Responses Of Olfactory Sensory Neurons Research Articles

Amplification of olfactory transduction currents implements sparse stimulus encoding.

Sensory systems must perform the dual and opposing tasks of being sensitive to weak stimuli while also maintaining information content in dense and variable sensory landscapes. This occurs in the olfactory system, where OSNs are highly sensitive to low concentrations of odors and maintain discriminability in complex odor environments. How olfactory sensory neurons (OSNs) maintain both sensitivity and sparsity is not well understood. Here, we investigated whether the calcium-activated chloride channel, TMEM16B, may support these dual roles in OSNs. We used multiphoton microscopy to image the stimulus-response density of OSNs in the olfactory epithelium. In TMEM16B knockout mice, we found that sensory representations were denser, and the magnitude of OSN responses was increased. Behaviorally, these changes in sensory representations were associated with an increased aversion to the odorant trimethylamine, which switches perceptual valence as its concentration increases, and a decreased efficiency of olfactory-guided navigation. Together, our results indicate that the calcium-activated chloride channel TMEM16B sparsens sensory representations in the peripheral olfactory system and contributes to efficient integrative olfactory-guided behaviors.

The Ca2+-activated Cl- channel TMEM16B shapes the response time course of olfactory sensory neurons.

Mammalian olfactory sensory neurons (OSNs) generate an odorant-induced response by sequentially activating two ion channels, which are in their ciliary membranes. First, a cationic, Ca2+-permeable cyclic nucleotide-gated channel is opened following odorant stimulation via a G protein-coupled transduction cascade and an ensuing rise in cAMP. Second, the increase in ciliary Ca2+ opens the excitatory Ca2+-activated Cl- channel TMEM16B, which carries most of the odorant-induced receptor current. While the role of TMEM16B in amplifying the response has been well established, it is less understood how this secondary ion channel contributes to response kinetics and action potential generation during single as well as repeated stimulation and, on the other hand, which response properties the cyclic nucleotide-gated (CNG) channel determines. We first demonstrate that basic membrane properties such as input resistance, resting potential and voltage-gated currents remained unchanged in OSNs that lack TMEM16B. The CNG channel predominantly determines the response delay and adaptation during odorant exposure, while the absence of the Cl- channels shortens both the time the response requires to reach its maximum and the time to terminate after odorant stimulation. This faster response termination in Tmem16b knockout OSNs allows them, somewhat counterintuitively despite the large reduction in receptor current, to fire action potentials more reliably when stimulated repeatedly in rapid succession, a phenomenon that occurs both in isolated OSNs and in OSNs within epithelial slices. Thus, while the two olfactory ion channels act in concert to generate the overall response, each one controls specific aspects of the odorant-induced response. KEY POINTS: Mammalian olfactory sensory neurons (OSNs) generate odorant-induced responses by activating two ion channels sequentially in their ciliary membranes: a Na+, Ca2⁺-permeable cyclic nucleotide-gated (CNG) channel and the Ca2⁺-activated Cl⁻ channel TMEM16B. The CNG channel controls response delay and adaptation during odorant exposure, while TMEM16B amplifies the response and influences the time required for the response to reach its peak and terminate. OSNs lacking TMEM16B display faster response termination, allowing them to fire action potentials more reliably during rapid repeated stimulation. The CNG and TMEM16B channels have distinct and complementary roles in shaping the kinetics and reliability of odorant-induced responses in OSNs.

Distinct information conveyed to the olfactory bulb by feedforward input from the nose and feedback from the cortex

Sensory systems are organized hierarchically, but feedback projections frequently disrupt this order. In the olfactory bulb (OB), cortical feedback projections numerically match sensory inputs. To unravel information carried by these two streams, we imaged the activity of olfactory sensory neurons (OSNs) and cortical axons in the mouse OB using calcium indicators, multiphoton microscopy, and diverse olfactory stimuli. Here, we show that odorant mixtures of increasing complexity evoke progressively denser OSN activity, yet cortical feedback activity is of similar sparsity for all stimuli. Also, representations of complex mixtures are similar in OSNs but are decorrelated in cortical axons. While OSN responses to increasing odorant concentrations exhibit a sigmoidal relationship, cortical axonal responses are complex and nonmonotonic, which can be explained by a model with activity-dependent feedback inhibition in the cortex. Our study indicates that early-stage olfactory circuits have access to local feedforward signals and global, efficiently formatted information about odor scenes through cortical feedback.

