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      Effect of Renal Ischemia/Reperfusion on Gene Expression of a pH-Sensitive K + Channel

      , , ,

      Nephron Physiology

      S. Karger AG

      Kir4.1, K+ channel, ATP depletion

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          Abstract

          Background: Sodium reabsorption depends on the Na/K/ATPase activity coupled to basolateral K<sup>+</sup> recycling through K<sup>+</sup> channels. ATP depletion reduces pump activity and increases K<sup>+</sup> leak resulting in transport dysfunction. Kir4.1 is a pH-sensitive K<sup>+</sup> channel expressed in the basolateral membrane of distal tubules. In this study, we evaluated whether Kir4.1 is also expressed in proximal tubules (PTs) and whether renal ischemia alters Kir4.1 mRNA expression levels. Methods: The presence of Kir4.1 mRNA was evaluated in PTs microdissected from collagenase-treated rat kidneys. Kir4.1 expression levels were estimated in the renal cortex by multiplex RT-PCR after 30 or 60 min of renal ischemia followed by 1, 24, 48 or 72 h of reperfusion. Results: The PCR product obtained from isolated tubules was sequenced and showed ∼98% homology with rat Kir4.1 cDNA. Ischemia/reperfusion for 30 min induced a time-dependent reduction in Kir4.1 mRNA expression in parallel with plasma creatinine, however recovery was delayed after 60 min of ischemia, remaining reduced after 72 h of reperfusion when plasma creatinine was already normalized. Conclusion: Kir4.1 mRNA expression was decreased by renal ischemia. The ischemia-induced cellular K<sup>+</sup> loss may be minimized by Kir4.1 downregulation and may contribute to the mechanism by which cellular acidification induces cell protection against ATP depletion.

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          Most cited references 15

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          An inward rectifier K(+) channel at the basolateral membrane of the mouse distal convoluted tubule: similarities with Kir4-Kir5.1 heteromeric channels.

          In this study, K(+) channels present in the basolateral membrane of the distal convoluted tubule (DCT) were investigated using patch-clamp methods. In addition, Kir4.1, Kir4.2 and Kir5.1 inward rectifier channels were investigated using RT-PCR and immunohistochemistry (Kir4.1). DCTs were microdissected from collagenase-treated mouse kidneys. One type of K(+) channel was detected in about 50 % of cell-attached patches from the DCT basolateral membrane; this channel was inwardly rectifying and had an inward conductance (g(in)) of approximately 40 pS at an external [K(+)] of 145 mM. The current-voltage relationship was linear when inside-out patches were exposed to a Mg(2+)-free medium. Mg(2+) at a concentration of 1.2 mM considerably reduced the outward conductance (g(out)), yielding a g(in)/g(out) ratio of approximately 4.7. The polycation spermine (5 x 10(-7) M) reduced the open probability (P(o)) by 50 %. Channel activity was dependent upon the intracellular pH, with acid pH decreasing, and basic pH increasing, P(o). Internal ATP (2 mM) and Ca(2+) (up to 10(-3) M) had no effect. Channel activity declined irreversibly when the inner side of the patch was exposed to Mg(2+). Kir4.1, Kir4.2 and Kir5.1 mRNAs were all detected in the DCT. The Kir4.1 protein co-localised with the Na(+)-Cl(-) cotransporter, which is specific to the DCT, and was located on basolateral membranes. The DCT K(+) channel differs from other functionally identified renal K(+) channels with regard to its inhibition by spermine and insensitivity to internal ATP and Ca(2+). At the current state of knowledge, the channel is similar to Kir4.1-Kir5.1 and Kir4.2-Kir5.1 heteromeric channels, but not to Kir4.1 or Kir4.2 homomeric channels.
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            pH dependence of the inwardly rectifying potassium channel, Kir5.1, and localization in renal tubular epithelia.

            The physiological role of the inwardly rectifying potassium channel, Kir5.1, is poorly understood, as is the molecular identity of many renal potassium channels. In this study we have used Kir5.1-specific antibodies to reveal abundant expression of Kir5.1 in renal tubular epithelial cells, where Kir4.1 is also expressed. Moreover, we also show that Kir5.1/Kir4.1 heteromeric channel activity is extremely sensitive to inhibition by intracellular acidification and that this novel property is conferred predominantly by the Kir5.1 subunit. These findings suggest that Kir5.1/Kir4.1 heteromeric channels are likely to exist in vivo and implicate an important and novel functional role for the Kir5.1 subunit.
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              In vivo formation of a proton-sensitive K+ channel by heteromeric subunit assembly of Kir5.1 with Kir4.1.

              Kir5.1 is an inwardly rectifying K+ channel (Kir) subunit, whose physiological function is unknown. Human embryonic kidney HEK293T cells co-transfected with rat Kir5.1 and Kir4.1 cDNA expressed a functional K+ channel, whose properties were significantly different from those of the homomeric Kir4.1 channel. Formation of a Kir4. 1/Kir5.1 assembly in HEK293T was confirmed biochemically. We found that heteromeric Kir4.1/Kir5.1 channel activity was affected by internal pH levels between 6.0 and 8.0, when the homomeric Kir4.1 channel activity was relatively stable. Changing external pH in this range had no effect on either Kir channel. Western blot analysis using specific antibodies revealed that Kir4.1 and Kir5.1 proteins were expressed in kidney and brain, but co-immunoprecipitated only from kidney. These results indicate that the co-assembly of Kir5.1 with Kir4.1 occurs in vivo, at least in kidney. The heteromeric Kir4. 1/Kir5.1 channel may therefore sense intracellular pH in renal epithelium and be involved in the regulation of acid-base homeostasis.
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                Author and article information

                Journal
                NEP
                Nephron Physiol
                10.1159/issn.1660-2137
                Nephron Physiology
                S. Karger AG
                1660-2137
                2007
                May 2007
                02 April 2007
                : 106
                : 1
                : p1-p7
                Affiliations
                Disciplina de Nefrologia, Universidade Federal de São Paulo, São Paulo, Brasil
                Article
                101486 Nephron Physiol 2007;106:p1–p7
                10.1159/000101486
                17406122
                © 2007 S. Karger AG, Basel

                Copyright: All rights reserved. No part of this publication may be translated into other languages, reproduced or utilized in any form or by any means, electronic or mechanical, including photocopying, recording, microcopying, or by any information storage and retrieval system, without permission in writing from the publisher. Drug Dosage: The authors and the publisher have exerted every effort to ensure that drug selection and dosage set forth in this text are in accord with current recommendations and practice at the time of publication. However, in view of ongoing research, changes in government regulations, and the constant flow of information relating to drug therapy and drug reactions, the reader is urged to check the package insert for each drug for any changes in indications and dosage and for added warnings and precautions. This is particularly important when the recommended agent is a new and/or infrequently employed drug. Disclaimer: The statements, opinions and data contained in this publication are solely those of the individual authors and contributors and not of the publishers and the editor(s). The appearance of advertisements or/and product references in the publication is not a warranty, endorsement, or approval of the products or services advertised or of their effectiveness, quality or safety. The publisher and the editor(s) disclaim responsibility for any injury to persons or property resulting from any ideas, methods, instructions or products referred to in the content or advertisements.

                Page count
                Figures: 5, References: 23, Pages: 1
                Categories
                Original Paper

                Cardiovascular Medicine, Nephrology

                ATP depletion, K+ channel, Kir4.1

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