Abstract The phospholipid PI(3,4,5)P3 works as a second messenger in PI3K signaling pathway. The PI3K signaling pathway is involved in insulin stimulated nitric oxide (NO) production in vascular endothelium, leading to vasodilation and increased blood flow. However, the production of NO also has been reported in neurons as a neurotransmitter and in nucleus tractus solitarii (NTS), NO plays a role in central cardiovascular regulation. Previously, microinjection of insulin into the NTS of rats produces prominent depressor and bradycardic and activates the PI3K downstream Akt. Therefore, to investigate the detail downstream signaling of insulin stimulated NO production in NTS, the effects of PI(3,4,5)P3 on NO production were determined in neuronal cell lines PC12 and GH3 and in NTS of SD rats. The GH3 and differentiated PC12 exposed to 10