional characterization of an organic anion transporting polypeptide cloned from human liver.

BACKGROUND & AIMS: Based on a recently cloned rat liver organic anion transporter, we attempted to clone the corresponding human liver organic anion transporting polypeptide. METHODS: A human liver complementary DNA library was screened with a specific rat liver complementary DNA probe. The human liver transporter was cloned by homology with the rat protein and functionally characterized in Xenopus laevis oocytes. RESULTS: The cloned human liver organic anion transporting polypeptide consists of 670 amino acids and shows a 67% amino acid identity with the corresponding rat liver protein. Injection of in vitro transcribed complementary RNA into frog oocytes resulted in the expression of sodium-independent uptake of [35S]bromosulfophthalein (Michaelis constant [Km], approximately 20 mumol/L), [3H]cholate (Km, approximately 93 mumol/L), [3H]taurocholate (Km, approximately 60 mumol/L), [14C]glycocholate, [3H]taurochenodeoxycholate, and [3H]tauroursodeoxycholate (Km, approximately 19 mumol/L). Northern blot analysis showed cross-reactivity with messenger RNA species from human liver, brain, lung, kidney, and testes. Polymerase chain reaction analysis of genomic DNA from a panel of human-rodent somatic cell hybrids mapped the cloned human organic anion transporter to chromosome 12. CONCLUSIONS: These studies show that the cloned human liver organic anion transporter is closely related to, but probably not identical to, the previously cloned rat liver transporter. Furthermore, its additional localization in a variety of extrahepatic tissues suggests that it plays a fundamental role in overall transepithelial organic anion transport of the human body.

Amino Acid Sequence↗
PubMed1994

Hepatocellular transport of bile acids. Evidence for distinct subcellular localizations of electrogenic and ATP-dependent taurocholate transport in rat hepatocytes.

To investigate whether electrogenic and ATP-dependent taurocholate transport activities are both mediated by the same bile acid-transporting polypeptide in rat liver, we further purified isolated canalicular membrane vesicles by free flow electrophoresis. Removal of most of the contaminating endoplasmic reticulum resulted in a complete loss of electrogenic taurocholate transport from an ecto-ATPase-enriched canalicular membrane subfraction. In contrast, ATP-dependent taurocholate transport remained associated with both an ecto-ATPase-enriched and an ecto-ATPase-free canalicular membrane subfraction. Microsomes containing 64% of total endoplasmic reticulum exhibited saturable electrogenic (Km approximately 270 microM), but no ATP-dependent taurocholate uptake. Golgi membrane vesicles were devoid of any taurocholate transport activity. These results indicate that electrogenic taurocholate transport resides entirely in the endoplasmic reticulum, whereas ATP-dependent bile acid transport is an intrinsic function of the canalicular membrane as well as of a so far unidentified intracellular membrane bound compartment. Hence, the two transport activities are most probably mediated by two different bile acid transporting polypeptides. Furthermore, the finding of ATP-dependent taurocholate transport in virtually ecto-ATPase-free vesicles argues against the concept of primary active bile acid transport being exclusively mediated by the canalicular ecto-ATPase.

Adenosine Triphosphatases↗
PubMed1994

Functional expression cloning of the canalicular sulfate transport system of rat hepatocytes.

