title: Functional distinctions in cytosolic calcium regulation between cells of the glomerular filtration barrier.
abstract: The importance of intracellular calcium ([Ca(2+)]i) regulation in the glomerular filtration barrier (GFB) has recently been highlighted by mutations in the cation channel TRPC6, resulting in a renal-specific phenotype. We examined the effects of FFA, a tool that can activate TRPC6, on [Ca(2+)]i in human conditionally immortalised glomerular endothelial cells (ciGEnC) and human podocytes (ciPod) that form the GFB. Changes in [Ca(2+)]i stimulated by FFA were measured in Fura 2-AM loaded cells. In GEnC, cell activation by FFA was dependent on external Ca(2+), yet in ciPod it was not. Depletion of internal Ca(2+) stores with thapsigargin did not affect cell activation by FFA in ciGEnC, but inhibited it in ciPod in a nephrin-dependent manner, demonstrated using nephrin deficient (ND) ciPod in conjunction with nephrin rescue experiments. FFA induced [Ca(2+)]i store release in ciPod, but not in ciGEnC or ND ciPod. In parallel, there were differences in the localisation of overexpressed TRPC6 between ciGEnC and ciPod. Furthermore, co-transfection of nephrin with TRPC6 in HEK293 cells reduced the FFA-induced increase in [Ca(2+)]i and nephrin clustering altered TRPC6 distribution. In conclusion, cell activation by FFA in podocytes stimulates the opening of a Ca(2+) channel, probably TRPC6, in a nephrin-dependent manner with a different activation profile to GEnC.
authors.affiliation: The Academic Renal Unit, Southmead Hospital, University of Bristol, BS10 5NB, UK. becky.foster@bristol.ac.uk
authors.forename: Rebecca Rachael
authors.initials: RR
authors.lastname: Foster
authors.forename: Gavin I
authors.initials: GI
authors.lastname: Welsh
authors.forename: Simon C
authors.initials: SC
authors.lastname: Satchell
authors.forename: Robin D
authors.initials: RD
authors.lastname: Marlow
authors.forename: Mathew D
authors.initials: MD
authors.lastname: Wherlock
authors.forename: Debora
authors.initials: D
authors.lastname: Pons
authors.forename: Peter W
authors.initials: PW
authors.lastname: Mathieson
authors.forename: David O
authors.initials: DO
authors.lastname: Bates
authors.forename: Moin A
authors.initials: MA
authors.lastname: Saleem
mesh_terms: D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism*; D042783:Endothelial Cells / Q000187:drug effects / Q000502:physiology; D005439:Flufenamic Acid / Q000494:pharmacology; D057809:HEK293 Cells; D006801:Humans; D007678:Kidney Glomerulus / Q000166:cytology / Q000378:metabolism*; D008565:Membrane Proteins / Q000502:physiology; D050199:Podocytes / Q000378:metabolism; D050052:TRPC Cation Channels / Q000502:physiology*; D000074287:TRPC6 Cation Channel
chemical_list: D008565:Membrane Proteins; D050052:TRPC Cation Channels; D000074287:TRPC6 Cation Channel; C117032:TRPC6 protein, human; C113267:nephrin; D005439:Flufenamic Acid; D002118:Calcium