Características de carcaça, órgão vomeronasal, concentrações séricas de testosterona e IGF-I em machos Nelore com ductos incisivos bloqueados / Characteristics of carcass, vomeronasal organ, serum testosterone andIGF-I concentrations in Nellore males with blocked incisor ducts

Objetivou-se avaliar características quali - quantitativas da carcaça em machos Nelore, submetidos ao bloqueio dos ductos incisivos no período pré puberal. Além disso, objetivou-se avaliar as concentrações séricas de testosterona e do hormônio do crescimento semelhante a insulina do tipo I (IGF-I), e a arquitetura histológica do órgão vomeronasal (OVN). Trinta e quatro machos, no período pré puberal, foram divididos em três grupos experimentais: inteiros – animais controle sem qualquer procedimento (n = 11); bloqueados - animais com os ductos incisivos bilateralmente obstruídos (n = 10); castrados - animais com orquiepididectomia bilateral (n = 13). O OVN foi obtido no abate, ao final do experimento, para avaliação histomorfométrica. As coletas de sangue foram realizadas a cada três meses, em dois turnos (manhã e tarde), totalizando cinco avaliações. Observaram-se maiores (P0,05). Registrou-se redução da altura do epitélio neuro sensitivo do OVN nos bloqueados e castrados (P<0,05). Foi registrada menor concentração de testosterona nos bloqueados nas coletas quatro e cinco à tarde (P<0,05). Conclui-se que a obstrução dos ductos incisivos reduziu os estímulos sensoriais para o OVN, que por sua vez, promoveu alteração na concentração sérica de testosterona, sem alterar o ganho de peso dos animais, porém, não promoveu melhoria na qualidade da carcaça nas condições deste estudo.

A total of 34 prepuberal Nellore males were divided into three groups: control – animals without any procedure (n = 11); blocked - with bilaterally incisors ducts obstructed (n = 10); castrated - with bilateral orchiectomy (n = 13). This study aimed to evaluate the effect of blocking the vomeronasal organ (VNO) in qualitative and quantitative characteristics of the carcass. Also, this study aimed to evaluate testosterone and insulin-like growth factor (IGF-I) serum concentration, and the histological architecture of the OVN. Blood samples were taken every three months during two daily collections (morning and afternoon), totalizing five evaluations. It was observed higher (P 0.05). It was observed a reduction (P<0.05) of the VNO sensory epithelium height in blocked and castrated groups compared with control group. It was registered lower (P<0.05) serum testosterone concentration in blocked group at the fourth and fifth blood collection (afternoon). It was concluded that the obstruction of the incisive ducts reduced the sensorial stimuli for the OVN, which, in turn, promoted a change in the serum concentration of testosterone, without altering the weight gain of the animals, but did not promote improvement in the quality of the carcass under the conditions of this study.

Immunolocalization of VEGF/VEGFR system in human fetal vomeronasal organ during early development.

The vomeronasal system (VNS) is an accessory olfactory structure present in most mammals adhibited to the detection of specific chemosignals implied in social and reproductive behavior. The VNS comprises the vomeronasal organ (VNO), vomeronasal nerve and accessory olfactory bulb. VNO is characterized by a neuroepithelium constituted by bipolar neurons and supporting and stem/progenitor cells. In humans, VNO is present during fetal life and is supposed to possess chemoreceptor activity and participate in gonadotropin-releasing hormone neuronal precursor migration toward the hypothalamus. Instead, the existence and functions of VNO in postnatal life is debated. Vascular endothelial growth factor (VEGF) and its receptors (VEGFRs) have been demonstrated to play fundamental roles in various neurogenic events. However, there are no data regarding the localization and possible function of VEGF/VEGFRs in human fetal VNO. Therefore, this study was conceived to investigate the expression of VEGF/VEGFRs in human VNO in an early developmental period (9-12 weeks of gestation), when this organ appears well structured. Coronal sections of maxillofacial specimens were subjected to peroxidase-based immunohistochemistry for VEGF, VEGFR-1 and VEGFR-2. Double immunofluorescence for VEGF, VEGFR-1 or VEGFR-2 and the neuronal marker protein gene product 9.5 (PGP 9.5) was also performed. VEGF expression was evident in the entire VNO epithelium, with particularly strong reactivity in the middle layer. Strongly VEGF-immunostained cells with aspect similar to bipolar neurons and/or their presumable precursors were detected in the middle and basal layers. Cells detaching from the basal epithelial layer and detached cell groups in the surrounding lamina propria showed moderate/strong VEGF expression. The strongest VEGFR-1 and VEGFR-2 expression was detected in the apical epithelial layer. Cells with aspect similar to bipolar neurons and/or their presumable precursors located in the middle and basal layers and the detaching/detached cells displayed a VEGFR-1 and VEGFR-2 reactivity similar to that of VEGF. The basal epithelial layer exhibited stronger staining for VEGFRs than for VEGF. Cells with morphology and VEGF/VEGFR expression similar to those of the detaching/detached cells were also detected in the middle and basal VNO epithelial layers. Double immunofluorescence using anti-PGP 9.5 antibodies demonstrated that most of the VEGF/VEGFR-immunoreactive cells were neuronal cells. Collectively, our findings suggest that during early fetal development the VEGF/VEGFR system might be involved in the presumptive VNO chemoreceptor activity and neuronal precursor migration.

