scholarly journals Mg2+ transport in plasma membrane vesicles of renal epithelium of the Mozambique tilapia (Oreochromis mossambicus)

1997 ◽  
Vol 200 (13) ◽  
pp. 1931-1939 ◽  
Author(s):  
M Bijvelds ◽  
Z Kolar ◽  
S Bonga ◽  
G Flik
Keyword(s):  
Plasma Membrane ◽  
Brush Border ◽  
Transport Mechanism ◽  
Apical Membrane ◽  
Basolateral Membrane ◽  
Membrane Vesicles ◽  
Oreochromis Mossambicus ◽  
Renal Epithelium ◽  
Mozambique Tilapia ◽  
Vesicular Membrane

To elucidate the mechanisms involved in Mg2+ transport at the apical and basolateral poles of the renal tubular epithelium, apical and basolateral plasma membrane vesicle preparations were derived from kidney tissue of freshwater- and seawater-adapted Mozambique tilapia Oreochromis mossambicus. Brush-border preparations were enriched 15.8-fold in alkaline phosphatase activity and consisted almost exclusively of right-side-out membrane vesicles. Basolateral membrane preparations were enriched 7.5-fold in Na+/K+-ATPase activity and contained resealed vesicles and leaky membrane fragments. Mg2+ association with brush-border and basolateral plasma membranes, traced using radioactive 27Mg, occurred in an osmotically active space. In all instances, Mg2+ binding to the vesicular membrane was low compared with the vesicular uptake. Mg2+ equilibration across the vesicular membrane of brush-border preparations was rapid and sensitive to the presence of extravesicular Ca2+, suggesting that the apical membrane of the renal epithelium contains a transport pathway for divalent cations. Application of various ionic gradients did not affect vesicular Mg2+ transport in apical and basolateral membrane preparations, suggesting the presence of an ion-coupled transport mechanism. ATP or ATP--S did not stimulate Mg2+ fluxes, indicating that Mg2+ transport does not proceed via an ATP-driven or activated transporter. In these aspects, vesicular Mg2+ transport was similar in seawater and freshwater preparations. These results suggest that the apical membrane of renal epithelial cells lacks an active secretory Mg2+ transport mechanism. We propose that the Mg2+ conductivity of the apical membrane reflects a route for downhill Mg2+ entry and is involved in renal Mg2+ reabsorption.

scholarly journals Calcium pump activities in the kidneys of Oreochromis mossambicus

1995 ◽  
Vol 198 (6) ◽  
pp. 1351-1357
Author(s):  
M Bijvelds ◽  
A Heijden ◽  
G Flik ◽  
P Verbost ◽  
Z Kolar ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Membrane Fraction ◽  
Transport Mechanism ◽  
Physiological Role ◽  
Membrane Vesicles ◽  
Oreochromis Mossambicus ◽  
High Affinity ◽  
Pump Activity ◽  
Basolateral Plasma Membrane ◽  
Euryhaline Teleost

The mechanism that underlies transcellular Ca2+ reabsorption in the kidney of the euryhaline teleost Oreochromis mossambicus was studied. Preparations of membrane vesicles made from the kidneys of freshwater- and seawater-adapted fish were more than sevenfold enriched in the basolateral plasma membrane marker Na+/K+-ATPase. Significant recovery of NADH­ cytochrome c reductase enzyme activity and of oxalate-stimulated Ca2+ pump activities in the membrane preparations indicated that the membrane fraction was of endoplasmic reticular origin. Indeed, thapsigargin specifically inhibited Ca2+ pump activity that could be attributed to oxalate-permeable endoplasmic reticular fragments. Kinetic analysis of thapsigargin-insensitive Ca2+ pump activity indicated the existence of a homogeneous, high-affinity, ATP-driven Ca2+ pump. No Na+-driven Ca2+ transport mechanism could be demonstrated. Plasma membrane Ca2+ pump activity was 56 % lower in preparations from seawater-adapted fish than in preparations from freshwater-adapted fish, suggesting a physiological role for this Ca2+ pump activity in renal Ca2+ handling by euryhaline species, with an involvement in the regulation of Ca2+ reabsorption.

