Intravesicular Space Research Articles

Mechanism of neomycin stimulation of d-glucose uptake in rabbit intestinal brush border membrane

In order to study the effect of the antibiotic neomycin on the intestinal epithelium, d-glucose was used as a probe molecule and its transport into rabbit brush border membrane vesicles was measured by a rapid filtration method. Treatment of the epithelium with neomycin sulfate prior to the preparation of the brush border membrane enhanced the d-glucose uptake, whereas neutral N-acetylated neomycin did not. This action of neomycin was related to its polycationic character and not to its bactericidal action. No significant difference could be demonstrated between the protein content or disaccharidase-specific activities of the brush border fractions from treated or non-treated intestines. Electrophoretic protein patterns of SDS-solubilized membrane were not significantly different after neomycin treatment. To gain more information on the mechanism involved in the stimulation of d-glucose transport, experiments were conducted on phosphatidyl glycerol artificial membranes and the results compared with those obtained with brush border membrane. At a concentration of 10 −7 M, neomycin decreased the nonactin-induced K + conductance by a factor of approx. 100. The membrane conductance was linearly dependent on the neomycin concentration and the conductance in 10 −2 M KCl was 10 times that in 10 −3 M KCl. The valence of neomycin was estimated, from the slope of these curves, to be between 6 and 4. In contrast, acetylated neomycin had no effect on the nonactin-induced K + membrane conductance. Therefore, the effect of neomycin on artificial membrane is related to its 4 to 6 positive charges. It is proposed that the stimulation of sugar transport in brush border membrane is related to screening of the membrane negative charges by the positively-charged neomycin. Accumulation of anions at the membrane surface then occurs and their diffusion into the intravesicular space would increase the transmembrane potential which, in turn, stimulates the entry of d-glucose.

Sodium ion-coupled uptake of taurocholate by rat-liver plasma membrane vesicles.

Uptake of taurocholate into plasma membrane vesicles isolated from rat liver was investigated. In the presence of an extra- to intravesicular gradient of Na+ ions, a typical "overshoot" phenomenon in the accumulation pattern was observed. Osmotic manipulation of the incubation medium indicated that the transport of this bile acid occurs into an osmotically active intravesicular space. Uptake of taurocholate as measured after 1 min was specifically stimulated by Na+ ions: NaNO3 and NaCl were capable of supporting accumulation, whereas KNO3 was not. Na+-coupled uptake of taurocholate showed saturation kinetics and was inhibited by other bile acids or by preloading the vesicles with Na+. Our observations support the idea of a carrier-mediated bile-acid uptake system, as suggested previously for the intact rat liver and isolated rat hepatocytes. When the electrical potential difference across the vesicle membrane was changed by inducing different diffusion potentials (anion replacement), a more negative potential inside stimulated Na+-dependent taurocholate transport. The results demonstrate that rat-liver plasma membrane vesicles possess an electrogenic Na+-coupled transport system for taurocholate.

Transport of 5-oxoproline into rabbit renal brush border membrane vesicles

5-Oxoproline has been shown to be transported intact into rabbit renal brush border membrane vesicles by a Na + -gradient dependent mechanism. The transport occurs against a concentration gradient. The equilibrium transport of 5-oxoproline decreases with increasing medium osmolarity. The predominant portion of transport occurs into an osmotically responsive intravesicular space, the non-specific surface binding being negligible. Competition experiments show that transport of 5-oxoproline is inhibited by most of the neutral amino acids whereas acidic, basic and imino acids do not have any effect.

