Bilitranslocase localization and function in basolateral plasma membrane of renal proximal tubule in rat

1990 ◽  
Vol 259 (4) ◽  
pp. F559-F564 ◽  
Author(s):  
M. M. Elias ◽  
G. C. Lunazzi ◽  
S. Passamonti ◽  
B. Gazzin ◽  
M. Miccio ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Competitive Inhibition ◽  
Membrane Vesicles ◽  
Plasma Membrane Vesicles ◽  
Order Of Magnitude ◽  
Single Positive ◽  
Basolateral Plasma Membrane ◽  
Renal Tubular ◽  
And Function ◽  
Monospecific Antibodies

Bilirubin and phthalein dyes are taken up by the liver via a carrier-mediated mechanism operated at least in part by bilitranslocase (BTL). Because they also undergo renal transport, the presence and function of BTL was investigated in rat renal tubular plasma membrane vesicles. Transport of sulfobromophthalein (BSP) was enriched in basolateral domain of plasma membrane and followed the distribution pattern of Na(+)-K(+)-ATPase but not of gamma-glutamyltransferase. BSP uptake was inhibited by addition of monospecific antibodies raised against hepatic BTL. As in liver vesicles, BSP transport was electrogenic, being greatly accelerated by addition of valinomycin in presence of an inwardly directed K+ gradient. Apparent Km of BSP transport was 17 +/- 2 microM (n = 3 expts), one order of magnitude higher than that measured in liver; however, Vmax was similar to that described in liver vesicles (429 +/- 18 nmol BSP.mg protein-1.min-1, n = 3 expts). Competitive inhibition was observed with both unconjugated bilirubin (Ki, 2.9 +/- 0.2 microM) and rifamycin SV (Ki, 76 +/- 10 microM), known competitors for hepatic BTL-mediated transport of BSP. Immunoblotting studies with anti-BTL monospecific antibodies revealed presence of a single positive band only in basolateral-enriched membrane fraction; its apparent molecular mass was 37 kDa, virtually identical to that of hepatic protein. Immunohistochemistry confined presence of BTL to renal proximal tubules (RPT) We conclude that BTL is present in basolateral plasma membrane of RPT cells. Lower affinity of renal, compared with hepatic protein, for substrates might explain the marginal role of kidney in plasma clearance of bilirubin and cholephilic dyes.

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 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

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)

Multispecific anion exchange in basolateral (sinusoidal) rat liver plasma membrane vesicles

1986 ◽  
Vol 251 (5) ◽  
pp. G656-G664 ◽  
Author(s):  
G. Hugentobler ◽  
P. J. Meier
Keyword(s):  
Plasma Membrane ◽  
Rat Liver ◽  
Driving Forces ◽  
Competitive Inhibition ◽  
Membrane Vesicles ◽  
Ph Gradient ◽  
Disulfonic Acid ◽  
Sulfate Uptake ◽  
Plasma Membrane Vesicles ◽  
Liver Plasma Membrane

The mechanisms and driving forces for hepatic uptake of sulfate were investigated in basolateral (sinusoidal) rat liver plasma membrane vesicles. A transmembrane pH difference (pH 8.0 inside, 6.0 outside) stimulated sulfate uptake above equilibrium (“overshoot”). This pH gradient-stimulated sulfate uptake was saturable with increasing concentrations of sulfate and could be inhibited by probenecid, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS), carbonyl cyanide p-(trifluoromethoxy)-phenylhydrazone, and nigericin. At low buffer concentrations and pH 6.0 an inwardly directed sodium gradient also stimulated sulfate uptake. This sodium-dependent sulfate uptake could be inhibited by amiloride and DIDS, indicating indirect coupling of sodium and sulfate flux through concomitant sodium-proton and sulfate-hydroxyl exchange. Cisinhibition of initial pH gradient-stimulated sulfate uptake, as well as transstimulation of sulfate uptake under pH-equilibrated conditions (pH 7.5 inside and outside), were observed with sulfate, thiosulfate, oxalate, and succinate, but not with chloride, bicarbonate, acetate, lactate, pyruvate, p-aminohippurate, citrate, glutamate, aspartate, and taurocholate. Furthermore, cholate and sulfobromophthalein exhibited competitive inhibition of pH gradient-stimulated sulfate uptake. In addition, an inside-to-outside hydroxyl gradient also stimulated uptake of cholate and this pH gradient-sensitive portion of cholate uptake was inhibited by extravesicular sulfate. In contrast to basolateral membranes, no evidence for multispecific sulfate-hydroxyl exchange was found in canalicular plasma membrane vesicles.

scholarly journals Calcium transport mechanisms in basolateral plasma membrane-enriched vesicles from rat parotid gland

1985 ◽  
Vol 227 (1) ◽  
pp. 239-245 ◽  
Author(s):  
T Takuma ◽  
B L Kuyatt ◽  
B J Baum
Keyword(s):  
Plasma Membrane ◽  
Parotid Gland ◽  
Gradient Centrifugation ◽  
Membrane Vesicles ◽  
Transport Systems ◽  
Transport Mechanisms ◽  
Plasma Membrane Vesicles ◽  
Rat Parotid Gland ◽  
Basolateral Plasma Membrane ◽  
Rat Parotid

Ca2+ transport was studied by using basolateral plasma membrane vesicles from rat parotid gland prepared by a Percoll gradient centrifugation method. In these membrane vesicles, there were two Ca2+ transport systems; Na+/Ca2+ exchange and ATP-dependent Ca2+ transport. An outwardly directed Na+ gradient increased Ca2+ uptake. Ca2+ efflux from Ca2+-preloaded vesicles was stimulated by an inwardly directed Na+ gradient. However, Na+/Ca2+ exchange did not show any ‘uphill’ transport of Ca2+ against its own gradient. ATP-dependent Ca2+ transport exhibited ‘uphill’ transport. An inwardly directed Na+ gradient also decreased Ca2+ accumulation by ATP-dependent Ca2+ uptake. The inhibition of Ca2+ accumulation was proportional to the external Na+ level. Na+/Ca2+ exchange was inhibited by monensin, tetracaine and chlorpromazine, whereas ATP-dependent Ca2+ transport was inhibited by orthovanadate, tetracaine and chlorpromazine. Oligomycin had no effect on either system. These results suggest that in the parotid gland cellular free Ca2+ is extruded mainly by an ATP-dependent Ca2+ transport system, and Na+/Ca2+ exchange may modify the efficacy of that system.

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