Calcium imaging of adult olfactory epithelium reveals amines as important odor class in fish.

The odor space of aquatic organisms is by necessity quite different from that of air-breathing animals. The recognized odor classes in teleost fish include amino acids, bile acids, reproductive hormones, nucleotides, and a limited number of polyamines. Conversely, a significant portion of the fish olfactory receptor repertoire is composed of trace amine-associated receptors, generally assumed to be responsible for detecting amines. Zebrafish possess over one hundred of these receptors, but the responses of olfactory sensory neurons to amines have not been known so far. Here we examined odor responses of zebrafish olfactory epithelial explants at the cellular level, employing calcium imaging. We report that amines elicit strong responses in olfactory sensory neurons, with a time course characteristically different from that of ATP-responsive (basal) cells. A quantitative analysis of the laminar height distribution shows amine-responsive cells undistinguishable from ciliated neurons positive for olfactory marker protein. This distribution is significantly different from those measured for microvillous neurons positive for transient receptor potential channel 2 and basal cells positive for proliferating cell nuclear antigen. Our results suggest amines as an important odor class for teleost fish.

Functional Interaction Between Drosophila Olfactory Sensory Neurons and Their Support Cells.

Insects detect volatile chemicals using antennae, which house a vast variety of olfactory sensory neurons (OSNs) that innervate hair-like structures called sensilla where odor detection takes place. In addition to OSNs, the antenna also hosts various support cell types. These include the triad of trichogen, tormogen, and thecogen support cells that lie adjacent to their respective OSNs. The arrangement of OSN supporting cells occurs stereotypically for all sensilla and is widely conserved in evolution. While insect chemosensory neurons have received considerable attention, little is known about the functional significance of the cells that support them. For instance, it remains unknown whether support cells play an active role in odor detection, or only passively contribute to homeostasis, e.g., by maintaining sensillum lymph composition. To investigate the functional interaction between OSNs and support cells, we used optical and electrophysiological approaches in Drosophila. First, we characterized the distribution of various supporting cells using genetic markers. By means of an ex vivo antennal preparation and genetically-encoded Ca2+ and K+ indicators, we then studied the activation of these auxiliary cells during odor presentation in adult flies. We observed acute responses and distinct differences in Ca2+ and K+ fluxes between support cell types. Finally, we observed alterations in OSN responses upon thecogen cell ablation in mature adults. Upon inducible ablation of thecogen cells, we notice a gain in mechanical responsiveness to mechanical stimulations during single-sensillum recording, but a lack of change to the neuronal resting activity. Taken together, these results demonstrate that support cells play a more active and responsive role during odor processing than previously thought. Our observations thus reveal that support cells functionally interact with OSNs and may be important for the extraordinary ability of insect olfactory systems to dynamically and sensitively discriminate between odors in the turbulent sensory landscape of insect flight.

Response of Olfactory Sensory Neurons to Mercury Ions in Zebrafish: An Immunohistochemical Study.

Olfactory sensory neurons (OSNs) of fish belong to three main types: ciliated olfactory sensory neurons (cOSNs), microvillous olfactory sensory neurons (mOSNs), and crypt cells. Mercury is a toxic metal harmful for olfaction. We exposed the olfactory epithelium of zebrafish to three sublethal Hg2+ concentrations. Molecular markers specific for the different types of OSNs were immunohistochemically detected. Image analysis of treated sections enabled counting of marked cells and measurement of staining optical density indicative of the response of OSNs to Hg2+ exposure. The three types of OSNs reacted to mercury in a different way. Image analysis revealed that mOSNs are more susceptible to Hg2+ exposure than cOSNs and crypt cell density decreases. Moreover, while the ratio between sensory/nonsensory epithelium areas is unchanged, epithelium thickness drops, and dividing cells increase in the basal layer of the olfactory epithelium. Cell death but also reduction of apical processes and marker expression could account for changes in OSN immunostaining. Also, the differential results between dorsal and ventral halves of the olfactory rosette could derive from different water flows inside the olfactory chamber or different subpopulations in OSNs.

