An In Situ Kidney Slice Model for Studying Angiotensin II‐ and TRPC5‐Mediated Calcium Signaling

Abstract

Current approaches to study calcium signaling in glomeruli use isolated glomeruli or cultured cells, however the structure and physiology of the tissue is likely to be perturbed by the isolation process in both models. We used kidney slices maintained in physiological solution to better preserve the macroscopic structure of glomeruli and measured the impact of angiotensin II receptor activation and blockade of Transient Potential Receptor Canonical 5 (TRPC5) to examine their possible interaction in a number of pre‐ and intraglomerular cell types using spinning‐disk confocal calcium imaging.Calcium transients and tissue distortion in intact kidney slices of mice expressing genetically modified calcium‐sensors (GCaMP5) in Cx40‐positive cells were recorded and analyzed using particle analysis.We observed spontaneous calcium transients in preglomerular cells located at the vascular pole that were blocked by the SERCA pump inhibitor, cyclopiazonic acid (CPA). Stimulation with angiotensin II increased calcium activity and contraction in these cells and induced calcium transients in distinct populations of intraglomerular cells that were also blocked with CPA. While removal of external calcium did not change angiotensin II‐induced calcium transients in intraglomerular cells, the frequency of calcium transients in preglomerular cells was significantly reduced. In angiotensin II‐preincubated kidney slices, spontaneous calcium transients in the preglomerular area were diminished by the specific TRPC4/5 inhibitor, ML204. If given alone, ML204 completely inhibited all preglomerular calcium transients.Thick kidney slices maintained in vitro appear to better preserve the structure and responses of different cell types in glomeruli as measured using calcium transients and contractions. In our experimental set‐up, angiotensin II induced calcium transients in intraglomerular cells, while its impact on preglomerular cells appeared more indirect. As the TRPC5 antagonist was more effective at inhibiting calcium transients in preglomerular cells when the tissue was not prestimulated with angiotensin II, we suggest that TRPC5‐mediated calcium signaling is at least party modulated by angiotensin II.Support or Funding InformationSupported by Deutsche Forschungsgemeinschaft (DFG)This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.