The vomeronasal organ and adjacent glands express components of signaling cascades found in sensory neurons in the main olfactory system

The vomeronasal organ (VNO) is a sensory organ that influences social and/or reproductive behavior and, in many cases, the survival of an organism. The VNO is believed to mediate responses to pheromones; however, many mechanisms of signal transduction in the VNO remain elusive. Here, we examined the expression of proteins involved in signal transduction that are found in the main olfactory system in the VNO. The localization of many signaling molecules in the VNO is quite different from those in the main olfactory system, suggesting differences in signal transduction mechanisms between these two chemosensory organs. Various signaling molecules are expressed in distinct areas of VNO sensory epithelium. Interestingly, we found the expressions of groups of these signaling molecules in glandular tissues adjacent to VNO, supporting the physiological significance of these glandular tissues. Our finding of high expression of signaling proteins in glandular tissues suggests that neurohumoral factors influence glandular tissues to modulate signaling cascades that in turn alter the responses of the VNO to hormonal status.     

Molecular biology of pheromone perception in mammals

In mammals, olfactory sensory perception is mediated by two anatomically and functionally distinct sensory organs: the main olfactory epithelium (MOE) and the vomeronasal organ (VNO). Pheromones activate the VNO and elicit a characteristic array of innate reproductive and social behaviors, along with dramatic neuroendocrine responses. Recent approaches have provided new insights into the molecular biology of sensory transduction in the vomeronasal organ. Differential screening of cDNA libraries constructed from single sensory neurons from the rat VNO has led to the isolation of a family of genes which are likely to encode mammalian pheromone receptors. The isolation of these receptors from the vomeronasal organ might permit the analysis of the molecular events which translate the bindings of pheromones into innate stereotypic behaviors and help to elucidate the logic of pheromone perception in mammals.     

The effects of sex on chemosensory communication in a terrestrial salamander (Plethodon shermani)

Although much evidence reveals sexually dimorphic processing of chemosensory cues by the brain, potential sex differences at more peripheral levels of chemoreception are understudied. In plethodontid salamanders, the volume of the vomeronasal organ (VNO) is almost twice as large in males as compared to females, both in absolute and relative size. To determine whether the structural sexual dimorphism in VNO volume is associated with sex differences in other peripheral aspects of chemosensation, we measured sex differences in chemo-investigation and in responsiveness of the VNO to chemosensory cues. Males and females differed in traits influencing stimulus access to VNO chemosensory neurons. Males chemo-investigated (“nose tapped”) neutral substrates and substrates moistened with female body rinses more than did females. Compared to females, males had larger narial structures (cirri) associated with the transfer of substrate-borne chemical cues to the lumen of the VNO. These sex differences in chemo-investigation and narial morphology likely represent important mechanisms for regulating sex differences in chemical communication. In contrast, males and females did not differ in responsiveness of VNO chemosensory neurons to male mental gland extract or female skin secretions. This important result indicates that although males have a substantially larger VNO compared to females, the male VNO was not more responsive to every chemosensory cue that is detected by the VNO. Future studies will determine whether the male VNO is specialized to detect a subset of chemosensory cues, such as female body rinses or female scent marks.     

Sexual dimorphism of the vomeronasal organ and the accessory olfactory bulb of the mandarin vole<Emphasis Type="Italic">Microtus mandarinus</Emphasis> and the reed vole<Emphasis Type="Italic">M. fortis</Emphasis>

Sexual dimorphisms of the vomeronasal organ (VNO) and the accessory olfactory bulb (AOB) of the mandarin voleMicrotus mandarinus Milne-Edwards, 1871 and reed voleM. fortis Büchner, 1889 are reported for the first time in the present work. The thickness and length of the vomeronasal epithelium (VE) and the nuclear size of the receptor cells, the width and length of the granule cell zone, the width and length of the mitral cell zone, and the density of the mitral cells were surveyed. The thickness and length of the vomeronasal epithelium (VE), the length of the granule cell zone and the mitral cell zone, and the densities of mitral cells were significantly different between male and female reed voles. Male and female mandarin voles had no significant differences in any of these parameters. Polygamous reed voles had a greater degree of sexual dimorphism in VNO and AOB than did monogamous mandarin voles. The present results provide evidence to the hypothesis that the degree of sexual dimorphism may be related to the mating system.     

The vomeronasal organ of the male ferret.