We have cloned a single cDNA encoding the canalicular sulfate transporter of rat liver using Xenopus laevis oocytes as a functional expression system. The cloned cDNA sulfate anion transporter-1 (sat-1) expresses saturable Na(+)-independent sulfate uptake (Km approximately 0.14 mM) that can be inhibited by 4,4'-diisothiocyano-2,2'-disulfonic acid stilbene (DIDS, IC50 = 28 microM) and oxalate, but not by succinate or cholate. These properties are very similar to sulfate uptake expressed in oocytes injected with total rat liver mRNA and to the bicarbonate/sulfate exchange system previously characterized in canalicular rat liver plasma membrane vesicles. The cloned sat-1 cDNA has a total length of 3726 base pairs (bp) with an open reading frame encompassing 2109 bp, a 5'-untranslated region of 367 bp, and a 3'-untranslated region of 1250 bp. The coding region predicts a protein of 703 amino acids with a calculated molecular mass of 75.4 kDa. Computer-based hydrophobicity analysis suggests the presence of 12 putative transmembrane spanning domains. Furthermore, three potential glycosylation sites are detected (Asn-158, Asn-163, Asn-587). Northern blot analysis indicates that similar sulfate anion transporters are also present in the kidney, muscle, and brain of rat and in the liver of the mouse. Using antisense oligonucleotides the mRNA-species of the sat-1 analogue in rat kidney has been characterized by hybrid depletion experiments (Markovich, D., Bissig, M., Sorribas, V., Hagenbuch, B., Meier, P. J., and Murer, H. (1994) J. Biol. Chem. 269, 3022-3026).

Amino Acid Sequence↗
PubMed1994

Expression cloning of a rat liver Na(+)-independent organic anion transporter.

Using expression cloning in Xenopus laevis oocytes, we have isolated a cDNA encoding a rat liver organic anion-transporting polypeptide (oatp). The cloned oatp mediated Na(+)-independent uptake of sulfobromophthalein (BSP) which was Cl(-)-dependent in the presence of bovine serum albumin (BSA) at low BSP concentrations (e.g., 2 microM). Addition of increasing amounts of BSA had no effects on the maximal velocity of initial BSP uptake, but it increased the Km value from 1.5 microM (no BSA) to 24 microM (BSA/BSP molar ratio, 3.7) and 35 microM (BSA/BSP ratio, 18.4). In addition to BSP, the cloned oatp also mediated Na(+)-independent uptake of conjugated (taurocholate) and unconjugated (cholate) bile acids. Sequence analysis of the cDNA revealed an open reading frame of 2010 nucleotides coding for a protein of 670 amino acids (calculated molecular mass, 74 kDa) with four possible N-linked glycosylation sites and 10 putative transmembrane domains. Translation experiments in vitro indicated that the transporter was indeed glycosylated and that its polypeptide backbone had an apparent molecular mass of 59 kDa. Northern blot analysis with the cloned probe revealed crossreactivity with several mRNA species from rat liver, kidney, brain, lung, skeletal muscle, and proximal colon as well as from liver tissues of mouse and rabbit, but not of skate (Raja erinacea) and human.

Amino Acid Sequence↗
PubMed1994

In situ localization of the hepatocytic Na+/Taurocholate cotransporting polypeptide in rat liver.

BACKGROUND/AIMS: An Na+/taurocholate cotransporting polypeptide (Ntcp) has recently been cloned from rat liver. The aim of this study was to directly characterize the native Ntcp on the protein level and study its in situ distribution in rat liver. METHODS: A rabbit antiserum was raised against a fusion protein containing the maltose-binding protein and the C terminus of Ntcp. Native Ntcp was localized in situ by immunofluorescent techniques. Expression of Ntcp was directly correlated with taurocholate uptake measurements in stably transfected Chinese hamster ovary cells. RESULTS: Native Ntcp showed an apparent molecular weight of 51,000. After deglycosylation of isolated basolateral rat liver plasma membranes, the apparent molecular weight of Ntcp decreased to 33,500. In intact rat liver, Ntcp was selectively localized at the basolateral surface domain of hepatocytes. In short-term cultured hepatocytes, a positive surface immunoreaction was only obtained in detergent-permeabilized cell cultures. In stably transfected Chinese hamster ovary cells, the surface expression of immunopositive Ntcp was associated with Na(+)-dependent taurocholate uptake activity. CONCLUSIONS: Native Ntcp represents a glycoprotein of the basolateral hepatocyte plasma membrane with its C-terminal end facing the intracellular compartment. Furthermore, surface expression of Ntcp is a prerequisite for Na(+)-dependent taurocholate uptake to occur, thus providing further proof for its bile acid transport function in rat liver.

