The mammalian vomeronasal organ (VNO) represents the site for peripheral receptors of the vomeronasal (accessory olfactory) system. Situated at the anterior base of the nasal septum (Fig. 1), and within a cartilaginous or osseous capsule, the VNO receives nonvolatile chemical stimuli from the nasal cavity, from the oral cavity, or both (Wysocki and Meredith, 1987). A link between these stimuli and sociosexual behaviors has been established in certain species of mammals (e.g., various rodents; Wysocki and Meredith, 1987), including some primates (Barret et al., 1993; Aujard, 1997). Chemical stimuli received by the mammalian VNO are most often considered pheromones, which are chemical signals between members of the same species that convey behavioral or neuroendocrine information (see Meredith, 2001, for a discussion of the definition of pheromones). Although much literature has focused on the importance of the VNO to pheromone reception, it should be understood that the VNO may not be limited to this function and that the olfactory system also can detect pheromones (Kelliher et al., 2001). The most voluminous literature on primate VNO function has accumulated on adult humans (Moran et al., 1991; Berliner et al., 1996; Jahnke and Merker, 1998; Monti-Bloch et al., 1998; Grosser et al., 2000). Ironically, the human VNO has often been considered absent (Crosby and Humphrey, 1939) or vestigial (Johnson et al., 1985) in adults, although a comparative basis for such statements is far from complete.
Synchronization of menstrual cycles provides the most convincing evidence for human pheromonal communication (Jacob and McClintock, 2000), but there are no data linking this phenomena to the VNO. The human VNO has been described to mediate changes in mood or physiology (e.g., body temperature; Monti-Bloch et al., 1998; Grosser et al., 2000). This mediation may not qualify as pheromonal communication (Meredith, 2001). Functionality of the VNO in nonhuman primates is far more clear. For example, experimental removal of the VNO or lesions of the vomeronasal nerves have reduced male-male aggression, anogenital investigations, or mounts in mouse lemurs (Aujard, 1997) as reported for other mammals (Wysocki and Lepri, 1991).
The primate VNO is traditionally understood to be functional in New World monkeys and strepsirhine primates (see Table 1 for primate taxa) but absent or vestigial in catarrhine primates, at least postnatally (Schilling, 1970; Hunter et al., 1984; Ankel-Simons, 2000; Maier, 1997, 2000). This view correlates with data that indicate the accessory olfactory bulb (AOB), the central connection of the vomeronasal system, is absent in catarrhines (Crosby and Humphrey, 1939; Stephan et al., 1981; Meisami and Bhatnagar, 1998). Among primates, the AOB is best developed in strepsirhines, followed by New World monkeys (Stephan et al., 1981), whereas in catarrhine primates, it may be present only transiently during fetal development, as noted for humans (Humphrey, 1940). AOB size differences seem to relate strongly to VNO morphology. For example, strepsirhines such as Microcebus murinus have proportionately very large AOBs (Stephan et al., 1981), as well as VNOs with receptor-dense sensory epithelia (SE) (Schilling, 1970). New World monkeys have much variation in both AOB size and VNO epithelial morphology (see below). Unfortunately, the correlation between VNO and AOB size has not been studied in primates. A positive correlation is indeed suggested if one compares the so-called “microsmatic” and “macrosmatic” taxa, but this relationship is not perfect—most notably because adult humans possess a vomeronasal organ (Johnson et al., 1985; Smith et al., 1998; Trotier et al., 2000; Bhatnagar and Smith, 2001) but no trace of the AOB (Bhatnagar et al., 1987). How can the human VNO persist without a central connection? This question has been debated for more than 60 years, and may be pertinent to other primates as well.
Modified after Nowak, 1999; terms from text in bold).
Tarsius spp. are still variously classified within the suborder Haplorhini, with humans, apes, and New World and Old World monkeys, or in the suborder Prosimii, with lemurs and lorises (Ankel-Simons, 2000).
Haplorhine primates may be divided into New World taxa (platyrrhines—New World monkeys) and Old World taxa (catarrhines—Old World monkeys, apes, and humans).
From a phylogenetic standpoint, the existence of a VNO is a primitive trait among primates (Maier, 1980). The best developed VNOs seem to exist in most strepsirhine species. The VNO of mouse lemurs provides an excellent example (Fig. 1C). They are tubular epithelial structures with a central lumen. In all strepsirhines and New World Monkeys examined thus far, a dual communication has been described between the VNO lumen and the environment (Fig. 1A–D). That is, the duct leading to the VNO (vomeronasal duct) joins the nasopalatine duct of each side, which runs obliquely from the nasal cavity to the oral cavity (Hunter et al., 1984). Thus, chemical stimuli could potentially be accessed from either cavity in such primates.
In coronal section (Fig. 1H,I), the “chemosensory” VNO (see Table 2 for terminology) generally has a thin, laterally situated, nonsensory epithelium (receptor-free epithelium; Breipohol et al., 1979) and a markedly thicker SE on its medioventral border. These two distinct epithelial types enclose the lumen, which provides the environment in which chemical stimuli are accessed by the microvillar processes of VNO receptors. The SE consists of three cell types: basal cells that provide a stem cell population, bipolar receptor cells that respond to chemical stimuli, and supporting cells that provide a structural integrity to the epithelium. The terminal processes of the receptor cells are fine, thread-like microvilli that extend into the lumen (Ciges et al., 1977). The nonsensory part of the VNO consists of a simpler epithelium (e.g., no receptors) that does not participate in chemoreception (Breipohl et al., 1979).
Table 2. Terminology related to the VNO1 (all structures are situated bilaterally, unless stated otherwise)
All structures are situated bilaterally, unless otherwise stated.
There is unfortunate confusion, because the vomeronasal duct is the accepted veterinary terminology for the VNO (International Committee on Veterinary Gross Anatomical Nomenclature).
VNO Jacobson's organ Organon vomeronasale2 Organum vomeronasal3 Duct of Ruysch Vomeronasal duct
Bilateral epithelial tubes situated in the anteroventral nasal septum of most terrestrial vertebrates (Wysocki and Meredith, 1987). Bats and primates have highly variant forms ranging from total absence of the tubular structure to a well-developed chemoreceptor organ (Bhatnagar and Meisami, 1998). The presence of VN epithelial tube disguised in any form has been erroneously considered to be chemosensory. Such is not true for mammals with variant forms of the VNO (e.g., in humans, great apes, and bats). We propose the terms, chemosensory VNO (where at least part of the epithelial tube comprises neuroreceptors) and the nonchemosensory VNO (where VN neuroreceptors are lacking).
Vomeronasal organ complex (Cooper and Bhatnagar, 1976)
Includes the vomeronasal duct, the epithelial tube with cellular elements, septal glands, blood vessels (a large venous sinus situated laterally in most mammals, but absent in humans and chimpanzees) which are not called vomeronasal arteries, veins or sinuses, associated nerves, a cartilaginous (syn. paraseptal cartilage; Huschka's cartilage) or a bony capsule enclosing the VNO, and an accessory olfactory bulb (AOB) situated posterodorsally within the main olfactory bulb.
Vomeronasal primordium (VNP)
That medial invagination of the nasal sac growing medially toward the nasal septum, soon breaking off in a circular to oval tube-like structure with a lumen. This VNP further develops into the particular type of the VNO. In some species of catarrhine primates, the VNP does not form a tube. Instead, it remains as an epithelial thickening, and apparently regresses (Hendrickx, 1971; Zingeser, 1984).
Vomeronasal duct (VND)
A short, stratified squamous epithelium-lined duct connecting the VNO either with the nasal cavity (as in humans, but where it may be more apparent in the fetus [Smith and Bhatnagar, 2000] than in the adult), or with the oral cavity through its opening into the nasopalatine duct. In a recent imaging study (Abolmaali et al., 2001), the entire VNO was called the VND.
A long, stratified squamous epithelium-lined duct connecting the nasal cavity with the oral cavity. In humans, it is not patent.
Incisive canal (IC)
The IC is a skeletal feature. It is a passageway within the maxillary bone (or between the maxillary and premaxillary bone, in most mammals) which houses the nasopalatine vessels, nerves, and duct (the duct is obliterated or blind-ended in adult great apes and humans) as they pass from the nasal cavity to the oral cavity.
An O-, U-, J-, or C-shaped cartilage enveloping the VNO tube. In animals its bar shape signifies either an absent or nonchemosensory VNO, as the case in some primates and bats (Bhatnagar and Meisami, 1998). A bony capsule develops in voles and in rodents (Wysocki and Meredith, 1987). Strictly speaking, humans (Bhatnagar and Smith, 2001) and chimpanzees (Smith et al., 2001d), lack a vomeronasal cartilage, but instead have the homologous paraseptal cartilage. A critical difference is that the paraseptal cartilage has no direct association with the VNO (see Fig. 6A). The vomeronasal cartilage is erroneously accepted as a valid term in humans (see Nomina Anatomica,1989; FCAT, 1998).
Vomeronasal epithelial tube (Wible and Bhatnagar, 1996)
Usually dorso-ventrally elongated and compressed laterally, it shows differentiation into thick, medial chemosensory epithelium and thin, single layered, ciliated, respiratory epithelium designated receptor-free epithelium (Breipohl et al., 1979). It should be noted that the adult human and the chimpanzee VNO is quite unlike the types described above.
This term is not in use and we are not proposing that the terms vomeronasal arteries and vomeronasal veins be used. However, the presence of a large venous sinus in many bat species (e.g., Artibeus, Bhatnagar and Kallen, 1974), and other mammals (Schilling, 1970) is qualified to be called a vomeronasal sinus, a new term that we propose.
A thick bed of unmyelinated nerve fila underlying the chemosensory epithelium course superiorly on the nasal septum. These separate bundles eventually unite into a single bundle (the vomeronasal nerve, VNN), pierce the cribriform plate on its medial aspect, traverse the olfactory bulb and ramify over the posterodorsally situated accessory olfactory bulb (AOB) (Bhatnagar and Kallen, 1974). Again it should be noted that adult humans lack the AOB (Bhatnagar et al., 1987; Meisami and Bhatnagar, 1998).
Accessory olfactory bulb
While there is extensive literature on this central terminal for the vomeronasal nerve, in humans it is lacking (Bhatnagar et al., 1987; Meisami and Bhatnagar, 1998). Structurally it is a miniature, but disorganized olfactory bulb. The presence of an AOB is a clear and unequivocal evidence of the presence of chemoreceptors in the VN epithelium, whereas its absence also clearly denotes the nonchemosensory nature of the VNO. The nonchemosensory VNO can exist without an AOB (as in humans, the chimpanzee, and certain bat species (Meisami and Bhatnagar 1998), but the reverse has never been reported.