An l-proline-dependent proton flux is located at the apical membrane level of the eel enterocytes

2000 ◽  
Vol 279 (5) ◽  
pp. R1619-R1624 ◽  
Author(s):  
L. Ingrosso ◽  
S. Marsigliante ◽  
V. Zonno ◽  
C. Storelli ◽  
S. Vilella
Keyword(s):  
Brush Border ◽  
Apical Membrane ◽  
Basolateral Membrane ◽  
Membrane Vesicles ◽  
Proton Flux ◽  
Coupling Mechanism ◽  
Acetoxymethyl Ester ◽  
Extracellular Medium ◽  
Imino Acid ◽  
Membrane Level

This study has demonstrated the existence of an l-proline-dependent (Na independent) proton flux at the apical membrane level of the eel intestinal absorbing cells. Using isolated eel enterocytes and the pH-sensitive fluorescent dye 2′,7′-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein, acetoxymethyl ester (BCECF), it was shown that a 20 mM concentration of the imino acid l-proline in the extracellular medium determined an intracellular acidification of ∼0.28 pH units. However, neither sucrose nor other amino acids were able to significantly acidify the resting intracellular pH. A hyperbolic relationship between extracellular proline concentration and intracellular proton accumulation was observed. Using both isolated brush-border and basolateral membrane vesicles, it was demonstrated that this proline-proton cotransport mechanism was located at the apical membrane level only. In addition, the existence of a coupling mechanism between proline and proton fluxes was demonstrated by the observation that, in brush-border membrane vesicles, the presence of a pH gradient (pHin > pHout) stimulated the uptake ofl-proline.

Electrogenic ATP-dependent Cl- transport by plasma membrane vesicles from Aplysia intestine

1988 ◽  
Vol 254 (1) ◽  
pp. R127-R133 ◽  
Author(s):  
G. A. Gerencser
Keyword(s):  
Plasma Membrane ◽  
Transport Process ◽  
Transport Mechanism ◽  
Plasma Membranes ◽  
Membrane Vesicles ◽  
Sucrose Density Gradient ◽  
Differential Centrifugation ◽  
Plasma Membrane Vesicles ◽  
Cl Transport ◽  
Vesicular Membrane

A Cl--stimulated adenosinetriphosphatase (ATPase) activity and an ATP-dependent Cl- transport process were found in Aplysia enterocyte plasma membranes. In an attempt to further elucidate this transport process plasma membrane vesicles from Aplysia enterocytes were prepared utilizing differential centrifugation and sucrose density gradient techniques. Electrogenicity of the ATP-dependent Cl- transport was confirmed in three ways. First, an inwardly directed valinomycin-induced K+ diffusion potential, making the vesicle interior electrically positive, enhanced ATP-driven Cl- uptake compared with vesicles lacking the ionophore. Second, ATP plus Cl- increased intravesicular negativity measured by lipophilic triphenylmethylphosphonium distribution across the vesicular membrane. Third, both vanadate and thiocyanate inhibited the ATP plus Cl--dependent intravesicular negativity. These results are consistent with the hypothesis that the active electrogenic Cl- transport mechanism in Aplysia intestine could be a Cl--stimulated ATPase found in the enterocyte plasma membrane.

Basolateral dipeptide transport by the intestine of the teleost Oreochromis mossambicus

1996 ◽  
Vol 270 (5) ◽  
pp. R948-R954 ◽  
Author(s):  
M. Thamotharan ◽  
V. Zonno ◽  
C. Storelli ◽  
G. A. Ahearn
Keyword(s):  
Brush Border ◽  
Specific Activity ◽  
Gradient Centrifugation ◽  
Basolateral Membrane ◽  
Specific Inhibitor ◽  
Membrane Vesicles ◽  
Oreochromis Mossambicus ◽  
Peptide Transport ◽  
Transport Systems ◽  
Uptake Kinetic