Na +-independent l-arginine transport in rabbit renal brush border membrane vesicles

Na +-independent l-arginine uptake was studied in rabbit renal brush border membrane vesicles. The finding that steady-state uptake of l-arginine decreased with increasing extravesicular osmolality and the demonstration of accelerative exchange diffusion after preincubation of vesicles with l-arginine, but not d-arginine, indicated that the uptake of l-arginine in brush border vesicles was reflective of carrier-mediated transport into an intravesicular space. Accelerative exchange diffusion of l-arginine was demonstrated in vesicles preincubated with l-lysine and l-ornithine, but not l-alanine or l-proline, suggesting the presence of a dibasic amino acid transporter in the renal brush border membrane. Partial saturation of initial rates of l-arginine transport was found with extravesicular [arginine] varied from 0.005 to 1.0 mM. l-Arginine uptake was inhibited by extravesicular dibasic amino acids unlike the Na +-independent uptake of l-alanine, l-glutamate, glycine or l-proline in the presence of extravesicular amino acids of similar structure. l-Arginine uptake was increased by the imposition of an H + gradient (intravesicular pH<extravesicular pH) and H + gradient stimulated uptake was further increased by FCCP. These findings demonstrate membrane-potential-sensitive, Na +-independent transport of l-arginine in brush border membrane vesicles which differs from Na +-independent uptake of neutral and acidic amino acids. Na +-independent dibasic amino acid transport in membrane vesicles is likely reflective of Na +-independent transport of dibasic amino acids across the renal brush border membrane.

Characterisation of medium-inaccessible adenine nucleotides of rabbit skeletal muscle sarcoplasmic reticulum

ATP and ADP associate irreversibly with isolated sarcoplasmic reticulum vesicles in the presence of cytoplasmic concentrations of ATP, Mg 2+, and Ca 2+ (Aderem, A. A., McIntosh, D. B., and Berman, M. C., 1979, Proc. Nat. Acad. Sci. USA 76, 3622–3626). The nature of these medium-inaccessible adenine nucleotides has been studied from the kinetics of appearance and disappearance of these species in relation to nucleotides in the medium. Incubation of SR vesicles in ATP for 2 min at 30 °C results in an ATP concentration-dependent increase in nucleotide binding which was resistant to treatment with anion-exchange resin and activated charcoal and inaccessible to enzymes in the medium. The process is saturable at 5 nmol ATP + ADP/mg sarcoplasmic reticulum protein and half-maximal binding occurred with 5 m m ATP. The time dependence of increase of binding showed an initial rapid phase ( t 1 2 ∼- 0.10 min ), followed by a slower phase ( t 1 2 ∼-9 min ). The rapid phase of binding was Ca 2+-dependent with [Ca 2+] 0.5 = 3.5 × 10 −7 m. The slow phase was Ca 2+ independent. An Arrhenius plot of the temperature dependence of the rapid phase of binding showed a break at 24 °C with activation energies of 12.0 and 4.9 kcal mol −1 above and below this point, respectively. The data are consistent with the postulate that the rapid phase of binding of ATP, resulting in a medium-inaccessible species, represents tight binding of ATP to the (Ca 2+, Mg 2+)-ATPase, while the slow nonsaturable process represents leakage into the intravesicular space.

Iron uptake by rabbit intestinal mucosal membrane vesicles

Iron uptake by brush border membrane vesicles isolated from rabbit small intestine was studied using a rapid filtration technique. Special attention was directed at avoiding hydrolysis of iron at Fe(III) and oxidation of iron at Fe(II). Uptake of Fe(III) was studied from media containing ferric citrate with a 1000-fold molar excess of citrate to iron, calculated to be sufficient to prevent simple hydrolysis and polynuclear complex formation. Fe(III) bound to the mucosal membranes in a temperature-dependent and saturable fashion. Transport into the intravesicular space, however, could not be demonstrated. Fe(III) uptake was almost identical after heat denaturation of membrane proteins, which probably rules out the involvement of a specific protein receptor. Membrane binding of Fe(III) was higher when media with a lower pH were used and when there was only a 20-fold molar excess of citrate to iron. The major part of the 55Fe(III) is bound to the membranes very strongly, and cannot be removed by washing with excess unlabeled Fe(III). Under similar experimental conditions uptake of Fe(II), from a medium with a 20-fold molar excess of ascorbate to iron, was several hundred times higher than that of Fe(III). Again, there was no difference between the uptake of Fe(II) by normal or boiled vesicles. In contrast to results with Fe(III), however, uptake of Fe(II) into an osmotic space could be demonstrated. The large amount of Fe(II) transported may be explained by assuming an intravesicular iron trap, possibly provided by oxidation and hydrolysis in the vesicular lumen. Our findings indicate that the uptake of iron by microvillous membrane vesicles probably entails both binding and simple diffusion.