Paravomeronasal ganglion (PVNG)
Seen and reported for the first time in the bat Artibeus (Bhatnagar and Kallen, 1974), it is a collection of nerve cell bodies underneath the chemosensory vomeronasal epithelium. It has been observed in other bat species also. Likewise, intraepithelial neurons have also been reported in the vomeronasal epithelium of Artibeus. PVNG-like cells have been reported in humans (Smith and Bhatnagar, 2000).
Vomeronasal glands (VNG)
Glands that usually surround the chemosensory VNO on its dorsolateral aspects. These are multicellular, seromucus, tubuloacinar or simple acinar glands (Roslinski et al., 2000). In humans, these glands differ histochemically compared to other mammals, and by opening into the VNO lumen through all sides of the vomeronasal epithelium, thus discharging into the nasal vestibule or anterior nasal cavity (Roslinski et al., 2000).
Gray's Anatomy (Goss, 1972:1123) describes it as follows: “immediately over the incisive canal at the inferior edge of the cartilage of the septum is a depression, the nasopalatine recess. In the septum close to this recess a minute orifice may be discerned; it leads posteriorly into a blind pouch, the rudimentary vomeronasal organ of Jacobson, which is supported by a strip of cartilage, the vomeronasal cartilage.” Thus, the nasopalatine recess and a fossa within it has been erroneously been considered the vomeronasal pit by some workers (see Jacob et al., 2000 and Bhatanagar and Smith, 2001 for further discussion), even though the VNO actually opens directly on the nasal septum superior to the paraseptal cartilage and the nasopalatine recess. To complicate matters, other authors have clearly used the term VNO pit to denote the true opening of the VNO (Johnson et al., 1985). Therefore, the use of the term vomeronasal pit has created much confusion, and it is suggested here that the term vomeronasal duct opening be substituted to denote the true opening of the human VNO, which is found from 6 to 11 mm above the floor of the nasal cavity, or palate (Bhatnagar and Smith, 2001; Smith et al., 2001a).
There are at least several morphologic departures from this “typical” VNO among primates. The vomeronasal duct is typically lined with stratified squamous epithelium but has numerous glandular “crypts” in Otolemur (Galago) crassicaudatus (Hedewig, 1980). One strepsirhine (Lemur catta) and several New World monkeys (Callithrix jacchus, Cebuella pygmaea, Saguinus fuscicollis, and Saimiri sciureus) have been reported to exhibit SE on all sides of the VNO (Maier, 1980; Evans, 1984; Taniguchi et al., 1992; Evans and Grigorieva, 1994; Mendoza et al., 1994). In contrast, Aotus trivergatus and Arctocebus calabarensis each seem to have both SE and nonsensory epithelium, whereas the VNO epithelium of Ateles geoffroyi was described as “similar to that lining the nasopalatine ducts” (Hunter et al., 1984). A most unusual variation was described in Saguinus labiatus, where receptor cells were described outside of the VNO epithelium (Evans and Grigorieva, 1994). (This finding may be similar to populations of neurons, termed the paravomeronasal ganglia, found outside the VNO of the bat Artibeus by Bhatnagar and Kallen, 1974). The VNO of Tarsius spectrum was described as exhibiting uniform thickness but with a ciliated inner sensory epithelium (Woollard, 1925). This finding would be an unusual state, if accurate, because cilia are not associated with the SE of the VNO in mammals (Ciges et al., 1977), except perhaps in small transitional zones between SE and nonsensory VNO epithelia.
Thus, haplorhines have extremely variable VNOs; if catarrhine primates truly lack a VNO during postnatal ontogeny (Ankel-Simons, 2000), “VNO absence” would be yet another character state. However, an atypical VNO has been reported numerous times in just one catarrhine species—Homo sapiens.
THE HUMAN VNO
The human VNO has been the subject of conflicting reports since Ruysch's description of an infant nasal septum (see Bhatnagar and Reid, 1996, for translation); a consensus on its existence in the adults has been reached only recently (Johnson et al., 1985; Smith et al., 1998; Bhatnagar and Smith, 2001). Studies on VNO incidence have provided ranges from 6% (Zbar et al., 2000) to 100% (Moran et al., 1991), and VNO length has been reported to vary from 2 mm (Smith et al., 1998; Bhatnagar and Smith, 2001) to 62 mm (Mangakis, 1901). Furthermore, a clear description of the location and microanatomy of the human VNO has remained elusive. Early reports detailed gross findings, consistently locating the opening of the VNO along the anteroinferior surface of the nasal septal mucosa (e.g., Gegenbaur, 1886; Kölliker, 1877) (Fig. 2A,B). Some texts and articles suggest a VNO position that is closely adjacent to the palate (Goss, 1972; Moran et al., 1991), whereas histologic studies demonstrate that the VNO is approximately 6 to 11 mm above the palate (Smith et al., 1998; Trotier et al., 2000; Bhatnagar and Smith, 2001).
Light microscopic studies indicate that the VNO epithelium in humans resembles that which lines most of the nasal cavity in general—a ciliated, pseudostratified, columnar epithelium (Johnson et al., 1985; Jahnke and Merker, 1998; Smith et al., 1998; Roslinski et al., 2000; Bhatnagar and Smith, 2001). Other descriptions indicate stratified squamous epithelium (Gaafar et al., 1998) and some specifically state that cilia are lacking (Gaafar et al., 1998) or sparse (Moran et al., 1991). Several electron microscopical studies have described receptor-like cells with short microvilli in the VNO epithelium (e.g., Moran et al., 1991; Jahnke and Merker, 1998), but their specific function has not been clarified.
