Age-dependent decrease in glomeruli and receptor cells containing α1-2 fucose glycan in the mouse main olfactory system but not in the vomeronasal system.

Receptor cells of the olfactory epithelium (OE) and vomeronasal organ (VNO) project axons to glomeruli in the main olfactory bulb (MOB) and accessory olfactory bulb (AOB), respectively and undergo continuous turnover throughout life. Alpha1-2 fucose (α1-2Fuc) glycan mediates neurite outgrowth and synaptic plasticity and plays important roles in the formation of the olfactory system during development. We previously confirmed the localization of α1-2Fuc glycan in the olfactory system of 3- to 4-month-old mice but whether such localization persists throughout life remains unknown. Here, the MOB, AOB, OE and VNO of 1-, 3- and 8-month-old mice were histochemically examined using Ulex europaeus agglutinin-I (UEA-I) that specifically binds to α1-2Fuc glycan. Binding sites for UEA-I in the MOB were similar among all age groups but the ratio of UEA-I-positive glomeruli significantly decreased with aging. The frequency of UEA-I-positive receptor cells in the OE of the two older groups was also significantly lower than that of 1-month-old mice. On the other hand, UEA-I binding in the AOB and VNO did not significantly differ among all three groups. These findings suggest that the primary pathway of the main olfactory system requires the role of α1-2Fuc glycan in young mice rather than old mice, while the vomeronasal pathway equally requires this glycan in both young and old mice.

Structural and ultrastructural studies on the developing vomeronasal sensory epithelium in the grass snake Natrix natrix (Squamata: Colubroidea).

The sensory olfactory epithelium and the vomeronasal sensory epithelium (VSE) are characterized by continuous turnover of the receptor cells during postnatal life and are capable of regeneration after injury. The VSE, like the entire vomeronasal organ, is generally well developed in squamates and is crucial for detection of pheromones and prey odors. Despite the numerous studies on embryonic development of the VSE in squamates, especially in snakes, an ultrastructural analysis, as far as we know, has never been performed. Therefore, we investigated the embryology of the VSE of the grass snake (Natrix natrix) using electron microscopy (SEM and TEM) and light microscopy. As was shown for adult snakes, the hypertrophied ophidian VSE may provide great resolution of changes in neuron morphology located at various epithelial levels. The results of this study suggest that different populations of stem/progenitor cells occur at the base of the ophidian VSE during embryonic development. One of them may be radial glia-like cells, described previously in mouse. The various structure and ultrastructure of neurons located at different parts of the VSE provide evidence for neuronal maturation and aging. Based on these results, a few nonmutually exclusive hypotheses explaining the formation of the peculiar columnar organization of the VSE in snakes were proposed.

Unique blood vasculature and innervation in the cavernous tissue of murine vomeronasal organs.

The vomeronasal organ (VNO) is an accessory olfactory device related to reproductive behavior. The soft tissue of the tubular organ is composed of sensory/non-sensory epithelia and a highly developed vasculature, which in the latter the dilation and contraction of blood vessels are thought to contribute to pumping in and out luminal fluid or air, like penile erectile tissue. The present histological observation of the murine VNO revealed a more complicated vasculature than previously evaluated ones with large differences along the rostro-caudal axis. An immunohistochemical study for vasoactive substances displayed extremely dense innervation by cholinergic nerves containing nitric oxide synthase and VIP/PHI in the thick smooth muscle layer surrounding venous sinuses at light and electron microscopic levels. Furthermore, the differential distribution of cholinergic nerves and adrenergic nerves may provide a novel insight into the pumping mechanism of VNO.

Vomeronasal neurons promote synaptic formation on dendritic spines but not dendritic shafts in primary culture of accessory olfactory bulb neurons.

