Onset of odorant receptor gene expression during olfactory sensory neuron regeneration

Individual olfactory sensory neurons are thought to express only one odorant receptor gene from a repertoire of hundreds to thousands of genes. How do these sensory neurons choose just one specific odorant receptor to express during their differentiation? As an initial attempt toward understanding the process of odorant receptor gene regulation, we studied when odorant receptor expression is activated during sensory neuron regeneration. We find that receptor gene expression is activated in postmitotic neurons and can occur in the absence of the olfactory bulb. These results suggest that receptor expression is restricted to the terminal stages of olfactory neuron differentiation, and sensory neurons do not simply inherit the odorant receptor that is already expressed in mitotic precursor cells. Our results also support a model in which odorant receptor gene expression occurs independent of the olfactory bulb.     

Interchromosomal interactions and olfactory receptor choice

The expression of a single odorant receptor (OR) gene from a large gene family in individual sensory neurons is an essential feature of the organization and function of the olfactory system. We have used chromosome conformation capture to demonstrate the specific association of an enhancer element, H, on chromosome 14 with multiple OR gene promoters on different chromosomes. DNA and RNA fluorescence in situ hybridization (FISH) experiments allow us to visualize the colocalization of the H enhancer with the single OR allele that is transcribed in a sensory neuron. In transgenic mice bearing additional H elements, sensory neurons that express OR pseudogenes also express a second functional receptor. These data suggest a model of receptor choice in which a single trans-acting enhancer element may allow the stochastic activation of only one OR allele in an olfactory sensory neuron.     

The molecular logic of olfaction in Drosophila

Drosophila fruit flies display robust olfactory-driven behaviors with an olfactory system far simpler than that of vertebrates. Endowed with 1300 olfactory receptor neurons, these insects are able to recognize and discriminate between a large number of distinct odorants. Candidate odorant receptor molecules were identified by complimentary approaches of differential cloning and genome analysis. The Drosophila odorant receptor (DOR) genes encode a novel family of proteins with seven predicted membrane-spanning domains, unrelated to vertebrate or nematode chemosensory receptors. There are on the order of 60 or more members of this gene family in the Drosophila genome, far fewer than the hundreds to thousands of receptors found in vertebrates or nematodes. DOR genes are selectively expressed in small subsets of olfactory neurons, in expression domains that are spatially conserved between individuals, bilaterally symmetric and not sexually dimorphic. Double in situ RNA hybridization with a number of pairwise combinations of DOR genes fails to reveal any overlap in gene expression, suggesting that each olfactory neuron expresses one or a small number of receptor genes and is therefore functionally distinct. How is activation of such a subpopulation of olfactory receptor neurons in the periphery sensed by the brain? In the mouse, all neurons expressing a given receptor project with precision to two of 1800 olfactory bulb glomeruli, creating a spatial map of odor quality in the brain. We have employed DOR promoter transgenes that recapitulate expression of endogenous receptor to visualize the projections of individual populations of receptor neurons to subsets of the 43 glomeruli in the Drosophila antennal lobe. The results suggest functional conservation in the logic of olfactory discrimination from insects to mammals.     

Taste perception and coding in Drosophila

BACKGROUND: Discrimination between edible and contaminated foods is crucial for the survival of animals. In Drosophila, a family of gustatory receptors (GRs) expressed in taste neurons is thought to mediate the recognition of sugars and bitter compounds, thereby controlling feeding behavior. RESULTS: We have characterized in detail the expression of eight Gr genes in the labial palps, the fly's main taste organ. These genes fall into two distinct groups: seven of them, including Gr66a, are expressed in 22 or fewer taste neurons in each labial palp. Additional experiments show that many of these genes are coexpressed in partially overlapping sets of neurons. In contrast, Gr5a, which encodes a receptor for trehalose, is expressed in a distinct and larger set of taste neurons associated with most chemosensory sensilla, including taste pegs. Mapping the axonal targets of cells expressing Gr66a and Gr5a reveals distinct projection patterns for these two groups of neurons in the brain. Moreover, tetanus toxin-mediated inactivation of Gr66a- or Gr5a-expressing cells shows that these two sets of neurons mediate distinct taste modalities-the perception of bitter (caffeine) and sweet (trehalose) taste, respectively. CONCLUSION: Discrimination between two taste modalities-sweet and bitter-requires specific sets of gustatory receptor neurons that express different Gr genes. Unlike the Drosophila olfactory system, where each neuron expresses a single olfactory receptor gene, taste neurons can express multiple receptors and do so in a complex Gr gene code that is unique for small sets of neurons.     