For decades, such conflicting information has frustrated authors, leading some to question the functionality of the adult human VNO (e.g., Johnson, 1998; Wysocki and Preti, 2000), and even to doubts regarding the homology of the human VNO to that of other mammals (see Smith et al., 1998; Smith and Bhatnagar, 2000). Prenatal development of human VNO provides the answer to the question of homology. The first embryonic evidence for the human VNO is seen at approximately 33 days fertilization age, as a thickening (vomeronasal primordium) situated along the medial surface of the invaginating nasal sac (Bossy, 1980; Smith and Bhatnagar, 2000) (Fig. 3B). At first, this primordium is closely adjacent to the developing olfactory epithelium, and connects to vomeronasal nerves that course toward the roof of the nasal cavity (Bossy, 1980; Smith and Bhatnagar, 2000). This early stage of VNO morphogenesis can be demonstrated in rodents (see Garrosa et al., 1998 for review), bats (Bhatnagar et al., 1996), and even other primates (Smith and Bhatnagar, 2000) (Fig. 3A,C). The VNO primordium subsequently invaginates, in a posteroanterior sequence, and then “pinches” off from the epithelial lining of the nasal septum, leaving only an anterior communication with the nasal cavity (Kreutzer and Jafek, 1980). The result is an anteroposteriorly oriented epithelial tube (Figs. 3E, 4B) that increases in length and volume during the second and third trimesters of fetal development (Smith et al., 1997). Before elevation of the palatal shelves, nearly all mammals seem to possess this anteroposterior tube that communicates with the nasal cavity (Wöhrmann-Repenning and Ciba, 1989; Smith and Bhatnagar, 2000). However, position of the VNO differs markedly between humans and most other mammals after this point of development.
Conflicting information has frustrated authors, leading some to question the functionality of the adult human VNO, and even to doubts regarding the homology of the human VNO to that of other mammals.
In prenatal strepsirhines and New World monkeys (Figs. 3D, 4C), the VNO remains intimately associated with the vomeronasal cartilage throughout late embryonic and fetal development (Frets, 1914; Smith and Bhatnagar, 2000). In contrast, the human VNO becomes displaced during late embryonic development, until it is spatially separated from the paraseptal cartilages (vomeronasal cartilage homolog) superolateral to the bulbous inferior tip of the nasal septum (Smith and Bhatnagar, 2000). A correlate of this relatively more superior VNO position in fetal and adult humans is the retained communication with the nasal cavity (Fig. 4), a reflection of the primitive embryologic state in all primates (Fig. 4C; see Smith and Bhatnagar, 2000). In strepsirhines and New World monkeys, the formation of the primary and secondary palate changes this relationship. During palatogenesis, the nasopalatine duct also forms and merges with the VNO opening (Fig. 4D; Shimp et al., 2001). Smith and Bhatnagar (2000) attributed the contrast in VNO position to heterochronic and heterotropic differences in midfacial growth between humans and other mammals. It may not be a “superior shift” of the VNO as much as relatively rapid downward midfacial growth in humans (Burdi, 1969).
Because most investigators have studied fetal and/or adult human VNOs (Jordan, 1972; Johnson et al., 1985; Smith et al., 1998), a stark contrast has been seen compared with most other mammals (including many primates), where the VNO undergoes a mediolateral differentiation into distinct nonsensory epithelium and SE during late embryonic development (Garrosa et al., 1998). No doubt, the varied and incongruous position, in combination with the atypical cellular characteristics of human fetuses and adults, was the primary factor in arguments against homology of the human VNO to that of other mammals (Gegenbaur, 1886). However, there are greater cellular similarities in the VNOs of humans and other mammals during embryogenesis. Human embryos from approximately 37 days to 8 weeks fertilization age have dense populations of cells in the epithelium, and some human embryos seem to have mediolateral differences in epithelial thickness (Smith and Bhatnagar, 2000). During early fetal development, the epithelium becomes relatively simplified, with only two or three rows of nuclei from base to apex. A more unusual change occurs at the apical border, where cilia begin to appear superiorly at 10 weeks, and progressively become distributed on all sides of the VNO by 14 weeks (Boehm and Gasser, 1993; Smith and Bhatnagar, 2000). Thus, the human VNO undergoes several stages of development; only a comparison of embryos before palatogenesis reveals the homology of the human VNO.
IS THE VNO PRESENT OR ABSENT IN OTHER CATARRHINE PRIMATES?
Aside from studies on humans, the most thorough studies of catarrhine primate VNOs have concerned prenatal macaques and baboons (Hendrickx, 1971; Wilson and Hendrickx, 1977; Zingeser, 1984). In these primates, a tubular VNO does not form, although the vomeronasal primordium is easily observed in embryos. The only remnant of the VNO is a small epithelial thickening on the septal mucosa (Fig. 3F), to which presumptive vomeronasal nerves can be observed to connect. The position of the septal epithelial thickenings is entirely similar to the tubular VNOs observed in human fetuses, except for the more superficial location.
Maier (1997, Maier 2000) found no VNO-like structures in fetal gorillas, orangutans, or gibbons. However, Starck (1960) described structures resembling human VNOs in a fetal chimpanzee—i.e., simple epithelial tubes, parallel but separated from the paraseptal cartilage. At the time, these structures represented the only link to the incongruous human VNO. Even in this light, a troubling question remained: why is a similar VNO not present in any other adult catarrhine primate (Preti et al., 1997)? Until recently, all studies on postnatal nonhuman catarrhine primates had failed to locate the VNO, except in humans (Frets, 1914; Jordan, 1972; Loo, 1973; Zingeser, 1984).
