Induction of astrocytic Slc22a3 regulates sensory processing through histone serotonylation.

Neuronal activity drives alterations in gene expression within neurons, yet how it directs transcriptional and epigenomic changes in neighboring astrocytes in functioning circuits is unknown. We found that neuronal activity induces widespread transcriptional up-regulation and down-regulation in astrocytes, highlighted by the identification of Slc22a3 as an activity-inducible astrocyte gene that encodes neuromodulator transporter Slc22a3 and regulates sensory processing in the mouse olfactory bulb. Loss of astrocytic Slc22a3 reduced serotonin levels in astrocytes, leading to alterations in histone serotonylation. Inhibition of histone serotonylation in astrocytes reduced the expression of γ-aminobutyric acid (GABA) biosynthetic genes and GABA release, culminating in olfactory deficits. Our study reveals that neuronal activity orchestrates transcriptional and epigenomic responses in astrocytes while illustrating new mechanisms for how astrocytes process neuromodulatory input to gate neurotransmitter release for sensory processing.

Genetic variation in the human olfactory receptor OR5AN1 associates with the perception of musks.

Humans have significant individual variations in odor perception, derived from their experience or sometimes from differences in the olfactory receptor (OR) gene repertoire. In several cases, the genetic variation of a single OR affects the perception of its cognate odor ligand. Musks are widely used for fragrance and are known to demonstrate specific anosmia. It, however, remains to be elucidated whether the OR polymorphism contributes to individual variations in musk odor perception. Previous studies reported that responses of the human musk receptor OR5AN1 to a variety of musks in vitro correlated well with perceptual sensitivity to those odors in humans and that the mouse ortholog, Olfr1440 (MOR215-1), plays a critical role in muscone perception. Here, we took advantage of genetic variation in OR5AN1 to examine how changes in receptor sensitivity are associated with human musk perception. We investigated the functional differences between OR5AN1 variants in an in vitro assay and measured both perceived intensity and detection threshold in human subjects with different OR5AN1 genotypes. Human subjects homozygous for the more sensitive L289F allele had a lower detection threshold for muscone and found macrocyclic musks to be more intense than subjects homozygous for the reference allele. These results demonstrate that the genetic variation in OR5AN1 contributes to perceptual differences for some musks. In addition, we found that the more functional variant of OR5A1, a receptor involved in ß-ionone perception, is associated with the less functional variant of OR5AN1, suggesting that the perceived intensities of macrocyclic musks and ß-ionone are inversely correlated.

Identification and characterization of olfactory receptor genes and olfactory perception in rapa whelk Rapana venosa (Valenciennes, 1846) during larval settlement and metamorphosis.

The rapa whelk Rapana venosa, an economically important marine fishery resource in China but a major invader all over the world, changes from a phytophagous to a carnivorous form following settlement and metamorphosis. However, the low settlement and metamorphosis rates (<1%) of larvae limit the abundance of R. venosa. This critical step (settlement and metamorphosis) remains poorly characterized but may be related to how larvae perceive the presence of shellfish, their new source of food. Here, we report that larvae may use olfactory perception to sense shellfish. Olfactory receptor (OR) genes are involved in odor sensing in animals. We identified a total of 463 OR genes, which could be grouped into nine clades based on phylogenetic analysis. When assessing the attraction of larvae at different developmental stages to oyster odor, R. venosa showed active settlement and metamorphosis behavior only at the J4 stage (competent larva, 1000-1500 µm shell length) and in the presence of shellfish odor at the same time. Expression of OR gene family members differed between stage 2 (four-spiral whorl stage) and stage 1 (single- to three-spiral whorl stage), indicating significant changes in the olfactory system during larval development. These findings broaden our understanding of olfactory perception, settlement, and metamorphosis in gastropods and can be used to improve R. venosa harvesting, as well as the sustainable development and utilization of this resource.

From musk to body odor: Decoding olfaction through genetic variation.