Odorant receptor gene choice in olfactory sensory neurons: the one receptor-one neuron hypothesis revisited

Designed for general chemical recognition, the mammalian olfactory system shares many similarities with the immune system. Among these is the popular notion that a single olfactory sensory neuron expresses a single odorant receptor gene, while all other approximately 1000 genes of this type remain silent. Here, I examine the evidence supporting the one receptor-one neuron hypothesis. I conclude that, contrary to widespread belief, it is far from being proven. I propose an hypothesis of a developmental phase of oligogenic expression that is followed by positive and negative selection resulting usually in cells with one expressed receptor. Curiously, selective processes are well established and widely accepted for lymphocytes, but these concepts are essentially ignored for olfactory sensory neurons, despite the analogies that are frequently made between these two systems. More attention must be paid to odorant receptor gene choice and expression during development and neuronal differentiation.     

Identification of olfactory receptor genes in Atlantic salmon Salmo salar

It has been hypothesized that salmonids use olfactory cues to return to their natal rivers and streams. The key components of the molecular pathways involved in imprinting and homing, however, are still unknown. Aquatic chemical cues are received through the nares and into the nasal cavity that contains a single olfactory organ, the olfactory rosette. The olfactory rosette contains sensory neurons, each of which is thought to express only one olfactory receptor. If odorants are involved in salmonid homing migration then olfactory receptors should play a critical role in the dissipation of information from the environment to the fish. Therefore, to understand the molecular basis for imprinting and homing in Atlantic salmon Salmo salar it is important to identify and characterize the repertoire of olfactory receptors in this species. The first public assembly of the S. salar genome was searched for genes encoding three of the superfamilies of fish olfactory receptors: V2R-like (olfc), V1R-like (ora) and main olfactory receptor (mor). A further six ora genes were added to ora1 and ora2, which had been described previously. In addition, 48 putative mors were identified, 24 of which appear to be functional based on their gene structures and predicted amino-acid sequences. Phylogenetic analyses were then used to compare these S. salar olfactory receptor genes with those of zebrafish Danio rerio, two pufferfish species Takifugu rubripes and Tetraodon nigroviridis, medaka Oryzias latipes and three-spined stickleback Gasterosteus aculeatus.     

Advantage of the Highly Restricted Odorant Receptor Expression Pattern in Chemosensory Neurons of Drosophila

A fundamental molecular feature of olfactory systems is that individual neurons express only one receptor from a large odorant receptor gene family. While numerous theories have been proposed, the functional significance and evolutionary advantage of generating a sophisticated one-receptor-per neuron expression pattern is not well understood. Using the genetically tractable Drosophila melanogaster as a model, we demonstrate that the breakdown of this highly restricted expression pattern of an odorant receptor in neurons leads to a deficit in the ability to exploit new food sources. We show that animals with ectopic co-expression of odorant receptors also have a competitive disadvantage in a complex environment with limiting food sources. At the level of the olfactory system, we find changes in both the behavioral and electrophysiological responses to odorants that are detected by endogenous receptors when an olfactory receptor is broadly misexpressed in chemosensory neurons. Taken together these results indicate that restrictive expression patterns and segregation of odorant receptors to individual neuron classes are important for sensitive odor-detection and appropriate olfactory behaviors.     

