Abstract: The present study underlines the importance of p21^ras in regulating the inducible nitric oxide synthase (iNOS) in primary astrocytes. Bacterial lipopolysaccharides induced the GTP loading of p21^ras, and the expression of a dominant‐negative mutant of p21^ras (Δp21^ras) inhibited lipopolysaccharide‐induced GTP loading in rat primary astrocytes. To delineate the role of p21^ras in the induction of iNOS, we examined the effect of Δp21^ras on the expression of iNOS and the production of nitric oxide. It is interesting that expression of Δp21^ras markedly inhibited the production of nitric oxide and the expression of iNOS in lipopolysaccharide‐ and proinflammatory cytokine (tumor necrosis factor‐α, interleukin‐1β; interferon‐γ)‐stimulated rat and human primary astrocytes. Inhibition of iNOS promoter‐derived chloramphenicol acetyltransferase activity by Δp21^ras suggests that p21^ras is involved in the transcription of iNOS. As activation of nuclear factor‐κB (NF‐κB) is necessary for the transcription of iNOS, we examined the effect of Δp21^ras on the activation of NF‐κB. Expression of Δp21^ras inhibited the DNA binding as well as the transcriptional activity of NF‐κB in activated astrocytes, suggesting that Δp21^ras inhibits the expression of iNOS by inhibiting the activation of NF‐κB. These studies also suggest that inhibitors of p21^ras may be used as therapeutics in nitric oxide‐ and cytokine‐mediated neuroinflammatory diseases.