RECENT FINDINGS: THE VNO OF THE CHIMPANZEE
Evidence that the human VNO forms in a similar embryonic position to New World monkeys or strepsirhines (Smith and Bhatnagar, 2000) and may be present in fetal chimpanzees (Starck, 1960), strongly indicates that at least some other catarrhine primates retain an epithelial tube that is the true homolog of the mammalian VNO. The reported absence of the VNO in catarrhine primates may have been artifactual.
The search for the VNO in postnatal catarrhine primates may have been inadequate in several ways. First, some investigators may have had the expectation that the VNOs of catarrhine species would be located in a position similar to those of strepsirhines and New World species (i.e., near the palate, surrounded by vomeronasal cartilages), whereas the VNO of humans and chimpanzees occupies a superiorly displaced position in the septal mucosa (Smith et al., 2001d). Second, it is clear that studies based on discontinuous portions of the nasal septum (e.g., Johnson et al., 1985) yielded a lower success rate in finding the minute (∼2 to 11 mm long) adult human VNO compared with studies based on serial sections (Bhatnagar and Smith, 2001; Smith et al., 1998). Some previous studies of nonhuman catarrhine primates may have missed the VNO in this way (Frets, 1914). Third, very few species of catarrhine primates have been carefully examined. Therefore, demonstrating postnatal “absence” of the VNO in catarrhine primates requires a thorough examination by using complete histologic series of nasal septal tissue in multiple taxa.
If prenatal retrogression of the vomeronasal primordium in macaques and baboons is taken as one course of VNO development among primates, then there are at least two character states among catarrhines: postnatal VNO absence, or presence of a simplified, tubular VNO as in humans (Jordan, 1972; Zingeser, 1984; Smith et al., 2001d). Recently, Smith et al. (2001d; authors' new data) attempted to find VNOs in several catarrhine primates, including Colobus guereza, Macaca fascicularis, Macaca nemestrina, and Pan troglodytes. Epithelial tubes similar to the human VNOs were found in the chimpanzee, but no VNO-like structures were found in any other species (Figs. 5, 6). These epithelial tubes of the juvenile and adult chimpanzee were entirely similar to that of humans at similar ages, positionally and histologically (Fig. 6A–F). Therefore, they were considered VNO homologs (Smith et al., 2001d), although an embryologic study of these structure is still needed (Maier, 2000). The VNOs of the chimpanzee and human are located relatively high up along the nasal septum, approximately 1 to 5 mm (in either species) from the paraseptal cartilages (Bhatnagar and Smith, 2001; Smith et al., 2001d). The anterior VNO opening communicates with the nasal cavity alone (not the nasopalatine duct) and is found in a small depression which, when macroscopically visible, represents the only surface landmark for the VNO (Fig. 2C, D).
The postnatal epithelial morphology of the VNO is identical in Pan and Homo, i.e., a ciliated, pseudostratified columnar epithelium (Smith et al., 2001d). The precise nature of the cilia has not been adequately clarified at the ultrastructural level. However, indications of basal bodies (Fig. 6C,F) and the similar appearance of the VNO cilia compared with that of adjacent respiratory epithelia, strongly suggests that VNO cilia may be motile, unlike other cell processes visible at the light microscopic level (stereocilia, olfactory cilia, or long microvilli). These processes could be seen on all apical surfaces (medially, laterally, superiorly, and inferiorly) of the VNO epithelium in nearly all chimpanzees and humans examined (Roslinski et al., 2000; Smith et al., 2001d; Bhatnagar and Smith, 2001). They were found throughout all anteroposterior percentiles of VNO length, but with some apparently nonciliated regions. Further examination, particularly by using electron microscopy, will be required to clarify whether all human and chimpanzee VNOs are ciliated and whether other cellular processes may be found.
The similarities to respiratory mucosa also extend to the underlying lamina propria. In both chimpanzees (unpublished data) and humans (Roslinski et al., 2000), the glands and secretions found in the VNO are very similar to those of the nasal cavity in general. This is in contrast to most other mammals in which the VNO secretions are histochemically different from that of the remainder of the nasal cavity. Based on the characteristics noted above, Roslinski et al. (2000) and Bhatnagar and Smith (2001) have suggested that an altered or accessory function, glandular secretion, and transport, is possible for the human VNO (and perhaps the chimpanzee).
These recent findings show that previous studies on adult catarrhine primates missed the VNO in at least some species; a thorough reevaluation of presence versus absence of the VNO in this group is clearly needed. The VNO was probably overlooked because of unique characteristics that distinguish the catarrhine vomeronasal complex compared with other primates: (1) spatial separation of the VNO or fetal VNO vestiges from the paraseptal cartilage (Smith et al., 2001d; Bhatnagar and Smith, 2001) and (2) VNOs with a greatly simplified epithelial structure.
DEFINING THE HUMAN/CHIMPANZEE VNO
As discussed above, the VNO in humans or chimpanzees has marked differences with that of strepsirhine primates. A clear description of the position and morphology of the human/chimpanzee VNO is needed to extend investigations to other catarrhine primates. It has been suggested recently that the ciliated nature of the human (and chimpanzee) VNO may render it unique among tubular septal structures and should be part of the definition of the human VNO (Roslinski et al., 2000; Smith and Bhatnagar, 2000; Smith et al., 2001d). There is one complication with this definition. Gegenbaur (1886) found the human VNO to be similar to the septal gland ducts seen in some strepsirhine primates; such a duct was recently found (authors' new data) in a neonatal mongoose lemur (Eulemur mongoz). This duct was seen unilaterally, rostral (anterior) to the nasopalatine duct and to the VNO itself. Moreover, it was located superiorly (dorsally) to the laminae transversalis anterior, paired cartilages that are continuous posteriorly with the vomeronasal cartilages. Certain portions of this tube were ciliated. Thus, in relative position and histology, this duct bore some resemblance to the human/chimpanzee VNO (Fig. 6H).
