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. 2007 Dec;211(6):754–768. doi: 10.1111/j.1469-7580.2007.00826.x

Skeletal morphology and development of the olfactory region of Spea (Anura: Scaphiopodidae)

L A Pugener 1, A M Maglia 1
PMCID: PMC2375839  PMID: 18045351

Abstract

The nasal capsules of anurans are formed by an intricate set of sac-like cavities that house the olfactory organ and constitute the beginning of the respiratory system. In tadpoles, nasal capsules do not have a respiratory function, but each is composed of a single soft tissue cavity lined with olfactory epithelium. Our study has revealed that in Spea the nasal cartilages and septomaxillae are de novo adult structures that form dorsal to the larval skeleton of the ethmoid region. The only element of the adult nasal capsule that is partially derived from the larval skeleton is the solum nasi. Development of the nasal skeleton begins at about Gosner Stage 31, with chondrification of the septum nasi and lamina orbitonasalis. The alary cartilage and superior prenasal cartilage are the first of the anterior nasal cartilages to chondrify at Gosner Stage 37. By Gosner Stages 40/41, the ethmoid region is composed of the larval structures ventrally and the adult structures dorsally. By Stage 44, the larval structures have eroded. The adult nasal capsule is characterized by: (1) a septum nasi that projects ventrally beyond the plane of the nasal floor; (2) a paranasal commissure that forms the ventral margin of the fenestra nasolateralis; and (3) a large skeletal support for the eminentia olfactoria formed by the nasal floor and vomer. The timing of chondrification of the anterior nasal cartilages and the development of the postnasal wall, inferior prenasal cartilage, fenestra nasolateralis, and paranasal commissure are discussed and compared with those of other anuran species. This study also includes a discussion of the morphology of the skeletal support for the eminentia olfactoria, a structure best developed in distinctly ground-dwelling frogs such as spadefoot toads. Finally, we propose a more precise restriction of the terminology that is used to designate the posterior structures of the olfactory region of anurans.

Keywords: Anura, morphology, olfactory region, Scaphiopodidae, Spea, skeletal development

Introduction

Spadefoot toads of the genus Spea are extraordinary anurans having small, stout bodies and short limbs. These fossorial frogs are found in a wide range of arid and semi-arid habitats in the Great Plains regions of Canada, the USA, and Mexico (Wiens, 1989). Their hind limbs bear keratin-covered metatarsal spades, which are used for excavation underground, where they may spend up to 8–10 months aestivating in soil-filled burrows (Ruibal et al. 1969). Natural populations of spadefoot toads are characterized by the occurrence of two tadpole morphologies, carnivorous morphs and omnivorous morphs (Pomeroy, 1981; Farrar & Hay, 1997, 2005; Morey, 2005).

Several authors have described the skeletal anatomy and developmental morphology of Spea. Wiens (1989) published a description of the skeletal development of Spea bombifrons, and Hall & Larsen (1998) and Banbury & Maglia (2006) conducted similar studies on the development of Spea intermontana and Spea multiplicata, respectively. In addition, references to the development and larval and adult characters of several species of Speacan be found in a number of publications in the context of comparative investigations (e.g. Zweifel, 1956; Sokol, 1981; Cannatella, 1985; Henrici, 1994; Maglia, 1998; Pugener, 2002; Haas, 2003). The nasal capsules of this genus, on the other hand, have been described in detail only for S. intermontana. Jurgens (1971) provided a description of the adult nasal capsules and Hall & Larsen (1998) provided an account of the development of the nasal cartilages. In addition, Wiens (1989) briefly described the development of the nasal cartilages of S. bombifrons and Maglia (1999) provided a succinct description of the adult morphology of the nasal cartilages of S. multiplicata.

Here we describe the adult skeletal morphology of the olfactory region of S. bombifrons and S. multiplicata and the development of the nasal capsules of S. multiplicata, and comment on the nasal capsule morphology of S. intermontana. In addition to providing baseline descriptive data, we also offer new interpretative insights into the processes by which certain structures develop.

Materials and methods

The description of the morphology and development of the olfactory region of Spea(Cope, 1866) is based on the study of 65 specimens (60 cleared and double-stained tadpoles, 3 cleared and double-stained juveniles, and 2 cleared and double-stained adults) of S. multiplicata and three specimens (1 serially cross-sectioned premetamorphic, 1 cleared and double-stained juvenile, and 1 serially cross-sectioned adult) of S. bombifrons. The results obtained from this study are complemented with those presented by Wiens (1989) for S. bombifrons and Hall & Larsen (1998) for S. intermontana (Scaphiopus intermontanus).

Tadpoles were lab-raised and staged according to the developmental table of Gosner (1960); the sample represents omnivorous morph tadpoles between Stages 30 and 46. The technique for clearing and double-staining follows a method adapted from Taylor & van Dyke (1985), which differentially stains cartilage blue with Alcian Blue and bone red with Alizarin Red. [See Trueb (1970) for a detailed description of the histological preparation of the serial sections.] Catalogue number, stage, and preparation methods of the specimens examined are presented in Table 1. Specimens are deposited in the herpetological collection of the Natural History Museum and Biodiversity Research Center at the University of Kansas (KU). In all, 314 cross-sections were used in the generation of a three-dimensional visualization of the adult olfactory region of S. bombifrons using the software packages imagej© (Able Software Corp., Lexington, MA, USA) and 3d doctor© (National Institute of Mental Health, Bethesda, MD, USA). The 3D image is deposited in the web-based anatomical library MorphologyNet (http://www.morphologynet.org; Leopold et al. 2005) with catalog number MN009. All larval specimens were used previously in endocrinology investigations (Buchholtz, 1999) and studies of skeletal development (Banbury & Maglia, 2006). Adult specimens were included in studies by Banbury & Maglia (2006), Maglia (1998, 1999), and Wiens (1989). Observations and illustrations were made with the use of an Olympus SZX12 stereoscope (Olympus America Inc., Pennsylvania, USA) with an attached camera lucida and 5-megapixel digital camera.

Table 1.

Specimens examined

Species KU # n Stage Preparation
Spea bombifrons 90044 1 premetamorphic sectioned
Spea bombifrons 205038 1 juvenile C&S
Spea bombifrons 18450 1 adult sectioned
Spea multiplicata 292081 3 Gosner 30 C&S
Spea multiplicata 292082 3 Gosner 31 C&S
Spea multiplicata 292083 3 Gosner 32 C&S
Spea multiplicata 292084 2 Gosner 33 C&S
Spea multiplicata 292085 3 Gosner 34 C&S
Spea multiplicata 292086 2 Gosner 35 C&S
Spea multiplicata 292087 2 Gosner 36 C&S
Spea multiplicata 292088 3 Gosner 37 C&S
Spea multiplicata 292089 3 Gosner 38 C&S
Spea multiplicata 292090 4 Gosner 39 C&S
Spea multiplicata 292091 3 Gosner 40 C&S
Spea multiplicata 292092 7 Gosner 41 C&S
Spea multiplicata 292093 3 Gosner 42 C&S
Spea multiplicata 292094 5 Gosner 43 C&S
Spea multiplicata 292095 5 Gosner 44 C&S
Spea multiplicata 292096 4 Gosner 45 C&S
Spea multiplicata 292097 5 Gosner 46 C&S
Spea multiplicata 49468 1 juvenile C&S
Spea multiplicata 97355 1 juvenile C&S
Spea multiplicata 106255 1 juvenile C&S
Spea multiplicata 86662 1 adult C&S
Spea multiplicata 86664 1 adult C&S
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C&S, clear and stained; KU, Natural History Museum and Biodiversity Research Left, The University of Kansas, USA; n, number of specimens.

