共查询到20条相似文献,搜索用时 15 毫秒
1.
S Sennoune A Gerbi M J Duran J P Grillasca E Compe S Pierre R Planells M Bourdeaux P Vague G Pieroni J M Maixent 《European journal of biochemistry》2000,267(7):2071-2078
Na+/K+-ATPase during diabetes may be regulated by synthesis of its alpha and beta subunits and by changes in membrane fluidity and lipid composition. As these mechanisms were unknown in liver, we studied in rats the effect of streptozotocin-induced diabetes on liver Na+/K+-ATPase. We then evaluated whether fish oil treatment prevented the diabetes-induced changes. Diabetes mellitus induced an increased Na+/K+-ATPase activity and an enhanced expression of the beta1 subunit; there was no change in the amount of the alpha1 and beta3 isoenzymes. Biphasic ouabain inhibition curves were obtained for diabetic groups indicating the presence of low and high affinity sites. No alpha2 and alpha3 isoenzymes could be detected. Diabetes mellitus led to a decrease in membrane fluidity and a change in membrane lipid composition. The diabetes-induced changes are not prevented by fish oil treatment. The results suggest that the increase of Na+/K+-ATPase activity can be associated with the enhanced expression of the beta1 subunit in the diabetic state, but cannot be attributed to changes in membrane fluidity as typically this enzyme will increase in response to an enhancement of membrane fluidity. The presence of a high-affinity site for ouabain (IC50 = 10-7 M) could be explained by the presence of (alphabeta)2 diprotomeric structure of Na+/K+-ATPase or an as yet unknown alpha subunit isoform that may exist in diabetes mellitus. These stimulations might be related, in part, to the modification of fatty acid content during diabetes. 相似文献
2.
Localization of Na+,K+-ATPase alpha-subunit to the sinusoidal and lateral but not canalicular membranes of rat hepatocytes 总被引:4,自引:2,他引:2
下载免费PDF全文
![点击此处可从《The Journal of cell biology》网站下载免费的PDF全文](/ch/ext_images/free.gif)
E S Sztul D Biemesderfer M J Caplan M Kashgarian J L Boyer 《The Journal of cell biology》1987,104(5):1239-1248
Controversy has recently developed over the surface distribution of Na+,K+-ATPase in hepatic parenchymal cells. We have reexamined this issue using several independent techniques. A monoclonal antibody specific for the endodomain of alpha-subunit was used to examine Na+,K+-ATPase distribution at the light and electron microscope levels. When cryostat sections of rat liver were incubated with the monoclonal antibody, followed by either rhodamine or horseradish peroxidase-conjugated goat anti-mouse secondary, fluorescent staining or horseradish peroxidase reaction product was observed at the basolateral surfaces of hepatocytes from the space of Disse to the tight junctions bordering bile canaliculi. No labeling of the canalicular plasma membrane was detected. In contrast, when hepatocytes were dissociated by collagenase digestion, Na+,K+-ATPase alpha-subunit was localized to the entire plasma membrane. Na+,K+-ATPase was quantitated in isolated rat liver plasma membrane fractions by Western blots using a polyclonal antibody against Na+,K+-ATPase alpha-subunit. Plasma membranes from the basolateral domain of hepatocytes possessed essentially all of the cell's estimated Na+,K+-ATPase catalytic activity and contained a 96-kD alpha-subunit band. Canalicular plasma membrane fractions, defined by their enrichment in alkaline phosphatase, 5' nucleotidase, gamma-glutamyl transferase, and leucine aminopeptidase had no detectable Na+,K+-ATPase activity and no alpha-subunit band could be detected in Western blots of these fractions. We conclude that Na+,K+-ATPase is limited to the sinusoidal and lateral domains of hepatocyte plasma membrane in intact liver. This basolateral distribution is consistent with its topology in other ion-transporting epithelia. 相似文献
3.