These findings have two important implications. First, the VNO may not be unique in location and histology among other septal structures. This can potentially create confusion in defining the human VNO, although it does not discount the evidence for homology of the human VNO to that of other mammals. An ontogenetic transformation of a VNO containing receptors to a ciliated duct is, therefore, an important part of the definition of the human VNO (and perhaps that of other hominoids). This finding also means that the human/chimpanzee VNO acquires characteristics that are analogous to septal ducts of some strepsirhines. Second, if other primates possess structures that resemble the human VNO (in addition to a functional VNO), it strongly suggests that ciliated septal ducts may have unique, perhaps glandular functions. In this light, an altered function for the human/chimpanzee VNO should be strongly considered. The ciliated tube observed in Eulemur mongoz present an interesting analogy, and may indicate that some gland ducts are ciliated to facilitate transport.
IS THE VNO A PHEROMONE RECEPTOR ORGAN IN CATARRHINES?
Although the presence of a VNO homolog in chimpanzees suggests that VNOs may have been overlooked in at least some catarrhines, it seems clear that the VNO is postnatally absent in adult macaques and baboons. Among all catarrhine primates, VNO functionality has only been considered in humans. The most debated questions are whether these structures are responsive to putative human pheromones and whether they have the neural apparatus necessary for generating a response (i.e., receptor cells). Unfortunately, evidence for the presence of receptor cells in human VNOs is unclear, in contrast to strepsirhines in which receptor cells are easily found and can be quite dense in population (Schilling, 1970).
VNO size has been used as an indirect measure of receptor cell populations in the mammalian VNO (Dawley, 1998). Data on VNO length in selected species of primates are shown in Figure 7. Although VNO length may not provide the most accurate estimate of VNO receptor population size in mammalian species, it can provide a reasonable preliminary comparison of VNO size (Dawley, 1998; Smith et al., 2001c). These data (Fig. 7), all acquired by using one measurement technique, indicate that VNOs of humans and chimpanzees are proportionately small compared with strepsirhine primates. It has also been demonstrated that size differences can be seen prenatally, and are quite profound when comparing VNOs of humans and mouse lemurs (Smith et al., 2001b). The nature of VNO function in humans and chimpanzees is far less certain than what is known of strepsirhines (Aujard, 1997). If the human VNO is involved in pheromonal communication (Monti-Bloch et al., 1998), it is remarkably small in absolute and proportional size compared with that of other primates.
If the human VNO is involved in pheromonal communication, it is remarkably small in absolute and proportional size compared with that of other primates.
Among catarrhines, direct evidence for VNO function has only been sought in humans. In a series of studies, a team of investigators administered steroidal compounds derived from the skin to the putative opening of the VNO in adult humans (Berliner et al., 1996; Monti-Bloch et al., 1994, 1998; Grosser et al., 2000). The results have provided highly interesting electrophysiological data, derived from an electrode positioned within the same “mini-probe” used to administer the chemical stimuli, termed “vomeropherins”. These vomeropherins comprise at least six compounds, some of which have not been identified, pending patent approval (Monti-Bloch et al., 1994). In summary, vomeropherins seemed to stimulate the human vomeronasal epithelium but not olfactory epithelium. Conversely, olfactory stimulants generated electrical potentials from the olfactory epithelium but not the vomeronasal epithelium (Monti-Bloch et al., 1994). Additionally, these compounds appeared to have some sex-specific effects (Monti-Bloch et al., 1994, 1998; Grosser et al., 2000). Changes in serum levels of steroidal hormones in men were also reported (Monti-Bloch et al., 1998), and it was suggested that the hypothalamus may be targeted by vomeronasal input (Berliner et al., 1996).
Such highly specific responses from the human VNO epithelium would seem to indicate postnatal functionality. However, there is cause to reserve judgment on this question. Preti et al. (1997) suggested alternate explanations for the findings of Monti-Bloch et al. (1994, 1998). For example, the electrical potentials might be explained as stimulation of trigeminal nerve endings in the nasal mucosa. A recent review by Meredith (2001) provides further discussion on the possible basis (whether chemosensory or artifactual) for electrical potentials reported in the VNO epithelium.
A second troubling consideration is the lack of definitive evidence for vomeronasal receptor cells, vomeronasal nerves that pass from the VNO to the forebrain, or an AOB in adult humans (Bhatnagar and Meisami, 1998). Much more work is needed to characterize the cells of the human VNO that have been identified as receptor candidates, especially because they only show nonspecific similarities to the vomeronasal receptors of other mammals (Trotier et al., 2000). Meredith (2001) considered this population of cells to be too sparse to generate a detectable electrical potential, suggesting that other possibilities must be investigated.
The question of precise anatomic location also may have a profound importance to studies on human VNO function. The results of several reports (Jacob et al., 2000; Smith et al., 2001a; Bhatnagar and Smith, 2001) have suggested that not all investigators are examining the identical structure—some studies may have inadvertently identified gland ducts or a remnant of the nasopalatine duct as the human VNO. It seems that histologic verification, by using adjacent structures for orientation (e.g., paraseptal cartilages), is necessary for positive identification of the VNO. Gross indicators alone are unreliable. Monti-Bloch et al. (1994, 1998) provided rather vague descriptions of the location for the VNO opening, but the possibility that they successfully found it can by no means be ruled out.