The vomeronasal organ (VNO) is known to play a major role in sexual behavior in many mammals. This study is the first report that the adult male ferret has a VNO, which is considerably smaller and morphologically different from the usually crescent-shaped epithelium in several mammalian species, particularly rodents. There were no differences in the size or structure of the ferret VNO between the mating season in spring and the sexually quiescent season in autumn, although plasma testosterone, testis size and brain size are dramatically increased in spring and behavior changes significantly. The histological data suggest that the VNO might be not as important a structure in male ferret sexual behavior as in rodents.     

The vomeronasal organ and associated structures of the fetal African elephant, Loxodonta africana (Proboscidea, Elephantidae)

The vomeronasal organ (VNO) is a chemosensory structure of the nasal septum found in most tetrapods. Although potential behavioural correlates of VNO function have been shown in two of the three elephant species, its morphology in Loxodonta africana has not been studied. The development of the VNO and its associated structures in the African elephant are described in detail using serially sectioned material from fetal stages. The results show that many components of the VNO complex (e.g. neuroepithelium, receptor‐free epithelium, vomeronasal nerve, paravomeronasal ganglia, blood vessels, vomeronasal cartilage) are well developed even in a 154‐day‐old fetus, in which the VNO opens directly into the oral cavity with only a minute duct present. However, the vomeronasal glands and their ducts associated with the VNO were developed only in the 210‐day‐old fetus. Notably, in this fetus, the vomeronasal–nasopalatine duct system had acquired a pathway similar to that described in the adult Asian elephant; the VNOs open into the oral cavity via the large palatal parts of the nasopalatine ducts, which are lined by a stratified squamous epithelium. The paired palatal ducts initially coursed anteriorly at an angle of 45° from the oral recess and/or the oral cavity mucosa, and merged into the vomeronasal duct. This study confirms the unique characteristics of the elephant VNO, such as its large size, the folded epithelium of the VNO tube, and the dorsomedial position of the neuroepithelium. The palatal position and exclusive communication of the VNO with the oral cavity, as well as the partial reduction of the nasopalatine duct, might be related to the unique elephantid craniofacial morphogenesis, especially the enormous growth of the tusk region, and can be seen as autapomorphies.     

Timing of neuronal intermediate filament proteins expression in the mouse vomeronasal organ during pre- and postnatal development. An immunohistochemical study

Several types of intermediate filament proteins are expressed in developing and mature neurons; they cooperate with other cytoskeletal components to sustain neuronal function from early neurogenesis onward. In this work the timing of expression of nestin, peripherin, internexin, and the neuronal intermediate filament triplet [polypeptide subunits of low (NF-L), medium (NF-M), and high (NF-H) molecular weight] was investigated in the developing fetal and postnatal mouse vomeronasal organ (VNO) by means of immunohistochemistry. The results show that the sequence of expression of intermediate filament proteins is internexin, nestin, and NF-M in the developing vomeronasal sensory epithelium; internexin, peripherin, and NF-M in the developing vomeronasal nerve; and nestin, internexin and peripherin, NF-L, and NF-M in the nerve supply to accessory structures of the VNO. At sexual maturity (2 months) NF-M is only expressed in vomeronasal neurons and NF-M, NF-L and peripherin are expressed in extrinsic nerves supplying VNO structures. The differential distribution of intermediate filament proteins in the vomeronasal sensory epithelium and nerve is discussed in terms of the cell types present therein. It is concluded that several intermediate filament proteins are sequentially expressed during intrauterine development of the VNO neural structures in a different pattern according to the different components of the VNO.     

Are Pheromones Detected Through the Main Olfactory Epithelium?

A major sensory organ for the detection of pheromones by animals is the vomeronasal organ (VNO). Although pheromones control the behaviors of various species, the effect of pheromones on human behavior has been controversial because the VNO is not functional in adults. However, recent genetic, biochemical, and electrophysiological data suggest that some pheromone-based behaviors, including male sexual behavior in mice, are mediated through the main olfactory epithelium (MOE) and are coupled to the type 3 adenylyl cyclase (AC3) and a cyclic nucleotide-gated (CNG) ion channel. These recent discoveries suggest the provocative hypothesis that human pheromones may signal through the MOE.     

