eNOS Mediates l -Arginine–Induced Inhibition of Thick Ascending Limb Chloride Flux

Publisher Website
Google Scholar
Abstract
—We recently reported that the rat thick ascending limb (THAL) possesses an active isoform of nitric oxide synthase (NOS) that is substrate-limited in vitro. NO produced by THAL NOS inhibits chloride flux. Protein and transcript for each of the primary NOS isoforms−endothelial (eNOS), inducible (iNOS), and neuronal (nNOS)−have been demonstrated in THALs. However, the NOS isoform that mediates NO-induced inhibition of chloride flux is unknown. We hypothesized that NO produced from eNOS in the THAL inhibits NaCl transport. THALs from male eNOS, iNOS, and nNOS knockout mice and C57BL/6J wild-type controls were perfused in vitro and the response of transepithelial chloride flux (J Cl ) to l -arginine (L-Arg), the substrate for NOS, and spermine NONOate (SPM), an NO donor was measured. We first tested whether isolated mouse THALs could synthesize NO and whether this NO inhibits transport. Addition of 0.5 mmol/L L-Arg to the bath decreased J Cl from 105.8±17.5 to 79.2±15.8 pmol/mm per minute ( P Cl. In contrast, addition of 0.5 mmol/L L-Arg to the bath did not alter J Cl of THALs from eNOS knockout mice. When 10 μmol/L SPM was added to the bath of eNOS knockout THALs, J Cl decreased from 89.1±8.6 to 74.8±7.5 pmol/mm/min ( P Cl in THALs from iNOS and nNOS knockout mice by 37.7±6.4% ( P P Cl .