Effect of papain treatment on dipeptide transport into rabbit intestinal brush border vesicles

Purified rabbit intestinal brush border membrane vesicles transport glycyl-L-proline into an osmotically responsive intravesicular space by a Na +- independent, carrier-mediated process. With short incubation, transport occurs mostly as the intact dipeptide, followed by hydrolysis. Pretreatment of the vesicles with papain results in a 60% reduction of L-alanine transport while glycyl-L-proline transport is stimulated by 40%. Papain treatment does not change the intravesicular volume, nor does it increase membrane permeability. Dipeptide transport into papain treated vesicles remains completely Na +- independent as it is in the control vesicles. Many dipeptides inhibit glycyl-L-proline transport into papain treated vesicles both in the presence and absence of a Na + gradient.

Characteristics of dipeptide transport in normal and papain-treated brush border membrane vesicles from mouse intestine I. Uptake of glycyl- l-phenylalanine

Papain treatment of isolated brush border membrane vesicles was carried out to study peptide transport in the absence of hydrolytic events associated with the brush border membrane. Such a treatment allowed a 70% decrease in the activity of membrane-associated oligopeptidases and the study of peptide transport in the complete absence of free amino acids up to 1 min of incubation. A comparison between the time course curves of glycyl- l-phenylalanine uptake by normal and papain-treated vesicles showed that the overshoots seen in the presence of Na + and K + gradients (extravesicular intravesicular) when using normal vesicles were no longer evident after papain treatment. This result, together with the demonstration of uptake into an osmotically reactive intravesicular space and the analysis of uptake of free phenylalanine, allowed the coclusion that peptide transport was the result of two complementary mechanisms, uptake of free amino acids following hydrolysis by the membrane-bound oligopeptidases, and intact peptide transport down a concentration gradient by a non-Na + (and non-K +)-dependent process. These results also showed the non-involvement of γ-glutamyltransferase and the γ-glutamyl cycle in peptide absorption. A linear relationship has been established between initial dipeptide uptake and glycyl- l-phenylalanine concentration for the intact peptide transport process. However, this process can be inhibited to various extents by other di- and tripeptides but the inhibition never exceeded 43%. These results are consistent with both passive and facilitated diffusion mechanisms of intact peptide transport, the latter occuring by either a low affinity-high capacity or a high affinity-low capacity system.

Interaction of NH4+ and Li+ with the renal microvillus membrane Na+-H+ exchanger.

We have examined the interaction of NH4+ and Li+ with the Na+-H+ exchanger in rabbit renal microvillus membrane vesicles. Alkalinization of the intravesicular space, monitored with [14C]DMO uptake by flow dialysis, resulted from the addition of Cl- salts of Na+, Li+, or NH4+ to the external medium. The 22Na+ uptake rate, assayed by a rapid filtration technique, was saturable with respect to external Na+ concentration (KT 6 mM Na+) and was competitively inhibited by external Li+ (Ki 1.9 mM) and NH4+ (Ki 4.3 mM). Efflux by 22Na+ from vesicles preloaded with Na+ was stimulated by external Na+ and NH4+ but inhibited by external Li+. We conclude that the renal microvillus membrane Na+-H+ exchanger has affinity for Li+ and NH4+ and can mediate the exchange not only of Na+ for H+ but also of Na+ for Na+, Na+ for Nh4+, Li+ for H+, possibly NH4+ for H+, and perhaps Li+ for Na+. The physiological significance of exchange modes other than Na+-H+ exchange is not certain at present, but Na+-NH4+ exchange could play a role in the proximal tubular acidification process.