The anatomical lexicon used herein conforms to the current understanding of homology of the nasal capsule elements and follows, in general, that of Jurgens (1971) and the Amphibian Anatomical Ontology (http://www.amphibanat.org; Leopold et al. 2007; Maglia et al. 2007a). Alternative terms to those used in the descriptions are indicated throughout the text and each is followed by one or more reference citations. The description of the olfactory region of the adult is followed by a detailed description of the ethmoid region of a Gosner Stage-30 larva, the stage immediately prior to the appearance of the first nasal capsule structures. The Stage-30 description provides a reference to facilitate descriptions of the developmental patterns that unfold during subsequent stages. In addition to the 3D reconstruction of the nasal capsules of an adult Spea, the adult olfactory region was illustrated based on a cleared and double-stained metamorphosing tadpole because the skeleton is fully developed, but the intramembranous investing bones exhibit comparatively less ossification, and thus do not conceal the underlying structures.

Results

Adult skeletal morphology

The paired nasal (olfactory) capsules have both respiratory and olfactory functions, and are located in the foremost section of the cranium, the olfactory region, which composes the anterior fourth of the skull. The nasal capsules lie anterior to the braincase, within an area enclosed by the premaxillae, maxillae, nasals, and vomers (Figs 1–4). Each nasal capsule has two openings, the apertura nasalis externa and the apertura nasalis interna. The apertura nasalis externa (= external nostril sensuStephenson, 1951; Roček, 1981; = external naris sensuTrueb, 1970; Baldauf & Tanzer, 1965; Wiens, 1989; = nostril sensuStebbins & Cohen, 1997) lies on the anterodorsal surface of the cranium and opens through the fenestra endonarina communis into the main cavity of the nasal capsule (Fig. 5A). The apertura nasalis interna (= choana sensuStephenson, 1951; Baldauf, 1958; Michael, 1961; Baldauf & Tanzer, 1965; Roček, 1981; Trueb, 1993; = internal naris sensuTrueb, 1970; Hall & Larsen, 1998; = internal nostril sensuStebbins & Cohen, 1997) opens through the fenestra endochoanalis into the buccal cavity (Fig. 5B). Internally, each nasal capsule is formed by a series of sac-like cavities. The largest nasal cavity (= cavitas nasalis or cavitas capsulae nasalis sensuRoček, 2003), which is also the most dorsal, is the cavum principale. In addition, two accessory cavities, the cavum inferius and the cavum medium, are located ventrally and laterally, respectively (Fig. 6). All of these cavities are lined by respiratory and sensory epithelium and glandular tissue (Duellman & Trueb, 1994). The skeleton of the nasal capsules is mostly cartilaginous, and remains cartilaginous throughout life. The nasal cartilages and the septomaxillae – the only bony elements internal to the nasal capsules – form a structural framework that provides support for the nasal sacs and ducts.

Fig. 1.

Fig. 1

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Three-dimensional reconstruction of the olfactory region of the adult S. bombifrons (KU 18450) in anterior view. Blue denotes nasal cartilage; pink denotes endochondral bones and septomaxillae; red denotes intramembranous bone (right elements omitted; represented as transparent to show underlying structures). ant, anterior; c, cartilage; inf, inferior; p, process; pren, prenasal; sup, superior.

Fig. 4.

Fig. 4

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Lateral view of left nasal capsule of Stage-46 S. multiplicata(KU 292097). Blue denotes nasal cartilage; red denotes bone (represented as transparent to show underlying structures); black denotes fenestrae. ant, anterior; c, cartilage; inf, inferior; p, process; pren, prenasal; sup, superior.

Fig. 5.

Fig. 5

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Schematic representation of the nasal fenestrae of Spea in lateral (A) and ventral (B) views. Gray denotes cartilage and black denotes fenestrae.

Fig. 6.

Fig. 6

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Serial cross-sections of the adult olfactory region of S. bombifrons (KU 18450). Sections at the level of the anterior nasal wall (A,B); alary cartilage (C,D); crista intermedia (E,F); anterior margin of septomaxilla (G,H). Blue denotes nasal cartilage and red denotes bone. c, cartilage; cav, cavum; cr, crista; inf, inferior; int, intermedia; lam, lamina; med, medium; pren, prenasal; septomax, septomaxilla; sup, superior.

The septum nasi (= planum verticale sensuHiggins, 1921; = nasal septum sensuHall & Larsen, 1998) is a vertical plate that separates the nasal capsules throughout their entire length (Figs 1, 6, 7). The septum nasi is formed by spongy bone. Anteriorly, the septum nasi protrudes through the anterior nasal wall and forms a triangular anterior median prenasal process (= processus praenasalis medius sensuStephenson, 1951; Roček, 1981); its anteriormost tip is cartilaginous. The posterior margins of the septum nasi, formed by the olfactory foramina, are clearly visible. Dorsally, the septum nasi is continuous with the tectum nasi. The septum nasi projects ventrally beyond the plane of the nasal floor to the level of the fenestra nasobasalis; posterior to the fenestra, the ventral margin of the septum is continuous with the solum nasi. The thickness and height of the septum decrease posteriorly; the thickness at the level of the anterior nasal wall is approximately four to five times as wide and about one-third taller than that at the level of the postnasal wall. In addition, it is evident in serial sections that, from the level of the anterior nasal wall to the level of the posterior margin of the lamina inferior, the septum projects ventrally beyond the plane of the solum nasi (Fig. 6). In metamorphic individuals the septum is completely cartilaginous, but in juveniles (e.g. KU 49468, KU 97355, KU 106255, KU 205038) the posterior half is replaced by sphenethmoid ossification.

Fig. 7.

Fig. 7

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Serial cross-sections of the adult olfactory region of S. bombifrons (KU 18450). Sections at the level of posterior margin of lamina inferior (A,B); anterior end of skeletal support for eminentia olfactoria (C,D); central area of skeletal support for eminentia olfactoria (E,F); and posterior end of skeletal support for eminentia olfactoria (G,H). Right side omitted in (F) and (H). Blue denotes nasal cartilage and red denotes bone. c, cartilage; fen, fenestra; inf, inferior; lam, lamina; nasobas, nasobasalis; pl, planum; pren, prenasal; term, terminale.

The tectum nasi (= planum tectale sensuHiggins, 1921; = nasal roof sensuHall & Larsen, 1998) is an unpaired, horizontal plate that forms the roof of the nasal capsules (Figs 1, 2, 6, 7). Each half of the tectum nasi (when divided mediolaterally) is crescent-shaped and covers the olfactory capsule posteromedially. The tectum nasi becomes co-ossified with the overlying, intramembranously-derived nasal at about the level of the anterior tip of the eminentia olfactoria. The anterior part of the plate, formed by the combined tectum nasi–nasal bone, is thin and is formed by compact bone; posterior to the eminentia olfactoria the plate thickens and is formed by spongy bone. In metamorphic specimens, the tectum nasi is completely cartilaginous.