To investigate the morphological changes of accessory olfactory bulb (AOB) neurons arising from pheromonal signals, a coculture system of AOB neurons and vomeronasal (VN) neurons had been established. Our previous study indicates that under coculture condition, the density of dendritic spines of an AOB neuron is less and the individual spine-head volume is larger than those under monoculture condition. In this study, to determine whether these differences in the dendrites of AOB neurons reflect the differences in synapse formation and synaptic properties, we observed these cultured cells by electron microscopy. Various synapses were observed under each culture condition. Synapses were classified on the basis of their postsynaptic structure and the size of postsynaptic density (PSD) was measured. Under the coculture condition with VN neurons, synapses on dendritic spines, which formed between AOB neurons, were observed frequently. In contrast, many synapses were formed on dendritic shafts under monoculture condition. The PSD of asymmetrical synapses on the spines under coculture condition was larger than that under monoculture condition. Moreover, some dendrodendritic reciprocal synapses were found only in coculture. We confirmed synapse formation between VN axons and AOB dendrites by immunohistochemical electron microscopy; thus, the characteristics of synapses between AOB neurons are considered to be modified by the synaptic contacts with VN axons.

Localization of N-acyl phosphatidylethanolamine phospholipase D (NAPE-PLD) expression in mouse brain: A new perspective on N-acylethanolamines as neural signaling molecules.

N-acylethanolamines (NAEs) are membrane-derived lipids that are utilized as signaling molecules in the nervous system (e.g., the endocannabinoid anandamide). An N-acyl phosphatidylethanolamine phospholipase D (NAPE-PLD) that catalyzes formation of NAEs was recently identified as a member of the zinc metallohydrolase family of enzymes. NAPE-PLD(-/-) mice have greatly reduced brain levels of long-chain saturated NAEs but wild-type levels of polyunsaturated NAEs (e.g., anandamide), suggesting an important role for NAPE-PLD in the biosynthesis of at least a subset of endogenous NAEs in the mammalian nervous system. To provide a neuroanatomical basis for investigation of NAPE-PLD function, here we have analyzed expression of NAPE-PLD in the mouse brain using mRNA in situ hybridization and immunocytochemistry. NAPE-PLD(-/-) mice were utilized to establish the specificity of probes/antibodies used. The most striking feature of NAPE-PLD expression in the brain was in the dentate gyrus, where a strong mRNA signal was detected in granule cells. Accordingly, immunocytochemical analysis revealed intense NAPE-PLD immunoreactivity in the axons of granule cells (mossy fibers). Intense NAPE-PLD immunoreactivity was also detected in axons of the vomeronasal nerve that project to the accessory olfactory bulb. NAPE-PLD expression was detected in other brain regions (e.g., hippocampus, cortex, thalamus, hypothalamus), but the intensity of immunostaining was weaker than in mossy fibers. Collectively, the data obtained indicate that NAPE-PLD is expressed by specific populations of neurons in the brain and targeted to axonal processes. We suggest that NAEs generated by NAPE-PLD in axons may act as anterograde synaptic signaling molecules that regulate the activity of postsynaptic neurons.

Distribution of olfactory marker protein on a tissue section of vomeronasal organ measured by AFM.

Distribution of olfactory marker protein (OMP) on a tissue section of vomeronasal organ (VNO) was successfully measured by atomic force microscopy (AFM). Anti-OMP antibodies were covalently crosslinked with the tip of the AFM and were used as a probe to observe the distribution of OMP on a tissue section. First, force measurements were performed using a glass surface on which OMP was covalently immobilized to verify the success of tip modification. Clear differences of interaction forces were observed between a specific pair and the control experiments, indicating that the tip preparation succeeded. Next, distributions of OMP on the tissue section were observed by AFM and were compared with immunohistochemical observations. For large scale observation, a microbead was used as a probe in the AFM measurements. The results of the AFM measurements were well overlapped with that of immunohistochemistry, confirming the reliability of our method. A mapping of the AFM measurement with high resolution was also successfully obtained, which showed an advantage of the application of the AFM measurement in analysis of proteins on the tissue section.

Diversity of the vomeronasal system in mammals: the singularities of the sheep model.

