Cytokines such as interleukin-1beta (IL-1beta) stimulate inducible nitric oxide synthase (iNOS) expression and nitric oxide overproduction leading to beta-cell damage. Meanwhile, glucagon-like peptide-1 (GLP-1) and its potent analog exendin-4 (EX-4) were well known for beta-cell proliferation. However, the protective mechanisms of GLP-1 in beta-cells exposed to cytokines were not fully elucidated. Therefore, the effects of EX-4 on the IL-1beta-induced iNOS gene expression were investigated employing RINm5F beta-cells. EX-4 inhibited IL-1beta-induced iNOS protein expression and nitrite production. However, northern blot and promoter analyses showed that EX-4 failed to inhibit IL-1beta-induced iNOS mRNA expression and iNOS promoter activity. By electrophoretic mobility shift assay (EMSA), EX-4 did not alter the binding activity of NF-kappaB to the iNOS promoter. Consistent with the EMSA result, EX-4 did not inhibit nuclear translocation of p65. We also tested the effect of EX-4 on iNOS mRNA stability. Actinomycin D chase experiments showed that EX-4 did not affect the decay rate of iNOS mRNA and the promoter assay using the construct containing 3'-untranslated region of iNOS showed that EX-4 did not alter the stability of iNOS mRNA. Meanwhile, forskolin significantly inhibited IL-1beta-induced iNOS protein, which was reversed by H-89, a protein kinase A (PKA) inhibitor. Moreover, EX-4 pretreatment restored IL-1beta-induced decrease in cAMP toward control level. Additionally, the cycloheximide chase study demonstrated that EX-4 significantly accelerated iNOS protein degradation. We therefore concluded that EX-4 inhibited IL-1beta-induced iNOS protein and nitrite production via cAMP/PKA system irrespective of both transcriptional and posttranscriptional mechanisms of iNOS gene, and this inhibitory effect of EX-4 appears to be regulated at posttranslational level.