Odorant Receptor Inhibition is Fundamental to Odor Encoding

Abstract

Most natural odors are complex mixtures of many volatile components, competing to bind odorant receptors (ORs) expressed in olfactory sensory neurons (OSNs) of the nose. To date surprisingly little is known about how OR antagonism shapes neuronal representations in the periphery of the olfactory system. Here, we investigated its prevalence, the degree to which it disrupts OR ensemble activity, and its conservation across related ORs. Calcium imaging microscopy of dissociated OSNs revealed significant inhibition, often complete attenuation, of responses to indole, a commonly occurring volatile associated with both floral and fecal odors, by a set of 36 tested odorants. To confirm an OR mechanism for the observed inhibition, we performed single-cell transcriptomics on OSNs that exhibited specific response profiles to a diagnostic panel of odorants and identified the receptor Olfr743 which, when tested in vitro, recapitulated ex vivo responses. We screened ten ORs from the Olfr743 clade with 800 perfumery-related odorants spanning a range of chemical scaffolds and functional groups, over half of which (430) antagonized at least one of the ten ORs. Furthermore, OR activity outcomes were divergent rather than redundant, even for the most closely related paralogs. OR activity fitted a mathematical model of competitive receptor binding and suggests that normalization of OSN ensemble responses to odorant mixtures is the rule rather than the exception. In summary, we observed OR antagonism, inverse agonism and partial agonism occurring frequently and in a combinatorial manner. Thus, extensive receptor-mediated computation of mixture information appears to occur in the olfactory epithelium prior to transmission of odor information to the olfactory bulb. 2 3 heterologous expression systems and the lack of a protein structure [41] have impeded systematic exploration of the full range of possible receptor-ligand interactions for ORs. As such, it is not well understood to what extent classical GPCR interactions, like antagonism, partial agonism and inverse agonism shape odor representations before they are propagated to the olfactory bulb.This lack of information on the prevalence of non-agonist interactions, and on correlations between ORs severely limits our ability to understand fundamental aspects of mixture encoding by the olfactory system. For example, infrequent, but strong antagonism would simply prevent the activation of a limited number of ORs in the presence of an antagonist. Alternatively, widespread antagonism could lead to extensive normalization of inputs [29] and decoupling of similarity between mixture compositions and their resulting OSN activity ensembles. With the rise of mathematical models of olfaction demonstrating wide varieties of possible encoding strategies, it has become important to provide data which can help test parameter values and rule out certain classes of model.In this study, we investigated the prevalence of antagonism, quantified its ability to disrupt OSN activity, and examined its cons...

Antagonistic odor interactions in olfactory sensory neurons are widespread in freely breathing mice

“…Beyond our theoretical observations, there are other more recent experimental studies (in bioRxiv preprints) that report related effects (Inagaki et al, 2019;Pfister et al, 2019;Xu et al, 2019). Our work is unique in the following regard:…”

“…Recent theoretical work has shown that a variety of non-linear interactions among multiple ligands at the same receptor can readily arise with a simple two-step model of receptor activation (Reddy et al, 2018). Experimental work in vitro in OSNs has suggested the existence of non-linear interactions, especially antagonism (Firestein and Shepherd, 1992;Kurahashi et al, 1994;Mathis et al, 2016;Rospars et al, 2008Rospars et al, , 2000Singh et al, 2019) (also bioRxiv preprints; Inagaki et al, 2019;Pfister et al, 2019;Xu et al, 2019). However, the prevalence of these interactions, especially in living animals within the constraints of natural sniffing dynamics, has not been explored.…”

Odor mixtures: A chord with silent notes

“…All these lines of evidence support competitive antagonism as the most dominant form of modulation. In addition to partial agonism and synergetic and/or allosteric modulation we and others have reported, inverse antagonism in which high baseline activity in certain OSNs could be reduced by odorants (de March et al, 2020;Inagaki et al, 2020;Pfister et al, 2020;Zak et al, 2020).…”

“…Consistent with our observations with SCAPE microscopy, several recent studies also demonstrated widespread modulation among odorants at the receptor level using various other techniques. These approaches include in vivo studies of dorsal OSNs (and glomeruli) in GCaMP3 mice with two-photon microscopy (Inagaki et al, 2019;Zak et al, 2020), identification of odorant receptors displaying odorant modulatory effects first in freely behaving mice (de March et al, 2020;McClintock et al, 2020) or in dissociated OSNs (Pfister et al, 2020) and then verification of the modulation effects in some of these odorant receptors with heterologous expression assays, as well as theoretical modeling predictions (Marasco et al, 2016;Reddy et al, 2018;Singh et al, 2019).…”

Finding the Brain in the Nose

“…In the context of mixtures, monomolecular odorants can act as agonists, antagonists, and partial agonists (Oka et al 2004, Rospars et al 2008, Takeuchi et al 2009) (Figure 2a). Indeed, most OR-ligand interactions are likely influenced by other monomolecular odors present in natural mixtures (Pfister et al 2019, Xu et al 2020. Furthermore, population-level OSN responses to natural stimuli are at least partially normalized in magnitude by competitive interactions between mixture constituents (Reddy et al 2018).…”