Glycan diversity in the vomeronasal organ of the Korean roe deer, Capreolus pygargus: A lectin histochemical study.

Glycans in the epithelium play an important role in cell-to-cell communication and adhesion. No detailed evaluation of glycoconjugates in the vomeronasal organs (VNO) of the roe deer has been published previously. The aim of this study was to characterize glycan epitopes in the vomeronasal sensory epithelium (VSE) and non-sensory epithelium (VNSE) using lectin histochemistry. Glycan epitopes identified by lectin histochemistry were grouped as follows: N-acetylglucosamine (s-WGA, WGA, BSL-II, DSL, LEL, STL), mannose (Con A, LCA, PSA), galactose (RCA120, BSL-I, Jacalin, PNA, ECL), N-acetylgalactosamine (VVA, DBA, SBA, and SJA), fucose (UEA-I) and complex type N-glycan (PHA-E and PHA-L) groups. The free border of the VSE was positive for all 21 lectins, and 18 of the lectins (excluding DBA, SJA, and PHA-L) showed weak and/or moderate staining in the receptor cells. The supporting cells were weakly positive for 19 lectins (excluding PNA and SJA). Moreover, 17 lectins (excluding BSL-II, Jacalin, PNA, and SJA) were expressed in the basal cells. In the VNSE of roe deer, the free border showed staining for all 21 lectins examined. The ciliated cells were positive for 16 lectins (excluding BSL-II, DSL, PNA, VVA, and SJA). Furthermore, 15 lectins (excluding DSL, LEL, ECL, UEA-I, PHA-E, and PHA-L) were expressed in goblet cells. Twenty lectins (excluding SJA) were expressed in the acini of the vomeronasal glands. Collectively, both VSE and VNSE were rich in N-acetylglucosamine, mannose, galactose, N-acetylgalactosamine, fucose, and complex-type N-glycans, although the different cell types of the VSE and VNSE expressed different glycoconjugates of varying intensities, suggesting that these carbohydrate residues may be involved in odor perception as well as cell-to-cell communication in the VNO.

The rat vomeronasal organ is a vitamin D target.

We studied the expression of vitamin D receptor and of vitamin D binding protein in the rat vomeronasal organ. With immunofluorescence, in situ hybridization and with reverse transcriptase PCR we found both proteins in sensory as well as in non-sensory cells. Sensory neurons contained immunoreactivity for vitamin D3 receptor in nuclei, in portions of the cytoplasm, and in apical dendrites and their microvilli. Vitamin D binding protein was observed in sensory neuron axons and cytoplasm, mostly confined to dendrites. Colocalization appeared in the contact zone of supporting cells and sensory dendrites. Both proteins were also found in single ciliated cells within the non-sensory epithelium. Vitamin D binding protein was also localized in secretory vesicles in a portion of the vomeronasal glands. Our findings suggest that the rat vomeronasal organ is a vitamin D target.

Elevated Cytosolic Cl- Concentrations in Dendritic Knobs of Mouse Vomeronasal Sensory Neurons.

