title: Interaction of O(2)(-) and NO in the thick ascending limb.
abstract: Nitric oxide (NO) is an important regulator of NaCl absorption by the thick ascending limb of the loop of Henle (THAL). The free radical superoxide (O(2)(-)) reacts with NO, decreasing its bioavailability. O(2)(-) is produced by mitochondria and various oxidases, some of which are present in the THAL. However, the ability of the THAL to produce O(2)(-) and its interaction with NO have not been studied. We hypothesized that NO bioavailability is decreased by O(2)(-). THALs were isolated and perfused and NO production was measured with an NO-selective microelectrode. Addition of L-Arg (250 micromol/L), but not D-arginine, to the bath increased NO release by 34.8 +/- 11.8 pA (n=7). The response to L-Arg was completely abolished by the NO synthase inhibitor L-NAME (n=7). Scavenging THAL O(2)(-) with the superoxide dismutase (SOD) mimetic Tempol (50 micromol/L) increased L-Arg-induced NO release. At all concentrations of L-Arg tested (50, 100, 250, 500, and 750 micromol/L), further addition of Tempol to the bath significantly increased NO release by THALs. Addition of SOD (300 U/mL) to the bath increased L-Arg-induced NO levels by 82% (n=5; P<0.02). Pretreatment of THALs with the SOD inhibitor diethyl-dithiocarbamate (250 micromol/L) blunted L-Arg-induced NO release by 63% compared with untreated tubules (n=5; P<0.05). Finally, we tested the effect of Tempol on NO-induced inhibition of THAL chloride transport. Addition of L-Arg decreased THAL Cl(-) absorption by 35%. Subsequent addition of Tempol (50 micromol/L) to the bath further decreased Cl(-) absorption by 35% (n=6; P<0.05). We conclude that NO bioavailability in the THAL is decreased by O(2)(-). In addition, we believe our studies are the first to show that endogenous O(2)(-) may act as a physiological regulator of nephron NaCl transport.
authors.affiliation: Division of Hypertension and Vascular Research, Department of Internal Medicine, Henry Ford Hospital, Detroit, MI 48202, USA.
authors.forename: Pablo A
authors.initials: PA
authors.lastname: Ortiz
authors.forename: Jeffrey L
authors.initials: JL
authors.lastname: Garvin
mesh_terms: D000818:Animals; D001692:Biological Transport; D008138:Loop of Henle / Q000378:metabolism*; D008297:Male; D009569:Nitric Oxide / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism*
chemical_list: D017382:Reactive Oxygen Species; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase