Both the cAMP and the phosphoinositide (PI) second messenger systems have been implicated in olfactory signal transduction. We have developed a primary culture system of mammalian olfactory receptor neurons (ORNs; Ronnett et al., 1991a) to permit analysis of odorant- induced second messenger system activation in the intact ORN. The ability of a series of odorants to stimulate PI turnover and adenylyl cyclase was examined. All odorants stimulated both second messenger systems, although with differential potencies. Stimulation of PI turnover desensitized upon reexposure of cultures to odorant. The enhancement by single odorants of both adenylyl cyclase and PI turnover, but to varying degrees, affords a mechanism for increased specificity in olfactory signal transduction.