In rodents, the vomeronasal system controls social and sexual behavior. However, several mechanistic aspects of sensory signaling in the vomeronasal organ remain unclear. Here, we investigate the biophysical basis of a recently proposed vomeronasal signal transduction component-a Ca(2+)-activated Cl(-) current. As the physiological role of such a current is a direct function of the Cl(-) equilibrium potential, we determined the intracellular Cl(-) concentration in dendritic knobs of vomeronasal neurons. Quantitative fluorescence lifetime imaging of a Cl(-)-sensitive dye at the apical surface of the intact vomeronasal neuroepithelium revealed increased cytosolic Cl(-) levels in dendritic knobs, a substantially lower Cl(-) concentration in vomeronasal sustentacular cells, and an apparent Cl(-) gradient in vomeronasal neurons along their dendritic apicobasal axis. Together, our data provide a biophysical basis for sensory signal amplification in vomeronasal neuron microvilli by opening Ca(2+)-activated Cl(-) channels.

Immunohistochemical and lectin histochemical studies on the developing olfactory organs of fetal camel.

Little is known about the development of the olfactory organs of camel. In this study, prenatal development and neuronal differentiation of the vomeronasal organ (VNO) and the olfactory epithelium (OE) of the one-humped camel were studied by immunohistochemistry and lectin histochemistry. A neuronal marker, protein gene product (PGP) 9.5, but not a marker of fully differentiated olfactory receptor cells, olfactory marker protein, intensely labeled the olfactory receptor cells of the VNO and OE at 395 mm, 510 mm, and 530 mm fetal ages, indicating that the olfactory receptor cells are differentiated, but not fully matured both in the VNO and the OE. In 187 mm and 190 mm fetuses, PGP 9.5 yielded faint immunoreactive signals in the VNO, but not in the OE, although the presence of olfactory receptor cells were demonstrated in both tissues by intense WGA and LEL stainings. We conclude that the camel VNO and OE bear differentiated, but still immature receptor cells; in addition, the onset of neuronal differentiation seems to be somewhat earlier in the VNO than in the OE till half of the prenatal life.

Molecular evolutionary characterization of a V1R subfamily unique to strepsirrhine primates.

Vomeronasal receptor genes have frequently been invoked as integral to the establishment and maintenance of species boundaries among mammals due to the elaborate one-to-one correspondence between semiochemical signals and neuronal sensory inputs. Here, we report the most extensive sample of vomeronasal receptor class 1 (V1R) sequences ever generated for a diverse yet phylogenetically coherent group of mammals, the tooth-combed primates (suborder Strepsirrhini). Phylogenetic analysis confirms our intensive sampling from a single V1R subfamily, apparently unique to the strepsirrhine primates. We designate this subfamily as V1Rstrep. The subfamily retains extensive repertoires of gene copies that descend from an ancestral gene duplication that appears to have occurred prior to the diversification of all lemuriform primates excluding the basal genus Daubentonia (the aye-aye). We refer to the descendent clades as V1Rstrep-α and V1Rstrep-ß. Comparison of the two clades reveals different amino acid compositions corresponding to the predicted ligand-binding site and thus potentially to altered functional profiles between the two. In agreement with previous studies of the mouse lemur (genus, Microcebus), the majority of V1Rstrep gene copies appear to be intact and under strong positive selection, particularly within transmembrane regions. Finally, despite the surprisingly high number of gene copies identified in this study, it is nonetheless probable that V1R diversity remains underestimated in these nonmodel primates and that complete characterization will be limited until high-coverage assembled genomes are available.

The electrovomeronasogram: field potential recordings in the mouse vomeronasal organ.

Mammalian vomeronasal neurons (VSNs) located in the sensory epithelium of the vomeronasal organ (VNO) detect and transduce molecular cues emitted by other individuals and send this information to the olfactory forebrain. The initial steps in the detection of pheromones and other chemosignals by VSNs involve interaction of a ligand with a G protein-coupled receptor and downstream activation of the primary signal transduction cascade, which includes activation of ion channels located in microvilli and the dendritic tip of a VSN. The electrovomeronasogram (EVG) recording technique provides a sensitive means through which ligand-induced activation of populations of VSNs can be recorded from the epithelial surface using an intact, ex vivo preparation of the mouse VNO. We describe methodological aspects of this preparation and the EVG recording technique which, together with single-cell recordings, contributed significantly to our understanding of mammalian vomeronasal function, the identification of pheromonal ligands, and the analysis of mice with targeted deletions in specific signal transduction molecules such as Trpc2, Gαo, V1R, or V2R receptors.