Fig. 2.

Fig. 2

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Dorsal view of right nasal capsule of Stage-46 S. multiplicata(KU 292097). Blue denotes nasal cartilage; red denotes bone (represented as transparent to show underlying structures); black denotes fenestrae. c, cartilage; cr, crista; intermed, intermedia; inf, inferior; pl, planum; pren, prenasal; sup, superior.

The solum nasi (= planum basale sensuHiggins, 1921; = nasal floor sensuHall & Larsen, 1998) is an unpaired, horizontal plate that forms the floor of the nasal capsules (Figs 1, 3). In metamorphic individuals, the solum nasi is completely cartilaginous, but in juveniles and adults, the cartilage posterior and medial to the fenestra endochoanalis is replaced by spongy bone. Medially, the solum nasi is continuous with the septum nasi; the anteriormost part is connected to the mid-portion of the septum and the posteromedial portion is attached to the ventral margin of the septum (Figs 6, 7). The posterior margin of the solum nasi is continuous with the postnasal wall. Each half of the solum nasi (when divided mediolaterally) is pierced by the fenestra nasobasalis, the fenestra endochoanalis, and an opening anterior to the fenestra endochoanalis. Medial to the fenestra endochoanalis, the dorsal surface of the solum nasi is elevated significantly, forming a skeletal support for the eminentia olfactoria (= solar ossification of eminentia sensuRamaswami, 1937; = turbinal fold sensuEstes, 1970; Hall & Larsen, 1998); these combined structures occlude most of the posterior nasal cavity (Fig. 7E,F). The skeletal support for the eminentia olfactoria appears after metamorphosis, and extends from a level just posterior to the posterior ramus of the septomaxilla to the posterior region of the nasal capsule, anterior to the olfactory foramen. It is extensively ossified by spongy bone, except for the anterior terminus, which remains cartilaginous even in large individuals. The apical portion of the skeletal support for the eminentia olfactoria is slightly wider than the base and is medially concave throughout its entire length. In addition, the skeletal support is relatively narrow anteriorly and projects forward as a process (= capsular process sensuEstes, 1970), whereas its posterior portion widens to about twice the width of the anterior end. In addition to the thickening of the solum nasi, the eminentia olfactoria is also supported by the upturned lateral margin of the vomer. Most of the solum nasi is underlain by the large vomer.

Fig. 3.

Fig. 3

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Ventral view of left nasal capsule of Stage-46 S. multiplicata(KU 292097). Blue denotes nasal cartilage; red denotes bone (represented as transparent to show underlying structures); black denotes fenestrae/foramina. ant, anterior; c, cartilage; f, foramen; inf, inferior; lat, lateral; max, maxillary; med, medial; orbiton, orbitonasal; p, process; pl, planum; post, posterior; pren, prenasal; sup, superior; triang, triangulare.

The anterior nasal walls are paired, vertical, and relatively straight plates that form the anterior limit of the nasal capsules (Figs 1, 6A,B). Each anterior nasal wall (= solum anterius sensuHiggins, 1921; = prenasal wall sensuRoček, 1989) is continuous with the septum nasi medially, the tectum nasi dorsomedially, the solum nasi ventrally, and the crista subnasalis posterolaterally. The ventromedial margin of the anterior nasal wall adjacent to the septum forms the anterior edge of the fenestra nasobasalis. Most of the anterior nasal wall is cartilaginous, although in large individuals (e.g. KU 18450), ossification from the septum nasi may invade the medial portion of the wall.

The paired postnasal walls are anteriorly concave structures that form the posterior limit of the nasal capsules. Each postnasal wall (= pars plana nasi sensuGaupp, 1896; Michael, 1961) consists of two parts: a ventral piece, the lamina orbitonasalis, and a dorsal plate that is part of the tectum nasi. Adjacent to the septum nasi, the postnasal wall is pierced by a large medial orbitonasal foramen for the medial branch (= ramus medialis nasi) of the ophthalmic nerve. Dorsally, the crista supraorbitalis extends posteriorly as a short, blunt process from the posteromedial margin of the postnasal wall, on the anterodorsal corner of the orbit above the medial orbitonasal foramen. The crista supraorbitalis is clearly discernible in postmetamorphic and juvenile individuals, but is obscured in large adults by the frontoparietal. In metamorphic individuals the postnasal wall is cartilaginous (Figs 1–4). Distal to the postnasal wall is the cartilaginous planum triangulare (= pars plana sensuParker, 1876; = lamina externa sensuHiggins, 1921; = processus antorbitalis sensuRoček, 1981; = triangular plane sensuHall & Larsen, 1998). The planum triangulare invests the anteromedial portion of the pars facialis of the maxilla and bears both an anterior and a posterior maxillary process (Figs 2–4). The anterior maxillary process is small, projects anteriorly to the level of the anterior margin of the fenestra nasolateralis, and invests the medial face of the pars facialis of the maxilla. The posterior maxillary process extends posteriorly, investing the maxilla laterally and the anterior ramus of the pterygoid medially. The posterior end of this process is synchondrotically fused to the pterygoid process. Proximally, the planum triangulare is pierced by the lateral orbitonasal foramen housing the lateral ramus (= ramus lateralis nasi proper) of the ophthalmic nerve.

The alary cartilages are paired, cup-shaped structures (Figs 1, 2, 4, 6C,D). Each alary cartilage (= cartilago alaris sensuGaupp, 1896; Stadtmüller, 1936; Baldauf, 1958; Baldauf & Tanzer, 1965; Roček, 1981, 2003; = cartilago alaris nasi sensude Beer, 1985; = alinasal cartilage sensuHiggins, 1921; = processus alaris sensuJarvik, 1942) provides support to the anterior rim of the apertura nasalis externa and closes the apertura during the breathing cycle. In addition, this cartilage serves as the anterodorsal margin of the fenestra endonarina. The alary cartilage is medial and dorsal to the septomaxilla and anterolateral to the oblique cartilage. The dorsal margin of the alary cartilage extends posteriorly as a rectangular plate. The anterior, convex face of the alary cartilage is synchondrotically fused to the superior prenasal cartilage. A cartilaginous bridge connects the ventromedial edge of the alary cup to the anterior nasal wall, and the dorsomedial margin of the alary cartilage is fused to the anterolateral edge of the oblique cartilage.

The paired superior prenasal cartilages are rod-like structures, round in cross-section (Figs 1, 2, 4, 6C,D). Each superior prenasal cartilage (= processus prenasalis superior lateralis sensuStadtmüller, 1936; Jarvik, 1942; de Beer, 1985; = cartilago prenasalis superior sensuBaldauf & Tanzer, 1965; Roček, 1981; 2003) extends anteroventrally to abut the dorsomedial margin of the posterior face of the alary process of the premaxilla. The function of this cartilage is to transmit the movement of the premaxilla to the alary cartilage during the closure of the apertura nasalis externa. [See Jurgens (1971) and Gans & Pyles (1983) for a detailed description of the mechanism.]

