Production and actions of superoxide in the renal medulla

Abstract

The present study characterized the biochemical pathways responsible for superoxide (O2-) production in different regions of the rat kidney and determined the role of O2- in the control of renal medullary blood flow (MBF) and renal function. By use of dihydroethidium/DNA fluorescence spectrometry with microtiter plates, the production of O2- was monitored when tissue homogenate from different kidney regions was incubated with substrates for the major O2- -producing enzymes, such as NADH/NADPH oxidase, xanthine oxidase, and mitochondrial respiratory chain enzymes. The production of O2- via NADH oxidase was greater (P<0.05) in the renal cortex and outer medulla (OM) than in the papilla. The mitochondrial enzyme activity for O2- production was higher (P<0.05) in the OM than in the cortex and papilla. Compared with NADH oxidase and mitochondrial enzymes, xanthine oxidase and NADPH oxidase produced much less O2-in the kidney under this condition. Overall, the renal OM exhibited the greatest enzyme activities for O2- production. In anesthetized rats, renal medullary interstitial infusion of a superoxide dismutase inhibitor, diethyldithiocarbamate, markedly decreased renal MBF and sodium excretion. Diethyldithiocarbamate (5 mg/kg per minute by renal medullary interstitial infusion [R1])reduced the renal medullary laser-Doppler flow signal from 0.6±0.04 to 0.4±0.03 V, a reduction of 33%, and both urine flow and sodium excretion decreased by 49%. In contrast, a membrane-permeable superoxide dismutase mimetic, 4-hydroxytetramethyl-piperidine-l-oxyl (TEMPOL, 30 μmol/kg per minute RI) increased MBF and sodium excretion by 34% and 69%, respectively. These effects of TEMPOL on renal MBF and sodium excretion were not altered by pretreatment with NG-nitro-L-arginine methyl ester (10/μg/kg per minute RI). We conclude that (1) renal medullary, O2- is primarily produced in the renal OM: (2) both NADH oxidase and mitochondrial enzymes are responsible for the O2- production in this kidney region: and (3) O2- exerts a tonic regulatory action on renal MBF.