Animals↗
PubMed1994

Effect of obstructive cholestasis on membrane traffic and domain-specific expression of plasma membrane proteins in rat liver parenchymal cells.

We investigated the effect of bile duct ligation and its release on membrane traffic and plasma membrane protein distribution in rat hepatocytes. Immunofluorescence studies with monoclonal antibodies against six domain-specific surface antigens revealed that bile duct ligation leads to an accumulation of pericanalicular vesicles containing canalicular antigens. All apical antigens could be demonstrated in the basolateral plasma membrane, whereas only one out of three basolateral antigens redistributed to the canalicular plasma membrane. After release of bile duct ligation, the accumulated pericanalicular vesicles disappeared within minutes, whereas the plasma membrane polarity was not restored within 1 hr. Monitoring secretion of polymeric IgA and horseradish peroxidase into bile demonstrated that bile duct ligation also inhibits the transcytotic vesicle pathway and severely impairs the function of tight junctions. In contrast, bile duct ligation appears not to affect the endoplasmic reticulum to basolateral membrane traffic as assessed by determination of newly synthesized albumin and transferrin in serum nor does it influence receptor mediated endocytosis at the basolateral plasma membrane.

Animals↗
PubMed1994

Expression and characterization of a functional rat liver Na+ bile acid cotransport system in COS-7 cells.

A cDNA for the rat liver sodium-dependent bile acid cotransporter was expressed in COS-7 cells to study the functional properties of the translated protein in a mammalian cell line. A 1.2-kb insert was ligated into a pMAMneo vector and transiently transfected using electroporation. After optimal conditions were established, the transiently transfected COS cells were screened with fluorescent-conjugated labeled bile acids for evidence of expression of the cotransporter after 48 h. The uptake of [3H]taurocholate ([3H]TC) was then determined in cells transfected with or without the bile acid insert. Progressive uptake of [3H]TC (0.45 microM) was observed for 30 min in the presence of sodium. In contrast, no uptake of [3H]TC was observed in the absence of sodium, in nontransfected COS cells, or in COS cells transfected with the empty plasmid. Kinetic studies revealed a Michaelis constant (Km) of 29 microM, essentially identical to the Km of this cotransporter described in intact rat hepatocytes and membrane vesicles. Uptake of [3H]TC (5.0 microM) at 5 min (n = 3-6) was inhibited by 100 microM taurochenodeoxycholic acid (81%), tauroursodeoxycholic acid (77%), cholic acid (55%), chenodeoxycholic acid (74%), and ursodeoxycholic acid (56%) but not by 100 microM taurodehydrocholate, 1 mM probenecid, or 100 microM bilirubin. In contrast, bumetanide (500 microM) inhibited [3H]TC uptake by 52%. These studies indicate that the isolated cDNA codes for a physiological bile acid transporter present in rat hepatocytes and that posttranslational factors present in mammalian cells may not be as important in defining properties of this cotransport system.

Animals↗
PubMed1994

Apical and basolateral parathyroid hormone receptors in rat renal cortical membranes.

