Nitric oxide (NO) has been implicated as an autocrine modulator of active sodium transport. To determine whether tonic exposure to NO influences active sodium transport in epithelial cells, we established transfected medullary thick ascending limb of Henle (MTAL) cell lines that overexpressed NO synthase-2 (NOS2) and analyzed the effects of deficient or continuous NO production [with or without N^G-nitro-L-arginine methyl ester (L-NAME) in the culture medium, respectively] on Na⁺-K⁺-ATPase function and expression. The NOS2-transfected cells exhibited high-level NOS2 expression and NO generation, which did not affect cell viability or cloning efficiency. NOS2-transfected cells were grown in the presence of vehicle, N^G-nitro-D-arginine methyl ester (D-NAME), or L-NAME for 16 h, after which ⁸⁶Rb⁺ uptake assays, Northern analysis, or nuclear run-on transcription assays were performed. The NOS2-transfected cells allowed to produce NO continuously (vehicle or D-NAME) exhibited lower rates of ouabain-sensitive ⁸⁶Rb⁺ uptake (~65%), lower levels of Na⁺-K⁺-ATPase 1-subunit mRNA (~60%), and reduced rates of de novo Na⁺-K⁺-ATPase 1-subunit transcription compared with L-NAME-treated cells. These results have uncovered a novel effect of NO to inhibit transcription of the Na⁺-K⁺-ATPase 1-subunit gene.

sodium pump; nitric oxide synthase; gene expression; kidney; sodium transport

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