Sex- and strain-specific expression and vomeronasal activity of mouse ESP family peptides

Male mice secrete exocrine-gland-secreting peptide 1 (ESP1) from the extraorbital lacrimal gland into tear fluid [1]. Other mice detect ESP1 through sensory neurons in the vomeronasal organ (VNO), a secondary olfactory system that senses pheromonal information, including sex, strain, and species. ESP1 is now known to be a member of a multigene family that encodes peptides of various lengths. We herein performed genomic and expression analyses of the ESP family. The ESP family consists of 38 members in mice and 10 members in rat but is absent from the human genome, suggesting rapid molecular evolution. In addition to the male-specific ESP1, we discovered one, which we designated ESP36, that, in adult BALB/c mice, is expressed only in the female extraorbital lacrimal gland. The sexually dimorphic expression is ensured by the release of testosterone after puberty. However, we observed dramatic differences in the expression levels of ESPs between strains. Finally, all ESPs elicited an electrical response in the vomeronasal epithelium but not in the main olfactory epithelium. Multielectrode recording of VNO activity demonstrated that ESP1 induces action potentials in vomeronasal neurons, leading to an increase in the spike firing rate, and that ESP1 is recognized by narrowly tuned vomeronasal sensory neurons. Sexual dimorphism and strain differences of ESPs and their reception in the VNO suggest that the ESP family can convey information about sex and individual identity via the vomeronasal system. The chemosensation of this nonvolatile peptide family by direct contact appears to be one of strategies for sociosexual communication in rodent species.     

Discrimination of conspecific sex and reproductive condition using chemical cues in axolotls (<Emphasis Type="Italic">Ambystoma mexicanum</Emphasis>)

Chemosensory cues play an important role in the daily lives of salamanders, mediating foraging, conspecific recognition, and territorial advertising. We investigated the behavioral effects of conspecific whole-body odorants in axolotls, Ambystoma mexicanum, a salamander species that is fully aquatic. We found that males increased general activity when exposed to female odorants, but that activity levels in females were not affected by conspecific odorants. Although males showed no difference in courtship displays across testing conditions, females performed courtship displays only in response to male odorants. We also found that electro-olfactogram responses from the olfactory and vomeronasal epithelia were larger in response to whole-body odorants from the opposite sex than from the same sex. In males, odorants from gravid and recently spawned females evoked different electro-olfactogram responses at some locations in the olfactory and vomeronasal epithelia; in general, however, few consistent differences between the olfactory and vomeronasal epithelia were observed. Finally, post hoc analyses indicate that experience with opposite-sex conspecifics affects some behavioral and electrophysiological responses. Overall, our data indicate that chemical cues from conspecifics affect general activity and courtship behavior in axolotls, and that both the olfactory and vomeronasal systems may be involved in discriminating the sex and reproductive condition of conspecifics.Abbreviations EOG electro-olfactogram - VNO vomeronasal organ     

Vomeronasal neuroepithelium and forebrain Fos responses to male pheromones in male and female mice

Male urinary pheromones modulate behavioral and neuroendocrine function in mice after being detected by sensory neurons in the vomeronasal organ (VNO) neuroepithelium. We used nuclear Fos protein immunoreactivity (Fos-IR) as a marker of changes in neuronal activity to examine the processing of male pheromones throughout the VNO projection pathway to the hypothalamus. Sexually naive male and female Balb/c mice were gonadectomized and treated daily with estradiol benzoate (EB) or oil vehicle for 3 weeks. Subjects were then exposed to soiled bedding from gonadally intact Balb/c males or to clean bedding for 90 min prior to sacrifice and processing of their VNOs and forebrains for Fos-IR. Male pheromones induced similar numbers of Fos-IR cells in the VNO neuroepithelium of oil-treated male and female subjects; however, EB-treated females had significantly more Fos-IR neurons in the VNO than any other group. There was an equivalent neuronal Fos response to male odors in the mitral and granule cells of the anterior and posterior accessory olfactory bulb of males and females, regardless of hormone treatment. In central portions of the VNO projection pathway (i.e., bed nucleus of the stria terminalis, medial preoptic area) neuronal Fos responses to male pheromones were present in female but absent in male subjects, regardless of hormone treatment. In a separate experiment, mating induced neuronal Fos-IR in these brain regions at levels in gonadally intact male subjects which were equal to or greater than those seen in ovariectomized females primed with estrogen and progesterone. This suggests that neurons in the central portions of the male's VNO pathway are capable of expressing Fos. Our results suggest that sexually dimorphic central responses to pheromones exist in mice that may begin in the VNO neuroepithelium.     