The enormous morphological diversity and heterogeneity of the vomeronasal system (VNS) in mammals--as well as its complete absence in some cases--complicates the extrapolation of data from one species to another, making any physiological and functional conclusions valid for the whole Mammalian Class difficult and risky to draw. Some highly-evolved macrosmatic mammals, like sheep, have been previously used in interesting behavioral studies concerning the main and accessory olfactory systems. However, in this species, certain crucial morphological peculiarities have not until now been considered. Following histological, histochemical and immunohistochemical procedures, we have studied the vomeronasal organ (VNO) and the accessory olfactory bulb (AOB) of adult sheep. We have determined: (1) that all structures which classically define the VNO in mammals are present and well developed, providing the morphological basis for functional activity. (2) that, conversely, there is only a scant population of scattered mitral/tufted cells. One morphological consequence of both details is that the strata of the AOB in adult sheep are not as sharply defined as in other species; moreover, the small number of the mitral/tufted cells in the AOB may imply that the VNS of adult sheep is not capable of functioning in the way a well-developed VNS does in other species. (3) the zone to zone projection from the apical and basal sensory epithelium of the VNO to the anterior and posterior part of the AOB, respectively, typical in rodents, lagomorphs and marsupials, is not present in adult sheep.

Distribution of olfactory marker protein in the rat vomeronasal organ.

Olfactory marker protein (OMP) may act as a modulator within the olfactory signal-transduction cascade. It has also been shown to have some importance in development of olfactory sensory organs. Here we used high resolution immunocytochemistry to localize OMP in the rat vomeronasal organ (VNO). Immunofluorescence for OMP was abundant in cilia and in apical dendrites of sensory cells, mostly associated with intraepithelial capillaries. Perikarya were stained to a lesser extent while intense OMP immunoreactivity was seen in axons of sensory neurons. Single cells within the non-sensory portion of the VNO exhibited intense OMP immunofluorescence in apical cilia and weak cytoplasmic staining. Some of the exocrine cells in the vomeronasal glands contained OMP positive secretory granules. Electron microscopy revealed that non-sensory ciliated cells had short rod like kinocilia as well as microvilli. These cells contained secretory vesicles. Their basal portion was in close apposition to nerve endings. Our findings suggest that the sensory part of the VNO contains OMP positive sensory neurons and that the non-sensory epithelium may contain secondary sensory cells. In addition OMP may be liberated from secretory glands into vomeronasal secretions.

Histomorphology of vomeronasal organ in Scincella tsinlingensis / Histomorfología del órgano vomeronasal en Scincella tsinlingensis

Due to the great change in the morphology of squamate vomeronasal organ (VNO), the histomorphology characteristics of VNO in Scincella tsinlingensis were studied by light and electronic microscopy. The results indicated that the VNO of S. tsinlingensis was located at the base of nasal cavity and consisted of a mushroom body situated anteroventrally and a sensory epithelium (SE) situated dorsocaudally. SE was composed of supporting cells, receptor cells and basal cells, and the supporting cells contained secretory granules near the surface membrane. Most of receptor cells were irregular in shape with long cytoplasmic extensions and characterized by microtubules, vesicles, and mitochondria. The basal cells with long cytoplasmic extensions were also irregular in shape and appeared a greater electron density than others. The thick nerve bundles were found on the dorsomedial area of VNO, and the surface of mushroom body was non-sensory epithelium consisting of ciliated and basal cells, without goblet cells. Epithelial cells were arranged in irregular, with many cilia and microvilli distributed on its free surface. Cells on the basal layer were irregularly circular in shape and arranged sparsely. Taken together, the results indicated that the fine structure of VNO in S. tsinlingensis was similar to other species from scincomorphs.

Debido al gran cambio en la morfología del órgano vomeronasal (OVN), se estudiaron las características histomorfológicas en la Scincella tsinlingensis por microscopías de luz y electrónica. Los resultados indicaron que el OVN de S. tsinlingensis se localizaba en la base de la cavidad nasal y consistía en un cuerpo como hongo situado anteroventralmente y un epitelio sensorial (ES) situado dorso caudamente. El ES estaba compuesto de células de soporte, células receptoras y células basales, y las células de soporte contenían gránulos secretores cerca de la membrana superficial. En gran parte de la mayoría de las células receptoras se observó una forma irregular con largas extensiones citoplasmáticas, caracterizadas por microtúbulos, vesículas y mitocondrias. Las células basales con extensiones citoplasmáticas también tenían forma irregular y algunas parecían tener una mayor densidad de electrones. Los haces gruesos nerviosos se encontraron en el área dorsomedial del OVN, la superficie del cuerpo de estaba compuesto de epitelio no sensorial y consistía de células ciliadas y basales, sin células caliciformes. Las células epiteliales estaban dispuestas de manera irregular, con muchos cilios y microvellosidades distribuidas en su superficie libre. Las células en la capa basal eran escasas y de forma circular irregular. Tomados en conjunto, los resultados indicaron que la estructura fina del OVN en S. tsinlingensis era similar a otras especies de scincomorpha.

