Supplementary Material for: Fluorescence Imaging of Intracellular Calcium Signals in Intact Kidney Tissue
Peppiatt-Wildman C.M.
Crawford C.
Hall A.M.
10.6084/m9.figshare.5468665.v1
https://karger.figshare.com/articles/dataset/Supplementary_Material_for_Fluorescence_Imaging_of_Intracellular_Calcium_Signals_in_Intact_Kidney_Tissue/5468665
<p><b><i>Background:</i></b> 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. <b><i>Methods:</i></b> 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. <b><i>Results:</i></b> 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. <b><i>Conclusion:</i></b> 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.</p>
2017-10-04 10:52:27
Calcium
Fluorescence imaging
Intact kidney