Electrical recordings from the accessory olfactory bulb in VNO-AOB ex vivo preparations.

Electrical recordings from individual accessory olfactory bulb neurons allow exploration of the functional properties of this important pheromonal processing circuit. Several approaches to performing such recordings have been used. Here, we describe ex vivo methods that we have found useful for recording from accessory olfactory bulb neurons using simple extracellular glass electrodes.

Nasal passages of Göttingen minipigs from the neonatal period to young adult.

Histopathological examination of the nasal passages requires a standardized approach for recording lesion distribution patterns. Nasal diagrams provide guidance to map the lesions. Information on lesions exists for rodents, dogs, and monkeys, which all have been used in inhalation studies. Recently, minipigs have garnered interest as an inhalation model because minipigs resemble humans in many features of anatomy, physiology, and biochemistry and may be a good alternative to monkeys and dogs. The present work explored the microanatomy and histology of the nasal passages of Göttingen minipigs from postnatal day 1 until 6 months of age. Six nasal levels were selected, which allow examination of the squamous, transitional (nonciliated) and ciliated respiratory, and olfactory epithelia; the nasopharynx; and relevant structures such as the vomeronasal organ, olfactory bulb, and nasal/nasopharynx-associated lymphoid tissue.

Seasonal changes in the histochemical properties of the olfactory epithelium and vomeronasal organ in the Japanese striped snake, Elaphe quadrivirgata.

Seasonal changes in the histochemical properties of the vomeronasal and olfactory epithelia of the Japanese striped snake were examined in four seasons, viz. the reproductive, pre-hibernating, hibernating and post-hibernating seasons. In the vomeronasal and olfactory supporting cells, secretory granules were much more abundant in the hibernating season than in the other seasons. In the vomeronasal and olfactory receptor cells, the lipofuscin granules were much fewer in the post-hibernating season than in the other seasons. In histochemical studies with 21 lectins, several lectins stained the vomeronasal and olfactory epithelia (receptor cells, supporting cells and free border) more weakly in the hibernating season than in the reproductive season. However, all lectins stained both epithelia in the hibernating season after sialic acid removal in a similar manner as in the reproductive season after sialic acid removal. These lectin histochemical studies indicate that sialic acid residues in the vomeronasal and olfactory epithelia are more numerous in the hibernating season than in the reproductive season. The results suggest that during hibernation, the vomeronasal and olfactory receptor cells possibly undergo rapid cell turnover, and that during this time, the vomeronasal and olfactory epithelia are securely protected from pathogens by an innate immune defence system.

Properties of mouse vomeronasal receptor and assessment of its role in pheromone signalling.

Vomeronasal type 2 receptor (V2Rx) from Swiss mouse (Mus musculus (L.)) was analyzed by high-resolution ion-exchange chromatography, reversed-phase high-performance liquid chromatography (RP-HPLC), matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS), Ion Spray tandem mass spectrometry (MS/MS), 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and 1-aminoanthracene (1-AMA) fluorometric assay. Vomeronasal sensory neuronal cell bound proteins were resolved into major protein peaks. Several proteins were identified and subsequently purified as the V2Rx receptor on 10% SDS-PAGE with trace amounts of other protein bands. The molecular weight of the identified V2Rx was 109 kDa. MALDI-TOF and micro-sequencing experiments demonstrated that the identified V2Rx receptor shared considerable sequence similarity with vomeronasal receptor type 2 (NCBI Accession Number AB267725), which is a seven transmembrane peptide with 912 amino acid residues. The molecular characterization revealed that the N-terminus of the V2Rx receptor contained the 11GAEAAE16 domain involved in pheromone signalling. The biometric assay (octanamine-V2Rx binding) showed the identified V2Rx receptor and mouse sex pheromone to 2-octanamine (methyl heptyl) in a 1:1 ratio. Uptake of odourants determined in physiological condition showed enhanced V2Rx receptors as volatile hydrophobic pheromone receptors in the vomeronasal neuron of the Swiss mouse.

Lectin histochemical studies on the olfactory epithelium and vomeronasal organ in the Japanese striped snake, Elaphe quadrivirgata.

