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The thick ascending limb of the loop of Henle (THAL) plays an essential role in the regulation of sodium and water homeostasis by the kidney. l-Arginine, the substrate for nitric oxide synthase (NOS), decreases NaCl absorption by THALs. We hypothesized that eNOS produces the NO that regulates THAL NaCl transport and that selective expression of eNOS in the THAL of eNOS knockout(-/-) mice would restore the effects of l-arginine on NaCl absorption. eNOS-/- mice were anesthetized, the left kidney was exposed, and the renal interstitium was injected with recombinant adenoviral vectors that expressed green fluorescent protein (GFP) or eNOS driven by the promoter of the Na/K/2Cl cotransporter Ad-NKCC2GFP and Ad-NKCC2eNOS, respectively. In Ad-NKCC2eNOS-transduced kidneys, eNOS expression was detected 7 days after injection but was absent in Ad-NKCC2GFP-transduced kidneys. In THALs from eNOS-/- mice transduced with Ad-NKCC2eNOS, adding L-arginine increased DAF-2DA fluorescence, a measure of NO production, by 9.1+/-1.1% (P&lt;0.05; n=5), but not in THALs transduced with Ad-NKCC2GFP. In THALs from eNOS-/- mice transduced with Ad-NKCC2eNOS, Cl absorption averaged 85.9+/-11.8 pmol/min per millimeter. Adding l-arginine (1 mmol/L) to the bath decreased Cl absorption to 59.7+/-11.0 pmol/min per millimeter (P&lt;0.05; n=6). In THALs transduced with Ad-NKCC2GFP, Cl absorption averaged 96.0+/-21.0 pmol/min per millimeter. Adding L-arginine to the bath did not significantly affect Cl absorption (100.6+/-20.6 pmol/min per millimeter; n=4). We concluded that gene transfer of eNOS to the THAL of eNOS-/- mice restores L-arginine-induced inhibition of NaCl transport and NO production. These data indicate that eNOS is essential for the regulation of THAL NaCl transport by NO.

Gene Transfer of eNOS to the Thick Ascending Limb of eNOS-KO Mice Restores the Effects of l -Arginine on NaCl Absorption