Brush border (BBM) and basolateral membranes (BLM) of rat renal cortical cells separated by free flow electrophoresis revealed two distinct peaks of BBM-specific leucine aminopeptidase and Na+/K(+)-ATPase for BLM. PTH/PTH-related protein (PTHrP) receptors were identified in BBM and BLM. Specific binding of 125 pM [125I]chicken [Tyr36]-PTHrP-(1-36)amide [chPTHrP-(1-36)] to individual fractions of membranes separated by free flow electrophoresis overlapped with the leucine aminopeptidase and Na+/K(+)-ATPase profiles. Binding to pooled BBM was 53 +/- 5% (mean +/- SEM) of that to BLM (P < 0.01). In BBM and BLM, half-maximal inhibition of binding was obtained with 0.4-0.9 nM chPTHrP-(1-36) and 0.2-0.6 nM rat PTH-(1-34). Guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S; 100 microM) lowered chPTHrP-(1-36) binding to 50% of control levels, and half-maximal inhibition of binding was obtained with 480 and 8 nM GTP gamma S in BBM and BLM, respectively. Cross-linking of the PTH/PTHrP receptors with [125I]chPTHrP-(1-36) modified with N-hydroxysuccinimidyl-4-azidobenzoate revealed indistinguishable doublets of 83 and 73 kilodaltons in both BBM and BLM. Adenylyl cyclase was stimulated 6- and 10-fold by chPTHrP-(1-36) and GTP gamma S, respectively, in BLM and 1.3- and 1.9-fold in BBM. In conclusion, PTH receptors were recognized in both the basolateral and brush border membranes. Different receptor coupling to G-proteins and minimal cAMP stimulation in BBM provide evidence for PTH/PTHrP receptor isotypes and/or different postreceptor activation in BBM and BLM.

Adenylyl Cyclases↗
PubMed1994

Functional characterization of the basolateral rat liver organic anion transporting polypeptide.

To characterize the transport functions of a recently cloned basolateral organic anion transporting polypeptide of rat hepatocytes we performed further kinetic transport and substrate cis-inhibition studies in organic anion-transporting polypeptide-cRNA injected Xenopus laevis oocytes. The studies demonstrate saturable Na(+)-independent sulfobromophthalein (Michaelis-Menten constant, 1.5 mumol/L) and taurocholate (Michaelis-Menten constant, 50 mumol/L) uptake by organic anion-transporting polypeptide. Sulfobromophthalein uptake was inhibited by the following organic anions: 0.01 mmol/L bilirubin (43%), 0.1 mmol/L indocyanine green (81%), 0.1 mmol/L 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS; 52%) and 1 mmol/L probenecid (74%). Competitive inhibition was shown for indocyanine green (inhibition constant about 1.3 mumol/L). Sulfobromophthalein and taurocholate uptakes were also inhibited by cholate, chenodeoxycholate, deoxycholate and ursodeoxycholate, as well as their glycine and taurine conjugates. Organic anion-transporting polypeptide also mediated uptake of glycocholate, tauroursodeoxycholate and taurochenodeoxycholate. No cis-inhibition of sulfobromophthalein uptake was seen in the presence of ATP, para-aminohippuric acid, bumetanide, digitoxin, reduced glutathione, leukotriene C4, nicotinic acid, ouabain, oxalate, rifampicin, succinate or sulfate. Furthermore, radioactively labeled para-aminohippuric acid, alpha-ketoglutarate and reduced glutathione were not taken up by organic anion-transporting polypeptide in cRNA-injected frog oocytes. These data confirm that organic anion-transporting polypeptide represents a novel hepatocellular organic anion uptake system that can mediate Na(+)-independent transport of monovalent (e.g., bile acids) and divalent (e.g., sulfobromophthalein and indocyanine green) cholephilic organic anions.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Evidence for the presence of a phosphatidylcholine translocator in isolated rat liver canalicular plasma membrane vesicles.