The olfactory epithelium and the vomeronasal organ of the Japanese striped snake were examined by lectin histochemistry. Of the 21 lectins used in the study, all lectins except succinylated-wheat germ agglutinin (s-WGA) showed similar binding patterns in the vomeronasal receptor cells and the olfactory receptor cells with varying intensities. The binding patterns of s-WGA varied among individuals in the vomeronasal and olfactory receptor cells, respectively. Four lectins, Bandeiraea simplicifolia lectin-II (BSL-II), Dolichos biflorus agglutinin (DBA), Sophora japonica agglutinin (SJA), and Erythrina cristagalli lectin (ECL) stained secretory granules and the organelles in the olfactory supporting cells and did not stain them in the vomeronasal supporting cells. These results suggest that the glycoconjugate moieties are similar in the vomeronasal and olfactory receptor cells of the Japanese striped snake.

Anatomical, immnunohistochemical and physiological characteristics of the vomeronasal vessels in cows and their possible role in vomeronasal reception.

The general morphology of the vomeronasal vessels in adult cows was studied following a classic protocol, including optical, confocal and ultrastructural approaches. This anatomical work was completed immunohistochemically. The vomeronasal organ in cows is well developed, and its vessels are considerable in size. This fact allowed some functional properties of the vomeronasal arteries to be evaluated and, for the first time, their isometric tension to be recorded. Our functional studies were in agreement with the immunohistochemistry, and both corroborated the morphological data on the similarity between the vomeronasal vessels and those of the typical erectile tissue. In consequence, the vasoconstriction and vasodilation of the vomeronasal vessels would facilitate an influx and outflow of fluids in the vomeronasal organ, that is to say, this organ in cows would be able to work as a pump mechanism to send chemical signals to the vomeronasal receptor neurones.

Receptores de feromonas de mamíferos: supervivencia y sexualidad / Mammalian pheromone receptors: survival and sexuality

En 1995 Catherine Dulac y Richard Axel publicaron la existencia de una nueva familia de genes que codificaban los posibles receptores de feromonas, pertenecientes a la amplia familia de los de siete hélices transmembranares y acoplados a proteínas G. Desde entonces se han clonado nuevos genes que han sido agrupados en dos familias, los receptores vomeronasales tipo 1 y 2, V1R y V2R, con diferente estructura y situados con diferente distribución en el órgano vomeronasal. La naturaleza química de las feromonas y de las proteínas que las asocian y transportan conocidas como lipocalinas es otro de los aspectos de los que se dispone de abundante información. Los mecanismos de transducción de la señal mediada por feromonas sobre los receptores V1R y V2R implican la activación de la fosfolipasa C tipo β2, PLCβ2, generando el fosfatidilinositol trifosfato y el diacilglicerol en la cara interna de la membrana neuronal. El diacilglicerol es un ligando endógeno, que permite la apertura del canal de la familia TRPC (Transient Receptor Potential Channel) denominado TRPC2 que se abre y deja pasar iones Ca2+ y Na+ al interior de la neurona sensorial, iniciando la despolarización de la membrana y originando el potencial de acción. La señal eléctrica es conducida al bulbo olfativo auxiliar por axones que llegan de modo disperso y establecen conexión con las células mitrales, las cuales envían sus prolongaciones hasta el sistema límbico y otras estructuras cerebrales, donde influencian o provocan las respuestas de supervivencia de la especie, entre ellas las de apareamiento y agresividad. Un aspecto relevante desde el punto de vista evolutivo es que en primates el gen TRPC2 es un pseudogen sin funcionalidad y por lo tanto el órgano vomeronasal es un vestigio carente de función. Recientes estudios indican que la captación de feromonas en primates se realiza a través del epitelio olfativo y el bulbo olfativo principal e incluso en otros mamíferos esta estructura parece mediar en algunas respuestas especie específicas

