Inhibition of Amino Acid Transport in Rabbit Intestine by p-Chloromercuriphenyl Sulfonic Acid

Schaeffer, John F.; Preston, Robert L.; Curran, Peter F.

doi:10.1085/jgp.62.2.131

Cited by 45 publications

(15 citation statements)

References 24 publications

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“…Kinetic analysis of the PCMBS-treated cells indicates that the modified protein exhibits decreased affinity for the substrate amino acid as well as a slower rate of translocation. Similar changes in Km and Vma, (Schaeffer, et al, 1973;Batt et al, 1976) values for other transporters have been observed following protein modification. These results, in conjunction with the specificity of the protection by amino acids, suggest that the site of chemical modification is localized within or near the amino acid binding site.…”

Section: Discussionsupporting

confidence: 57%

System a transport activity in normal rat hepatocytes and transformed liver cells: Substrate protection from inactivation by sulfhydryl‐modifying reagents

Chiles

Kilberg

1986

Journal Cellular Physiology

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The transport of amino acids by normal rat hepatocytes and several hepatoma cell lines has been examined for inactivation by various protein-modifying reagents, including the sulfhydryl-preferring reagents N-ethylmaleimide (NEM) and p-chloromercuribenzene sulfonate (PCMBS). Uptake of 2-aminoisobutyric acid (AIB), a specific probe for hepatic System A-mediated transport, was equally sensitive to inhibition by the organic mercurial PCMBS in each of the cell types tested. In contrast, the sensitivity of System A to inactivation by NEM was substantially different among the five cell types. Normal hepatocytes showed the greatest sensitivity, while the hepatoma cells varied in their responsiveness from moderate to no inhibition. PCMBS inactivated greater than 85% of the System A activity in rat H4 hepatoma cells within 10 min (t1/2 = 3 min). The inhibition by PCMBS was rapidly reversed by treatment of the cells with dithiothreitol. Amino acids showing a high affinity for System A protected the transport system from inactivation, whereas non-substrates produced little or no protection. Amino acid-dependent protection was stereospecific and system-specific. L-norleucine competitively inhibited AIB uptake (Ki = 1.9 +/- 0.1 mM) in H4 cells and also protected System A from PCMBS-dependent inactivation (half-maximal protection occurred at an amino acid concentration of 0.6 +/- 0.1 mM). N-bromosuccinimide was completely ineffective as an inhibitor of System A activity in hepatocytes, whereas treatment of H4 rat hepatoma cells with this reagent resulted in greater than 95% inhibition.

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Section: Discussionsupporting

confidence: 57%

System a transport activity in normal rat hepatocytes and transformed liver cells: Substrate protection from inactivation by sulfhydryl‐modifying reagents

Chiles

Kilberg

1986

Journal Cellular Physiology

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“…Certain of the changes seen in the ability of glucose to support sodium transport across intestines taken from suckled animals resembled those induced artificially in the rabbit intestine through the use of sulphydryl blocking agents (Schaefer, Preston & Curran, 1973). A final part of the present work describes attempts to mimic, with the use of a sulphydryl blocking agent, some of the changes in microvillar membrane function seen to occur normally as a result of suckling.…”

Section: Introductionmentioning

confidence: 71%

“…Signs and symbols as for a. dependent increase in short-circuit current. One conclusion formed from recent work using PCMPS on rabbit ileum was that this sulphydryl blocking agent acted to inhibit the normal amino acid-sodium interaction at the microvillar membrane by reducing the affinity of sodium for the ternary carrier (Schaefer et al 1973). This had the effect of causing an apparent decrease in the affinity of amino acid for its carrier, a situation rather similar to that seen for glucose in the pig ileum.…”

Section: Resultsmentioning

confidence: 97%

“…Both inhibit the ability of glucose to cause a rapid increase in short-circuit current and both result in a situation where the continued presence of glucose causes a secondary slow increase in current, an increase which continues for at least 10 min. The known action of PCMPS is to block sulphydryl groups and this has been equated, in the rabbit ileum, with an ability to inhibit the binding of sodium to an amino acid carrier in the microvillar membrane (Schaefer et al 1973). If the binding of an amino acid or sugar to its carrier unmasks a sodium binding site by changing the conformation around the carrier, in the manner suggested by Schultz (1969), then PCMPS could well be responsible for inhibiting this change.…”