The olfactory system combines input from multiple receptor types to represent odor information, but there are few explicit examples relating olfactory receptor (OR) activity patterns to odor perception. To uncover these relationships, we performed genome-wide scans on odor-perception phenotypes for ten odors in 1000 Han Chinese and validated results for six of these odors in an ethnically diverse population (n = 364). In both populations, consistent with previous studies, we replicated three previously reported associations (ß-ionone/OR5A1, androstenone/OR7D4, cis-3-hexen-1-ol/OR2J3 LD-band), but not for odors containing aldehydes, suggesting that olfactory phenotype/genotype studies are robust across populations. Two novel associations between an OR and odor perception contribute to our understanding of olfactory coding. First, we found a SNP in OR51B2 that associated with trans-3-methyl-2-hexenoic acid, a key component of human underarm odor. Second, we found two linked SNPs associated with the musk Galaxolide in a novel musk receptor, OR4D6, which is also the first human OR shown to drive specific anosmia to a musk compound. We noticed that SNPs detected for odor intensity were enriched with amino acid substitutions, implying functional changes of odor receptors. Furthermore, we also found that the derived alleles of the SNPs tend to be associated with reduced odor intensity, supporting the hypothesis that the primate olfactory gene repertoire has degenerated over time. This study provides information about coding for human body odor, and gives us insight into broader mechanisms of olfactory coding, such as how differential OR activation can converge on a similar percept.

BarH1 regulates odorant-binding proteins expression and olfactory perception of Monochamus alternatus Hope.

Insect odorant-binding proteins (OBPs) are a class of small soluble proteins that can be found in various tissues wherein binding and transport of small molecules are required. Thus, OBPs are not only involved in typical olfactory function by specific activities with odorants but also participate in other physiological processes in non-chemosensory tissues. To better understand the complex biological functions of OBPs, it is necessary to study the transcriptional regulation of their expression patterns. In this paper, an apparent gradient expression pattern of Obp19, that was highly and specifically expressed in antennae and played an essential role in the detection of camphene, was defined in the antennae of the Japanese pine sawyer. Further, the transcription factor BarH1, that also presented gradient expression pattern in antennae, was found to regulate expression of Obp19 directly through binding to its upstream DNA sequence. The condition of BarH1 gene silence, the gene expression levels of Obp19 significantly decreased. At the same time, additional olfactory genes also were regulated and thus influence camphene reception. These findings provide us an opportunity to incorporate Obps in the gene regulatory networks of insects, which contribute to a better understanding of the multiplicity and diversity of OBPs and the olfactory mediated behaviors.

Sensory processing during sleep in Drosophila melanogaster.

During sleep, most animal species enter a state of reduced consciousness characterized by a marked sensory disconnect. Yet some processing of the external world must remain intact, given that a sleeping animal can be awoken by intense stimuli (for example, a loud noise or a bright light) or by soft but qualitatively salient stimuli (for example, the sound of a baby cooing or hearing one's own name1-3). How does a sleeping brain retain the ability to process the quality of sensory information? Here we present a paradigm to study the functional underpinnings of sensory discrimination during sleep in Drosophila melanogaster. We show that sleeping vinegar flies, like humans, discern the quality of sensory stimuli and are more likely to wake up in response to salient stimuli. We also show that the salience of a stimulus during sleep can be modulated by internal states. We offer a prototypical blueprint detailing a circuit involved in this process and its modulation as evidence that the system can be used to explore the cellular underpinnings of how a sleeping brain experiences the world.

CRISPR/Cas9 mutagenesis abolishes odorant-binding protein BdorOBP56f-2 and impairs the perception of methyl eugenol in Bactrocera dorsalis (Hendel).

Olfaction underpins many insect behaviors, such as foraging, host location, mating, and predator avoidance. In the first step of insect olfaction, odorant-binding proteins (OBPs) bind hydrophobic odorants and transport them to odorant receptors. Methyl eugenol (ME) is a powerful attractant for mature males of the oriental fruit fly Bactrocera dorsalis (Hendel), one of the most destructive fruit pests. The underlying molecular mechanism is unclear, but there is in vitro evidence that BdorOBP56f-2 is involved in ME perception. We used microscale thermophoresis to confirm that BdorOBP56f-2 directly binds ME with strong affinity in vitro. We then used CRISPR/Cas9 to knock out the BdorOBP56f-2 gene, allowing us to establish a homozygous mutant B. dorsalis line. The electroantennogram response and behavioral attraction to ME were significantly reduced in the mutant, providing in vivo evidence that BdorOBP56f-2 is necessary for efficient ME perception. Our results offer insight into the molecular mechanism of ME perception in B. dorsalis and provide a theoretical basis for the functional analysis of other OBPs.

