Taurocholate transport by basolateral plasma membrane vesicles isolated from developing rat liver

1985 ◽  
Vol 248 (6) ◽  
pp. G648-G654
Author(s):  
F. J. Suchy ◽  
S. M. Courchene ◽  
B. L. Blitzer
Keyword(s):  
Plasma Membrane ◽  
Rat Liver ◽  
Basolateral Membrane ◽  
Membrane Vesicles ◽  
Marker Enzyme ◽  
Marker Enzymes ◽  
Plasma Membrane Vesicles ◽  
Adult Rat ◽  
Basolateral Plasma Membrane ◽  
Taurocholate Transport

Taurocholate transport was characterized in basolateral plasma membrane vesicles prepared from the livers of 14-day-old Sprague-Dawley rats using a self-generating Percoll gradient method. Liver plasma membrane protein yield, intravesicular volume, and enrichments of various marker enzymes were similar to those obtained for vesicles from adult rat liver. The basolateral marker enzyme Na+-K+-ATPase was enriched 26-fold in the suckling rat basolateral membrane fraction while the bile canalicular marker enzymes alkaline phosphatase and Mg2+-ATPase were enriched only 3- and 5-fold, respectively. The activities of marker enzymes for endoplasmic reticulum, mitochondria, or lysosomes were not enriched compared with homogenate. In the presence of an inwardly directed 100 mM Na+ gradient, vesicle accumulation of taurocholate transiently reached a concentration 1.5- to 2-fold higher than that at equilibrium ("overshoot") in suckling and adult membrane vesicles, but the initial rate of taurocholate entry and peak intravesicular accumulation were markedly decreased in suckling compared with adult membrane vesicles. In the presence of an inwardly directed 100 mM K+ gradient, the rate of uptake was slower, and no overshoot occurred in either suckling or adult rat vesicles. The decreased rate of Na+-coupled taurocholate uptake by membrane vesicles from suckling rat liver could not be explained on the basis of more rapid dissipation of the transmembrane Na+ gradient. Kinetic studies demonstrated saturable, Na+-dependent taurocholate uptake for both suckling and adult vesicles. However, the Vmax for taurocholate uptake in suckling rat vesicles was less than half of the adult rate (2.46 +/- 0.13 vs. 5.25 +/- 0.22 nmol X mg prot-1 X min-1, respectively, P less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

Na(+)-H+ exchange in basolateral plasma membrane vesicles from neonatal rat liver

1990 ◽  
Vol 259 (3) ◽  
pp. G334-G339 ◽  
Author(s):  
A. L. Goodrich ◽  
F. J. Suchy
Keyword(s):  
Plasma Membrane ◽  
Rat Liver ◽  
Basolateral Membrane ◽  
Membrane Vesicles ◽  
Neonatal Rat ◽  
Plasma Membrane Vesicles ◽  
Adult Rat ◽  
Age Related ◽  
Basolateral Plasma Membrane ◽  
22Na Uptake

Basolateral plasma membrane vesicles (BLMV) were used to determine the activity of the Na(+)-H+ exchanger in neonatal compared with adult rat liver. Imposition of an outwardly directed H+ gradient (pH 5.5in/7.5out) markedly stimulated the uptake of 1 mM 22Na+ by adult and neonatal BLMV above rates observed under pH-equilibrated conditions (pH 7.5in/7.5out) and resulted in a transient uphill accumulation ("overshoot") of Na+ to levels 1.5- to 2-fold higher than at equilibrium. 22Na+ uptake via Na(+)-H+ exchange (identified as the amiloride-sensitive component of total Na+ flux) was greater in BLMV from neonatal compared with adult liver under both pH-gradient and -equilibrated conditions. These age-related differences in Na+ uptake could not be explained by membrane potential effects or by differences in membrane permeability to H+. Kinetic analysis revealed a saturable process with a similar Km for Na+ (8.6 +/- 1.6 vs. 8.1 +/- 1.5 mM) but with a 2.5-fold higher Vmax (8.06 +/- 0.67 vs. 3.20 +/- 0.40 nmol.mg protein-1.min-1; P less than 0.01) in neonatal vs. adult vesicles. We conclude from these studies that basolateral membrane vesicles from neonatal rat liver exhibit enhanced Na(+)-H+ exchange as a result of an increased number or translocation rate of carriers in the plasma membrane.

scholarly journals Albumin binding of unconjugated [3H]bilirubin and its uptake by rat liver basolateral plasma membrane vesicles

1996 ◽  
Vol 316 (3) ◽  
pp. 999-1004 ◽  
Author(s):  
Lorella PASCOLO ◽  
Savino DEL VECCHIO ◽  
Ronald K. KOEHLER ◽  
J. Enrique BAYON ◽  
Cecile C. WEBSTER ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Rat Liver ◽  
Basolateral Membrane ◽  
Membrane Vesicles ◽  
Plasma Membrane Vesicles ◽  
Experimental Conditions ◽  
Saturation Kinetics ◽  
Basolateral Plasma Membrane ◽  
Component C ◽  
Uptake Studies