Searching for the Ligands of Odorant Receptors

Through the sense of smell mammals can detect and discriminate between a large variety of odorants present in the surrounding environment. Odorants bind to a large repertoire of odorant receptors located in the cilia of olfactory sensory neurons of the nose. Each olfactory neuron expresses one single type of odorant receptor, and neurons expressing the same type of receptor project their axons to one or a few glomeruli in the olfactory bulb, creating a map of odorant receptor inputs. The information is then passed on to other regions of the brain, leading to odorant perception. To understand how the olfactory system discriminates between odorants, it is necessary to determine the odorant specificities of individual odorant receptors. These studies are complicated by the extremely large size of the odorant receptor family and by the poor functional expression of these receptors in heterologous cells. This article provides an overview of the methods that are currently being used to investigate odorant receptor–ligand interactions.     

Information processing in mammalian olfactory system

In recent years, considerable progress has been made in understanding how the olfactory system uses neural space to encode sensory information. In this review, we focus on recent studies aimed at understanding the organizational strategies used by the mammalian olfactory system to encode information. The odorant receptor gene family is discussed in the context of its genomic organization as well as the specificity of olfactory sensory neurons. These data have important consequences for the mechanisms of odorant receptor gene choice by a given sensory neuron. Division of the olfactory epithelium into zones that express different sets of odorant receptors is the first level of input organization. The topographical relationship between periphery and olfactory bulb represents a further level of processing of information and results in the formation of a highly organized spatial map of information in the olfactory bulb. There, local circuitry refines the sensory input through various lateral interactions. Finally, the factors that may drive the development of such a spatial map are discussed. The onset of expression and the establishment of the zonal organization of odorant receptor genes in the epithelium are not dependent upon the presence of the olfactory bulb, suggesting that the functional identity of olfactory sensory neurons is determined independently of target selection. © 1996 John Wiley & Sons, Inc.     

Excitatory interactions between olfactory processing channels in the Drosophila antennal lobe

Each odorant receptor gene defines a unique type of olfactory receptor neuron (ORN) and a corresponding type of second-order neuron. Because each odor can activate multiple ORN types, information must ultimately be integrated across these processing channels to form a unified percept. Here, we show that, in Drosophila, integration begins at the level of second-order projection neurons (PNs). We genetically silence all the ORNs that normally express a particular odorant receptor and find that PNs postsynaptic to the silent glomerulus receive substantial lateral excitatory input from other glomeruli. Genetically confining odor-evoked ORN input to just one glomerulus reveals that most PNs postsynaptic to other glomeruli receive indirect excitatory input from the single ORN type that is active. Lateral connections between identified glomeruli vary in strength, and this pattern of connections is stereotyped across flies. Thus, a dense network of lateral connections distributes odor-evoked excitation between channels in the first brain region of the olfactory processing stream.     

Prominent roles for odorant receptor coding sequences in allelic exclusion

Mammalian odorant receptors (ORs) are crucial for establishing the functional organization of the olfactory system, but the mechanisms controlling their expression remain largely unexplained. Here, we utilized a transgenic approach to explore OR gene regulation. We determined that although olfactory sensory neurons (OSNs) are capable of supporting expression of multiple functional ORs, several levels of control ensure that each neuron normally expresses only a single odorant receptor. Surprisingly, this regulation extends beyond endogenous ORs even preventing expression of transgenes consisting of OR-coding sequences driven by synthetic promoters. Thus, part of the intrinsic feedback system must rely on elements present in the OR-coding sequence. Notably, by expressing the same transgenic ORs precociously in immature neurons, we have overcome this suppression and established a generic method to express any OR in approximately 90% of OSNs. These results provide important insights into the hierarchy of OR gene expression and the vital role of the OR-coding sequence in this regulation.     

Local and cis effects of the H element on expression of odorant receptor genes in mouse

From the approximately 1,200 odorant receptor (OR) genes in the mouse genome, an olfactory sensory neuron is thought to express only one gene. The mechanisms of OR gene choice are not understood. A 2.1 kilobase region (the H element) adjacent to a cluster of seven OR genes has been proposed as a trans- and pan-enhancer for OR gene expression. Here, we deleted the H element by gene targeting in mice. The deletion abolishes expression of a family of three OR genes proximal to H, and H operates in cis on these genes. Deletion of H has a graded effect on expression of a distal group of four OR genes, commensurate with genomic distance. There is no demonstrable effect on expression of OR genes located outside the cluster. Our findings are not consistent with the hypothesis of H as an essential trans-acting enhancer for genome-wide regulation of OR gene expression.     