The unique ultrastructure of secretory membranes in gastric parietal cells depends upon the presence of H+, K+ -ATPase 总被引:1,自引:0,他引:1
Miller ML Judd LM Van Driel IR Andringa A Flagella M Bell SM Schultheis PJ Spicer Z Shull GE 《Cell and tissue research》2002,309(3):369-380
Ion transporters play a central role in gastric acid secretion. To determine whether some of these transporters are necessary for the normal ultrastructure of secretory membranes in gastric parietal cells, mice lacking transporters for H+, K+, Cl-, and Na+ were examined for alterations in volume density (Vd) of basolateral, apical, tubulovesicular and canalicular membranes, microvillar dimensions, membrane flexibility, and ultrastructure. In mice lacking Na+/H+ exchanger 1 (NHE1) or the Na+-K+-2Cl- cotransporter (NKCC1), the ultrastructure and Vd of secretory membranes and the secretory canalicular to tubulovesicular membrane ratio (SC/TV), a morphological correlate of secretory activity, were similar to those of wild-type mice. In mice lacking Na+/H+ exchanger 2 (NHE2) or gastric H+, K+ -ATPase alpha- or beta-subunits, the SC/TV ratio and Vd of secretory membranes were decreased, though canaliculi were often dilated. In H+, K+ -ATPase-deficient parietal cells, canalicular folds were decreased, normally abundant tubulovesicles were replaced with a few rigid round vesicles, and microvilli were sparse, stiff and short, in contrast to the long and flexible microvilli in wild-type cells. In addition, microvilli of the H+, K+ -ATPase-deficient parietal cells had centrally bundled F-actin filaments, unlike the microvilli of wild-type cells, in which actin filaments were peripherally positioned concentric to the plasmalemma. Data showed that the absence of H+, K+ -ATPase produced fundamental changes in parietal cell membrane ultrastructure, suggesting that the pump provides an essential link between the membranes and F-actin, critical to the gross architecture and suppleness of the secretory membranes. 相似文献
4.
Localization of the ecto-ATPase (ecto-nucleotidase) in the rat hepatocyte plasma membrane. Implications for the functions of the ecto-ATPase 总被引:3,自引:0,他引:3
S H Lin 《The Journal of biological chemistry》1989,264(24):14403-14407
The surface distribution of the plasma membrane Ca2+ (Mg2+)-ATPase (ecto-ATPase) in rat hepatocytes was determined by several methods. 1) Two polyclonal antibodies specific for the ecto-ATPase were used to examine the distribution of the enzyme in frozen sections of rat liver by immunofluorescence. Fluorescent staining was observed at the bile canalicular region of hepatocytes. 2) Plasma membranes were isolated from the canalicular and sinusoidal regions of rat liver. The specific activity of ecto-ATPase in the canalicular membranes was 22 times higher than that of sinusoidal membranes. The enrichment of the ecto-ATPase activity in the canalicular membrane is closely parallel to that of two other canalicular membrane markers, gamma-glutamyltranspeptidase and leucine aminopeptidase. 3) By immunoblots with polyclonal antibodies against the ecto-ATPase and the Na+,K+-ATPase, it was found that the ecto-ATPase protein was only detected in canalicular membranes and not in sinusoidal membranes, while the Na+,K+-ATPase protein was only detected in sinusoidal membranes and not in canalicular membranes. These results indicate that the ecto-ATPase is enriched in the canalicular membranes of rat hepatocytes. 相似文献
5.