In the present study we used the water-soluble short chain phosphatidylcholine analogue L-alpha-dibutyryl-glycero-3-phosphatidylcholine (diC4PC) to investigate the mechanism involved in the canalicular secretion of phospholipids in rat liver. Uptake of 14C-labeled di-C4PC was studied in isolated microsomes as well as in basolateral (sinusoidal) and canalicular plasma membrane vesicles. Saturable uptake of diC4PC into an osmotically active space was observed in microsomes and canalicular membrane vesicles. In contrast, diC4PC uptake into basolateral membrane vesicles could be accounted for by cross-contamination with endoplasmic reticulum and canalicular membrane vesicles. Whereas the Km values for diC4PC uptake (37 degrees C) were similar in microsomes (7.4 +/- 2.6 mM) and canalicular membrane vesicles (8.2 +/- 2.0 mM), the Vmax values were approximately 2-fold higher in canalicular membrane vesicles (29.6 +/- 2.7 nmol/mg of protein x min) than in microsomes (16.7 +/- 2.1 nmol/mg of protein x min). Furthermore, Pronase treatment of the membrane vesicles reduced diC4PC uptake by 34-54% in both subfractions, whereas the D-[14C]glucose-accessible water space was only reduced by approximately 20%. These data provide direct evidence for the presence of a protein-mediated phosphatidylcholine translocating activity in the canalicular membrane of rat hepatocytes. This canalicular "flippase" has kinetic properties similar to those described previously in microsomes and provides a potential pathway for the translocation of bile salt dissolvable biliary phospholipids to the exoplasmic leaflet of the canalicular membrane.

Animals↗
PubMed1993

Phylogenic and ontogenic expression of hepatocellular bile acid transport.

The phylogenic and ontogenic expression of mRNA for the Na+/bile acid cotransporter was determined by Northern analysis utilizing a full-length cDNA probe recently cloned from rat liver. mRNA was detected in several mammalian species, including rat, mouse, and man, but could not be found in livers from nonmammalian species, including chicken, turtle, frog, and small skate. When expression of the bile acid transporter in developing rat liver was studied, mRNA was detected between 18 and 21 days of gestation, at the time when Na(+)-dependent bile acid transport is first detected. Two hepatoma cell lines (HTC and HepG2), the latter of which is known to have lost the Na+/bile acid cotransport system, also did not express mRNA for this transporter. Finally, when mRNA from the lower vertebrate (the small skate) was injected into Xenopus oocytes, only a sodium-independent, chloride-dependent transport system for bile acids was expressed, confirming the integrity of the mRNA and consistent with prior functional studies of bile acid transport in this species. These findings establish that the Na+/bile acid cotransport mRNA is first transcribed in mammalian species, a process that is recapitulated late during mammalian fetal development in rat liver, and that this mRNA is lost in dedifferentiated hepatocytes. In contrast, the mRNA for a multispecific Na+/independent organic anion transport system is transcribed earlier in vertebrate evolution.

Aging↗
PubMed1993

Ethinylestradiol treatment induces multiple canalicular membrane transport alterations in rat liver.

We investigated the effects of 17 alpha-ethinylestradiol treatment of rats on various transport functions in isolated basolateral and canalicular liver plasma membrane vesicles. Both membrane subfractions were purified to a similar degree from control and cholestatic livers. Although moderate membrane lipid alterations were predominantly observed in basolateral vesicles, no change in basolateral Na+/K(+)-ATPase activity was found. Furthermore, while Na(+)-dependent taurocholate uptake was decreased by approximately 40% in basolateral vesicles, the maximal velocity of ATP-dependent taurocholate transport was decreased by 63% in canalicular membranes. In contrast, only minimal changes or no changes at all were observed for electrogenic taurocholate transport in "cholestatic" canalicular membranes and total microsomes, respectively. However, canalicular vesicles from cholestatic livers also exhibited marked reductions in ATP-dependent transport of S-(2,4-dinitrophenyl)glutathione and in Na(+)-dependent uptake of adenosine, while in the same vesicles HCO3-/SO4- exchange and Na+/glycine cotransport activities were markedly stimulated. These data show that in addition to the previously demonstrated sinusoidal transport abnormalities ethinylestradiol-induced cholestasis is also associated with multiple canalicular membrane transport alterations in rat liver. Hence, functional transport alterations at both polar surface domains might ultimately be responsible for the inhibitory effects of estrogens on the organic anion excretory capacity and on bile formation in rat liver.

Adenosine↗