THE present study examined changes in the angiotensin type 2 (AT2) receptor mRNA level after carbachol exposure in PC12 cells. The AT2 receptor mRNA level increased 2-to 3-fold after 12–18 h of carbachol exposure (100 μM). The up-regulation of AT2 receptor mRNA was antagonized by atropine, which is a muscarinic receptor antagonist, thus suggesting that the increase in AT2 receptor mRNA is mediated via muscarinic acetylcholine receptors. This increase is blocked by NG-nitro-L-arginine-methylester, nitric oxide (NO) synthase inhibitor, and hemoglobin NO trap. Protein kinase C (PKC) inhibitors, H-7 and calphostin C, inhibited the carbachol-induced upregulation. In addition, a G kinase inhibitor, ODQ (1H-[1,2,4] oxadiazolo[4,3-a] quinoxalin-1-one), inhibited this increase. These findings suggest that the carbachol-induced increases in AT2 receptor mRNA is regulated by the activation of NO-cGMP and/or the PKC pathway.