Fujii T Takahashi Y Itomi Y Fujita K Morii M Tabuchi Y Asano S Tsukada K Takeguchi N Sakai H 《The Journal of biological chemistry》2008,283(11):6869-6877
Gastric parietal cells migrate from the luminal to the basal region of the gland, and they gradually lose acid secretory activity. So far, distribution and function of K+-Cl(-) cotransporters (KCCs) in gastric parietal cells have not been reported. We found that KCC3a but not KCC3b mRNA was highly expressed, and KCC3a protein was predominantly expressed in the basolateral membrane of rat gastric parietal cells located in the luminal region of the glands. KCC3a and the Na+,K+-ATPase alpha1-subunit (alpha1NaK) were coimmunoprecipitated, and both of them were highly localized in a lipid raft fraction. The ouabain-sensitive K+-dependent ATP-hydrolyzing activity (Na+,K+-ATPase activity) was significantly inhibited by a KCC inhibitor (R-(+)-[(2-n-butyl-6,7-dichloro-2-cyclopentyl-2,3-dihydro-1-oxo-1H-inden-5-yl)oxy]acetic acid (DIOA)). The stable exogenous expression of KCC3a in LLC-PK1 cells resulted in association of KCC3a with endogenous alpha1NaK, and it recruited alpha1NaK in lipid rafts, accompanying increases of Na+,K+-ATPase activity and ouabain-sensitive Na+ transport activity that were suppressed by DIOA, whereas the total expression level of alpha1NaK in the cells was not significantly altered. On the other hand, the expression of KCC4 induced no association with alpha1NaK. In conclusion, KCC3a forms a functional complex with alpha1NaK in the basolateral membrane of luminal parietal cells, and it up-regulates alpha1NaK in lipid rafts, whereas KCC3a is absent in basal parietal cells. 相似文献
6.
Comparison of specific binding of human epidermal growth factor (EGF) to sinusoidal and bile canalicular membranes isolated from rat liver 总被引:2,自引:0,他引:2
K Yachi Y Sugiyama H Sato D C Kim T Fuwa T Iga M Hanano 《Journal of biochemistry》1988,103(3):448-451
The binding characteristics of human epidermal growth factor (EGF) were compared between highly purified canalicular (CMV) and sinusoidal (basolateral) rat liver plasma membrane (SMV) preparations. The dissociation constants (2-3 nM) for these membranes were comparable, while the binding capacity for CMV was approximately half that for SMV. The binding capacity for CMV was too high to be accounted for only by the contamination with sinusoidal membranes, since the measurements of specific activities of various enzymes (Na+,K+-ATPase, alkaline phosphatase, and leucine aminopeptidase) indicated that the extents of the cross contamination with other membrane fractions were at most 10%. Although the physiological function of specific binding of EGF to bile canalicular membrane domain remains to be determined, it may have a role in biliary excretion of EGF. The specific binding of EGF to bile canalicular membranes from rat liver was identified for the first time. 相似文献
7.
Gerbi A Zérouga M Maixent JM Debray M Durand G Bourre JM 《The Journal of nutritional biochemistry》1999,10(4):230-236
The effects of dietary (n-6)/(n-3) polyunsaturated fatty acid balance on fatty acid composition, ouabain inhibition, and Na(+) dependence of Na(+), K(+)-ATPase isoenzymes of whole brain membranes were studied in 60-day-old rats fed over two generations a diet either devoid of alpha-linolenic acid [18:3(n-3)] (sunflower oil diet) or rich in 18:3(n-3) (soybean oil diet). In the brain membranes, the sunflower oil diet led to a dramatic decrease in docosahexaenoic acid [22:6(n-3)] membrane content. The activities of Na(+), K(+)-ATPase isoenzymes were discriminated on the basis of their differential affinities for ouabain and their sensitivity to sodium concentration. The ouabain titration curve of Na(+), K(+)-ATPase activity displayed three inhibitory processes with markedly different affinity [i.e., low (alpha1), high (alpha2), and very high (alpha3)] for brain membranes of rats fed the sunflower oil diet, whereas the brain membranes of rats fed the soybean oil diet exhibited only two inhibitory processes, low (alpha1) and high (alpha2' = alpha2 + alpha3). Regardless of the diet, on the basis of the Na(+) dependence of Na(+), K(+)-ATPase activity, three isoenzymes were found: alpha1 form displaying an affinity 1.5- to 2-fold higher that of than alpha2 and 3-fold higher that of alpha3. In rats fed the sunflower oil diet, alpha2 isoenzyme exhibited higher affinity for sodium (Ka = 8.8 mmol/L) than that of rats fed the soybean oil diet (Ka = 11.7 mmol/L). These results suggest that the membrane lipid environment modulates the functional properties of Na(+), K(+)-ATPase isoenzymes of high ouabain affinity (alpha2). 相似文献
8.
