List of Abstracts from the 15th International Symposium on Olfaction and Taste

Abstract

Natural products have an illustrious history when it comes to deciphering basic cellular and molecular mechanisms that contribute to the detection or modulation of noxious (pain-producing) signals. We, too, have exploited the power of natural products and folk medicine to identify cellular signaling molecules that enable primary afferent sensory neurons to detect chemical and physical stimuli that elicit discomfort or pain. These include members of the excitatory TRP ion channel family that respond to plant-derived irritants such as capsaicin, menthol, and mustard oil. Using genetic, electro-physiological, and behavioral methods, we have tested roles for these channels in a variety of somatosensory modalities, including thermosensation and chemosensation, both in regard to acute nociception and pain hyper-sensitivity. We have also used natural products (including novel spider tox-ins) to elucidate mechanisms of TRP channel activation and modulation, with the goal of understanding how these channels integrate information from physical and chemical stimuli to regulate sensory neuron excitability. Recent findings from these studies will be presented and discussed. The Drosophila melanogaster odorant-binding protein (OBP) LUSH mediates olfactory responses to two different ligands: one, ethanol, is a nonspecific environmental cue; the other, 11-cis-vaccenyl acetate (cVA), is a species-specific aggregation and mating pheromone. Flies lacking lush are defective in their avoidance of high ethanol concentrations (Kim et al., Genetics 150:711-721, 1998), and also are not attracted to cVA (Xu et al. Neuron 45:193-200, 2000). These defects can be rescued by either expression of LUSH or addition of the protein exogenously. These studies suggest that LUSH may have a more complex role in olfaction than simply acting as a passive odorant carrier. We have solved the X-ray crystal structure of LUSH bound to cVA to 1.4 resolution. From this structure we have identified a key interaction between cVA and F121 in LUSH that we hypothesize induces an ''active'' conformation of the protein. Substitution of this residue with alanine diminishes both pheromone-induced and spontaneous depolarizations of cVA-sensitive olfactory receptor neurons (ORN) in vivo. Furthermore, mutation of a neighboring residue, D118, to alanine results in a protein that induces ORN depolarizations in the absence of cVA. The X-ray crystal structure of D118A-LUSH without cVA indicates that this substitution allows the protein to adopt the same conformation as the WT protein with cVA, thus providing a mechanistic explanation for the activating effect. These results provide strong support for the hypothesis that LUSH is the primary ligand for the olfactory receptor complex in cVA sensitive T1 olfactory sensilla. This data also explain how this single OBP can mediate recognition of both attractive and repulsive stimuli. Deciphering olfactory encoding requires a thorough description of the ligands that activate each odorant receptor. In mammalian systems, however , ligands are known for only a handful of over 1000 odorant receptors, greatly limiting our understanding of olfaction. We performed high-throughput screening of ligands for mammalian odorant receptors using a large repertoire of mouse and human odorant receptors expressed in het-erologous cells. We identified excitatory ligands for 52 mouse and 10 human odorant receptors, greatly expanding our knowledge of receptor-ligand interactions. We used the resulting interaction profiles to develop a predic-tive model relating physicochemical odorant properties, receptor sequences, and their interactions. Our model can predict a tested receptor's response to a novel odorant (dÕ = 0.63, p <0.001) and a novel receptor's response to a tested odorant (dÕ = 0.22, p <0.001). This provides a large dataset of mam-malian receptor-ligand interactions to constrain the search for rules underlying olfactory transduction and a framework for identifying active ligands for untested receptors in the mouse and human odorant receptor repertoire. Odorants initiate signal transduction in mammalian olfactory sensory neu-rons (OSNs) that leads to increased intracellular Ca 2+ from the opening of cyclic nucleotide gated channels. The Ca 2+ is then cleared preparing the cell for the next stimulation. Several candidates for Ca 2+ clearance include the Na + /Ca 2+ exchanger (NCX), plasma membrane calcium ATPases (PMCAs), and ER calcium pump (SERCA). We show here the significance of the PMCAs in Ca 2+ clearance from OSNs after stimulation and olfactory driven behavior. We know from immunofluorescence and RT-PCR that all 4 isoforms of the PMCAs are expressed in mouse OSNs. It matters very much