Gene therapy rescues olfactory perception in a clinically relevant ciliopathy model of Bardet-Biedl syndrome.

Bardet-Biedl syndrome (BBS) is a hereditary genetic disorder that results in numerous clinical manifestations including olfactory dysfunction. Of at least 21 BBS-related genes that can carry multiple mutations, a pathogenic mutation, BBS1M390R, is the single most common mutation of clinically diagnosed BBS outcomes. While the deletion of BBS-related genes in mice can cause variable penetrance in different organ systems, the impact of the Bbs1M390R mutation in the olfactory system remains unclear. Using a clinically relevant knock-in mouse model homozygous for Bbs1M390R, we investigated the impact of the mutation on the olfactory system and tested the potential of viral-mediated, wildtype gene replacement therapy to rescue smell loss. The cilia of olfactory sensory neurons (OSNs) in Bbs1M390R/M390R mice were significantly shorter and fewer than those of wild-type mice. Also, both peripheral cellular odor detection and synaptic-dependent activity in the olfactory bulb were significantly decreased in the mutant mice. Furthermore, to gain insight into the degree to which perceptual features are impaired in the mutant mice, we used whole-body plethysmography to quantitatively measure odor-evoked sniffing. The Bbs1M390R/M390R mice showed significantly higher odor detection thresholds (reduced odor sensitivity) compared to wild-type mice; however, their odor discrimination acuity was still well maintained. Importantly, adenoviral expression of Bbs1 in OSNs restored cilia length and re-established both peripheral odorant detection and odor perception. Together, our findings further expand our understanding for the development of gene therapeutic treatment for congenital ciliopathies in the olfactory system.

Optogenetic Manipulation of Olfactory Responses in Transgenic Zebrafish: A Neurobiological and Behavioral Study.

Olfaction is an important neural system for survival and fundamental behaviors such as predator avoidance, food finding, memory formation, reproduction, and social communication. However, the neural circuits and pathways associated with the olfactory system in various behaviors are not fully understood. Recent advances in optogenetics, high-resolution in vivo imaging, and reconstructions of neuronal circuits have created new opportunities to understand such neural circuits. Here, we generated a transgenic zebrafish to manipulate olfactory signal optically, expressing the Channelrhodopsin (ChR2) under the control of the olfactory specific promoter, omp. We observed light-induced neuronal activity of olfactory system in the transgenic fish by examining c-fos expression, and a calcium indicator suggesting that blue light stimulation caused activation of olfactory neurons in a non-invasive manner. To examine whether the photo-activation of olfactory sensory neurons affect behavior of zebrafish larvae, we devised a behavioral choice paradigm and tested how zebrafish larvae choose between two conflicting sensory cues, an aversive odor or the naturally preferred phototaxis. We found that when the conflicting cues (the preferred light and aversive odor) were presented together simultaneously, zebrafish larvae swam away from the aversive odor. However, the transgenic fish with photo-activation were insensitive to the aversive odor and exhibited olfactory desensitization upon optical stimulation of ChR2. These results show that an aversive olfactory stimulus can override phototaxis, and that olfaction is important in decision making in zebrafish. This new transgenic model will be useful for the analysis of olfaction related behaviors and for the dissection of underlying neural circuits.

The olfactory critical period is determined by activity-dependent Sema7A/PlxnC1 signaling within glomeruli.

In mice, early exposure to environmental odors affects social behaviors later in life. A signaling molecule, Semaphorin 7A (Sema7A), is induced in the odor-responding olfactory sensory neurons. Plexin C1 (PlxnC1), a receptor for Sema7A, is expressed in mitral/tufted cells, whose dendrite-localization is restricted to the first week after birth. Sema7A/PlxnC1 signaling promotes post-synaptic events and dendrite selection in mitral/tufted cells, resulting in glomerular enlargement that causes an increase in sensitivity to the experienced odor. Neonatal odor experience also induces positive responses to the imprinted odor. Knockout and rescue experiments indicate that oxytocin in neonates is responsible for imposing positive quality on imprinted memory. In the oxytocin knockout mice, the sensitivity to the imprinted odor increases, but positive responses cannot be promoted, indicating that Sema7A/PlxnC1 signaling and oxytocin separately function. These results give new insights into our understanding of olfactory imprinting during the neonatal critical period.