M Y Abeywardena E J McMurchie G R Russell W H Sawyer J S Charnock 《Biochimica et biophysica acta》1984,776(1):48-59
The effects of different dietary fat intake on the lipid composition and enzyme behaviour of sarcolemmal (Na+ + K+)ATPase and sarcoplasmic reticulum Ca2+-ATPase from rat heart were investigated. Rat diets were supplemented with either sunflower seed oil (unsatd./satd. 5.6) or sheep kidney fat (unsatd./satd. 0.8). Significant changes in the phospholipid fatty acid composition were observed in both membranes after 9 weeks dietary lipid treatment. For both membranes, the total saturated/unsaturated fatty acid levels were unaffected by the dietary lipid treatment, however the proportions of the major unsaturated fatty acids were altered. Animals fed the sunflower seed oil diet exhibited an increase in n-6 fatty acids, including linoleic (18:2(n-6] and arachidonic (20:4(n-6] while the sheep kidney fat dietary rats were higher in n-3 fatty acids, principally docosahexaenoic (22:6), with the net result being a higher n-6/n-3 ratio in the sunflower seed oil group compared to sheep kidney fat dietary animals. Fluorescence polarization indicated that the fluidity of sarcoplasmic reticular membrane was greater than that of sarcolemmal membrane, with a dietary lipid-induced decrease in fluidity being observed in the sarcoplasmic reticular membrane from sheep kidney fat dietary animals. Despite these significant changes in membrane composition and physical properties, neither the specific activity nor the temperature-activity relationship (Arrhenius profile) of the associated ATPases were altered. These results suggest that with regard to the parameters measured in this study, the two ion-transporting ATPases are not modulated by changes which occur in the membrane lipid composition as a result of the diet. 相似文献
9.
Coste T Pierlovisi M Leonardi J Dufayet D Gerbi A Lafont H Vague P Raccah D 《The Journal of nutritional biochemistry》1999,10(7):411-420
Metabolic and vascular abnormalities are implicated in the pathogenesis of diabetic neuropathy. Two principal metabolic defects are altered lipid metabolism resulting from the impairment of delta-6-desaturase, which converts linoleic acid (LA) into gamma linolenic acid (GLA), and reduced nerve Na+, K+ ATPase activity. This reduction may be caused by a lack of incorporation of (n-6) fatty acids in membrane phospholipids. Because this ubiquitous enzyme maintains the membrane electrical potential and allows repolarization, disturbances in its activity can alter the process of nerve conduction velocity (NCV). We studied the effects of supplementation with GLA (260 mg per day) on NCV, fatty acid phospholipid composition, and Na+, K+ ATPase activity in streptozotocin-diabetic rats. Six groups of 10 rats were studied. Two groups served as controls supplemented with GLA or sunflower oil (GLA free). Two groups with different durations of diabetes were studied: 6 weeks with no supplementation and 12 weeks supplemented with sunflower oil. To test the ability of GLA to prevent or reverse the effects of diabetes, two groups of diabetic rats were supplemented with GLA, one group for 12 weeks and one group for 6 weeks, starting 6 weeks after diabetes induction. Diabetes resulted in a 25% decrease in NCV (P < 0.0001), a 45% decrease in Na+, K+ ATPase activity (P < 0.0001), and an abnormal phospholipid fatty acid composition. GLA restored NCV both in the prevention and reversal studies and partially restored Na+, K+ ATPase activity in the preventive treatment group (P < 0.0001). These effects were accompanied by a modification of phospholipid fatty acid composition in nerve membranes. Overall, the results suggest that membrane fatty acid composition plays a direct role in NCV and confirm the beneficial effect of GLA supplementation in diabetic neuropathy. 相似文献
10.
