The long-term objective of this proposal is to understand in quantitative detail the cAMP- and Ca2+-signalings in sensory transduction by olfactory receptor neurons (ORNs). We shall focus on the canonical olfactory-transduction mechanism in the vertebrate main olfactory epithelium. This mechanism involves a cAMP-signaling cascade, leading to Na+ and Ca2+ influxes through a cyclic-nucleotide-gated (CNG), non-selective cation channel to depolarize the ORN to firing threshold. The Ca2+ influx leads to signal amplification via an inward Ca2+-activated Cl current, as well as olfactory adaptation via multiple Ca2+-activated negative-feedback pathways. Recently, however, the significance and performance of the negative-feedback pathways are thrown into doubt and confusion. In Aim 1, we propose to re-examine this question and to settle it once and for all. Most recently, by using M71-monoclonal-nose mouse ORNs, we have succeeded in quantifying the density of M71-OR molecules on the olfactory cilia membrane. We also found that an M71-OR, when liganded with acetophenone (among the most efficacious odorants for M71-OR), nonetheless still has a very low probability of success (nominally ~10-4) in activating a single downstream Golf/adenylyl cyclase III effector complex. This low probability is very different from the situation in rod phototransduction, about which we recently showed that one photoexcited rhodopsin activates 10-20 rod transducin/cGMP-phosphodiesterase effectors. Thus, a ligand-activated GPCR pathway may be quite different in signal amplification from a light-activated GPCR pathway. In Aims 2, we propose to study another mouse nasal chemoreceptor, TAAR4, which is exceedingly sensitive to 2-phenylethylamine, a predator odorant aversive to mouse. The objective is to compare the findings with those from M71-OR, to figure out whether TAAR4's molecular density on the cilia's surface membrane is very different from that of M71-OR, and to ask whether the amplification at the downstream G-protein/effector enzyme complex activation step is any different from the case of M71-OR. In Aim 3, as another comparison, we shall address the same questions for mOR256-17, an OR with one of the highest abundances known so far in the main olfactory epithelium and with an unusually broad odorant spectrum. Quantitatively elucidating the steps of olfactory transduction will provide great insight into normal olfactory functions, as well as malfunctions arising from genetic defects in the transduction pathway, as amply demonstrated by the huge success as such in the case of visual transduction.