The history of research on the human VNO is important to recall while considering functionality of the human VNO. Absence is difficult to demonstrate; this applies to the VNO itself and putative receptor cells. The debate regarding pheromonal function in the human VNO awaits further analysis, and accessory or alternate functions (e.g., glandular secretion/transport) must also be considered. It is highly beneficial to know that comparisons can now be made to a yet to be determined number of other catarrhine primates.
THE PHYLOGENETIC PICTURE
The phylogenetic distribution of the VNO among haplorhines is presently unclear, but one may speculate that the common ancestor of New World monkeys and catarrhines possessed a functional VNO. At present, existence of the chimpanzee VNO allows us to generate the hypothesis that VNO presence is characteristic of all hominoids, which is further supported by the presence of epithelial tubes similar to the human VNO in orangutan and gorilla fetuses (C.S. Evans, personal communication). One may only guess about its presence in catarrhines in general.
A useful analogy to the variability seen in the primate VNO is found in bats (Fig. 8). Bats (order Chiroptera, class Mammalia) comprise nearly 925 species, of which there are 42 genera and 166 species of Megachiroptera, and 135 genera and 756 species of Microchiroptera (see Table 1 in Wible and Bhatnagar, 1996). In ontogeny, the VNO does not progress beyond a primordial stage in the megachiropteran Rousettus (Bhatnagar et al., 1996), whereas in the Microchiroptera, there is the single New World family Phyllostomidae (with 49 genera and 143 species), which is endowed with a functional VNO with its full complement of associated structures. In the other 13 microchiropteran families, the VNO is lacking as a general rule. Primates apparently share the VNO traits exhibited by bats, from well-developed VNOs (strepsirhines, some New World monkeys), to reduced VNOs (at least some hominoids and some New World monkeys) to VNO absence (at least some Old World monkeys). As more and more species are examined (serial histology of the entire head is not an easy task), known variations are likely to increase. Even though an attempt was made in Bhatnagar and Meisami (1998, see Fig. 4 and Tables 2, 3 therein) to classify such variations, additional data may alter our understanding.
The known variations in primates and bats raise both functional and phylogenetic questions. For example, the VNO of humans, chimpanzees, and at least one bat (Eptesicus) shares similar characteristics relating to discharge of mucus in the nasal chamber. The similar duct-like VNOs in such disparate taxa suggests a degree of functional analogy (and convergence). It is also striking that some primate and bat VNOs are present and chemosensory, whereas VNOs are completely absent in others. The functional significance of VNO absence is completely unknown; is VNO loss inconsequential, or is its function taken over by another chemosensory system? Regarding phylogeny, primates and bats that lack receptors, or in which a VNO does not form at all, are derived compared with a primitive state in which a chemosensory VNO has been retained. Because VNO morphology among bats has important phylogenetic implications (Wible and Bhatnagar, 1996), questions related to the VNO in catarrhine primates may indeed be more complex than previously thought.
Therefore, the search for the VNO in catarrhines should be continued, keeping the known extent of VNO variation among haplorhines in mind. A standardized terminology for vomeronasal structures is also needed (Table 2). In histologic structure, VNOs may be assigned to one of five categories. These character states have been derived from the literature and must be reassessed and expanded as necessary in the future: (1) well-developed: a VNO that closely resembles the strepsirhine state, i.e., (a) partially surrounded by the vomeronasal cartilage, or (b) possessing a lateral, nonsensory epithelium, and a thicker, ventromedial, sensory epithelium; (2) sensory epithelium only: where no nonsensory epithelium is present, as reported for Saguinus fuscicollis (Mendoza et al., 1994); (3) sensory epithelium reduced where the sensory epithelium only occurs in reduced patches and nonsensory epithelium is found everywhere else, as seen in Saguinus geoffroyi (Fig. 1J; possibly a developmental stage; see Evans and Grigorieva, 1994); (4) displaced VNO with simplified, homogeneous epithelium, as in humans and chimpanzees, where it is spatially separated from the paraseptal cartilages (Smith et al., 1998, 2001d) with a potentially altered function; (5) absent: no structure resembling a well-developed or displaced VNO, either in morphology or location, is found in the mucosa of the nasal septum.
Even as histologic studies examine larger samples and a greater number of taxa, being aware of the variability and understanding the reason for it are quite different issues. The adaptive significance of a functioning VNO has yet to receive a thorough consideration in primates, and such an analysis (related to unique sociosexual and ecological aspects of primates) must await a reevaluation of VNO presence/absence among catarrhines. Two other issues need to be addressed: does the distribution of characters follow a pattern of phylogenetic relatedness (i.e., are New World monkeys consistently different from Old World monkeys), do the characters relate to specific other morphologies that cross-cut taxonomic boundaries (short faces vs. long faces), or both?
THE ONTOGENETIC PICTURE
Exceptions to consistent patterns occur not infrequently in the craniofacial region of primates (Ankel-Simons, 2000). Leaving out the often problematic Tarsius, we can find marked differences in the disposition of the snout (long vs. short, the anterior vs. downward trajectory of growth), and the configuration of the neurocranium (inferior vs. posterior location of the foramen magnum). The contrast here is vivid and the primitive mammalian pattern is evidenced more often in the predominantly “macrosmatic” strepsirhines. The exceptions however, are interesting. Some Old World monkeys (baboons, mandrills) exhibit a secondarily derived long snout but fail to evidence the concomitant enhanced olfaction/functioning VNO. We also find that some of the shortest faced primates (marmosets) are among the New World Monkeys and have chemosensory VNOs (Taniguchi et al., 1992).