A Role for STOML3 in Olfactory Sensory Transduction.

Stomatin-like protein-3 (STOML3) is an integral membrane protein expressed in the cilia of olfactory sensory neurons (OSNs), but its functional role in this cell type has never been addressed. STOML3 is also expressed in dorsal root ganglia neurons, where it has been shown to be required for normal touch sensation. Here, we extended previous results indicating that STOML3 is mainly expressed in the knob and proximal cilia of OSNs. We additionally showed that mice lacking STOML3 have a morphologically normal olfactory epithelium. Because of its presence in the cilia, together with known olfactory transduction components, we hypothesized that STOML3 could be involved in modulating odorant responses in OSNs. To investigate the functional role of STOML3, we performed loose patch recordings from wild-type (WT) and Stoml3 knock-out (KO) OSNs. We found that spontaneous mean firing activity was lower with additional shift in interspike intervals (ISIs) distributions in Stoml3 KOs compared with WT neurons. Moreover, the firing activity in response to stimuli was reduced both in spike number and duration in neurons lacking STOML3 compared with WT neurons. Control experiments suggested that the primary deficit in neurons lacking STOML3 was at the level of transduction and not at the level of action potential generation. We conclude that STOML3 has a physiological role in olfaction, being required for normal sensory encoding by OSNs.

Calmodulin regulates the olfactory performance in Drosophila melanogaster

Insect odorant receptors (ORs) detect volatile chemical cues with high sensitivity. These ORs operate as ligand-gated ion channels and are formed by heptahelical OrX and Orco (co-receptor) proteins. A highly conserved calmodulin (CaM) binding site (CBS) 336SAIKYWVER344 within the second intracellular loop of Drosophila melanogaster Orco constitutes a target for regulating OR performance. Here we asked how a point mutation K339N in this CBS affects the olfactory performance of Drosophila melanogaster. We first asked how this mutation would affect the odor responses of olfactory sensory neurons (OSNs). Using Ca2+ imaging in an ex-vivo antenna preparation, we activated all OR (OrX/Orco) expressing neurons using the synthetic agonist VUAA1. In a next attempt, we restricted the OR spectrum to Or22a expressing neurons (Or22a/Orco) and stimulated these OSNs with the ligand ethyl hexanoate. In both approaches, we found that flies carrying the K339N point mutation in Orco display a reduced olfactory response. We also found that the mutation abolishes the capability of OSNs to sensitize by repeated weak odor stimuli. Next, we asked whether OrcoK339N might affect the odor localization performance. Using a wind tunnel bioassay, we found that odor localization in flies carrying the OrcoK339N mutation was severely diminished.

Aldehyde-specific responses of olfactory sensory neurons in the praying mantis

Although praying mantises rely mainly on vision for predatory behaviours, olfaction also plays a critical role in feeding and mating behaviours. However, the receptive processes underlying olfactory signals remain unclear. Here, we identified olfactory sensory neurons (OSNs) that are highly tuned to detect aldehydes in the mantis Tenodera aridifolia. In extracellular recordings from OSNs in basiconic sensilla on the antennae, we observed three different spike shapes, indicating that at least three OSNs are housed in a single basiconic sensillum. Unexpectedly, one of the three OSNs exhibited strong excitatory responses to a set of aldehydes. Based on the similarities of the response spectra to 15 different aldehydes, the aldehyde-specific OSNs were classified into three classes: B, S, and M. Class B broadly responded to most aldehydes used as stimulants; class S responded to short-chain aldehydes (C3–C7); and class M responded to middle-length chain aldehydes (C6–C9). Thus, aldehyde molecules can be finely discriminated based on the activity patterns of a population of OSNs. Because many insects emit aldehydes for pheromonal communication, mantises might use aldehydes as olfactory cues for locating prey habitat.

The cyclic AMP signaling pathway in the rodent main olfactory system.