Odorant and pheromone receptor gene regulation in vertebrates

The largest mammalian gene family codes for odorant receptors and is exclusively devoted to the perception of the outside world. Its expression is very peculiar, since olfactory sensory neurons are only allowed to express a single of its numerous members, from a single parental allele. How this is achieved is unknown, but recent work points to multiple regulatory mechanisms, possibly shared by pheromone receptor genes, acting at (a) a general level, via the expression of the chemoreceptor itself and (b) a more restricted level, defined by activator elements.     

Specific projection of the sensory crypt cells in the olfactory system in crucian carp, Carassius carassius

To study the projection of a special type of sensory neuron called crypt cells in the olfactory system in crucian carp, Carassius carassius, we applied the neural tracer 1,1-dilinoleyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI) in the olfactory bulb (OB). Small crystals of DiI were applied in a small area at the synaptic region at the ventral part of the OB, where a population of secondary neurons specific for sex pheromones has been identified. In those samples (4 out of 24) where only axons in the lateral bundle of the medial olfactory tract were stained, the majority (50-66%) of olfactory sensory neurons stained were crypt cells situated in the peripheral layer of the olfactory epithelium. Because this bundle of the tract mediates reproductive behavior, it is conceivable that crypt cells express olfactory receptors for sex pheromones.     

A spatial map of olfactory receptor expression in the Drosophila antenna

Insects provide an attractive system for the study of olfactory sensory perception. We have identified a novel family of seven transmembrane domain proteins, encoded by 100 to 200 genes, that is likely to represent the family of Drosophila odorant receptors. Members of this gene family are expressed in topographically defined subpopulations of olfactory sensory neurons in either the antenna or the maxillary palp. Sensory neurons express different complements of receptor genes, such that individual neurons are functionally distinct. The isolation of candidate odorant receptor genes along with a genetic analysis of olfactory-driven behavior in insects may ultimately afford a system to understand the mechanistic link between odor recognition and behavior.     

Combinatorial coexpression of neural and immune multigene families in mouse vomeronasal sensory neurons

The vomeronasal organ (VNO) is a chemosensory organ specialized in the detection of pheromones in higher vertebrates. In mouse and rat, two gene superfamilies, V1r and V2r vomeronasal receptor genes, are expressed in sensory neurons whose cell bodies are located in, respectively, the apical and basal layers of the VNO epithelium. Here, we report that neurons of the basal layer express another multigene family, termed H2-Mv, representing nonclassical class I genes of the major histocompatibility complex. The nine H2-Mv genes are expressed differentially in subsets of neurons. More than one H2-Mv gene can be expressed in an individual neuron. In situ hybridization with probes for H2-Mv and V2r genes reveals complex and nonrandom combinations of coexpression. While neural expression of Mhc class I molecules is increasingly being appreciated, the H2-Mv family is distinguished by variegated expression across seemingly similar neurons and coexpression with a distinct multigene family encoding neural receptors. Our findings suggest that basal vomeronasal sensory neurons may consist of multiple lineages or compartments, defined by particular combinations of V2r and H2-Mv gene expression.     

Integrating the molecular and cellular basis of odor coding in the Drosophila antenna

We investigate how the molecular and cellular maps of the Drosophila olfactory system are integrated. A correspondence is established between individual odor receptors, neurons, and odors. We describe the expression of the Or22a and Or22b receptor genes, show localization to dendritic membranes, and find sexual dimorphism. Or22a maps to the ab3A neuron, which responds to ethyl butyrate. Analysis of a deletion mutant lacking Or22a, along with transgenic rescue experiments, confirms the mapping and demonstrates that an Or gene is required for olfactory function in vivo. Ectopic expression of Or47a in a mutant cell identifies the neuron from which it derives and its odor ligands. Ectopic expression in a wild-type cell shows that two receptors can function in a single cell. The ab3A neuron does not depend on normal odor receptor gene expression to navigate to its target in the CNS.