Transport characteristics of [14C]glycylsarcosine ([14C]Gly-Sar) were measured in herbivorous tilapi (Oreochromis mossambicus) intestinal basolateral membrane vesicles (BLMV) purified with Percoll gradient centrifugation. Specific activity of the vesicle Na(+)-K(+)-adenosinetriphos- phatase was increased 12-fold, whereas specific activity of the brush-border enzyme alkaline phosphatase was enriched only by 0.8-fold. [14C]Gly-Sar uptake was stimulated by increasing concentrations of extravesicular protons rather than by a transmembrane proton gradient. A transmembrane K+ diffusion potential (inside negative) did not stimulate [14C]Gly-Sar uptake above that observed with short-circuited vesicles. An inwardly directed Na+ gradient had no effect on peptide uptake. Kinetic analysis of basolateral transport rate revealed that the transport occurred by a saturable process conforming to Michaelis-Menten kinetics [Kt [concentration of [14C]Gly-Sar that yielded one-half of maximal influx (Jmax)] = 13.27 +/- 3.80 mM, Jmax = 15,155 +/- 3,096 pmol.mg protein-1.6 s-1]. The basolateral transporter was insensitive to diethylpyrocarbonate (DEP), a specific inhibitor of proton-coupled peptide transport systems. [14C]Gly-Sar influx into tilapia BLMV showed cis-inhibition by several other dipeptides, suggesting that the [14C]Gly-Sar transporter was shared by other peptides too. These observations strongly suggest that the basolateral intestinal dipeptide transporter in herbivorous fishes is distinctly different from either the high- or low-affinity brush-border transporter. It is proton dependent, electroneutral, sodium independent and accepts a wide variety of dipeptides.

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.

scholarly journals Characterization of sodium-dependent and sodium-independent nucleoside transport systems in rabbit brush-border and basolateral plasma-membrane vesicles from the renal outer cortex

1989 ◽  
Vol 264 (1) ◽  
pp. 223-231 ◽  
Author(s):  
T C Williams ◽  
A J Doherty ◽  
D A Griffith ◽  
S M Jarvis
Keyword(s):  
Brush Border ◽  
Basolateral Membrane ◽  
Membrane Vesicles ◽  
Nucleoside Transport ◽  
Transport Systems ◽  
Facilitated Diffusion ◽  
Basolateral Membrane Vesicles ◽  
Plasma Membrane Vesicles ◽  
Outer Cortex ◽  
Overshoot Phenomenon

The transport of uridine into rabbit renal outer-cortical brush-border and basolateral membrane vesicles was compared at 22 degrees C. Uridine was taken up into an osmotically active space in the absence of metabolism for both types of membrane vesicles. Uridine influx by brush-border membrane vesicles was stimulated by Na+, and in the presence of inwardly directed gradients of Na+ a transient overshoot phenomenon was observed, indicating active transport. Kinetic analysis of the saturable Na+-dependent component of uridine flux indicated that it was consistent with Michaelis-Menten kinetics (Km 12 +/- 3 microM, Vmax. 3.9 +/- 0.9 pmol/s per mg of protein). The sodium:uridine coupling stoichiometry was found to be consistent with 1:1 and involved the net transfer of positive charge. In contrast, uridine influx by basolateral membrane vesicles was not dependent on the cation present and was inhibited by nitrobenzylthioinosine (NBMPR). NBMPR-sensitive uridine transport was saturable (Km 137 +/- 20 microM, Vmax. 5.2 +/- 0.6 pmol/s per mg of protein). Inhibition of uridine flux by NBMPR was associated with high-affinity binding of NBMPR to the basolateral membrane (Kd 0.74 +/- 0.46 nM). Binding of NBMPR to these sites was competitively blocked by adenosine and uridine. These results indicate that uridine crosses the brush-border surface of rabbit proximal renal tubule cells by Na+-dependent pathways, but permeates the basolateral surface by NBMPR-sensitive facilitated-diffusion carriers.