Odor perception begins with the detection of odorant molecules by the main olfactory epithelium located in the nasal cavity. Odorant molecules bind to and activate a large family of G-protein-coupled odorant receptors and trigger a cAMP-mediated transduction cascade that converts the chemical stimulus into an electrical signal transmitted to the brain. Morever, odorant receptors and cAMP signaling plays a relevant role in olfactory sensory neuron development and axonal targeting to the olfactory bulb. This review will first explore the physiological response of olfactory sensory neurons to odorants and then analyze the different components of cAMP signaling and their different roles in odorant detection and olfactory sensory neuron development.

Heterogeneity of Antiviral Responses in the Upper Respiratory Tract Mediates Differential Non-lytic Clearance of Influenza Viruses

Influenza viruses initiate infection in the upper respiratory tract (URT), but early viral tropism and the importance of cell-type-specific antiviral responses in this tissue remain incompletely understood. By infecting transgenic lox-stop-lox reporter mice with a Cre-recombinase-expressing influenza B virus, we identify olfactory sensory neurons (OSNs) as a major viral cell target in the URT. These cells become infected, then eliminate the virus and survive in the host post-resolution of infection. OSN responses to infection are characterized by a strong induction of interferon-stimulated genes and more rapid clearance of viral protein relative to other cells in the epithelium. We speculate that this cell-type-specific response likely serves to protect the central nervous system from infection. More broadly, these results highlight the importance of evaluating antiviral responses across different cell types, even those within the same tissue, to more fully understand the mechanisms of viral disease.

Widespread Inhibition, Antagonism, and Synergy in Mouse Olfactory Sensory Neurons In Vivo

Sensory information is selectively or non-selectively enhanced and inhibited in the brain, but it remains unclear whether and how this occurs at the most peripheral level. Using invivo calcium imaging of mouse olfactory bulb and olfactory epithelium in wild-type and mutant animals, we show that odors produce not only excitatory but also inhibitory responses in olfactory sensory neurons (OSNs). Heterologous assays indicate that odorants can act as agonists to some but inverse agonists to other odorant receptors. We also demonstrate that responses to odor mixtures are extensively suppressed or enhanced in OSNs. When high concentrations of odors are mixed, widespread antagonism suppresses the overall response amplitudes and density. In contrast, a mixture of low concentrations of odors often produces synergistic effects and boosts the faint odor inputs. Thus, odor responses are extensively tuned by inhibition, antagonism, and synergy at the most peripheral level, contributing to robust sensory representations.

Odorant Receptor Inhibition Is Fundamental to Odor Encoding

Most natural odors are complex mixtures of 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 detection layer of the olfactory system. Here, we investigated its prevalence, the degree to which it disrupts OR ensemble activity, and its conservation across phylogenetically 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 exhibiting specific response profiles to a diagnostic panel of odorants and identified three paralogous receptors-Olfr740, Olfr741, and Olfr743-which, when tested invitro, recapitulated OSN responses. We screened ten ORs from the Olfr740 gene family with ∼800 perfumery-related odorants spanning a range of chemical scaffolds and functional groups. Over half of these compounds (430) antagonized at least one of the ten ORs. OR activity fitted a mathematical model of competitive receptor binding and suggests normalization of OSN ensemble responses to odorant mixtures is the rule rather than the exception. In summary, we observed OR antagonism occurred 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.

Rainbow trout (Oncorhynchus mykiss) chemosensory detection of and reactions to copper nanoparticles and copper ions