In 1995 Catherine Dulac and Richard Axel discovered a new gene family corresponding to the pheromone receptors. They were members of the seven transmembrane helix coupled to G proteins. Since then, new genes have been clonned and grouped according their sequence homology in two main families of vomeronasal receptors the V1R and the V2R. They exhibit different distribution pattern at the vomeronasal epithelium, where they are coupled to different G proteins. The chemical nature of the mammalian pheromones is very diverse and can associate with proteins called lipocalins to reach the vomeronasal organ. The transduction mechanisms of pheromone receptors, V1R and V2R, require respectively a Gi and a Go proteins, to further activate a phospholipase C, the PLCβ2. This enzyme hydrolyses the phosphatidyl inositol located at the plasma membrane originating phosphatidylinositol triphosphate and diacylglycerol. Diacylglycerol is an endogenous ligand that opens the TRPC2 channel (Transient Receptor Potential Channel), allowing the entrance of cations, mostly Ca2+ y Na+. The membrane depolarisation at the vomeronasal neuron originates the action potential that is sent to the accessory olfactory bulb by the axon, which in a different way as those from the main olfactory epithelia, do not organise the axonal prolongations and reach the mitral neurones in a disperse way, without forming a glomerular structure, afterwards the mitral cells send their axons to the limbic system and other cerebral structures related to aggressive behaviour and mating. It is relevant to underline that in monkeys from the old world and primates including humans, the vomeronasal organ is only a vestigial structure without function. The reason relies on the TRPC2 gene, which is a pseudo gene, without physiological function. Recent experimental approaches have demonstrated that the sensing of some pheromonal signals in these species, and also in mammals with a functional vomeronasal organ, can be carried out by the main olfactory epithelia through the main olfactory bulb. This structure being also connected to the hypothalamus, where neurones releasing LHRH can control sexual behaviour. These data confirm the broad possibilities of signalling through pheromones and that much effort is still required to fully understand their possibilities

Attenuation of the production of inositol 1,4,5-trisphosphate in the mouse vomeronasal organ by antibodies against the alphaq/11 subfamily of G-proteins.

The social and reproductive behaviors of most mammals are modulated by pheromones, which are perceived by the vomeronasal organ (VNO). Vomeronasal transduction in vertebrates is activated through G-protein-coupled receptors, which in turn leads to the generation of inositol 1,4,5-trisphosphate (IP(3)) and diacylglycerol (DAG) by the activity of phospholipase C. DAG has been shown to gate the transient receptor potential channel 2, whereas IP(3) may play a role in stimulating the release of calcium from the endoplasmic reticulum store. To investigate the role of the alpha subunits of G(q/11) in the transduction process, microvillar membranes from female mice VNO were preincubated with a selective C-terminal peptide antibody against Galpha(q/11) and then stimulated with adult male urine. Incubation of VNO membranes with antibodies against Galpha(q/11) blocked the production of IP(3) in a dose-dependent manner. We were also able to impair the production of IP(3) when we stimulated with 2-heptanone or 2,5-dimethylpyrazine in the presence of antibodies against the alpha subunit of G(q/11). 2-Heptanone is a known pheromone that has been linked to VIR receptors. Thus, our observations indicate that the alpha subunits of G(q/11) play a role in pheromonal signaling in the VNO.

Ultrastructural localization of alpha-galactose-containing glycoconjugates in the rat vomeronasal organ.

Binding sites of Griffonia simplicifolia I-B4 isolectin (GS-I-B4), which recognizes terminal alpha-galactose residues of glycoconjugates, were examined in the juxtaluminal region of the rat vomeronasal sensory epithelium and its associated glands of the vomeronasal organ, using a lectin cytochemical technique. Lowicryl K4M-embedded ultra-thin sections, which were treated successively with biotinylated GS-I-B4 and streptavidin-conjugated 10 nm colloidal gold particles, were observed under a transmission electron microscope. Colloidal gold particles, which reflect the presence of terminal alpha-galactose-containing glycoconjugates, were present in vomeronasal receptor neurons in the sensory epithelium and secretory granules of acinar cells of associated glands of the epithelium. Quantitative analysis demonstrated that the density of colloidal gold particles associated with sensory cell microvilli that projected from dendritic endings of vomeronasal neurons was considerably higher than that of microvilli that projected from neighboring sustentacular cells. The same was true for the apical cytoplasms of these cells just below the microvilli. These results suggest that of the sensory microvilli and dendritic endings contained a much larger amount of the alpha-galactose-containing glycoconjugates, compared with those in sustentacular microvilli. Further, biochemical analyses demonstrated several vomeronasal organ-specific glycoproteins with terminal alpha-galactose.

The vomeronasal organ of greater bushbabies (Otolemur spp.): species, sex, and age differences.

