Olfactory neurons transduce the binding of odorants into membrane depolarization. Two intracellular messengers, cyclic AMP (cAMP) and inositol trisphosphate (IP3), are thought to mediate this process, with cAMP generating responses to some odorants and IP3 mediating responses to others. cAMP causes membrane depolarization by activating a cation-selective cyclic nucleotide-gated (CNG) channel. We created a mutant "knockout" mouse lacking functional olfactory CNG channels to assess the roles of different second messenger pathways in olfactory transduction. Using an electrophysiological assay, we find that excitatory responses to both cAMP- and IP3-producing odorants are undetectable in knockout mice. Our results provide direct evidence that the CNG channel subserves excitatory olfactory signal transduction, and further suggest that cAMP is the sole second messenger mediating this process.