Copper is known to interfere with fish olfaction. Although the chemosensory detection and olfactory toxicity of copper ions (Cu2+) has been heavily studied in fish, the olfactory-driven detection of copper nanoparticles (CuNPs)—a rapidly emerging contaminant to aquatic systems—remains largely unknown. This study aimed to investigate the olfactory response of rainbow trout to equitoxic concentrations of CuNPs or Cu2+ using electro-olfactography (EOG, a neurophysiological technique) and olfactory-mediated behavioural assay. In the first experiment, the concentration of contaminants known to impair olfaction by 20% over 24 h (EOG-based 24-h IC20s of 220 and 3.5 μg/L for CuNPs and Cu2+, respectively) were tested as olfactory stimuli using both neurophysiological and behavioural assays. In the second experiment, to determine whether the presence of CuNPs or Cu2+ can affect the ability of fish to perceive a social cue (taurocholic acid (TCA)), fish were acutely exposed to one form of Cu-contaminants (approximately 15 min). Following exposure, olfactory sensitivity was measured by EOG and olfactory-mediated behaviour within a choice maze was recorded in the presence of TCA. Results of neurophysiological and behavioural experiments demonstrate that rainbow trout can detect and avoid the IC20 of CuNPs. The IC20 of Cu2+ was below the olfactory detection threshold of rainbow trout, as such, fish did not avoid Cu2+. The high sensitivity of behavioural endpoints revealed a lack of aversion response to TCA in CuNP-exposed fish, despite this change not being present utilizing EOG. The reduced response to TCA during the brief exposure to CuNPs may be a result of either olfactory fatigue or blockage of olfactory sensory neurons (OSNs) by CuNPs. The observed behavioural interference caused by CuNP exposure may indicate that CuNPs have the ability to interfere with other behaviours potentially affecting fitness and survival. Our findings also revealed the differential response of OSNs to CuNPs and Cu2+.

Tests of the chromatographic theory of olfaction with highly soluble odors: a combined electro-olfactogram and computational fluid dynamics study in the mouse.

ABSTRACTThe idea that the vertebrate nasal cavity operates like a gas chromatograph to separate and discriminate odors, referred to herein as the ‘chromatographic theory’ (CT), has a long and interesting history. Though the last decade has seen renewed interest in the notion, its validity remains in question. Here we examine a necessary condition of the theory: a correlation between nasal odor deposition patterns based on mucus solubility and the distribution of olfactory sensory neuron odotypes. Our recent work in the mouse failed to find such a relationship even across large sorption gradients within the olfactory epithelium (OE). However, these studies did not test extremely soluble odorants or low odor concentrations, factors that could explain our inability to find supporting evidence for the CT. The current study combined computational fluid dynamics (CFD) simulations of odor sorption patterns and electro-olfactogram (EOG) measurements of olfactory sensory neuron responses. The odorants tested were at the extremes of mucus solubility and at a range of concentrations. Results showed no relationship between local odor sorption patterns and EOG response maps. Together, results again failed to support a necessary condition of the CT casting further doubt on the viability of this classical odor coding mechanism.

Host Preference and Olfaction in Drosophila mojavensis.

Many organisms live in complex environments that vary geographically in resource availability. This environmental heterogeneity can lead to changes within species in their phenotypic traits. For example, in many herbivorous insects, variation in host plant availability has been shown to influence insect host preference behavior. This behavior can be mediated in part through the insect olfactory system and the odor-evoked responses of olfactory sensory neurons (OSNs), which are in turn mediated by their corresponding odorant receptor genes. The desert dwelling fly Drosophila mojavensis is a model species for understanding the mechanisms underlying host preference in a heterogeneous environment. Depending on geographic region, one to multiple host plant species are available. Here, we conducted electrophysiological studies and found variation in responses of ORNs to host plant volatiles both within and between 2 populations-particularly to the odorant 4-methylphenol. Flies from select localities within each population were found to lack a response to 4-methylphenol. Experiments then assessed the extent to which these electrophysiological differences were associated with differences in several odor-mediated behavioral responses. No association between the presence/absence of these odor-evoked responses and short range olfactory behavior or oviposition behavior was observed. However, differences in odor-induced feeding behavior in response to 4-methylphenol were found. Localities that exhibit an odor-evoked response to the odorant had increased feeding behavior in the presence of the odorant. This study sets the stage for future work examining the functional genetics underlying variation in odor perception.

Evolution of coeloconic sensilla in the peripheral olfactory system of Drosophila mojavensis

