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      Fluorescence Imaging of Intracellular Calcium Signals in Intact Kidney Tissue

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          Abstract

          Background: Intracellular calcium (Ca<sup>2+</sup>) plays an important role in normal renal physiology and in the pathogenesis of various kidney diseases; however, the study of Ca<sup>2+</sup> signals in intact tissue has been limited by technical difficulties, including achieving adequate loading of Ca<sup>2+</sup>-sensitive fluorescent dyes. The kidney slice preparation represents a model whereby three-dimensional tissue architecture is preserved and structures in both the cortex and medulla can be imaged using confocal or multiphoton microscopy. Methods: Ca<sup>2+</sup>-sensitive dyes Rhod-2, Fura-red and Fluo-4 were loaded into tubular and vascular cells in rat kidney slices using a re-circulating perfusion system and real-time imaging of Ca<sup>2+</sup> signals was recorded by confocal microscopy. Kidney slices were also obtained from transgenic mice expressing the GCaMP2 Ca<sup>2+</sup>-sensor in their endothelial cells and real time Ca<sup>2+</sup> transients stimulated by physiological stimuli. Results: Wide spread loading of Ca<sup>2+</sup> indicators was achieved in the tubular and vascular structures of both the medulla and cortex. Real time Ca<sup>2+</sup> signals were successfully recorded in different intracellular compartments of both rat and mouse cortical and medullary tubules in response to physiological stimuli (ATP and angiotensin II). Glomerular Ca<sup>2+</sup> transients were similarly recorded in kidney slices taken from the transgenic mouse expressing the GCaMP2 Ca<sup>2+</sup>-sensor. Conclusion: We present new approaches that can be adopted to image cytosolic and mitochondrial Ca<sup>2+</sup> signals within various cell types in intact kidney tissue. Moreover, techniques described in this study can be used to facilitate future detailed investigations of intracellular Ca<sup>2+</sup> homeostasis in renal health and disease.

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          Author and article information

          Journal
          NEE
          Nephron Exp Nephrol
          10.1159/issn.1660-2129
          Cardiorenal Medicine
          S. Karger AG
          1660-2129
          2012
          December 2012
          07 November 2012
          : 121
          : 1-2
          : e49-e58
          Affiliations
          aMedway School of Pharmacy, The Universities of Kent and Greenwich at Medway, Chatham Maritime, and bUCL Centre for Nephrology, Royal Free Hospital, London, UK
          Author notes
          *Dr. Claire M. Peppiatt-Wildman, Medway School of Pharmacy, The Universities of Kent and Greenwich at Medway, Anson Building, Central Avenue, Chatham Maritime ME4 4TB (UK), E-Mail C.M.Peppiatt@kent.ac.uk
          Article
          342812 Nephron Exp Nephrol 2012;121:e49–e58
          10.1159/000342812
          23147410
          c1bc2d0b-25c4-4d53-bdb5-c409d24af0bc
          © 2012 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: 4, Tables: 1, Pages: 10
          Categories
          Original Paper

          Cardiovascular Medicine, Nephrology

          Fluorescence imaging, Calcium, Intact kidney

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