Role of gp91phox-Containing NADPH Oxidase in Mediating the Effect of K Restriction on ROMK Channels and Renal K Excretion

Abstract

Previous study has demonstrated that superoxide and the related products are involved in mediating the effect of low K intake on renal K secretion and ROMK channel activity in the cortical collecting duct (CCD). This study investigated the role of gp91 phox -containing NADPH oxidase (NOXII) in mediating the effect of low K intake on renal K excretion and ROMK channel activity in gp91(Ϫ/Ϫ) mice. K depletion increased superoxide levels, phosphorylation of c-Jun, expression of c-Src, and tyrosine phosphorylation of ROMK in renal cortex and outer medulla in wild-type (WT) mice. In contrast, tempol treatment in WT mice abolished whereas deletion of gp91 significantly attenuated the effect of low K intake on superoxide production, c-Jun phosphorylation, c-Src expression, and tyrosine phosphorylation of ROMK. Patch-clamp experiments demonstrated that low K intake decreased mean product of channel number (N) and open probability (P) (NP o ) of ROMK channels from 1.1 to 0.4 in the CCD. However, the effect of low K intake on ROMK channel activity was significantly attenuated in the CCD from gp91(Ϫ/Ϫ) mice and completely abolished by tempol treatment. Immunocytochemical staining also was used to examine the ROMK distribution in WT, gp91(Ϫ/Ϫ), and WT mice with tempol treatment in response to K restriction. K restriction decreased apical staining of ROMK in WT mice. In contrast, a sharp apical ROMK staining was observed in the tempol-treated WT or gp91(Ϫ/Ϫ) mice. Metabolic cage study further showed that urinary K loss is significantly higher in gp91(Ϫ/Ϫ) mice than in WT mice. It is concluded that superoxide anions play a key role in suppressing K secretion during K restriction and that NOXII is involved in mediating the effect of low K intake on renal K secretion and ROMK channel activity.

NADPH Oxidases, Reactive Oxygen Species, and the Kidney

“…The effect of gp91phox deletion was mimicked by the administration of Tempol to wild-type mice on a low potassium diet. 37 These results are supported by the findings of Wei et al, 38 who observed that NAPDH oxidase mediated the effect of AngII on the reduction of ROMK channel activity in rats treated with a low potassium diet. Potassium restriction also inhibited protein phosphatase 2B (PP2B) in rat and mouse kidney; suppression of PP2B decreased ROMK channel activity in the cortical collecting duct.…”

NAD(P)H oxidase and renal epithelial ion transport

“…As urinary K ϩ excretion and plasma K ϩ decrease were slightly higher in gp91phox Ϫ/Ϫ mice compared with WT mice, these data indicate that O 2 Ϫ production is an adaptive mechanism for K ϩ conservation during K ϩ deprivation and that NOX2 is a source of O 2 Ϫ . Even though the increase in O 2 Ϫ production was less than that observed in WT mice, the observation that K ϩ deprivation was able to increase O 2 Ϫ levels in gp91phox Ϫ/Ϫ indicates a second source of ROS in addition to NOX2 (2). This secondary source of ROS remains undetermined.…”

“…Babilonia et al (2,3) found that in rats subject to a K ϩ -deficient (KD) diet for 7 days, enough to provoke hypokalemia and dramatically reduce urinary K ϩ excretion, O 2 Ϫ production, assessed by lucigenin chemiluminescence in homogenates of cortex and outer medulla, increased by 110% (3). Treatment of rats with Tempol to prevent the rise in renal O 2 Ϫ levels resulted in urinary K ϩ excretion levels above those of KD vehicle-treated animals, indicating that ROS were responsible for the reduction on K ϩ excretion (3).…”

Caveolin-1 Deficiency Inhibits the Basolateral K+ Channels in the Distal Convoluted Tubule and Impairs Renal K+ and Mg2+ Transport

“…44 Briefly, we used eight WT mice and eight cav-1 KO mice for the study, and both WT and KO mice were age and sex matched. Two mice were housed in a cage for the study, and they stayed in the cage for 3-4 days to allow them to be adapted with the environment.…”