The paired inferior prenasal cartilages are ventrally arcuate, forming an angle of about 90º (Figs 1, 3, 4). Proximally, each inferior prenasal cartilage (= processus prenasalis inferior lateralis sensuStadtmüller, 1936; Jarvik, 1942; de Beer, 1985; = cartilago prenasalis inferior sensuBaldauf, 1958; Baldauf & Tanzer, 1965; Roček, 1981, 2003; = inferior prenasal process sensuReiss, 1998) is flat in cross-section (Fig. 7A,B), and fuses to the ventral portion of the solum nasi at the level of the anterior margin of the paries nasi. This fusion is not restricted to the terminus of the inferior prenasal cartilage, but also involves a few millimeters of its lateral margin. Distally, the inferior prenasal cartilage is club-shaped, round in cross-section, and abuts the ventral three-fourths of the posterior aspect of the alary process of the premaxilla (Figs 1, 4, 6). The function of the inferior prenasal cartilage is to provide support to the premaxilla; in addition, it is used as a pivot during the closing/opening of the apertura nasalis externa. The inferior prenasal cartilages are about 2.5× longer than the superior prenasal cartilages.

The cristae intermediae are horizontal plates located ventral to the alary cartilage. Each crista intermedia serves as the skeletal boundary between the ventromedial part of the cavum principale and the cavum inferius (Figs 2, 4, 6E,F). The medial margin of the crista intermedia is fused to the mid-section of the septum nasi, just dorsal to, and ventrally continuous with, the solum nasi. The posterior margin of the crista intermedia is concave and located at a level posterior to the posterior margin of the alary cartilage. In metamorphic individuals (e.g. KU 292097), the crista intermedia is cartilaginous, but in adults it is ossified. The lateral margin of the crista intermedia bears two laminae: the lamina superior (= lamina superior cristae intermediae sensuStadtmüller, 1936; Roček, 1981, 2003) and the lamina inferior (= lamina inferior cristae intermediae sensuStadtmüller, 1936; Roček, 1981, 2003). Both laminae are unattached to the crista intermedia at the level of the transverse midline of the fenestra endonarina (Figs 2, 4), but are connected to each other almost to the level of the posterior margin of the crista subnasalis. The lamina superior serves as the roof of the cavum medium and the lamina inferior serves as its floor (Fig. 6E–H). The anterior ramus of the septomaxilla is attached to the posterior margin of the lamina superior. The lamina inferior lies ventral to the septomaxilla; its posterior margin extends more posteriorly than the posterior margin of the lamina superior, it is slightly emarginated, and is parallel to the septum nasi. Laterally, the lamina inferior curves dorsally, thus forming the lateral wall of the cavum medium. Anteriorly, the dorsal margin of the lateral wall of the lamina inferior is fused to the lateral edge of the lamina superior. Posteriorly, there is no connection with the lamina superior. Thus, the dorsal margin of the lamina inferior is free and ends in a medially concave terminus; the most posterior part of this terminus is fused to the planum terminale.

A semi-circular, relatively broad cartilaginous sheet lies laterally to each side of the septum nasi. This cartilage, called the oblique cartilage (= cartilago obliqua sensuGaupp, 1896; Stadtmüller, 1936; Baldauf, 1958; Baldauf & Tanzer, 1965; Jurgens, 1971; Roček, 1981, 2003; = cartilago obliqua nasi sensude Beer, 1985; = dorsal process sensuHiggins, 1921; = lamina obliqua sensuJarvik, 1942; = cartilago obliquo sensuTrueb, 1970), traverses from the ventrolateral to the dorsomedial aspect of the nasal capsule (Figs 1, 2, 4). The oblique cartilage complements the tectum nasi by forming a dorsal cover to the anterior nasal capsule. The anterior end of the oblique cartilage terminates in a small, free process. Lateral to this process, the anterior margin of the oblique cartilage forms the medial and posterior limits of the fenestra endonarina; the anterodorsal part of the margin provides support to a section of the rim of the external narial aperture. Medial to the anterior process, the margin of the oblique cartilage is fused to the anteriormost portion of the lateral margin of the tectum nasi. The posterolateral margin of the oblique cartilage forms the anterior margin of the fenestra nasolateralis. The ventrolateral portion of the oblique cartilage, the planum terminale (= pars terminalis sensuJarvik, 1942; = terminal plane sensuHall & Larsen, 1998), is a vertical plate of cartilage that forms the lateral wall of the nasal capsule. The planum terminale is fused anteriorly to the lamina inferior and posteriorly to the paranasal commissure (= processus lingularis sensuMichael, 1961; = connection between lamina nariochoanalis and postnasal wall sensuJurgens, 1971; = commissura lateralis nasi sensuHaas, 1996). The paranasal commissure is a cartilaginous bridge that extends posteriorly to the anterolateral corner of the posterior nasal wall (Figs 2, 4).

The parietes nasi are paired, elongate, ventrolaterally-oriented cartilaginous structures that extend to the premaxilla-maxilla articulation (Figs 1–4). Each paries nasi merges with the solum nasi medially, the anterior nasal wall anteriorly, and the lamina inferior dorsally. Ventral to the paries nasi, the round crista subnasalis (= subnasal crest sensuHall & Larsen, 1998) abuts the posterior end of the premaxilla anteroventrally and the medial wall of the pars facialis of the maxilla posteroventrally.

Each nasal capsule has five openings: fenestra endonarina communis, fenestra nasolateralis, fenestra nasobasalis, fenestra endochoanalis, and opening anterior to the fenestra endochoanalis (Fig. 5). The fenestra endonarina communis (= apertura nasalis externa sensuMichael, 1961; = fenestra narina sensude Beer, 1985; Haas, 1996) is located on the anterolateral aspect of the nasal capsule. This fenestra is delimited by the alary cartilage, the anterior nasal wall, the lateral wall of the lamina inferior, the oblique cartilage, and the tectum nasi. The fenestra endonarina communis opens into the cavum principale. The fenestra nasolateralis (= fenestra dorsalis sensuMichael, 1961; = lateral nasal fenestra sensuHall & Larsen, 1998) is an elongate opening located on the dorsal and lateral sides of the nasal capsule. This fenestra has a small, rostrally-directed indentation on its anterodorsal corner. The fenestra nasolateralis is delimited by the posterior margin of the oblique cartilage, the dorsal margin of the paranasal commissure, the posterolateral margin of the tectum nasi, and the anterior margin of the postnasal wall. The fenestra nasobasalis (= foramen apicale sensuGaupp 1896; Stephenson, 1951; = foramen epiphaniale sensuGaupp, 1896; = basal fenestra sensuHiggins, 1921; = nasobasal fenestra sensuHall & Larsen, 1998) is a small opening at the anterior tip of the nasal capsule, on the floor of the cavum inferius. This fenestra is lateral to the septum nasi, posterior to the anterior nasal wall, and anteromedial to the point of attachment of the inferior prenasal cartilage. The fenestra endochoanalis (= endochoanal fenestra sensuHall & Larsen, 1998) is located on the floor of the nasal capsule, and opens into the posterolateral area of the cavum principale. The anterior, medial, and posterior margins of the fenestra are formed by the solum nasi; the lateral margin is formed by the paranasal commissure. The anterior, medial, and posterior margins of the fenestra endochoanalis are underlain by the large vomer. Anteromedial to the fenestra endochoanalis, there is a small, diagonally-oriented fenestra in the nasal floor. The posteromedial margin of this fenestra is bony, whereas the anterolateral edge is cartilaginous and is covered ventrally by the anterior process of the vomer. This fenestra is present only in large adults, indicating that it may arise as the result of cartilage resorption.