Urate transport in the proximal tubule: in vivo and vesicle studies

1985 ◽  
Vol 249 (6) ◽  
pp. F789-F798 ◽  
Author(s):  
A. M. Kahn ◽  
E. J. Weinman
Keyword(s):  
Proximal Tubule ◽  
Brush Border ◽  
Anion Exchanger ◽  
Basolateral Membrane ◽  
Membrane Vesicles ◽  
Transport Processes ◽  
Basolateral Membrane Vesicles ◽  
Urate Transport ◽  
Rat Proximal Tubule

The transport of urate in the mammalian nephron is largely confined to the proximal tubule. Depending on the species, net reabsorption or net secretion is observed. The rat, like the human and the mongrel dog, demonstrates net reabsorption of urate and has been the most extensively studied species. The unidirectional reabsorption and secretion of urate in the rat proximal tubule occur via a passive and presumably paracellular route and by a mediated transcellular route. The reabsorption of urate, and possibly its secretion, can occur against an electrochemical gradient. A variety of drugs and other compounds affect the reabsorption and secretion of urate. The effects of these agents depend on their site of application (luminal or blood), concentration, and occasionally their participation in transport processes that do not have affinity for urate. Recent studies with renal brush border and basolateral membrane vesicles from the rat and brush border vesicles from the dog have determined the mechanisms for urate transport across the luminal and antiluminal membranes of the proximal tubule cell. Brush border membrane vesicles contain an anion exchanger with affinity for urate, hydroxyl ion, bicarbonate, chloride, lactate, p-aminohippurate (PAH), and a variety of other organic anions. Basolateral membrane vesicles contain an anion exchanger with affinity for urate and chloride but not for PAH. Both membrane vesicle preparations also permit urate translocation by simple diffusion. A model for the transcellular reabsorption and secretion of urate in the rat proximal tubule is proposed. This model is based on the vesicle studies, and it can potentially explain the majority of urate transport data obtained with in vivo techniques.

Na+-independent D-glucose transport in rabbit renal basolateral membranes

1988 ◽  
Vol 254 (5) ◽  
pp. F711-F718 ◽  
Author(s):  
P. T. Cheung ◽  
M. R. Hammerman
Keyword(s):  
Glucose Uptake ◽  
Glucose Transport ◽  
Brush Border ◽  
Glucose Transporter ◽  
Cytochalasin B ◽  
Basolateral Membrane ◽  
Membrane Vesicles ◽  
Rabbit Kidney ◽  
Basolateral Membrane Vesicles ◽  
Proximal Tubular

To define the mechanism by which glucose is transported across the basolateral membrane of the renal proximal tubular cell, we measured D-[14C]glucose uptake in basolateral membrane vesicles from rabbit kidney. Na+-dependent D-glucose transport, demonstrable in brush-border vesicles, could not be demonstrated in basolateral membrane vesicles. In the absence of Na+, the uptake of D-[14C]glucose in basolateral vesicles was more rapid than that of L-[3H]glucose over a concentration range of 1-50 mM. Subtraction of the latter from the former uptakes revealed a saturable process with apparent Km of 9.9 mM and Vmax of 0.80 nmol.mg protein-1.s-1. To characterize the transport component of D-glucose uptake in basolateral vesicles, we measured trans stimulation of 2 mM D-[14C]glucose entry in the absence of Na+. Trans stimulation could be effected by preloading basolateral vesicles with D-glucose, 2-deoxy-D-glucose, or 3-O-methyl-D-glucose, but not with L-glucose or alpha-methyl-D-glucoside. Trans-stimulated D-[14C]glucose uptake was inhibited by 0.1 mM phloretin or cytochalasin B but not phlorizin. In contrast, Na+-dependent D-[14C]glucose transport in brush-border vesicles was inhibited by phlorizin but not phloretin or cytochalasin B. Our findings are consistent with the presence of a Na+-independent D-glucose transporter in the proximal tubular basolateral membrane with characteristics similar to those of transporters present in nonepithelial cells.

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)

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