title: Angiotensin II activation of TRPC6 channels in rat podocytes requires generation of reactive oxygen species.
abstract: Angiotensin II (AII) plays a major role in the progression of chronic kidney diseases. Podocytes are essential components of the ultrafiltration apparatus, and are targets for AII signaling. AII has been shown to increase generation of reactive oxygen species (ROS) in podocytes. Canonical transient receptor potential-6 (TRPC6) channels stimulate Ca(2+) influx in podocytes, and have been implicated in glomerular disease. We observed that AII increased cationic currents in rat podocytes in an isolated glomerulus preparation in which podocytes are still attached to the underlying capillary. This effect was completely blocked by SKF-96365, by micromolar La(3+) , and by siRNA knockdown of TRPC6, indicating that TRPC6 is the primary source of Ca(2+) influx mobilized by endogenously expressed angiotensin II receptors in these cells. These responses were also blocked by the AT1R antagonist losartan, the phospholipase C inhibitor D-609, and by inhibition of G protein signaling. The pan-protein kinase C inhibitor chelerythrine had no effect. Importantly, pretreating podocytes with the ROS quencher manganese (III) tetrakis (4-benzoic acid) porphyrin chloride (MnTBAP) eliminated AII activation of TRPC6. Significant reductions of AII effects on podocyte TRPC6 were also observed after pretreatment with NADPH oxidase inhibitors apocynin or diphenylene iodonium (DPI). These data suggest that ROS production permits activation of TRPC6 channels by G protein and PLC-dependent cascades initiated by AII acting on AT1Rs in podocytes. This pathway also provides a basis whereby two forms of cellular stress-oxidative stress and Ca(2+) overload-converge on common pathways relevant to disease.
authors.affiliation: Department of Biology and Biochemistry, University of Houston, Houston, Texas.
authors.forename: Marc
authors.initials: M
authors.lastname: Anderson
authors.forename: Hila
authors.initials: H
authors.lastname: Roshanravan
authors.forename: Justin
authors.initials: J
authors.lastname: Khine
authors.forename: Stuart E
authors.initials: SE
authors.lastname: Dryer
mesh_terms: D000804:Angiotensin II / Q000494:pharmacology*; D000818:Animals; D005071:Evoked Potentials; D005786:Gene Expression Regulation / Q000502:physiology*; D007093:Imidazoles / Q000494:pharmacology; D050199:Podocytes / Q000502:physiology*; D034622:RNA Interference; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism*; D011945:Receptors, Angiotensin / Q000235:genetics / Q000378:metabolism; D050052:TRPC Cation Channels / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
chemical_list: D007093:Imidazoles; D017382:Reactive Oxygen Species; D011945:Receptors, Angiotensin; D050052:TRPC Cation Channels; C492145:Trpc6 protein, rat; D000804:Angiotensin II; C063159:1-(2-(3-(4-methoxyphenyl)propoxy)-4-methoxyphenylethyl)-1H-imidazole