Expression of vomeronasal receptor genes in Xenopus laevis.

In the course of evolution, the vomeronasal organ (VNO) first appeared in amphibians. To understand the relationship between the VNO and the vomeronasal receptors, we isolated and analyzed the expression of the vomeronasal receptor genes of Xenopus laevis. We identified genes of the Xenopus V2R receptor family, which are predominantly expressed throughout the sensory epithelium of the VNO. The G-protein Go, which is coexpressed with V2Rs in the rodent VNO, was also extensively expressed throughout the vomeronasal sensory epithelium. These results strongly suggest that the V2Rs and Go are coexpressed in the vomeronasal receptor cells. The predominant expression of the Xenopus V2R families and the coexpression of the V2Rs and Go imply that V2Rs play important roles in the sensory transduction of Xenopus VNO. We found that these receptors were expressed not only in the VNO, but also in the posterolateral epithelial area of the principal cavity (PLPC). Electron microscopic study revealed that the epithelium of the PLPC is more like that of the VNO than that of the principal and the middle cavity. These results suggest that in adult Xenopus the V2Rs analyzed so far are predominantly expressed in the vomeronasal and vomeronasal-like epithelium. The analysis of V2R expression in Xenopus larvae demonstrates that V2Rs are predominantly expressed in the VNO even before metamorphosis.

Association of ecto-5'-nucleotidase with specific cell types in the adult and developing rat olfactory organ.

A unique feature of the olfactory epithelium is its ability to give rise to new sensory neurons throughout life and also following injury. Cells at the basal side of the epithelium serve as neurogenic progenitor cells. The enzyme ecto-5'-nucleotidase is expressed at the surface of developing nerve cells and is regarded as a marker of neural development. To study the expression pattern of the enzyme, we analyzed its distribution in the adult and developing rat olfactory organ. Labeling is restricted to specific cell types and varies between the epithelia investigated. At the basal side of the olfactory epithelium, activity of 5'-nucleotidase is associated specifically with the dark/horizontal basal cells. Neither the light/globose basal cells, which are the immediate precursors of the sensory receptor cells, nor subsets of potentially immature olfactory receptor cells are labeled. On the other hand, microvillar cells dispersed at the lumenal side of the epithelium contain 5'-nucleotidase activity. The enzyme is also present at the inner lining of the ducts of Bowman's glands as they traverse the epithelium. Within the respiratory epithelium, activity of 5'-nucleotidase is associated with basal cells as well as with the epithelial surface. During development, 5'-nucleotidase is initially limited to the respiratory epithelium, including its basal cells. Dark/horizontal basal cells of the olfactory epithelium, which are positive for 5'-nucleotidase, first appear at the border of the respiratory epithelium, suggesting that they might originate from immigrating basal cells of the respiratory epithelium. Within the vomeronasal organ, labeling is largely restricted to the receptor-free epithelium. Although the functional role of 5'-nucleotidase in the olfactory system needs to be further defined, the distribution of the enzyme can be used successfully as a marker for defined cell types.

Ultrastructural localization of G-proteins and the channel protein TRP2 to microvilli of rat vomeronasal receptor cells.