Critical to an understanding of the VNO function and morphology is the variability of the subnasal region and the “trajectory” of facial growth. It should be noted that some specific characteristics of human facial development are found in other haplorhines (especially Old World species), such as the downward and forward midfacial growth (Enlow, 1990) and impingement of the incisive canal (and, thus, the nasopalatine duct) seen in the great apes (Jones et al., 1992). In most simple terms, human faces grow “downward” and other catarrhines lie along a continuum of reduced prognathism and increased inferior growth. Thus, VNO displacement may simply be a function of relocation of old structures as a result of a new facial growth pattern. The timing of the fusion of the premaxillary-maxillary suture is intimately related to the magnitude rather than direction of the growth of the snout and, therefore, may be free to vary across taxa. The disposition of the premaxilla (e.g., its contribution to mesial growth) can be related to the contribution of the nasoalveolar clivus to the snout and state of the incisive canal. One might question whether the small opening into the canal in the “impinged” condition found in orangutans relates to VNO function and is in opposition to the possible “primitive” condition of Old World monkeys, (i.e., “open”), and whether this impingement relates to the novel VNO communication to the nasal cavity alone seen in chimpanzees and humans. Such an ontogenetic “shift” in VNO location clearly must be investigated by using prenatal samples of hominoids, particularly in late embryonic/early fetal stages (Smith and Bhatnagar, 2000).
Is it plausible that the VNO undergoes a significant shift in location as a secondary consequence of major trends in primate facial growth? Such an alteration is entirely possible. The functional role of the VNO as a pheromone receptor may be either reduced, eliminated or subsumed by the olfactory sense in hominoids (and perhaps other catarrhines). The functional overlap of mammalian vomeronasal and olfactory systems (Kelliher et al., 2001) may render vomeronasal chemoreception expendable, and thus makes VNO position inconsequential. Even if hominoids do have a functional VNO, sole communication with the nasal cavity is compatible with VNO function in other mammals (Wysocki and Meredith, 1987).
CONCLUSIONS AND PERSISTING QUESTIONS
Ontogenetic data clearly show the homology of the human VNO to that of other mammals. However, its incidence in various catarrhines remains difficult to ascertain. Reliance on gross data has led to the misconception that the lack of visible VNO duct openings is tantamount to the absence of the entire VNO (Gaafar et al., 1998; Wöhrmann-Repenning, 2000) and has led to unreasonable length estimates (Mangakis, 1901). This finding indicates that serial histology is the most appropriate method to reexamine the belief that adult catarrhine primates lack a VNO. Quantitative and immunohistochemical studies should follow as appropriate.
Establishing the histologic definition of the human and chimpanzee VNO is also difficult. Nonetheless, the continual presence of the human VNO from embryos to adults is clear, and involves a transformation to a structure quite unlike the VNO of mammals in general. It is of immediate concern to establish whether this VNO morphology is only to be found in Homo sapiens and Pan troglodytes, or represents a synaptomorphic modification of the VNO in all hominoids and perhaps other catarrhines.
Regarding function, a parallel to another “vestigial” structure can be drawn. The function of the human appendix has frequently been questioned (see Fisher, 2000), and yet histologic similarities to the remainder of the digestive tract, along with lymphatic specializations, are clearly exhibited. The question with the appendix is not whether it functions, but how (e.g., immunologically) and when (subadult or adult ontogeny) does it function? Similar questions can be asked regarding the VNO of humans and chimpanzees (Schaal et al., 1998). The challenge of elucidating human VNO function is now more complex than ever, because the question may be applied to an as yet unknown number of close primate relatives.
The question with the appendix is not whether it functions, but how and when does it function? Similar questions can be asked regarding the VNO of humans and chimpanzees.
We thank K.L. Shimp, L.M. Maico, D.L. Roslinski, and T.A. Buttery for histologic processing of tissues used in this review. C.J. Bonar provided primate tissues that were critical to developing the topic. F.C. Kallen, A.M. Burrows, and K. Lal Gauri translated German literature. We thank A.M. Burrows, J.T. Laitman, M.P. Mooney, J.S. Reidenberg, C.F. Ross, and C.J. Wysocki for numerous helpful discussions and encouragement. F. Garza and K.L. Rogers provided valuable help in procuring archival literature for this review.
Dr. Smith is an Associate Professor in the School of Physical Therapy, Slippery Rock University, where he teaches histology, embryology, physiology, and pathology. He is also an Adjunct Research Associate Professor in the Department of Anthropology, University of Pittsburgh, where he received a Ph.D. in Physical Anthropology. His research interests include craniofacial growth and development, and he has a specific focus on functional histology and evolution of the mammalian vomeronasal organ. Dr. Siegel is a Professor in the Departments of Anthropology and Orthodontics, University of Pittsburgh. His research foci include craniofacial biology and computer imaging. Dr. Bhatnagar is a Professor in the Department of Anatomical Sciences and Neurobiology, University of Louisville. He has studied chemosensory systems for over 30 years with a special emphasis on the mammalian/human vomeronasal system. Both Dr. Smith and Dr. Bhatnagar are also Research Associates in the Carnegie Museum of Natural History, Pittsburgh.