Intramembranous osseous elements form directly from mesenchyme without a cartilaginous precursor, and often are labeled ‘investing bones’ (Maglia et al. 2007b). One pair of intramembranous bones, the septomaxillae, is located within the nasal capsules, embedded in the nasal cartilages. Other four pairs of intramembranous bones, i.e. the premaxillae, maxillae, nasals, and vomers, cover and partially shield the olfactory capsules. The function of the septomaxillae is to provide support to the anterior end of the nasolacrimal duct (Duellman & Trueb, 1994). Each septomaxilla (= turbinale sensuSarasin & Sarasin, 1890; = intranasale sensuGaupp, 1896; = nariale sensuWegner, 1922; Marcus et al. 1935; = internasale sensude Jongh, 1968) is located dorsal to the lamina inferior, posteriorly and ventrolaterally to the corresponding alary cartilage (Figs 2, 4). The septomaxilla is spiral shaped, with the anterior ramus being more ventral than the posterior one (Fig. 8). The posterior ramus is relatively straight, and its free medial terminus bears dorsal and ventral crests. The anterior ramus is more slender than the posterior ramus and its entire leading edge is fused to the lamina superior (Fig. 6G,H).

Fig. 8.

Fig. 8

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Dorsal view of the adult septomaxilla of S. multiplicata (KU 97355). Red denotes bone.

The premaxillae are paired, tooth-bearing bones that form the anterior margin of the maxillary arcade (Figs 1–4). They provide bony protection for the anterior portion of the nasal capsules and play a key role in the closing of the aperturae nasales externae (see de Jongh & Gans, 1969). The premaxillae are narrowly separated from one another medially, whereas laterally they are slightly overlapped by the maxillae. Each premaxilla (= praemaxilla sensuRoček, 2003) consists of three parts: (1) an anterolateral vertical plate, the pars dentalis, (2) a posterior lingual ledge, the pars palatina, and (3) a dorsal process, the alary process (= dorsal process sensuGaudin, 1969; = pars facialis sensuRoček, 1981). Two of the premaxillary parts, the pars palatina and the alary process, are in direct contact with the nasal cartilages. The pars palatina arises near the posteroventral margin of the pars dentalis and is narrower at its mid-region than at the posterolateral end of the premaxilla. Medially, the pars palatina bears a palatine process (= median palatal squame sensuJurgens, 1971). The posterolateral portion of the pars palatina is braced by the anteroventral end of the crista subnasalis. The well-developed alary processes extend vertically from the partes dentales, and their distal ends have a slightly lateral orientation. The posterior (inner) face of each alary process is concave and is invested dorsomedially by the superior prenasal cartilage and ventromedially by the inferior prenasal cartilage (Figs 1, 4, 6).

The maxillae are paired, tooth-bearing bones located on the lateral sides of the maxillary arcade, posterior to the premaxillae (Figs 1–4). These elements provide bony protection for the ventrolateral aspect of the nasal capsules. Like the premaxillae, each maxilla also has three distinct sections: (1) a dentate element, the pars dentalis (= crista dentalis sensuRoček, 2003), (2) a palatal ledge, the pars palatina, and (3) a dorsolateral facial flange, the pars facialis. Of the three sections, the pars facialis is most closely associated with the nasal cartilages. The pars facialis is well developed, lacks pre- and postorbital processes, and extends for approximately half the length of the maxilla, from the anterior margin of the bone to the level of the planum triangulare. The anterior margin of the pars facialis invests the entire posterior margin of the corresponding paries nasi (Figs 1, 3, 4). In addition, the processus frontalis of the maxilla (= processus palatinus sensuRoček, 1981, 2003; = palatine process of pars facialis of maxilla sensuCannatella, 1985; = preorbital process sensuHall & Larsen, 1998), a process that projects medially from the posterodorsal corner of the pars facialis, invests the planum triangulare and postnasal wall ventrally and articulates with the sphenethmoid.

The nasals are paired, trapezoidal bones that overlie the medial and posterolateral region of the olfactory capsules, covering the tectum nasi and fenestrae nasolaterales (Figs 1, 2, 4). Medially, the nasals are separated widely from one another; thus, in cleared-and-stained specimens, the septum nasi is obvious between them. The nasals overlap the sphenethmoid and postnasal walls, but do not come in contact with the premaxillae or the maxillae. A rostral process is absent, but a short, blunt maxillary process (= maxillary spine sensuGaudin, 1969) extends laterally toward the maxillary arcade.

The vomers (= prevomer sensuRamaswami, 1935; Baldauf, 1958; Trueb, 1970; 1973) are paired, palatal bones that lie ventral to the olfactory region and help to floor the nasal capsules (Fig. 3). In Spea, these bones underlie the sphenethmoid and are medially separated from one another. Each vomer is composed of five distinct portions, which are the anterior, dentigerous, prechoanal, postchoanal, and posteromedial processes. The anterior process of the vomer is flat and rectangular, and extends anterolaterally, reaching the premaxilla–maxilla articulation. The prechoanal and postchoanal processes (= circumchoanal processes sensuClarke, 1988) extend laterally from the anterior process; their bifurcation occurs at about the level of the dentigerous process. At their area of bifurcation, the lateral margins of the prechoanal and postchoanal processes are upturned to provide support to the eminentia olfactoria (Fig. 7). The prechoanal process is small and forms the medial portion of the anterior margin of the ovoid apertura nasalis interna. A thin sheet of bone extends between the posterolateral margin of the anterior process and the anterior margin of the prechoanal process. The postchoanal process is long and slender, and invests most of the ventral surface of the postnasal wall and planum triangulare; its lateral terminus rests at the level of the planum triangulare. The postchoanal process forms the entire posterior margin of the apertura nasalis interna. The transverse dentigerous process lies medial to the apertura nasalis interna and usually bears about five true teeth. The posteromedial process is short and extends posteriorly just beyond the level of the dentigerous process.

Larval skeletal morphology at Gosner Stage 30

At Gosner Stage 30 – a stage in which all the adult nasal capsule structures are still absent – the ethmoid region composes the anterior half of the chondrocranium and is formed by the trabecular plate and the trabecular horns (Fig. 9). The trabecular plate (= ethmoid plate sensuSokol, 1981; Wiens, 1989; Wild, 1997; = planum trabeculare anticum sensuHaas, 1996; = planum ethmoidale sensuSheil, 1999; Sheil & Alamillo, 2005) is located anterior to the braincase. The posteriormost end of the trabecular plate constitutes the planum internasalis (sensuBanbury & Maglia, 2006), which forms the anterior wall of the braincase. The dorsal margin of the planum internasalis is formed by the taenia ethmoidalis, which is slightly more elevated than the trabecular plate and describes an approximately 90° angle with the dorsal surface of the plate when observed in lateral profile. The taenia ethmoidalis is pierced by a pair of large olfactory foramina, which are widely separated from one another. Posterolaterally, the taenia ethmoidalis is continuous with the taenia tecti marginalis (= lateral roof sensuHall & Larsen, 1998). The planum internasalis is posterolaterally continuous with the lateral walls of the braincase (cartilago orbitalis).