Isoforms of Na+, K+-ATPase in human prostate; specificity of expression and apical membrane polarization 总被引:2,自引:0,他引:2
Mobasheri A Oukrif D Dawodu SP Sinha M Greenwell P Stewart D Djamgoz MB Foster CS Martín-Vasallo P Mobasheri R 《Histology and histopathology》2001,16(1):141-154
The cellular distribution of Na+, K+-ATPase subunit isoforms was mapped in the secretory epithelium of the human prostate gland by immunostaining with antibodies to the alpha and beta subunit isoforms of the enzyme. Immunolabeling of the alpha1, beta1 and beta2 isoforms was observed in the apical and lateral plasma membrane domains of prostatic epithelial cells in contrast to human kidney where the alpha1 and beta1 isoforms of Na+, K+-ATPase were localized in the basolateral membrane of both proximal and distal convoluted tubules. Using immunohistochemistry and PCR we found no evidence of Na+, K+-ATPase alpha2 and alpha3 isoform expression suggesting that prostatic Na+, K+-ATPase consists of alpha1/beta1 and alpha1/beta2 isozymes. Our immunohistochemical findings are consistent with previously proposed models placing prostatic Na+, K+-ATPase in the apical plasma membrane domain. Abundant expression of Na+, K+-ATPase in epithelial cells lining tubulo-alveoli in the human prostate gland confirms previous conclusions drawn from biochemical, pharmacological and physiological data and provides further evidence for the critical role of this enzyme in prostatic cell physiology and ion homeostasis. Na+, K+-ATPase most likely maintains an inwardly directed Na+ gradient essential for nutrient uptake and active citrate secretion by prostatic epithelial cells. Na+, K+-ATPase may also regulate lumenal Na+ and K+, major counter-ions for citrate. 相似文献
11.
S-adenosyl-L-methionine modulates Na+ + K+-ATPase activity in rat colonic basolateral membranes. 总被引:1,自引:0,他引:1
下载免费PDF全文
![点击此处可从《The Biochemical journal》网站下载免费的PDF全文](/ch/ext_images/free.gif)
Rat colonic basolateral membranes were incubated with S-adenosyl-L-[methyl-3H]methionine (0.3 mM) at 37 degrees C for 2 h at pH 9.0. This resulted in an increase in the specific activity of Na+ + K+-ATPase by 60%. Kinetic parameter analysis revealed a 2-fold increase in the Vmax. of this enzymatic activity, whereas the Km for ATP was unchanged. The methylation inhibitor S-adenosyl-L-homocysteine (2 mM) significantly reduced these S-adenosyl-L-methionine-stimulated increases in specific activity and the Vmax. of Na+ + K+-ATPase. S-Adenosyl-L-methionine treatment of basolateral membranes was also found to significantly increase the fluidity of these preparations, as assessed by steady-state fluorescence polarization techniques using the fluorophore 1,6-diphenyl-1,3,5-hexatriene; S-adenosyl-L-homocysteine (2 mM) again markedly reduced this S-adenosyl-L-methionine-induced increase in fluidity. While transmethylation reactions involving phospholipids, non-polar lipids and proteins were all found to exist in rat colonic basolateral membranes, based on a number of observations, the results of the present studies suggest that transmethylation of membrane phospholipids, but not membrane non-polar lipids or proteins, influenced the fluidity of basolateral membranes which, in turn, modified Na+ + K+-ATPase activity in these membranes. 相似文献
12.
Haag M Kearns SD Magada ON Mphata PR Claassen N Kruger MC 《Prostaglandins & other lipid mediators》2001,66(1):53-63
Duodenal ion transport processes are supported by ATPase enzymes in basolateral membranes of the enterocyte. In vivo studies have shown that long term n-6 poly-unsaturated fatty acid (PUFA) supplementation in rats causes increases in intestinal Ca absorption, coupled with a higher total calcium balance and bone calcium content. The present in vitro study was undertaken to test the effect of arachidonic acid (AA), a highly unsaturated (and thus physiologically potent) member of the n-6 PUFA family, on ATPases in enterocyte basolateral membranes isolated with a sorbitol density gradient procedure. This paper presents results which show that AA inhibits Na+,K+-ATPase in a dose-dependent manner (-67% of basal activity at a concentration of 30 microg/ml, P < 0.005) but that this effect is not mediated by protein kinase C, as shown by the use of the protein kinase C blocker calphostin (0.5 microM). Indomethacin (IDM) at 0.1 mM, a cyclo-oxygenase blocker, could also not reverse the inhibitory effect of AA on Na+,K+-ATPase. Ca2+-ATPase, on the other hand, is not affected significantly (-10%, P > 0.05) by arachidonic acid at 30 microg/ml. 相似文献
13.