Conditioned odor aversion induces social anxiety towards females in wild‐type and TrpC2 knockout male mice

Female‐emitted pheromonal inputs possess an intrinsic rewarding value for conspecific males, promoting approach and investigation of the potential mating partner. In mice these inputs are detected mainly by the vomeronasal organ (VNO) and the main olfactory epithelium (MOE). We investigated the role of VNO‐mediated inputs in experience‐dependent plasticity of reproductive responses. We applied a sex‐specific conditioned odor aversion (COA) paradigm on adult, wild‐type (WT) male mice and on male mice impaired in VNO‐mediated signal transduction (TrpC2^?/?). We found that WT males, which underwent COA to female‐soiled bedding, lost their innate preference to female odors and presented lower motivation to approach a sexually receptive female. COA also abolished the testosterone surge normally seen following exposure to female odors. Moreover, the conditioned males displayed impairments in copulatory behaviors, which lasted for several weeks. Surprisingly, these males also exhibited phobic behaviors towards receptive females, including freezing and fleeing responses. In contrast, WT males which underwent COA specifically to male pheromones showed no change in olfactory preference and only a marginally significant elevation in intermale aggression. Finally, we show that TrpC2^?/? males were able to acquire aversion to female‐soiled bedding and presented similar behavioral alterations following COA in their responses to female cues. Our results demonstrate that the intrinsic rewarding value of female pheromones can be overridden through associative olfactory learning, which occurs independently of VNO inputs, probably through MOE signaling.     

Goα expression in the vomeronasal organ and olfactory bulb of the tammar wallaby

The vomeronasal organ (VNO) detects pheromones via 2 large families of receptors: vomeronasal receptor 1, associated with the protein Giα2, and vomeronasal receptor 2, associated with Goα. We investigated the distribution of Goα in the developing and adult VNO and adult olfactory bulb of a marsupial, the tammar wallaby. Some cells expressed Goα as early as day 5 postpartum, but by day 30, Goα expressing cells were distributed throughout the receptor epithelium of the VNO. In the adult tammar, Goα appeared to be expressed in sensory neurons whose nuclei were mostly basally located in the vomeronasal receptor epithelium. Goα expressing vomeronasal receptor cells led to all areas of the accessory olfactory bulb (AOB). The lack of regionally restricted projection of the vomeronasal receptor cell type 2 in the tammar was similar to the uniform type, with the crucial difference that the uniform type only shows expression of Giα2 and no expression of Goα. The observed Goα staining pattern suggests that the tammar may have a third accessory olfactory type that could be intermediate to the segregated and uniform types already described.     

达乌尔黄鼠犁鼻器和副嗅球的组织结构及嗅球c-Fos表达的季节变化

啮齿动物的犁鼻器和副嗅球与社会通讯和生殖行为有关,主嗅球影响其觅食行为。达乌尔黄鼠（Spermophilus dauricus）是一种具有较低社会行为的储脂类冬眠动物。本研究用组织学和免疫组织化学方法探究了其犁鼻器和副嗅球的结构特点及嗅球神经元活动对季节变化的适应。结果发现,达乌尔黄鼠犁鼻器具有较大的血管,犁鼻器管腔外侧为非感觉性的呼吸上皮（Respiratory epithelium,RE）,内侧为感觉上皮（Sensory epithelium,SE）,RE较SE薄,靠近管腔处为假复层柱状上皮。选取犁鼻器中间部位比较,发现SE的厚度、长度及感觉细胞密度均无性别差异。副嗅球位于主嗅球后方背内侧,由6层细胞构成。侧嗅束穿过副嗅球,位于颗粒细胞层之上。雄性达乌尔黄鼠较雌性有更长的僧帽细胞层和颗粒细胞层。春季（3月）和冬季（1月）达乌尔黄鼠主嗅球的嗅小球层、僧帽细胞层和颗粒细胞层的c-Fos-ir神经元密度显著低于夏季（7月）和秋季（10月）,且冬季外网织层的c-Fos-ir神经元密度显著低于夏季和秋季,说明达乌尔黄鼠在冬季和春季的嗅觉神经活动较弱,呈现出对冬眠的生理性适应。这些结果丰富了动物犁鼻器和副嗅球的形态学资料,并有助于理解冬眠动物嗅觉系统对季节变化和冬眠的适应。     