Microvilli of vomeronasal organ (VNO) sensory epithelium receptor cells project into the VNO lumen. This lumen is continuous with the outside environment. Therefore, the microvilli are believed to be the subcellular sites of VNO receptor cells that interact with incoming VNO-targeted odors, including pheromones. Candidate molecules, which are implicated in VNO signaling cascades, are shown to be present in VNO receptor cells. However, ultrastructural evidence that such molecules are localized within the microvilli is sparse. The present study provides firm evidence that immunoreactivity for several candidate VNO signaling molecules, notably the G-protein subunits G(ialpha2) and G(oalpha), and the transient receptor potential channel 2 (TRP2), is localized prominently and selectively in VNO receptor cell microvilli. Although G(ialpha2) and G(oalpha) are localized separately in the microvilli of two cell types that are otherwise indistinguishable in their apical and microvillar morphology, the microvilli of both cell types are TRP2(+). VNO topographical distinctions were also apparent. Centrally within the VNO sensory epithelium, the numbers of receptor cells with G(ialpha2)(+) and G(oalpha)(+) microvilli were equal. However, near the sensory/non-sensory border, cells with G(ialpha2)(+) microvilli predominated. Scattered ciliated cells in this transition zone resembled neither VNO nor main olfactory organ (MO) receptor cells and may represent the same ciliated cells as those found in the non-sensory part of the VNO. Thus, this study shows that, analogous to the cilia of MO receptor cells, microvilli of VNO receptor cells are enriched selectively in proteins involved putatively in signal transduction. This provides important support for the role of these molecules in VNO signaling.

Supporting tissue and vasculature of the mammalian vomeronasal organ: the rat as a model.

The blood supply and osseocartilaginous support structures of the vomeronasal organ of the rat were studied. The study focused on adults, though 3- to 18-day-old animals were also examined. The techniques used included dissection and microdissection, injection of the vascular system with Araldite or with Indian ink in agar or gelatine, conventional histology, and scanning and transmission electron microscopy. The results indicated that blood reaches the vomeronasal organ via a branch of the sphenopalatine artery, and drains into an associated vein. Within the organ, one vein stood out by virtue of its size; this vein is accompanied by lesser veins, together with arterioles, capillaries, and lymphatic vessels. Connective tissue was readily apparent, though its distribution was heterogeneous. Analysis of series of transverse sections indicates that, in adults, the capsule that encases the vomeronasal organ is bony; in younger animals, the capsule is bony externally and cartilaginous internally; in very young animals, the capsule is entirely cartilaginous. However, it was noted that the change from cartilage to bone was due not to ossification of the existing cartilage, but to physical displacement of that cartilage by an extension of the vomer and incisive bones. Taken together, these results confirm the importance of considering the morphology of the vomeronasal organ as a whole, since there are major changes from rostral to caudal ends. Secondly, our findings regarding blood supply and the nature of the capsule support the view that the vomeronasal organ acts as a pump.

Vomeronasal organ and its associated structures in the opossum Monodelphis domestica.

The opossum Monodelphis domestica possesses a well-developed vomeronasal system. Uptake of chemical stimuli into the vomeronasal organ (VNO) involves a stereotypical "nuzzling" behavior. In the present study, ten animals were examined by light and electron microscopy. The peripheral oro-nasal structures that apparently enhance access of solutes into the VNO include: (1) two lateral grooves of the ventral rhinarium and a network of channels leading into them, (2) dental gap adjacent to the grooves, (3) butterfly-shaped incisive papilla, and (4) unique bristle/cup-shaped filiform papillae on the tongue. The longitudinal axis of the vomeronasal complex is composed of the VNO proper at its rostral end, and an extensive compound serous gland at its caudal end with a distinct transition zone in between. The transition zone is characterized by the following features: merging of the main excretory duct of the large vomeronasal gland with the VNO lumen and drainage of auxiliary glandular clusters into the lumen, irregularities in the sensory epithelium ("rosette" appearance), and the ending of the cartilaginous support surrounding the VNO. Multiple elongated bundles of smooth muscle are positioned between the sensory epithelium and the cartilaginous capsule and more caudally are intertwined with the glandular parenchyma. These bundles become more numerous at the transition zone. Contraction and extension of these muscles may function to enhance the flow of solutes and glandular secretion within the lumen. Two extensive venous sinuses are associated with the opossum VNO complex: the internal vein bordering the sensory epithelium at its rostral end, and the external vein alongside the nonsensory epithelium. This arrangement suggests a unique dual pumping mechanism.

Development of the vomeronasal receptor epithelium and the accessory olfactory bulb in sheep.