Fig. 9.

Fig. 9

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Dorsal view of anterior chondocranium of Stage-30 S. multiplicata(KU 292081). Gray denotes cartilage; black denotes foramina. artic, articularis; c, cartilage; f, foramen; l, ligament; lat, lateral; quad, quadrati; palatoquad, palatoquadrate; p, process.

A pair of rod-like, arcuate trabecular horns (= cornua trabecularum sensuRoček, 1981, 1989, 2003; de Sá & Trueb, 1991; Haas, 1996, 2003) project anterolaterally from the anterior margin of the trabecular plate. The angle of divergence of the trabecular horns from the midline of the trabecular plate is approximately 30°, describing a V-shape in dorsal view. The trabecular horns are medially connected by a thin, triangular sheet of cartilage, which in some specimens may bear a short medial process (= prenasal process sensuHall & Larsen, 1998). In some specimens, this sheet of cartilage is pierced by one or two small openings. The horns are relatively short (about one-third the length of the ethmoid region) and uniform in width throughout their length. In addition, they lack a prominent ventral curvature; hence, the position of the mouth is almost terminal. The anterior end of each trabecular horn is truncate and articulates with the corresponding suprarostral cartilage. In cross-section, the horn is crescentic with a convex dorsal surface. On the lateral margin of each horn, at about the level of fusion to the trabecular plate, it bears a small but distinct triangular process, the processus lateralis trabeculae, which serves as a site for the attachment of the quadratoethmoid ligament (= ligamentum quadratoethmoidale sensuSokol, 1975; Haas, 1996).

The wide quadratocranial commissure (= commissura quadratocranialis anterior sensuSokol, 1975; Haas, 1996; 2003; Wild, 1997) extends anterolaterally to the level of the posterior ethmoid region, connecting the quadrate (i.e. the anterolateral portion of the palatoquadrate cartilage sensuSokol, 1981) to the floor of the braincase. The anteromedial margin of the quadrate forms the pars articularis quadrati, which articulates with Meckel's cartilage. The quadratocranial commissure bears a low ridge across its median portion, the processus quadratoethmoidalis (= quadratoethmoid process sensuCannatella, 1999), to which the posterior end of the ligamentum quadratoethmoidale (= quadratoethmoid ligament sensuCannatella, 1999) is attached. The ligamentum cornu-quadratum (= ligamentum cornu-quadratum laterale sensuRoček, 2003; = ligamentum circumoralis sensuSokol, 1981; = lateral circumoral ligament sensuCannatella, 1999) extends from the lateral tip of the anterior end of the trabecular horn to the anterolateral margin of the pars articularis quadrati of the palatoquadrate. The upper jaw is supported by the suprarostral cartilage (= cartilago labialis superior sensuRoček, 1981, 2003; Haas, 1996, 2003), which is formed by two vertical, unipartite plates. At this stage, the suprarostral plates are unfused medially; they synchondrotically fuse to each other at about Stage 32 (Stage 34 fideBanbury & Maglia, 2006). Dorsally, the articular process of each suprarostral is synchondrotically connected to the flat anterior end of the corresponding trabecular horn.

Skeletal development

Although the components of the olfactory region are functionally adult structures, they commence formation early in development. The elements that form the nasal capsules in adults develop dorsal to the larval skeletal structures. In S. multiplicata, the initial nasal elements form at Gosner Stage 31, in S. intermontana at Stage 32, and in S. bombifrons at Stage 36. However, in each species most major skeletal developmental events occur during Stages 38–42, although the larval structures of the ethmoid region (e.g. trabecular horns, suprarostrals) are still present and functional during those stages. By metamorphosis, the nasal capsules attain a configuration that closely resembles the adult morphology; however, most structures are still cartilaginous, whereas in adults they are heavily ossified.

At Stage 31, the septum nasi appears as a low and thin vertical plate of newly formed cartilage between the olfactory foramina, dorsal to the midsection of the trabecular plate. The septum nasi is posteriorly continuous with the taenia ethmoidalis. The laminae orbitonasales also appear during Stage 31 as a pair of minute, triangular cartilages. Each arises at the posterolateral margin of the ethmoid plate, lateral to the olfactory foramen and above the area of attachment of the quadratocranial commissure. By Stage 36, each lamina orbitonasalis becomes synchondrotically fused to the lateral wall of the braincase, lateral to the olfactory foramen (Fig. 10A). In addition, the tectum nasi starts to chondrify, becoming continuous with the dorsomedial margin of each lamina orbitonasalis; these two structures, the ventral lamina orbitonasalis and the dorsal tectum nasi, thus form the adult postnasal wall.

Fig. 10.

Fig. 10

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Dorsal view of the right nasal region of S. multiplicata, Stages (A) 36 (KU 292087), (B) 37 (KU 292088), and (C) 38 (KU 292089). Blue denotes nasal cartilage; gray denotes cartilage not contributing to the nasal capsule; red denotes bone; black denotes foramina; white denotes ligaments. ant, anterior; c, cartilage; orbiton, orbitonasal; pren, prenasal; premax, premaxilla.

At Stage 37, the alary cartilages appear as small, triangular elements dorsal to each processus lateralis trabeculae (Fig. 10B). By early Stage 38, a minute superior prenasal cartilage is visible on the anterior margin of each of the small alary cartilages. The superior prenasal cartilages elongate, and by late Stage 38 their anterior ends are flanked dorsolaterally by a thin sliver of bone, the primordium of the alary process of the premaxilla (Fig. 10C).

The anterior nasal wall and paries nasi/crista subnasalis chondrify during early Stage 39 (Fig. 11A). The anterior nasal walls are medially continuous with the septum and tectum nasi and extend laterally. Each wall is ventral to the alary cartilage and superior prenasal cartilage, and reaches the level of the processus lateralis trabeculae. At this stage, the anterior nasal walls remain unfused to the trabecular plate. Each paries nasi appears as an expanded, small cartilaginous plate dorsal to the site of attachment of the quadratoethmoid ligament. Small, sickle-shaped nasal bone primordia with irregular margins are visible on the anterolateral margins of the tectum nasi. By the end of Stage 39, the anterior nasal wall ventrally fuses to the trabecular plate (Fig. 11B). Most of the trabecular plate becomes, at this stage, the solum nasi, although some lateral erosion and cartilage regrowth occurs. Also, a pair of V-shaped septomaxillae are visible just posterior to the lateral margin of each alary cartilage, and a pair of small, irregularly shaped primordia of the pars facialis of the maxillae appear dorsolateral to each quadratoethmoid ligament. In addition, the inferior prenasal cartilages have chondrified; each appears as a slightly arcuate rod of cartilage dorsal to the midline of the trabecular horn and ventral to the superior prenasal cartilage and alary process of the premaxilla. The posterior end of this cartilage is fused to the transitional zone between the anterior nasal wall and the solum nasi, whereas the anterior terminus is free. The alary process of each premaxilla widens and is invested by the anterior end of the superior prenasal cartilage. Also during this stage the anterior process of the postnasal wall develops on the anterolateral aspect of each lamina orbitonasalis; in the adult Spea, the anterior process of the postnasal wall is synchondrotically connected to the paranasal commissure. The distal end of the lamina orbitonasalis expands laterally and slightly anteriorly to form the planum triangulare.