Using highly purified unconjugated [3H]bilirubin (UCB), we measured UCB binding to delipidated human serum albumin (HSA) and its uptake by basolateral rat liver plasma membrane vesicles, in both the absence and presence of an inside-positive membrane potential. Free UCB concentrations ([Bf]) were calculated from UCB–HSA affinity constants (K´f), determined by five cycles of ultrafiltration through a Centricon-10 device (Amicon) of the same solutions used in the uptake studies. At HSA concentrations from 12 to 380 μM, K´f (litre/mol) was inversely related to [HSA], irrespective of the [Bt]/[HSA] ratio. K´f was 2.066×106+(3.258×108/[HSA]). When 50 mM KCl was iso-osmotically substituted for sucrose, the K´f value was significantly lower {2.077×106+(1.099×108/[HSA])}. The transport occurred into an osmotic-sensitive space. Below saturation ([Bf] ⩽ 65 nM), both electroneutral and electrogenic components followed saturation kinetics with respect to [Bf], with Km values of 28±7 and 57±8 nM respectively (mean±S.D., n = 3, P < 0.001). The Vmax was greater for the electrogenic than for the electroneutral component (112±12 versus 45±4 pmol of UCB·mg-1 of protein·15 s-1, P < 0.001). Sulphobromophthalein trans-stimulated both electrogenic (61%) and electroneutral (72%) UCB uptake. These data indicate that: (a) as [HSA] increases, K´f decreases, thus increasing the concentration of free UCB. This may account for much of the enhanced hepatocytic uptake of organic anions observed with increasing [HSA]. (b) UCB is taken up at the basolateral membrane of the hepatocyte by two systems with Km values within the range of physiological free UCB levels in plasma. The electrogenic component shows a lower affinity and a higher capacity than the electroneutral component. (c) It is important to calculate the actual [Bf] using a K´f value determined under the same experimental conditions (medium and [HSA]) used for the uptake studies.

Taurocholate transport and Na+-K+-ATPase activity in fetal and neonatal rat liver plasma membrane vesicles

1986 ◽  
Vol 251 (5) ◽  
pp. G665-G673 ◽  
Author(s):  
F. J. Suchy ◽  
J. C. Bucuvalas ◽  
A. L. Goodrich ◽  
M. S. Moyer ◽  
B. L. Blitzer
Keyword(s):  
Plasma Membrane ◽  
Atpase Activity ◽  
Rat Liver ◽  
Maximum Velocity ◽  
Membrane Vesicles ◽  
Neonatal Rat ◽  
Marker Enzyme ◽  
Marker Enzymes ◽  
Plasma Membrane Vesicles ◽  
Early Maturation

The ontogenesis of Na+-K+-ATPase activity and Na+-taurocholate cotransport was studied in basolateral plasma membrane vesicles from fetal and neonatal rat liver. Membrane vesicles from each age group were 30-fold enriched in the basolateral marker enzyme Na+-K+-ATPase, 4- to 7-fold enriched in the bile canalicular membrane marker enzymes alkaline phosphatase and Mg2+ ATPase, and not significantly enriched in activities of marker enzymes for intracellular organelles. Na+-K+-ATPase activity was significantly lower in basolateral membranes from late fetal (day 21–22) and neonatal (day 1) rat liver. Kinetic analysis of Na+-K+-ATPase activity at various concentrations of ATP revealed that the maximum velocity of enzyme reaction (Vmax) for Na+-K+-ATPase was 70 and 90% of adult activity in the fetus and the neonate, respectively. The ATP Km was significantly lower in the neonate than the adult, suggesting a higher affinity of the neonatal enzyme for ATP. In contrast to the early maturation of Na+-K+-ATPase, transport of taurocholate was markedly lower in both fetal and neonatal vesicles compared with the adult. Taurocholate uptake on day 19 of gestation did not differ in the presence of a Na+ or K+ gradient, and uphill transport, as indicated by an overshoot, did not occur. On day 20 taurocholate uptake was stimulated by a Na+ compared with a K+ gradient, and accumulation of isotope above equilibrium was demonstrated. Na+-dependent transport of taurocholate by late fetal (day 22) and neonatal vesicles was saturable but the Vmax at each age was significantly lower and the apparent Km higher in developing compared with adult membrane vesicles.(ABSTRACT TRUNCATED AT 250 WORDS)

Taurocholate transport by basolateral plasma membrane vesicles isolated from human liver

Hepatology ◽  
1989 ◽  
Vol 10 (4) ◽  
pp. 447-453 ◽  
Author(s):  
Donald A. Novak ◽  
Frederick C. Ryckman ◽  
Frederick J. Suchy
Keyword(s):  
Plasma Membrane ◽  
Human Liver ◽  
Membrane Vesicles ◽  
Plasma Membrane Vesicles ◽  
Basolateral Plasma Membrane ◽  
Taurocholate Transport