The morphological development of the vomeronasal organ (VNO) and accessory olfactory bulb (AOB) of the sheep from anlage to birth were studied by classical and histochemical methods using embryos and fetuses obtained from an abattoir with ages estimated from crown-to-rump length. Both VNO and AOB developed in a biologically logical sequence and completed their morphological development around day 98, at entry into the last third of the gestation period. A lectin with specificity for oligomeric N-acetylglucosamine labeled the sensory epithelium of the VNO, the vomeronasal nerves, and the nervous and glomerular layers of the AOB before birth. These results suggest that the vomeronasal system, which is well developed and functional in adult sheep, may be able to function at or even before birth in these animals (whereas in rodents, for example, this is precluded by the AOB not completing its development until after birth).

Vomeronasal organ in bats and primates: extremes of structural variability and its phylogenetic implications.

The mere appearance of a tubular, epithelially-covered, bilateral structure, no matter how minuscule, on the anteroventral nasal septum of tetrapods, is generally called the vomeronasal organ (of Jacobson). However, considering the functionality of this chemosensory structure, the presence of a non-cilated (microvillar) neuroepithelium (and not just any odd type of epithelium) encased in a variously shaped vomeronasal cartilage, along with vomeronasal nerve bundles and above all an accessory olfactory bulb connected to the limbic vomeronasal amygdala, are the absolute essential neurostructural characteristics and anatomic requirement for a functional VNO and the accessory olfactory system in any tetrapod. The distribution of the vomeronasal organ is reported here in two mammalian orders: Chiroptera and Primates. An impressive data pool on the vomeronasal organ of bats is now available, pointing to the fact that at this time bats may be the only group in which this organ system is extremely variable, ranging from total absence (even in the embryo) to spectacular development with numerous intervening stages in different chiropteran species. Of the eighteen bat families, only one family of New World leaf-nosed bats, family Phyllostomidae, exhibits functional vomeronasal organs. The vespertilionid bat Miniopterus, and the mormoopid bat Pteronotus, present exceptions to this rule. Among Primates, very few species have been rigorously studied. As a result, developmental variability of the vomeronasal organ is almost unknown; either the vomeronasal organ is well developed (such as in New World monkeys) or absent (as in Old World monkeys and great apes) in the adult. The concept whether adult humans or embryonic and fetal forms are endowed with this so-called sixth sense, is a controversial one and is under intense study in our laboratory and by others. The general phylogenetic implications based on our cladistic analysis of bats are that the vomeronasal organ complex has evolved several times. Among the prosimians and platyrrhine primates, the organ is well developed, although to a varying degree. Among catarrhine primates, its loss has occurred only once, as it is generally absent in the adult forms.

Fetal development of vomeronasal system in the goat.

Our previous study morphologically revealed that the adult goat vomeronasal (VN) system was different from the rodent and opossum one, and at least two types of VN systems exist in mammals. However, it remains unknown whether the developments in both types of VN systems are ontogenetically distinct and when the goat VN system is established. In this study, we morphologically observed the fetal development of the goat accessory olfactory bulb (AOB) and VN neuron. In the fetus, Gi2-expressing VN terminals terminated at glomeruli throughout the AOB, and no immunoreactivities for Go were detected in the nerve terminals reaching into AOB. The layer structure of AOB rapidly developed in the latter half of gestation. In the VN organ (VNO), at the middle stage of gestation, the dendritic processes of VN neuron were exposed in the VN lumen, and scattered and thin microvilli existed on the protrusion of the VN neuron. In the apical part of dendritic processes, no clear vesicle existed. However, the immunohistochemistry of an olfactory marker protein (OMP) revealed that a few VN neurons with OMP exist in VN sensory epithelium (VSE) before birth, although marked immunoreactivities were detected in adult VSE. Fetal VN neurons appeared to be underdeveloped. These results suggest that the goat VN system is ontogenetically distinct from the rodent and opossum VN systems, and is underdeveloped before birth. The goat VN system will develop and mature during the early postnatal period similar to the rodent and opossum VN systems.

Surface changes in the rat vomeronasal epithelium during degeneration and regeneration of sensory receptor cells.

