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      Fluorescence Imaging of Intracellular Calcium Signals in Intact Kidney Tissue


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          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

          Nephron Exp Nephrol
          Cardiorenal Medicine
          S. Karger AG
          December 2012
          07 November 2012
          : 121
          : 1-2
          : e49-e58
          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
          342812 Nephron Exp Nephrol 2012;121:e49–e58
          © 2012 S. Karger AG, Basel

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          Page count
          Figures: 4, Tables: 1, Pages: 10
          Original Paper

          Cardiovascular Medicine, Nephrology

          Fluorescence imaging, Calcium, Intact kidney


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