Effect of the rs2890498 polymorphism of the OBPIIa gene on the human ability to smell single molecules.

Most odors of foods and drinks are mixtures of molecules. By means of the coupled Gas Chromatography-Olfactometry (GC-O) technique, single components of flavor mixtures can be separated, identified and verbally evaluated by subjects. The number of single molecules smelled by subjects during GC-O analysis (i.e., the number of odor-active compounds) was previously found to be linearly correlated with odor Threshold (T) score. Using the "Sniffin' Sticks" test, the same subjects were classified as normosmic or hyposmic. Hydrophobic odorants are captured and transported through the mucus layer by the odorant binding proteins (OBPs), particularly expressed in the olfactory cleft and associated with the olfactory function. In this study, subjects were genotyped for the rs2590498 (A/G) polymorphism of the OBPIIa gene, whose major allele A is associated with a higher olfactory sensitivity as compared to the minor allele G. One-way ANOVA showed a significant effect of the genotype of the OBPIIa locus on the: a) T score; b) number of odor-active compounds smelled; c) intensity perceived when sniffing the complex odor of banana. In conclusion, the threshold olfactory performance, but also the individual ability to smell single molecules, can be attributed, partly at least, to the rs2590498 polymorphism of the OBPIIa gene.

Mutagenesis of the orco odorant receptor co-receptor impairs olfactory function in the malaria vector Anopheles coluzzii.

Mosquitoes rely heavily on their olfactory systems for host seeking, selection of oviposition sites, and avoiding predators and other environmental dangers. Of these behaviors, the preferential selection of a human blood-meal host drives the vectorial capacity of anthropophilic female Anopheles coluzzii mosquitoes. Olfactory receptor neurons (ORNs) are dispersed across several appendages on the head and express an obligate odorant receptor co-receptor (Orco) coupled with a "tuning" odorant receptor (OR) to form heteromeric, odor-gated ion channels in the membrane of these neurons. To examine the mechanistic and functional contributions of Orco/OR complexes to the chemosensory processes of An. coluzzii, we utilized CRISPR/Cas9 gene editing to create a line of homozygous, Orco-knockout, mutant mosquitoes. As expected, orco-/- ORNs across both adult and larval stages of An. coluzzii display significantly lower background activity and lack nearly all odor-evoked responses. In addition, blood-meal-seeking, adult female, orco-/- mutant mosquitoes exhibit severely reduced attraction to human- and non-human-derived odors while gravid females are significantly less responsive to established oviposition attractants. These results reinforce observations in other insects that Orco is crucial in maintaining the activity of ORNs. In that light, it significantly influences a range of olfactory-driven behaviors central to the anthropophilic host preference that is critical to the vectorial capacity of An. coluzzii as a primary vector for human malaria.

Sequence Variants in TAAR5 and Other Loci Affect Human Odor Perception and Naming.

Olfactory receptor (OR) genes in humans form a special class characterized by unusually high DNA sequence diversity, which should give rise to differences in perception and behavior. In the largest genome-wide association study to date based on olfactory testing, we investigated odor perception and naming with smell tasks performed by 9,122 Icelanders, with replication in a separate sample of 2,204 individuals. We discovered an association between a low-frequency missense variant in TAAR5 and reduced intensity rating of fish odor containing trimethylamine (p.Ser95Pro, pcombined = 5.6 × 10-15). We demonstrate that TAAR5 genotype affects aversion to fish odor, reflected by linguistic descriptions of the odor and pleasantness ratings. We also discovered common sequence variants in two canonical olfactory receptor loci that associate with increased intensity and naming of licorice odor (trans-anethole: lead variant p.Lys233Asn in OR6C70, pcombined = 8.8 × 10-16 and pcombined = 1.4 × 10-9) and enhanced naming of cinnamon (trans-cinnamaldehyde; intergenic variant rs317787-T, pcombined = 5.0 × 10-17). Together, our results show that TAAR5 genotype variation influences human odor responses and highlight that sequence diversity in canonical OR genes can lead to enhanced olfactory ability, in contrast to the view that greater tolerance for mutations in the human OR repertoire leads to diminished function.