Expression of candidate pheromone receptor genes in vomeronasal neurons

In mammals, olfactory sensory perception is mediated by two anatomically and functionally distinct organs: the main olfactory epithelium (MOE) and the vomeronasal organ (VON). Pheromones activate the VNO and elicit a characteristic array of innate reproductive and social behaviors, along with dramatic neuroendocrine responses. Recent approaches have provided new insights into the molecular biology of sensory transduction in the VNO. Differential screening of cDNA libraries constructed from single sensory neurons from the rat VNO has led to the isolation of a family of genes which are likely to encode mammalian pheromone receptors. The isolation of these receptors from the VNO might permit the analysis of the molecular events which translate the bindings of pheromones into innate stereotypic behaviors and help to elucidate the logic of pheromone perception in mammals.      

Identification of cytoskeletal markers for the different microvilli and cell types of the rat vomeronasal sensory epithelium

The vomeronasal organ (VNO) of the mammal nose is specialized to detect pheromones. The presumed site of the chemosensory signal transduction of pheromones is the vomeronasal brush border of the VNO sensory epithelium, which has been shown to contain two different sets of microvilli: (i) the tall microvilli of supporting cells and (ii) the short microvilli of the chemoreceptive VNO neurons that branch and intermingle with the basal portions of the longer supporting cell microvilli. A key problem when studying the subcellular distribution of possible VNO signal transduction molecules at the light microscope level is the clear discrimination of immunosignals derived from dendritic microvilli of the VNO neurons and surrounding supporting cell structures. In the present study we therefore looked for cytoskeletal marker proteins, that might help to distinguish at the light microscope level between the two sets of microvilli. By immunostaining we found that the VNO dendritic microvilli can be selectively labelled with antibodies to the calcium-sensitive actin filament-bundling protein villin, whereas supporting cell microvilli contain the actin filament cross-linking protein fimbrin, but not villin. Useful cytoplasmic marker molecules for cellular discrimination were cytokeratin 18 for supporting cells and β-tubulin for dendrites of VNO neurons. A further finding was that the non-sensory epithelium of the rat VNO contains brush cells, a cell type that appears to be involved in certain aspects of chemoreception in the gut. Brush cells or other structures of the vomeronasal brush border did not contain α-gustducin.     

Functional Architecture of the Vomeronasal Organ of the Frog (Genus Rana)

Abstract The vomeronasal organ in the frog, genus Rana, is composed of three interconnected cavities; superior, middle and inferior, which are separated from and anterior to the principal olfactory cavity. The superior cavity is found just underneath the external naris and forms a vestibule both for the principal olfactory organ and the vomeronasal organ. The vomeronasal sensory epithelium is located in the medial region of the inferior cavity and contains ciliated cells and microvillous receptor cells. Inspection of microscopic sections of frogs that had been swimming in fluorescent colorants revealed fluorescence on the surface of the vomeronasal organ, but not on that of the olfactory organ. Observations in vivo show that water enters via the external naris by two fissures, one on each side of the movable nasal lid, passes the middle cavity to flow via the sensory epithelium of the inferior cavity. The design of the frog nose makes it possible for this amphibious animal to sample the chemical composition of its environment; above water the frog can inhale air and expose its olfactory organ to volatile substances; in water the vomeronasal organ samples water-borne substances. These new findings are discussed in relation to the air/water interface and the position of the amphibians in the evolution of terrestrial vertebrates.     

Fine structure of the vomeronasal organ in the grass lizard, Takydromus tachydromoides

The squamates are composed of many taxa, among which there is morphological variation in the vomeronasal organ (VNO). To elucidate the evolution of chemoreception in squamate reptiles, morphological data from the VNO from a variety of squamate species is required. In this study, the morphology of the VNO of the grass lizard Takydromus tachydromoides was examined using light and electron microscopy. The VNO consists of a pair of dome-shaped structures, which communicate with the oral cavity. There are no associated glandular structures. Microvilli are present on the apical surfaces of receptor cells in its sensory epithelium, as well as on supporting cells, and there are centrioles and ciliary precursor bodies on the dendrites. In addition to ciliated cells and basal cells in the non-sensory epithelium, there is a novel type of non-ciliated cell in T. tachydromoides. They have constricted apical cytoplasm and microvilli instead of cilia, and are sparsely distributed in the epithelium. Based on these results, the variation in the morphology of the VNO in scincomorpha, a representative squamate taxon, is discussed.