Section: H De Jesus and M W Smithmentioning

confidence: 99%

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Protein and glucose‐induced changes in sodium transport across the pig small intestine

Jesus¹,

Smith²

1974

The Journal of Physiology

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SUMMARY1. The effect of glucose and of bovine y-globulin on short-circuit current and sodium transport was measured using new-born pig ileum before and after suckling.2. Glucose increased the short-circuit current with an apparent Km of 1-9 and 19-5 mm for new-born unsuckled and suckled intestines respectively. The Vmax was about 80 tA cm-2 in each case.3. Bovine y-globulin applied to new-born pig ileum caused a 20 % increase in sodium chloride influx without change of short-circuit current. The tissue conductance was also greater in the presence of globulin. These effects were not seen in ileum taken from suckled pigs.4. p-Chloromercuriphenyl sulphonic acid (PCMPS) inhibited the globulin induced increase in sodium influx in new-born pig ileum without changing the measured short-circuit current. 01 mM-PCMPS also changed the characteristics of the glucose-dependent increase in short-circuit current, so that this response came to resemble that seen in ileum taken from suckled pigs. Dithioerythritol reversed this effect of PCMPS.5. The suckling-dependent difference in results is explained in terms of a change in microvillar function taking place as a consequence of long-term pinocytosis in vivo. The ability of PCMPS to simulate some of these changes suggests that sulphydryl groups might be important regulators of microvillar membrane stability.

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“…Although Hg is known to induce toxic effects in different body organs and cellular components including jejunal epithelial cells [38, 39], limited or non existing data is available regarding the jejunal Na + ,K + -ATPase. In jejunum, Hg was associated with inhibition of amino acid and sugar transport [40,41,42] and the sites of Hg action were suggested to be located in the mucosal surface of jejunal slices but not in the basolateral cell border [43]. The results of the present study in HgCl 2 -treated rats showing that the jejunal Na + ,K + -ATPase activity changed from normal to low and to high values in parallel with changes in sodium balance further suggest that in jejunal epithelial cells the basolateral Na + ,K + -ATPase may escape Hg toxicity.…”

Section: Discussionmentioning

confidence: 99%

Jejunal Dopamine and Na+,K+-ATPase Activity in Nephrotic Syndrome

2005

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Background: The occurrence of complementary functions in sodium transport between the intestine and the kidney was suggested to occur when the renal function is immature or compromised and jejunal dopamine has been implicated in this renal-intestinal cross-talk. The jejunal sodium transport was not previously evaluated in the nephrotic syndrome. Methods: We examined the jejunal Na⁺,K⁺-ATPase activity and the role of dopamine in puromycin aminonucleoside (PAN) and HgCl₂-induced nephrotic syndrome rat models. Results: In both nephrotic syndrome rat models, the jejunal Na⁺,K⁺-ATPase activity was reduced during greatest sodium retention and ascites accumulation (PAN nephrosis, day 7; HgCl₂ nephrosis, day 14), whereas during enhanced sodium excretion and ascites mobilization the jejunal Na⁺,K⁺-ATPase activity was increased in HgCl₂ nephrosis (day 21) and was similar to controls in PAN nephrosis (day 14). In both PAN- and HgCl₂-induced nephrosis, the jejunal aromatic L-amino acid decarboxylase (AADC) activity, the enzyme responsible for the synthesis of jejunal dopamine, did not differ from controls. In addition, the jejunal Na⁺,K⁺-ATPase activity was not sensitive to inhibition by dopamine (1 µM) in both experimental groups throughout the study. Conclusions: In the nephrotic syndrome the jejunal Na⁺,K⁺-ATPase activity may respond in a compensatory way to changes in extracellular volume, through dopamine-independent mechanisms.

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Inhibition of Amino Acid Transport in Rabbit Intestine by p-Chloromercuriphenyl Sulfonic Acid

Cited by 45 publications

References 24 publications

System a transport activity in normal rat hepatocytes and transformed liver cells: Substrate protection from inactivation by sulfhydryl‐modifying reagents

System a transport activity in normal rat hepatocytes and transformed liver cells: Substrate protection from inactivation by sulfhydryl‐modifying reagents

Protein and glucose‐induced changes in sodium transport across the pig small intestine

Jejunal Dopamine and Na<sup>+</sup>,K<sup>+</sup>-ATPase Activity in Nephrotic Syndrome

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