Manipulating synthetic optogenetic odors reveals the coding logic of olfactory perception.

How does neural activity generate perception? Finding the combinations of spatial or temporal activity features (such as neuron identity or latency) that are consequential for perception remains challenging. We trained mice to recognize synthetic odors constructed from parametrically defined patterns of optogenetic activation, then measured perceptual changes during extensive and controlled perturbations across spatiotemporal dimensions. We modeled recognition as the matching of patterns to learned templates. The templates that best predicted recognition were sequences of spatially identified units, ordered by latencies relative to each other (with minimal effects of sniff). Within templates, individual units contributed additively, with larger contributions from earlier-activated units. Our synthetic approach reveals the fundamental logic of the olfactory code and provides a general framework for testing links between sensory activity and perception.

Interhemispheric asymmetry of c-Fos expression in glomeruli and the olfactory tubercle following repeated odor stimulation.

Odor adaptation allows the olfactory system to regulate sensitivity to different stimulus intensities, which is essential for preventing saturation of the cell-transducing machinery and maintaining high sensitivity to persistent and repetitive odor stimuli. Although many studies have investigated the structure and mechanisms of the mammalian olfactory system that responds to chemical sensation, few studies have considered differences in neuronal activation that depend on the manner in which the olfactory system is exposed to odorants, or examined activity patterns of olfactory-related regions in the brain under different odor exposure conditions. To address these questions, we designed three different odor exposure conditions that mimicked diverse odor environments and analyzed c-Fos-expressing cells (c-Fos+ cells) in the odor columns of the olfactory bulb (OB). We then measured differences in the proportions of c-Fos-expressing cell types depending on the odor exposure condition. Surprisingly, under the specific odor condition in which the olfactory system was repeatedly exposed to the odorant for 1 min at 5-min intervals, one of the lateral odor columns and the ipsilateral hemisphere of the olfactory tubercle had more c-Fos+ cells than the other three odor columns and the contralateral hemisphere of the olfactory tubercle. However, this interhemispheric asymmetry of c-Fos expression was not observed in the anterior piriform cortex. To confirm whether the anterior olfactory nucleus pars externa (AONpE), which connects the left and right OB, contributes to this asymmetry, AONpE-lesioned mice were analyzed under the specific odor exposure condition. Asymmetric c-Fos expression was not observed in the OB or the olfactory tubercle. These data indicate that the c-Fos expression patterns of the olfactory-related regions in the brain are influenced by the odor exposure condition and that asymmetric c-Fos expression in these regions was observed under a specific odor exposure condition due to synaptic linkage via the AONpE.

Estrés oxidativo y epigenética en la obesidad, el síndrome metabólico y la percepción olfativa / Oxidative stress and epigenetics in obesity, metabolic syndrome and olfactory perception

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Pheromonal activities of the bombykol isomer, (10E,12E)-10,12-hexadecadien-1-ol, in the pheromone gland of the silkmoth Bombyx mori.

Bombykol (EZ) is the single component of the female sex pheromone in the silkmoth Bombyx mori. EZ alone evokes full courtship behaviors from conspecific males; however, its geometric isomer (EE) was consistently detected in the pheromone glands (PG) of 16 B. mori strains and a field population of the wild silkmoth Bombyx mandarina, which also uses EZ as the single pheromone component. We investigated the pheromonal activities of EE using a commercial hybrid strain of B. mori, Kinshu × Showa. The behavioral assay demonstrated that a 104-105-fold higher dose of EE than EZ was able to elicit behavioral responses from males. To elucidate whether the trace contaminant of EZ in the EE standard is responsible for these responses, we examined the responses of male antennae to EE using a gas chromatograph-electroantennographic detector system (GC-EAD). The EE, at high doses elicited marginal responses from the male antennae. We next examined antennal and behavioral responses of B. mori whose BmOR1 gene, which is responsible for the reception of bombykol, was knocked out. The knockout of BmOR1 resulted in the complete loss of antennal and behavioral responses to EE and EZ, demonstrating that if EE itself is active, it induces these responses via the incidental stimulation of BmOR1, not via the stimulation of EE-specific receptors. The existence of EE in the PG of B. mori and B. mandarina is discussed from the viewpoints of pheromone biosynthesis and the evolution of pheromone communication systems.