The present study examined interspecies, intersexual, and age-related changes in size of the vomeronasal neuroepithelium (VNNE) of two species of greater bushbabies (genus Otolemur, Infraorder Lorisiformes, Suborder Strepsirrhini). Tissue blocks containing the vomeronasal organs of nine O. crassicaudatus (8 adults, 1 neonate) and ten O. garnettii (9 adults, 1 neonate) were studied by means of serial paraffin sectioning and computer-based reconstruction of VNNE volume. In addition, the immunoreactivity of the VNNE to two neuronal markers, neuron-specific beta tubulin (BT) and olfactory marker protein (OMP) was compared between species, sexes, and ages. Results indicated that a clear VNNE is present at birth in both species, and OMP immunoreactivity was verified in O. garnettii at birth. Male and female adults of both species showed OMP-immunoreactive and BT-immunoreactive neurons in the VNNE. Immunohistochemical findings indicated that all males and the youngest females had the thickest VNNE, especially at the marginal junctions with the receptor-free epithelium. Results of a 2-way Analysis of Variance (ANOVA, species x sex) revealed no significant differences in VNNE length or volume between species, but O. crassicaudatus had significantly (p < 0.05) greater palatal length. Significant (p < 0.05) differences also were found between sexes in VNNE volume, but no significant differences in palatal length or VNNE length. The distribution of VNNE volume against age indicated that the sex differences were more pronounced in O. crassicaudatus than O. garnettii. For both species and sexes, distribution of VNNE volume against age suggested an age-related reduction in volume. These findings demonstrate postnatal plasticity in VNNE size in Otolemur that is reminiscent of that found for olfactory structures in some rodents. Bushbabies or other strepsirrhine primates may offer an opportunity for further understanding of behavioral correlates of VNNE postnatal plasticity, which may represent primitive functional characteristics of the order Primates.

Maturation of vomeronasal receptor neurons in vitro by coculture with accessory olfactory bulb neurons.

To analyze the mechanisms of perception and processing of pheromonal signals in vitro, we previously developed a new culture system for vomeronasal receptor neurons (VRNs), referred to as the vomeronasal pocket (VN pocket). However, very few VRNs were found to express the olfactory marker protein (OMP) and to have protruding microvilli in VN pockets, indicating that these VRNs are immature and that VN pockets are not appropriate for pheromonal recognition. To induce VRN maturation in VN pockets, we here attempted to coculture VN pockets with a VRN target-accessory olfactory bulb (AOB) neurons. At 3 weeks of coculture with AOB neurons, the number of OMP-immunopositive VRNs increased. By electron microscopy, the development of microvilli in VRNs was found to occur coincidentally with OMP expression in vitro. These results indicate that VRN maturation is induced by coculture with AOB neurons. The OMP expression of VRNs was induced not only by AOB neurons but also by neurons of other parts of the central nervous system (CNS). Thus, VRN maturation requires only CNS neurons. Since the maturation of VRNs was not induced in one-well separate cultures, the nonspecific induction of OMP expression by CNS neurons suggests the involvement of a direct contact effect with CNS in VRN maturation.

Divergent V1R repertoires in five species: Amplification in rodents, decimation in primates, and a surprisingly small repertoire in dogs.

The V1R gene family comprises one of two types of putative pheromone receptors expressed in the mammalian vomeronasal organ (VNO). We searched the most recent mouse, rat, dog, chimpanzee, and human genome sequence assemblies to compile a near-complete repertoire of V1R genes for each species. Dog, human, and chimpanzee have very few intact V1Rs (8, 2, and 0, respectively) compared to more than a hundred intact V1Rs in each of the rat (106) and mouse (165) genomes. We also provide the first description of the diversity of V1R pseudogenes in these species. We identify at least 165 pseudogenes in mouse, 110 in rat, 102 in chimpanzee, 115 in human, and 54 in dog. Primate and dog pseudogenes are distributed among almost all V1R subfamilies seen in rodents, indicating that the common ancestor of these species had a diverse V1R repertoire. We find that V1R genes were subject to strikingly different fates in different species and in different subfamilies. In rodents, some subfamilies remained relatively stable or underwent roughly equivalent expansion in mouse and rat; other subfamilies expanded in one species but not the other. The small number of intact V1Rs in the dog genome is unexpected given the presumption that dogs, like rodents, have a functional VNO, and a complex system of pheromone-based behaviors. We identify an intact transient receptor potential channel 2beta in the dog genome, consistent with a functional VNO in dogs. The diminished V1R repertoire in dogs raises questions about the relative contributions of V1Rs versus other candidate pheromone receptor genes in the establishment of complex pheromone systems in mammals.