Fig. 11.

Fig. 11

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Dorsal view of the right nasal region of S. multiplicata, Stages (A) early 39 (KU 292090), (B) late 39 (KU 292090), and (C) 40 (KU 292091). Blue denotes nasal cartilage; gray denotes cartilage not contributing to the nasal capsule; red denotes bone (represented as transparent to show underlying structures); black denotes foramina; white denotes ligaments. ant, anterior; c, cartilage; inf, inferior; max, maxillary; p, process; pl, planum; post, posterior; postnas, postnasal; pren, prenasal; subn, subnasalis; triang, triangulare.

By Stage 40, all the remaining major elements of the nasal capsules have chondrified (i.e. crista intermedia, laminae superior and inferior, and oblique cartilage), and most structures have attained a morphology resembling in part that of the adult (Fig. 11C). The crista subnasalis slightly invests the pars facialis of the maxilla and the alary cartilage has a clear connection with the anterolateral margin of the tectum nasi and the anterior nasal wall. At this stage, the ventral margin of the planum terminale (i.e. the ventral portion of the oblique cartilage) ends freely. A thin sheet of cartilage connects the posterolateral margin of the oblique cartilage with the anterolateral margin of the tectum nasi. A small, anterior protuberance, which in the adult will become the anterior maxillary process, is visible on the anterior end of the developing planum triangulare. In addition, a minute posterior maxillary process is evident in the posterolateral margin of the planum triangulare. By Stage 41, the planum terminale attains its anterior synchondrotic connection to the lamina inferior. Also, the cartilaginous plate that connects the oblique cartilage and tectum nasi starts to erode behind the posterodorsal edge of the oblique cartilage, thus forming an incipient fenestra nasolateralis (Fig. 12A).

Fig. 12.

Fig. 12

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Dorsal view of the right nasal region of S. multiplicata, Stages (A) 41 (KU 292092), (B) 42 (KU 292093), and (C) 44 (KU 292095). Blue denotes nasal cartilage; gray denotes cartilage not contributing to the nasal capsule; red denotes bone (represented as transparent to show underlying structures); black denotes fenestrae/foramina; white denotes ligaments. ant, anterior; fac, facialis; fen, fenestra; max, maxillary; nasolat, nasolateralis; p, process; post, posterior.

During Stages 42 and 43, structural remodelling of the chondrocranium becomes more extensive (Fig. 12B). For example, the growing ossification of the septomaxilla starts to suggest a spiral shape. In some specimens, the developing anterior maxillary process of the planum triangulare invests the posterior margin of the pars facialis of the maxilla. Also, a more substantial synchondrotic connection forms between the developing posterior maxillary process and the anterior portion of the quadratocranial commissure (which will become the adult pterygoid process). The trabecular horns shift their orientation; their distal ends move to a more ventrolateral position. In addition, the suprarostral cartilages begin to regress medially. Also, by these stages the distal terminus of each inferior prenasal cartilage invests the alary process of the premaxilla. The posterior terminus of the inferior prenasal cartilage, in contrast, has begun to migrate posteriorly. The transfer of the posterior terminus of the inferior prenasal cartilage from an anterior position (seen in Stage 39) to a slightly more posterior position is the result of a differential growth of the anterior nasal wall. The pars dentalis of the premaxilla starts to develop during these stages as a thin and short sliver of bone ventral and lateral to the pars dorsalis. In addition, the plate of cartilage between the oblique cartilage and tectum nasi continues to erode, allowing for the expansion of the fenestra nasolateralis; only the ventral portion of the former cartilaginous plate remains chondrified, connecting the planum terminale with the anterior process of the postnasal wall. Herein, this cartilaginous connection, which forms the ventral margin of the fenestra nasolateralis, is called paranasal commissure (see above). The developing nasal, which by these stages has increased in size about three times that of its initial appearance, covers the dorsal portion of the fenestra.

By Stage 44, the cartilaginous skeleton of the olfactory region presents the adult morphology (Fig. 12C). The only modifications that occur after this stage and until adulthood are changes in the size and morphology of the intramembranous investing bones. For example, deposition of bone ventral and medial to the pars facialis of the maxilla results in the formation of the pars dentalis and pars palatina, respectively. Also, the processus frontalis starts to form as a small dorsal projection at the posterolateral corner of the pars facialis of the maxilla, parallel to the anterior margin of the planum triangulare and postnasal wall. Soon after appearing, the free end of the processus frontalis rotates about 45º and curves down ~30º; thus, the anterior side of the process becomes dorsal and the terminus faces medially (instead of dorsally). The maxillary process of the nasal bone arises after metamorphosis.

Discussion

The internal elements of the nasal capsules – nasal cartilages and septomaxillae – are de novo, functionally adult structures that form dorsal to the larval skeleton of the ethmoid region. The only larva-derived structures of the adult nasal capsule are the medial and posterior aspects of the solum nasi, which are formed by the trabecular plate. In the adult, the nasal capsules have a dual function – to house the olfactory organ and to form the beginning of the respiratory system. In the tadpoles, however, the nasal capsules do not have a respiratory function, although they seem to be lined with olfactory epithelium, which probably allows them to sample different olfactory environments, and in some cases, aids in the recognition of kin (Ultsch et al. 1999).

In S. multiplicata the chondrification process of the nasal cartilages begins during mid-premetamorphosis (sensuHall & Larsen, 1998). The septum nasi and lamina orbitonasalis start to form at Gosner Stage 31 and the tectum nasi at Stage 36. The alary cartilage and superior prenasal cartilage are the first elements of the anterior nasal cartilages to chondrify at Stage 37. These are followed by the anterior nasal wall and crista subnasalis by early Stage 39, and the inferior prenasal cartilage by late Stage 39. By Stages 40/41, all the remaining major elements of the nasal capsules are formed, and thus the ethmoid region is composed of the larval structures ventrally and the adult structures dorsally. These observations are comparable to those presented by Hall & Larsen (1998) for S. intermontana, but contrast with those reported for other species, such as Rana temporaria (de Jongh, 1968) and Pyxicephalus adspersus (Haas, 1999). For those species, the reported time of chondrification of the anterior nasal cartilages is around Gosner Stage 40 (de Jongh, 1968; Haas, 1999), but all elements chondrify almost simultaneously. This incongruence seems to reflect a precocious onset of chondrogenesis of the nasal skeleton of Spea when compared to other species.