To investigate cell turnover in the vomeronasal epithelium we used electron microscopy to obtain quantitative measurements of changes observed at the surface of the sensory epithelium. Receptor cell degeneration was induced by sensory nerve transection and animals were examined at postoperative recovery times of 2, 4, 6, 10, 15, 35 and 60 days. We measured the number and density of receptor and supporting cells, and membrane length at the surface of the sensory epithelium. The number of receptor cells rapidly decreased during the degeneration period, reaching a minimum at 6 days. After 15 days of recovery the number and density of receptor cells returned to control levels. The surface membrane length for regenerated receptor cells was similar to that of controls, however the morphological appearance was characteristic of immature cells. In contrast to the receptor cells, the number and density of supporting cells did not change during degeneration and regeneration. However, there was a significant increase in the length of supporting cell-surface membranes. These results suggest that during receptor cell degeneration, supporting cell membranes compensate for the loss of receptor cells by expanding their surface membrane length to help to maintain the continuity of the epithelial surface. Thus, an important role of vomeronasal supporting cells may be to maintain the structural integrity of the epithelium during turnover of the receptor cell population.

Early development of the olfactory organ in sturgeons of the genus Acipenser: a comparative and electron microscopic study.

Formation and morphology of the olfactory organ of vertebrates has been intensely studied in some taxa for more than a century. As a functionally important and complex sensory organ, its ontogenetic development has often been a matter of debate on higher-level craniate evolution. However, sufficient knowledge of structure and development of the olfactory organ in the crucial taxa needed for a serious phylogenetic reasoning is generally not available. This study aims at this essential primary data source, the detailed structure, morphogenesis, and character definition of the olfactory organ in more basal clades of jawed vertebrates (Gnathostomata). Sturgeon fishes (Acipenseriformes) as recent basal actinopterygians are expected to provide insight into archaic characters and character combinations in bony fishes. Thus, the development of the olfactory placodes of the sterlet, Acipenser ruthenus, and the Siberian sturgeon, Acipenser baerii, was followed histologically, by semi-thin serial sections, and by scanning and transmission electron microscopy. Except for the timing, virtually no differences were observed between the two species. The olfactory placodes become two-layered early in embryonic development. Both the superficial epidermal and the subepidermal layer can easily be distinguished and their development followed by ultrastructural properties. There are three different types of receptor cells: ciliated, microvillous, and crypt cells. The development of the ciliated and the less abundant microvillous receptor cells from the subepidermal layer of the placode is demonstrated. The non-sensory cells of the differentiated olfactory epithelium, i.e. ciliated non-sensory cells and supporting cells, exclusively derive from the superficial epidermal layer. In this respect, acipenserids clearly demonstrate close resemblance to the morphogenetic process found in the tetrapod Xenopus (Anura). The only other adequately described mode found in the actinopterygian zebrafish ( Danio rerio), is considered a derived character. In this case, all cells of the differentiated olfactory epithelium derive from one placodal cell layer. The mode of formation of the nasal sac and its ventilatory openings found in the acipenserids examined here, represents a widespread and probably a plesiomorphic condition of osteognathostomes. In both species, differentiation of the basic cellular composition of the olfactory epithelium is far advanced at the time of onset of extrinsic feeding.

The structure of the nasal chemosensory system in squamate reptiles. 2. Lubricatory capacity of the vomeronasal organ.

The vomeronasal organ is a poorly understood accessory olfactory organ, present in many tetrapods. In mammals, amphibians and lepidosaurian reptiles, it is an encapsulated structure with a central, fluid-filled lumen. The morphology of the lubricatory system of the vomeronasal organ (the source of this fluid) varies among classes, being either intrinsic (mammalian and caecilian amphibian vomeronasal glands) or extrinsic (anuran and urodele nasal glands). In the few squamate reptiles thus far examined, there are no submucosal vomeronasal glands. In this study, we examined the vomeronasal organs of several species of Australian squamates using histological, histochemical and ultrastructural techniques, with the goal of determining the morphology of the lubricatory system in the vomeronasal organ. Histochemically, the fluid within the vomeronasal organ of all squamates is mucoserous, though it is uncertain whether mucous and serous constituents constitute separate components. The vomeronasal organ produces few secretory granules intrinsically, implying an extrinsic source for the luminal fluid. Of three possible candidates, the Harderian gland is the most likely extrinsic source of this secretion.