Concentration-Dependent Recruitment of Mammalian Odorant Receptors

Abstract

A fundamental challenge in studying principles of organization used by the olfactory system to encode odor concentration information has been identifying comprehensive sets of activated odorant receptors (ORs) across a broad concentration range inside freely behaving animals. In mammals, this has recently become feasible with high-throughput sequencing-based methods that identify populations of activated ORs in vivo. In this study, we characterized the mouse OR repertoires activated by the two odorants, acetophenone and 2,5-dihydro-2,4,5trimethylthiazoline, from 0.01% to 100% (v/v) as starting concentrations using phosphorylated ribosomal protein S6 capture followed by RNA-Seq. We found Olfr923 to be one of the most sensitive ORs that is enriched by acetophenone. Using a mouse line that genetically labels Olfr923-positive axons, we provided evidence that acetophenone activates the Olfr923 glomeruli in the olfactory bulb. Through molecular dynamics stimulations, we identified amino acid residues in the Olfr923 binding cavity that facilitate acetophenone binding. This study sheds light on the active process by which unique OR repertoires may collectively facilitate the discrimination of odorant concentrations. 4 Significance Statement The ability of animals to discriminate odors over a range of odor concentrations while recognizing concentration-invariant odor identity presents an encoding challenge for the olfactory system. To further our understanding on how animals sense odors at different concentrations, it is important to describe how odor concentration information is represented at the receptor level. Here, we establish a sensitive in vivo approach to screen populations of odorant receptors enriched in the odor-activated sensory neurons in mice. We identified comprehensive lists of enriched odorant receptors against a 10,000-fold concentration range for two odorants. Describing the concentration-dependent activation for unique populations of odorant receptors is fundamental for future studies in determining how individual odorant receptors contribute to olfactory sensitivity and odor intensity coding.

Semiochemical responsive olfactory sensory neurons are sexually dimorphic and plastic

“…In situ hybridization probing for the expression of Olfr910/912 and immunostaining for pS6 showed OSNs expressing Olfr910/912 to display increasingly intense pS6 immunostaining following exposure to SBT in a concentration-dependent manner (0.01% SBT p < 0.05; 0.1% SBT p < 0.0001; 1% SBT p < 0.0001, one-way ANOVA with Dunnett’s multiple comparisons test correction). Further, in accordance with our previous findings, exposure to acetophenone did not lead to significant pS6 induction in OSNs expressing Olfr910/912 (Figure 4D-E) 11, 22 . Similarly, OSNs expressing Olfr1295 , identified by in situ hybridization, displayed increasingly intense pS6 immunostaining following exposure to MTMT in a concentration-dependent manner (100 μM MTMT p < 0.01; 10 mM MTMT p < 0.0001, one-way ANOVA with Dunnett’s multiple comparisons test correction).…”

Semiochemical responsive olfactory sensory neurons are sexually dimorphic and plastic

“…Similarly, OSNs expressing Olfr1295 , identified by in situ mRNA hybridization, displayed increasingly intense pS6 immunostaining following exposure to MTMT, again, in a concentration-dependent manner (100 μM MTMT p < 0.0001, 10 mM MTMT p < 0.0001, one-way ANOVA with Dunnett’s multiple comparisons test correction; Figure 4G , Figure 4—figure supplement 3C ). OSNs expressing Olfr910 , Olfr912 , or Olfr1295 also did not display significant pS6 signal intensity differences following exposure to acetophenone compared to control conditions (p > 0.05, one-way ANOVA with Dunnett’s multiple comparisons test correction; Figure 4E–G ), consistent with a lack of enrichment of Olfr910 , Olfr912 , and Olfr1295 by pS6-IP-Seq following exposure to 1% (v/v) acetophenone ( Jiang et al, 2015 ; Hu et al, 2020 ).…”

“…To test this hypothesis, we searched the literature for known sex-specific or sex-enriched odors and leveraged phosphorylated S6 ribosomal subunit capture (pS6-IP) as a means to determine the molecular identities of the OSNs activated by monomolecular odorants in vivo. Immunoprecipitation of phosphorylated ribosomes from activated neurons, followed by associated mRNA profiling by RNA-Seq (pS6-IP-Seq) and differential expression analysis, enabled us to perform an unbiased identification of the molecular profiles, including ORs expressed, of OSNs activated by specific odorants ( Figure 4A ; Jiang et al, 2015 ; Hu et al, 2020 ).…”

“…Despite the early discoveries of ORs being expressed in highly stereotyped locations, the functional consequences of these zonal expression distributions have remained unclear ( Ressler et al, 1993 ; Vassar et al, 1993 ). It is interesting to note, all of the identified sexually dimorphic populations of OSNs ( Olfr910 , Olfr912 , Olfr1295 , Olfr1437 , and Olfr235 ) were localized to the ventral region of the MOE ( Tan and Xie, 2018 ; Hu et al, 2020 ). These results are consistent with past work showing a robust response in the ventral region of the olfactory bulb to male mouse urine ( Lin et al, 2005 ; Xu et al, 2005 ).…”

The Mammalian Olfactory Bulb Contributes to the Adaptation of Odor Responses: A Second Perceptual Computation Carried Out by the Bulb

“…The ability to identify sensory information embedded in a complex environment requires the brain to be able to filter background information and attend to novel stimuli ( Linster et al, 2007 ; Gottfried, 2010 ). This is a challenging problem in the olfactory system because odor perception is thought to be related to the combination of activated olfactory receptors, yet different receptors can be activated by the same odor at different concentrations and the same receptor can be activated by perceptually different odors ( Duchamp-Viret et al, 1999 ; Wachowiak and Cohen, 2001 ; Hu et al, 2020 ). A new and potentially important odor stimulus can activate some of the same odor receptors as those activated by a background odor.…”