The protonmotive potential difference across the vacuo-lysosomal membrane of Hevea brasiliensis (rubber tree) and its modification by a membrane-bound adenosine triphosphatase

The vacuo-lysosomes of Hevea brasiliensis (rubber tree) constitute a suitable model system for the study of active transport and energization at the level of the membrane of plant vacuoles. The pH gradient (delta pH) and the membrane potential (delta psi) of vacuo-lysosomes were determined by means of the weak base methylamine and the lipophilic cation tetraphenylphosphonium. The values obtained depended strongly on the experimental conditions such as medium pH or K+ concentration. Under experimental conditions, i.e., pH 7.5 outside and low K+, the delta pH amounts to about 0.9 unit, interior acid, and the delta psi to -120 mV, interior negative. The delta psi is presumably caused by the imposed K+ gradient, and the internal acidification might be a consequence of the passive proton inflow along the electric field. This explanation is sustained by the ineffectiveness of carbonyl cyanide p-trifluoromethoxyphenylhydrazone in destroying the delta pH and delta psi, whereas higher K+ concentration decreased both. Under conditions existing in vivo, the membrane potential might be significantly lower. The presence of ATP increased the acidification of the intravesicular space by 0.5pH unit to a delta pH of up to 1.4 and shifts the membrane potential at least 60mV to a more positive value. The change of the protonmotive potential did not occur with ADP; the pH-dependence of the change was identical with the pH-dependence of a vacuo-lysosomal membrane-bound ATPase, and the effect of ATPase was prevented by the presence of the uncoupler carbonyl cyanide p-trifluoromethoxyphenylhydrazone. The change of protonmotive potential difference, brought about by the ATPase, was at least 90 mV. This is evidence that a vacuo-lysosomal ATPase in plants can function as an electrogenic proton pump that transfers protons into the vacuo-lysosomal space.

The kinetics of ionophore X-537A-mediated transport of manganese through dipalmitoylphosphatidylcholine vesicles A 1H-NMR study

We have studied the kinetics of ionophore X-537A-mediated transport of manganese ions into small unilamellar vesicles formed from dipalmitoylphosphatidylcholine. To follow the transport we used the paramagnetic effect of manganese on the 1H-NMR signal from choline trimethylammonium groups on the inner phospholipid monolayer. The transport of only one manganese ion produces an intravesicular concentration which is high enough (approx. 1 mM) to substantially broaden this signal. The observed signal thus arises predominantly from those vesicles which contain no manganese. Therefore, as manganese is transported into the vesicles the observed signal decreases in intensity, but does not broaden. The initial time-dependence of the intensity of the signal, S(t), can be approximated by the simple first-order rate law: S(t) = S (O) exp(−K′t) , where K′ is the probability per unit time for the transport of a manganese ion from the external medium to the intravesicular space. From the dependence of K′ on the ionophore X-537A concentration we conclude that manganese is transported into the vesicles via both 1 : 1 and 2 : 1 complexes with ionophore X-537A. At low ratios of ionophore X-537A to vesicles transport via the 1 : 1 complex predominates; at high ratios transport via the 2 : 1 complex predominates. From the dependence of K′ on manganese concentration we determined that under our conditions the equilibration of ionophore X-537A between vesicles is much faster than the transport of manganese through the vesicles. Lastly, from the dependence of K′ on temperature, we conclude that the ionophore X-537A-mediated transport of manganese into the dipalmitoylphosphatidylcholine vesicles is very sensitive to the gel-liquid crystalline phase transition.

Acetylcholine incorporation by cholinergic synaptic vesicles from Torpedo marmorata.