14.
We have applied free flow electrophoresis to separate the canalicular and basolateral (sinusoidal and lateral) domains of rat hepatocyte plasma membranes. Hepatocyte plasma membranes were prepurified by rat zonal and discontinous sucrose gradient centrifugation. In electrophoretic separation, the canalicular membranes were more deflected toward the anode than the basolateral membranes. Na+-dependent taurocholate uptake could be measured in both membrane fractions, transport activity being highest in fractions containing the highest specific activity in the basolateral marker enzyme Na+-K+-ATPase. Thus, differences in electrophoretic mobility permit the separation of functional intact plasma membrane vesicles derived from basolateral and canalicular plasma membrane domains of rat hepatocyte. 相似文献
15.
Oxidative stress reduces transintestinal transports and (Na+, K+) -ATPase activity in rat jejunum 总被引:2,自引:0,他引:2
Orsenigo MN Faelli A Porta C Sironi C Laforenza U Paulmichl M Tosco M 《Archives of biochemistry and biophysics》2007,466(2):300-307
Because oxidative stress is a component of gastrointestinal injury, we investigated the effect of H(2)O(2) on transintestinal transport using isolated rat jejunum incubated in vitro. Millimolar concentrations of H(2)O(2) inhibited all the tested parameters without inducing any cytotoxic effect. Electrophysiological experiments indicated that H(2)O(2) decreases significantly both short circuit current and transepithelial electrical potential difference without affecting transepithelial resistance. The possibility that H(2)O(2) could influence (Na+, K+) -ATPase activity was explored using isolated basolateral membranes. Besides H(2)O(2), free radicals (O(2)(*-), HO*) were generated using different iron-dependent and independent systems; (Na+, K+) -ATPase activity was inhibited after membrane exposure to all ROS tested. The inhibition was prevented by allopurinol, superoxide dismutase or desferrioxamine. Western blot analysis showed a decreased expression of the alpha(1)-subunit of (Na+, K+) -ATPase. We conclude that H(2)O(2) may be a modulator of jejunal ion and water transport by multiple mechanisms, among which a significant inhibition of the basolateral (Na+, K+) -ATPase. 相似文献
16.
The (Na+,K+)-ATPase ATP hydrolyzing activity from rabbit kidney medulla basolateral membrane vesicles was studied as a function of the cholesterol content of the basolateral membranes. The cholesterol content of the membranes was modified by incubation with phospholipid vesicles. When the cholesterol content was increased above that found in the native membrane, the (Na+,K+)-ATPase ATP hydrolyzing activity was inhibited. When the cholesterol content was decreased from that found in the native membranes, the (Na+,K+)-ATPase ATP hydrolyzing activity was inhibited. Analogous effects were found with the K+-activated phosphatase activity of the same membrane vesicles. Therefore, at low cholesterol contents, cholesterol was stimulatory, and at high cholesterol contents, cholesterol was inhibitory. The structural specificity of this effect was tested by introducing lanosterol and ergosterol as 50% of the membrane sterol. Ergosterol was the least effective at supporting (Na+,K+)-ATPase ATP hydrolyzing activity, while lanosterol was more effective, but still not as effective as cholesterol. 相似文献
17.
In interleukin-2 (IL-2)-induced human blood lymphocytes, the Na+/K+ pump function (assessed by ouabain-sensitive Rb+ influx), the abundance of Na+, K+-ATPase alpha1-subunit (determined by Western blotting) and the alpha1- and beta1-subunits mRNA of Na+, K+-ATPase (RT-PCR), as well as the phosphorylation of STAT5 and STAT3 family proteins and ERK1/2 kinase have been examined. A 3.5-4.0-fold increase in the expression of alpha1- and beta1-subunits mRNA of Na+, K+-ATPase was found at 24 h of IL-2 stimulation. The inhibitors of JAK3 kinase (B-42, WHI-P431) was shown to decrease both the phosphorylation of STATs and the rise in the oubain-sensitive rubidium influx as well as the increased abundance of Na+, K+-ATPase alpha1-subunit. The inhibition of the protein kinases ERK1/2 by PD98059 (20 microM) suppressed the alpha1-subunit accumulation. All the kinase inhibitors tested did not alter the intracellular content ofmonovalent cations in resting and IL-2-stimulated lymphocytes. It is concluded that MAPK and JAK/STAT signaling pathways mediate the IL-2-dependent regulation of the Na+, K+-ATPase expression during the lymphocyte transition from resting stage to proliferation. 相似文献
18.