Proximate causes and consequences of intergenerational influences of salient sensory experience.

Salient sensory environments experienced by a parental generation can exert intergenerational influences on offspring. While these data provide an exciting new perspective on biological inheritance, questions remain about causes and consequences of intergenerational influences of salient sensory experience. We previously showed that exposing male mice to a salient olfactory experience, like olfactory fear conditioning, resulted in offspring demonstrating a sensitivity to the odor used to condition the paternal generation and possessing enhanced neuroanatomical representation for that odor. In this study, we first injected RNA extracted from sperm of male mice that underwent olfactory fear conditioning into naïve single-cell zygotes and found that adults that developed from these embryos had increased sensitivity and enhanced neuroanatomical representation for the odor (Odor A) with which the paternal male had been conditioned. Next, we found that female, but not male offspring sired by males conditioned with Odor A show enhanced consolidation of a weak single-trial Odor A + shock fear conditioning protocol. Our data provide evidence that RNA found in the paternal germline after exposure to salient sensory experiences can contribute to intergenerational influences of such experiences, and that such intergenerational influences confer an element of adaptation to the offspring. In so doing, our study of intergenerational influences of parental sensory experience adds to existing literature on intergenerational influences of parental exposures to stress and dietary manipulations and suggests that some causes (sperm RNA) and consequences (behavioral flexibility) of intergenerational influences of parental experiences may be conserved across a variety of parental experiences.

The olfactory reception of acetic acid and ionotropic receptors in the Oriental armyworm, Mythimna separata Walker.

Various insect species including moths have shown significant foraging preference to acetic acid. However, the olfactory reception and behavioral outputs of acetic acid in moths remain unsolved. The female adults of Oriental armyworm, Mythimna separata, exhibit high preference to acetic acid enriched sweet vinegar solutions, making them good targets for exploration of acid reception and performance. We first proved that acetic acid is an essential component which elicited electrophysiological responses from volatiles of the sweet vinegar solution. Successive single sensillum recording tests showed that at least 4 types (as1, as2, as3, and as4) of sensilla were involved in acetic acid reception in the antennae. The low dosages of acetic acid elicited upwind flight and close search, and pre-contact proboscis extension responses of the fasted females, indicating it serves as a food related olfactory cue. In vivo optical imaging data showed that low dosages of acetic acid activated one ordinary glomerulus (DC3), and high dosages evoked additional two glomeruli (DC1 and AC1) in the antennal lobe. A systematic survey on olfaction related receptors in three related transcriptomes has yielded 67 olfactory receptors (ORs) and 19 ionotropic receptors (IRs). Among, MsepIR8a, MsepIR64a, MsepIR75q1, and MsepIR75q2 were chosen as putative acid receptors by blasting against known acid IRs in Drosophila and comparing essential amino acid residues which related to acid sensing. Later in situ hybridization revealed that MsepIr8a was co-expressed with each of the other 3 Irs, suggesting its putative co-receptor role. This study reveals olfactory reception of acetic acid as an attractant in M. separata, and it provides the solid basis for later deorphanization of relevant receptors.

Exploring the mechanism of olfactory recognition in the initial stage by modeling the emission spectrum of electron transfer.

Olfactory sense remains elusive regarding the primary reception mechanism. Some studies suggest that olfaction is a spectral sense, the olfactory event is triggered by electron transfer (ET) across the odorants at the active sites of odorant receptors (ORs). Herein we present a Donor-Bridge-Acceptor model, proposing that the ET process can be viewed as an electron hopping from the donor molecule to the odorant molecule (Bridge), then hopping off to the acceptor molecule, making the electronic state of the odorant molecule change along with vibrations (vibronic transition). The odorant specific parameter, Huang-Rhys factor can be derived from ab initio calculations, which make the simulation of ET spectra achievable. In this study, we revealed that the emission spectra (after Gaussian convolution) can be acted as odor characteristic spectra. Using the emission spectrum of ET, we were able to reasonably interpret the similar bitter-almond odors among hydrogen cyanide, benzaldehyde and nitrobenzene. In terms of isotope effects, we succeeded in explaining why subjects can easily distinguish cyclopentadecanone from its fully deuterated analogue cyclopentadecanone-d28 but not distinguishing acetophenone from acetophenone-d8.