The ability of cholinergic vesicles to incorporate acetylcholine (ACh) was studied using highly purified synaptic vesicles from Torpedo electric organ. Depleted vesicles were capable of rapidly taking up exogenous ACh. Evidence that this represented true incorporation was that labelled ACh comigrated with vesicular ATP on gel filtration and that vesicle-associated ACh was protected against enzymatic hydrolysis and was releasable under hypoosmotic conditions. The total amount of ACh incorporated depended on the ACh concentration up to 100 mM. A sudden fall in the external ACh concentration did not cause leakage of the ACh incorporated in vitro. Preliminary results indicated that retention of ACh inside the vesicle was pH-dependent. Choline was also taken up by vesicles, but the time pattern strongly suggested that it was not being retained. The magnitude of ach incorporation was estimated with respect to the intravesicular space.

Calcium uptake in isolated brush-border vesicles from rat small intestine

Ca2+ uptake in brush-border vesicles isolated from rat duodena was studied by a rapid-filtration technique. Ca2+ uptake showed saturation kinetics, was dependent on the pH and ionic strength of the medium and was independent of metabolic energy. Uptake activity was readily inhibited by Ruthenium Red, La3+, tetracaine, EGTA, choline chloride and Na+ or K+. The effect of variations in medium osmolarity on Ca2+ uptake and the ionophore A23187-induced efflux of the cation from preloaded vesicles indicated that the Ca2+-uptake process involved binding to membrane components, as well as transport into an osmotically active space. Scatchard-plot analyses of the binding data suggested at least two classes of Ca2+-binding sites. The high-affinity sites, Ka = (2.7 +/- 1.1) x 10(4) M-1 (mean +/- S.D.) bound 3.2 +/- 0.8 nmol of Ca2+/mg of protein, whereas the low-affinity sites (Ka = 60 +/- 6 M-1) bound 110 +/- 17 nmol of Ca2+/mg of protein. In the presence of 100 mM-NaCl, 1.7 and 53 nmol of Ca2+/mg of protein were bound to the high- and low-affinity sites respectively. Decreased Ca2+-uptake activity was observed in vesicles isolated from vitamin D-deficient as compared with vitamin D-replete animals and intraperitoneal administration of 1,25-dihydroxycholecalciferol to vitamin D-deficient rats 16 h before membrane isolation stimulated the initial rate of Ca2+ uptake significantly. The data indicated that Ca2+ entry and/or binding was passive and may involve a carrier-mediated Ca2+-uptake component that is associated with the brush-border membrane. Altering the electrochemical potential difference across the membrane by using anions of various permeability and selected ionophores appeared to increase primarily binding to the membrane rather than transport into the intravesicular space. Since there is considerable binding of Ca2+ to the vesicle interior, a comprehensive analysis of the transport properties of the brush-border membrane remains difficult at present.

Cystine uptake by rat renal brush-border vesicles.

Uptake of L-cystine by brush-border membrane vesicles isolated from rat renal-cortical tissue was time-dependent and occurred in the absence of cystine reduction. A significant capacity for vesicular binding of cystine was observed. The amount bound increased with time of incubation and could be displaced by thiol reagents. At early time points, cystine uptake measured the transport of cystine into the intravesicular space. Total cystine uptake was mediated by multiple transport systems, including a low-Km high-affinity component which was shared by lysine, arginine, ornithine and glutamine and on which hetero-exchange diffusion of lysine and cystine was demonstrated.