Effect of endotoxin on bile acid transport in rat liver: a potential model for sepsis-associated cholestasis

1996 ◽  
Vol 271 (1) ◽  
pp. G137-G146 ◽  
Author(s):  
R. H. Moseley ◽  
W. Wang ◽  
H. Takeda ◽  
K. Lown ◽  
L. Shick ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Bile Acid ◽  
Rat Liver ◽  
Membrane Vesicles ◽  
Tnf Alpha ◽  
Mrna Levels ◽  
Acid Transport ◽  
Plasma Membrane Vesicles ◽  
Bile Acid Transport ◽  
Taurocholate Transport

Intrahepatic cholestasis in the setting of extrahepatic bacterial infection has been attributed to the effects of endotoxin and cytokines such as tumor necrosis factor-alpha (TNF-alpha) on bile acid transport. To define the mechanism of sepsis-associated cholestasis, taurocholate transport was examined in basolateral (bLPM) and canalicular (cLPM) rat liver plasma membrane vesicles derived from control and endotoxin [lipopolysaccharide (LPS)]-treated animals and in plasma membrane vesicles prepared after TNF-alpha treatment. Na(+)-dependent [3H]taurocholate uptake and both membrane-potential-dependent and ATP-dependent [3H]taurocholate transport were reduced in bLPM and cLPM vesicles, respectively, after LPS treatment. In membrane vesicles from TNF-alpha-treated animals, Na(+)-dependent [3H]taurocholate uptake was also reduced. Northern blot hybridization, using cDNA probes for the putative sinusoidal bile acid transporter (Ntcp) and canalicular ecto-adenosinetriphosphatase, demonstrated decreased mRNA levels after LPS and TNF-alpha treatment. Immunoblot analysis of membrane extracts from LPS-treated animals revealed decreased levels of these putative bile acid transporters. Impaired bile acid transport at the sinusoidal and canalicular membrane domains by these and other mediators of the inflammatory response may account for sepsis-associated cholestasis.

scholarly journals Structural and functional polarity of canalicular and basolateral plasma membrane vesicles isolated in high yield from rat liver.

1984 ◽  
Vol 98 (3) ◽  
pp. 991-1000 ◽  
Author(s):  
P J Meier ◽  
E S Sztul ◽  
A Reuben ◽  
J L Boyer
Keyword(s):  
Plasma Membrane ◽  
Adenylate Cyclase ◽  
Rat Liver ◽  
High Speed ◽  
Membrane Vesicles ◽  
Fold Increase ◽  
High Yield ◽  
Marker Enzyme ◽  
Gamma Glutamyl Transpeptidase ◽  
Plasma Membrane Vesicles

A method has been developed for routine high yield separation of canalicular (cLPM) from basolateral (blLPM) liver plasma membrane vesicles of rat liver. Using a combination of rate zonal floatation (TZ-28 zonal rotor, Sorvall) and high speed centrifugation through discontinuous sucrose gradients, 9-16 mg of cLPM and 15-28 mg of blLPM protein can be isolated in 1 d. cLPM are free of the basolateral markers Na+/K+-ATPase and glucagon-stimulatable adenylate cyclase activities, but are highly enriched with respect to homogenate in the "canalicular marker" enzyme activities leucylnaphthylamidase (48-fold), gamma-glutamyl-transpeptidase (60-fold), 5'-nucleotidase (64-fold), alkaline phosphatase (71-fold), Mg++-ATPase (83-fold), and alkaline phosphodiesterase I (116-fold). In contrast, blLPM are 34-fold enriched in Na+/K+-ATPase activity, exhibit considerable glucagon-stimulatable adenylate cyclase activity, and demonstrate a 4- to 15-fold increase over homogenate in the various "canalicular markers." cLPM have a twofold higher content of sialic acids, cholesterol; and sphingomyelin compared with blLPM. At least three canalicular-(130,000, 100,000, and 58,000 mol wt) and several basolateral-specific protein bands have been detected after SDS PAGE of the two LPM subfractions. Specifically, the immunoglobin A-binding secretory component is restricted to blLPM as demonstrated by immunochemical techniques. These data indicate virtually complete separation of basolateral from canalicular LPM and demonstrate multiple functional and compositional polarity between the two surface domains of hepatocytes.

Uridine transport in basolateral plasma membrane vesicles from rat liver

1992 ◽  
Vol 128 (3) ◽  
Author(s):  
Bonaventura Ruiz-Montasell ◽  
F.Javier Casado ◽  
Antonio Felipe ◽  
Mar�al Pastor-Anglada
Keyword(s):  
Plasma Membrane ◽  
Rat Liver ◽  
Membrane Vesicles ◽  
Plasma Membrane Vesicles ◽  
Basolateral Plasma Membrane
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