Populations inhabiting habitats with different environmental conditions, such as climate, resource availability, predation, competition, can undergo selection for traits that are adaptive in one habitat and not the other, leading to divergence between populations. Changes in the olfactory systems of insects that rely on different host plants, for example, can occur in response to differences in sensory stimuli between habitats. In this study, we investigate the evolution of host preference by characterizing the coeloconic sensilla in Drosophila mojavensis, a species that breeds on different necrotic cacti across its geographic range. These cactus species differ in the volatile chemicals they emit, a primary sensory cue for host plant discrimination. Analysis of odor-evoked responses identified four coeloconic sensilla that were qualitatively similar to those of Drosophila melanogaster, but varied in the breadth and strength of their olfactory sensory neuron responses to some acids and amines. Variation in responses to certain odorants among D. mojavensis populations was also observed. Compared to D. melanogaster, there was a lack of sensitivity of antennal coeloconic type 3 (ac3) sensilla to primary ligands of OR35a across all populations. Consistent with this result was a lack of detectable Or35a gene expression. Using a comparative approach, we then examined odor specificity of ac3 sensilla for seven additional Drosophila species, and found that OR35a-like sensitivity may be limited to the melanogaster subgroup. The variation in specificity that was observed among species is not clearly attributable to the degree of ecological specialization, nor to the ecological niche.

Olfactory Sensory Neurons of the Asian Longhorned Beetle, Anoplophora glabripennis, Specifically Responsive to its two Aggregation-Sex Pheromone Components.

We performed single-sensillum recordings from male and female antennae of the Asian longhorned beetle, Anoplophora glabripennis, that included as stimuli the two components of this species' aggregation-sex pheromone in addition to various general odorants. We compared the aggregation-sex-pheromone-component responses of olfactory sensory neurons (OSNs) to those of OSNs that responded to a variety of plant-related odorants. In the smooth-tipped, tapered, trichoid sensilla on the most distal antennal flagellomeres nos. 10 or 11 of both males and females, we found OSNs with high-amplitude action potentials that were tuned to the aldehyde and alcohol pheromone components and that did not respond to various plant-related volatiles. Because this OSN type responded to both the alcohol and aldehyde components it cannot be considered to be specifically tuned to either component. These large-spiking OSNs were co-compartmentalized in these sensilla with a second, smaller-spiking OSN responding to plant-related volatiles such as geraniol, citronellal, limonene, 1-octanol, nerol and citral. The large-spiking OSNs thus appear to be a type that will be involved in aggregation-sex pheromone pathways targeting a specific glomerulus in the antennal lobe and in generating pheromone-related behavioral responses in A. glabripennis. In other sensilla located in these distal antennal flagellomeres as well as those located more proximally, i.e., mid-length along the antenna on flagellomere nos. 4-7, we found OSNs in blunt-tipped basiconic sensilla that were responsive to other plant-related volatiles, especially the terpenoids, (E,E)-alpha farnesene, (E)-β-farnesene, β-caryophyllene, and eugenol. Some of these terpenoids have been implicated in improving attraction to pheromone-baited traps. Some of these same OSNs responded additionally to either of the two sex pheromone components, but because these OSNs also responded to some of the above plant volatiles as shown by cross-adaptation experiments, these OSNs will not be the types that convey sex-pheromone-specific information to the antennal lobe.

Stable olfactory sensory neuron invivo physiology during normalaging.

Normal aging is associated with a number of smell impairments that are paralleled by age-dependent changes in the peripheral olfactory system, including decreases in olfactory sensory neurons (OSNs) and in the regenerative capacity of the epithelium. Thus, an age-dependent degradation of sensory input to the brain is one proposed mechanism for the loss of olfactory function in older populations. Here, we tested this hypothesis by performing invivo optical neurophysiology in 6-, 12-, 18-, and 24-month-old mice. We visualized odor-evoked neurotransmitter release from populations of OSNs into olfactory bulb glomeruli, and found that these sensory inputs are actually quite stable during normal aging. Specifically, the magnitude and number of odor-evoked glomerular responses were comparable across all ages, and there was no effect of age on the sensitivity of OSN responses to odors or on the neural discriminability of different sensory maps. These results suggest that the brain's olfactory bulbs do not receive deteriorated input during aging and that local bulbar circuitry might adapt to maintain stable nerve input.

Odor-Induced Electrical and Calcium Signals from Olfactory Sensory Neurons In Situ.

Electrophysiological recording and optical imaging enable the characterization of membrane and odorant response properties of olfactory sensory neurons (OSNs) in the nasal neuroepithelium. Here we describe a method to record the responses of mammalian OSNs to odorant stimulations in an ex vivo preparation of intact olfactory epithelium. The responses of individual OSNs with defined odorant receptor types can be monitored via patch-clamp recording or calcium imaging.