Sulfate-sodium cotransport by brush-border membrane vesicles isolated from rat ileum

Uptake of inorganic sulfate into brush-border membrane vesicles isolated by a calcium precipitation method from rat small intestine was investigated using a rapid filtration technique and 35Sulfur acid as tracer. Sulfate uptake by membrane vesicles was osmotically sensitive, suggesting transport into an intravesicular space rather than binding to or incorporation into the membrane. Transport of sulfate into brush-border vesicles isolated from rat ileum was only stimulated by sodium ions as compared with other monovalent cations. A typical “overshoot” phenomenon was observed in the presence of an inwardly directed NaCl gradient. Tracer sulfate exchange was faster in the presence of sodium than in the presence of potassium. Addition of the ionophores for monovalent cations, monactin, or gramicidin D, decreased the sodium gradient-driven sulfate uptake. Sulfate uptake showed a saturation phenomenon only in the presence of sodium. Transstimulation of sodium-dependent sulfate transport was shown with MoO42−, but not with PO42− and WO42−. Changing the electrical potential difference across the membrane vesicles by establishing different diffusion potentials (anion replacement; potassium gradient ± valinomycin) did not alter sodium-dependent sulfate uptake. Stimulation of sulfate transport by sodium was greater in membrane vesicles from ileal segments than from duodenum or jejunum. It is concluded that isolated brush-border membranes of rat ileum contain an electroneutral sodium-sulfate cotransport system.

Chloride uptake by brush border membrane vesicles isolated from rabbit renal cortex. Coupling to proton gradients and K+ diffusion potentials.

Brush border membrane vesicles were isolated from rabbit renal cortex by Mg(++)-precipitation and differential centrifugation. (36)Cl(-) and [(3)H]glucose uptakes were simultaneously determined by a rapid filtration technique. Lysis of the vesicles with distilled water abolished 90-95% of the radioactivity on the filters, suggesting that nearly all of the (36)Cl(-) and [(3)H]glucose counts represented uptake into an osmotically reactive intravesicular space. Inwardly directed K(+) gradients plus valinomycin stimulated (36)Cl(-) uptake, demonstrating a conductive pathway for chloride uptake into brush-border membrane vesicles. (36)Cl(-) uptake could also be stimulated by inwardly directed proton gradients (pH(outside) < pH(inside)). This effect was seen in the absence of sodium, as well as in the presence of valinomycin when the vesicles had equal K(+) concentrations inside and out. An "overshoot" phenomenon was observed when external (36)Cl(-) was 2 mM and the external pH was lowered from 7.5 to 6.0 or to 4.5. The effect of the proton gradient was presumed to be different from the conductive mechanism because (a) the stimulation of (36)Cl(-) uptake by inwardly directed K(+) diffusion potentials was additive to the proton gradient effect, and (b) competition studies revealed statistically significant effects of thiocyanate on the conductive pathway, but not on the proton-driven pathway.HCl cotransport or anion exchange are electrically neutral mechanisms which could couple (36)Cl(-) uptake to inwardly-directed proton gradients in a brush border membrane vesicle. If both electrically neutral and conductive path ways for chloride transport are present in the luminal membrane of the proximal tubule, then the mechanism as well as the direction of net chloride transport will be influenced by the nature of the accompanying cation transport process.

Transport of glycyl-L-proline into intestinal and renal brush border vesicles from rabbit.

Transport of labeled glycyl-L-proline has been shown to occur with highly purified brush border membrane vesicles from the epithelial cells of rabbit small intestine and renal cortex. With 1-min incubation, transport occurs mainly as the intact dipeptide since less than 10% of the dipeptide in the medium is hydrolyzed within the period. The properties of the dipeptide transport system are similar in both small intestinal and renal brush border membrane vesicles. The steady state transport varies inversely with medium osmolarity. Extrapolation to infinite medium osmolarity indicates that transport occurs predominantly into an osmotically reactive intravesicular space rather than binding to the membranes. The affinity constants (Kt) for glycyl-L-proline transport in small intestinal and renal brush border membrane vesicles are comparable (0.9 mM in intestine and 1.1 mM in kidney). Under conditions in which presence of a Na+ gradient between external and intravesicular media stimulated L-alanine transport, glycyl-L-proline transport remains unaffected. Other dipeptides strongly inhibit the transport of glycyl-L-proline but amino acids have no effect. The selective inhibition of glycyl-L-proline transport by other dipeptides is observed in the presence as well as in the absence of a Na+ gradient. Harmaline inhibits Na+-stimulated L-alanine transport but it has no effect on glycyl-L-proline transport even in the presence of Na+. In these respects, dipeptide transport seems to differ from amino acid transport. It is proposed that the Na+ gradient hypothesis of sugar and amino acid transport is not applicable for dipeptide transport. These data provide additional evidence for the distinct nature of amino acid and dipeptide transport systems.

