Objectives: Our purpose was to investigate the role of the nitric oxide (NO) signaling pathway in the regulation of ciliary beat frequency (CBF) in mouse nasal and tracheal epithelial cells. Methods: We studied the effects of the NO donor L-arginine (L-Arg) and specific inhibitors of the NO signaling pathway on CBF of both nasal and tracheal epithelial cells by using high-speed digital microscopy. We also examined eNOS, sGC beta, PKG I and acetylated a tubulin expression in native mouse nasal and tracheal epithelium using immunohistochemical methods. Results: L-Arg significantly increased CBF of cultured nasal and tracheal epithelial cells, and the effects were blocked by pretreatment with N-G-nitro-L-arginine methyl ester CL-NAME), a NOS inhibitor, with LY-83583, a sGC inhibitor, or with KT-5823, a PKG inhibitor. Positive immunostaining for NO signaling molecules including eNOS, sGC beta and PKG I was observed in either nasal or tracheal ciliated epithelium. Conclusion: NO plays a role in regulating CBF of mouse respiratory epithelial cells via a eN0S-NO-sGC beta-cGMP-PKG I pathway. (C) 2011 Elsevier Inc. All rights reserved.