The driving forces for L-glutamate transport were determined in purified canalicular (cLPM) and basolateral (i.e. sinusoidal and lateral; blLPM) rat liver plasma membrane vesicles. Initial rates of L-glutamate uptake in cLPM vesicles were stimulated by a Na+ gradient (Na+o greater than Na+i), but not by a K+ gradient. Stimulation of L-glutamate uptake was specific for Na+, temperature sensitive, and independent of nonspecific binding. Sodium-dependent L-glutamate uptake into cLPM vesicles exhibited saturation kinetics with an apparent Km of 24 microM, and a Vmax of 21 pmol/mg X min at an extravesicular sodium concentration of 100 mM. Specific anionic amino acids inhibited L-[3H]glutamate uptake and accelerated the exchange diffusion of L-[3H]glutamate. An outwardly directed K+ gradient (K+i greater than K+o) further increased the Na+ gradient (Na+o greater than Na+i)-dependent uptake of L-glutamate in cLPM vesicles, resulting in a transient accumulation of L-glutamate above equilibrium values (overshoot). The K+ effect had an absolute requirement for Na+. In contrast, in blLPM the initial rates of L-glutamate uptake were only minimally stimulated by a Na+ gradient, an effect that could be accounted for by contamination of the blLPM vesicles with cLPM vesicles. These results indicate that hepatic Na+ gradient-dependent transport of L-glutamate occurs at the canalicular domain of the plasma membrane, whereas transport of L-glutamate across sinusoidal membranes results mainly from passive diffusion. These findings provide an explanation for the apparent discrepancy between the ability of various in vitro liver preparations to transport glutamate and suggest that a canalicular glutamate transport system may serve to reabsorb this amino acid from bile. 相似文献
19.
In isolated basolateral and canalicular rat liver plasma membrane vesicles the membrane potential (measured with DiS-C2 (5] varied with transmembrane concentration gradients of Na+, K+ and Cl- revealing the following ion permeabilities: basolateral vesicles: PNa/PK: 0.76, PCl/PK: 0.45 and canalicular vesicles: PNa/PK: 0.69, PCl/PK: 0.56. The data indicate a permselectivity of PK greater than PNa greater than PCl for both membranes. 相似文献
20.
Two ATPase activities, a Na+-ATPase and a (Na+ + K+)-ATPase, have been found associated with sheets of basolateral plasma membranes from guinea-pig small intestinal epithelial cells. The specific activity of the former is 10-15% of the latter. The two ATPase activities differ in their affinity for Na+, their optimal pH, their K+ requirement and particularly in their behaviour in the presence of some inhibitors and of Ca2+. Thus the Na+-ATPase is refractory to ouabain but it is strongly inhibited by ethacrynic acid and furosemide, whilst the (Na+ + K+)-ATPase is totally suppressed by ouabain, partially by ethacrynic acid and refractory to furosemide. In addition, the Na+-ATPase is activated by micromolar concentrations of calcium and by resuspension of the membrane preparation at pH 7.8. The Na+-ATPase is only stimulated by sodium and to a lesser extent by lithium; however, this stimulation is independent of the anion accompanying Na+. The latter rules out the participation of an anionic ATPase. The relation between the characteristics of the sodium transport mechanism in basolateral membrane vesicles (Del Castillo, J.R. and Robinson, J.W.L. (1983) Experientia 39,631) and those of the two ATPase activities present in the same membranes, allow us to postulate the existence of two separate sodium pumps in this membranes. Each pump would derive the necessary energy for active ion transport from the hydrolysis of ATP, catalyzed by different ATPase systems. 相似文献