Carrier-mediated transport of glycyl-L-proline in renal brush border vesicles

Renal brush border vesicles prepared from rabbit were shown to transport intact glycyl-L-proline into the intravesicular space by a Na +-independent, carrier-mediated process. The kinetics of hydrolysis of glycyl-L-proline by the vesicles showed that this dipeptide was extremely resistant to hydrolysis. The intravesicular concentration of glycyl-L-proline was the same in both NaCl and KCl media. Dipeptide uptake into the vesicles appeared to be via carrier-mediated transport down a concentration gradient.

Calcium movements promoted by vesicles in a highly enriched sarcolemma preparation from canine ventricle. Calcium-calcium countertransport.

Calcium uptake by vesicles in a highly enriched sarcolemma preparation from canine ventricle was found to be markedly stimulated by intravesicular calcium. Stimulation of calcium uptake appeared to be a saturable function of intravesicular calcium. Calcium efflux from the vesicles was stimulated by calcium in the reaction medium. Calcium uptake, supported by intravesicular calcium, and calcium efflux, stimulated by extravesicular calcium, were found to correspond on a one-to-one basis. Only small changes in net uptake or efflux were observed to occur in response to chemical gradients of calcium across the membrane. It was concluded, therefore, that under certain conditions, the major means for calcium movement across vesicles in the preparation is via a one-to-one exchange of calcium. Sodium was found to stimulate calcium uptake when present in the intravesicular space and to stimulate calcium efflux when present in the extravesicular space, but the effects of calcium plus sodium were not additive with respect to stimulating either calcium uptake or efflux. The effects of unlabeled calcium, strontium, barium, and magnesium on calcium uptake stimulated by intravesicular calcium and by intravesicular sodium were similar though not identical. The temperature dependence for calcium-stimulated and sodium-stimulated calcium movements was characterized by Q10 values of 1.27 and 2.06, respectively. Previous work has associated the sodium-calcium exchange reaction with the sarcolemma. It is argued that the present study, in turn, provides evidence that the calcium-calcium exchange reaction is also associated with the sarcolemma. In addition, the results of the study are consistent with the hypothesis that one membrane system can promote the exchange of either calcium for calcium or calcium for sodium.

Polar distribution of sodium-dependent and sodium-independent transport system for L-lactate in the plasma membrane of rat enterocytes.

The uptake of L-lactate by rat small intestinal brush-border and basal-lateral plasma membrane vesicles has been studied. L-Lactate uptake by the isolated membrane vesicles is osmotically sensitive and represents predominantly transport into an intravesicular space and not binding to the membranes. The transport of L-lactate across the brush-border membrane is stimulated by sodium, whereas the transport across the basal-lateral plasma membrane is sodium-independent. In both types of membrane vesicles L-lactate is transported faster than D-lactate and L-lactate transport is inhibited by alpha-cyano-cinnamic acid. L-Lactate transport across basal-lateral membranes is inhibited by D-lactate and pyruvate and transstimulated by L-lactate and pyruvate. The polar distribution of transport system for L-lactate in the plasma membrane of rat enterocytes--a Na+/L-lactate cotransport system in the brush-border membrane and a facilitated diffusion system in the basal-lateral membrane--can explain the fact that in the intact epithelium L-lactate produced by cell metabolism is preferentially released on the serosal side and could enable the cell to perform vectorial, secondary active transport of L-lactate from the intestinal lumen to the serosal compartment.