Lack of a clear understanding of the homology of the wall that separates the olfactory region and orbit in anurans has resulted over time in a large and confusing collection of terms used to designate these anatomical structures, which includes both non-homologous structures with the same name and homologous structures with different names. For example, some authors (e.g. Trueb, 1970; Trueb & Hanken, 1992; da Silva, 1998; Sheil & Alamillo, 2005) refer to the entire wall as the planum antorbitale, whereas Roček (1981) used the term to designate the portion of the wall anterior to the orbit that reaches the maxilla. The same author (Roček, 1981), however, stated that the commonly used terms planum antorbitale (antorbital plane) and processus (cartilago) antorbitalis should be avoided because this wall is within the ethmoidal region, and therefore does not have any relation with the orbit. Furthermore, Roček (1981) suggested that the term lamina orbitonasalis (sensuSokol, 1981) should also be avoided because it designates only the larval structure that separates the nasal capsule and orbit, which he does not consider fully homologous with the structure of the same function in the adult.

Our observations show that in Spea the development of the entire posterior area of the olfactory region occurs in three main phases. The first phase includes the formation of the lamina orbitonasalis posterolateral to the ethmoid plate. The second phase is represented by the chondrification of the tectum nasi dorsal to the lamina orbitonasalis; thus, the tectum nasi and lamina orbitonasalis together form the compound postnasal wall. And finally, the third phase comprises the development of the cartilaginous planum triangulare. The planum triangulare, however, is not part of the postnasal wall because it does not form a hind wall for any portion of the nasal capsule. These results clearly indicate that in Spea the larval lamina orbitonasalis and the adult postnasal wall are partially homologous. They also suggest that the lamina orbitonasalis and the antorbital process (sensuTrueb, 1970; Sheil & Alamillo, 2005; and other authors) also are partially homologous structures. Moreover, based on developmental and morphological data, Reiss (1998) not only showed that the anuran lamina orbitonasalis and postnasal wall are partially homologous, but also suggested that the anuran lamina orbitonasalis is homologous with the urodele antorbital process.

To establish a more permanent terminology for this element, we propose a more precise use of existing terms in consideration of homology. The term ‘postnasal wall’ should designate the compound adult structure that, in anurans, forms the posterior limit of the nasal capsule. The term ‘planum triangulare’ should be used to designate the portion of the lamina orbitonasalis distal to the postnasal wall, which in the adult anuran invests the maxilla medially and bears anterior and posterior maxillary processes. And the terms ‘lamina orbitonasalis’ and ‘planum antorbitale’ should be considered synonyms, and their usage should be restricted to designate the horizontal bar that, in the tadpole, develops at the posterolateral margin of the ethmoid plate and, in the adult, forms the ventral aspect of the postnasal wall and the planum triangulare.

It has been reported that in some anurans (e.g. Leiopelma archeyi) the inferior prenasal cartilages are formed by the modified trabecular horns of the larva (Stephenson, 1951). In other taxa (e.g. pelobatids), the origin of the inferior prenasal cartilages has been suggested to be the suprarostral cartilages (Roček, 1981). In S. multiplicata, however, the inferior prenasal cartilages are de novo chondrifications (i.e. they appear dorsal to, and independent of, the larval skeleton). At the end of Gosner Stage 39, each inferior prenasal cartilage unmistakably is observed dorsal to the midline of the trabecular horn, and the suprarostral cartilages have not started to erode (erosion begins during Gosner Stage 42) and are still functioning as the skeletal support for the upper beak of the tadpole. Wiens (1989), who studied the development of S. bombifrons, could not confirm or refute Roček's (1981) observation, but Hall & Larsen (1998) reported that in S. intermontana the suprarostral cartilages do not contribute to the formation of the inferior prenasal cartilage. In addition, Hall & Larsen (1998) stated that in S. intermontana the remnants of each trabecular horn seem to be incorporated into the corresponding inferior prenasal cartilage by the end of Gosner Stage 43. Our data on S. multiplicata, however, do not corroborate their observation. An origin of the inferior prenasal cartilages similar to that described herein also has been observed in Gastrotheca marsupiata (Haas, 1996).

The nasal lateral wall of anurans usually bears two or three openings. The anterior fenestra endonarina communis forms as the result of the development of skeletal structures around the nostril area. The posterior fenestra(e) form through cartilage erosion prior to metamorphosis. Depending on the degree of cartilage resorption, the adult nasal capsule may possess a single, lateral fenestra – the fenestra nasolateralis – or two fenestrae – the fenestra dorsalis and fenestra lateralis – separated from each other by a medial cartilaginous bridge (Jurgens, 1971). A single fenestra is present in Acris crepitans (Maglia et al. 2007b), Ascaphus truei (Jurgens, 1971), Barbourula busuangensis (Jurgens, 1971), Bufo regularis (Michael, 1961), Hyla regilla (Jurgens, 1971), Leiopelma archeyi (Stephenson, 1951), Pelobates fuscus (Roček, 1981), Pyxicephalus adspersus (Sheil, 1999), and Rana temporaria (de Jongh, 1968), among others. Two fenestrae are present in Brachycephalus ephippium (McLachlan, 1943) and Gastrotheca riobambae (Haas, 1996). In Spea, the lateral walls of the nasal capsule are fairly complete in tadpoles of Gosner Stages 40 and 41. The tectum nasi, however, is resorbed between Stages 42 and 44, resulting in a single fenestra nasolateralis.

Ventrally, the fenestra nasolateralis of Spea is bound by a cartilaginous bridge, the paranasal commissure, which connects the planum terminale and the posterior nasal wall. A similar structure is present in Bufo regularis (Michael, 1961) and Pelobates fuscus (Roček, 1981). However, cartilage resorption may be so extensive in some cases that even the paranasal commissure disappears. For example, it is absent in Acris crepitans (Maglia et al. 2007b), Ascaphus truei (Jurgens, 1971), Barbourula busuangensis (Jurgens, 1971), Hyla regilla (Jurgens, 1971), Leiopelma archeyi (Stephenson, 1951), Pyxicephalus adspersus (Sheil, 1999), and Rana temporaria (de Jongh, 1968). In most of these species, nonetheless, vestiges of the former bridge [e.g. lingular process (= processus lingularis sensuBaldauf, 1955; Jurgens, 1971; = spina parachoanalis anterior sensuRoček, 1981) and triangular postnasal wall process (= anterior process of the postnasal wall sensuJurgens, 1971; Sheil & Alamillo, 2005; = spina parachoanalis posterior sensuRoček, 1981)] remain present throughout life.

The nasal floor of S. multiplicata and S. bombifrons has three pairs of fenestrae instead of the usual two pairs observed in most species. Two of these pairs of openings, the fenestrae endochoanales and fenestrae nasobasales, form as a result of the development of skeletal structures around the space occupied by each fenestra. The third pair of openings develops as a result of cartilage erosion after metamorphosis. However, although these species have extra fenestrae, their nasal floor is more complete than that of other taxa because all of the openings are rather small. A similar condition was observed by Jurgens (1971) in large adult specimens of S. intermontana.

Medial to the fenestra endochoanalis, the dorsal surface of the solum nasi of Spea is elevated, forming a large skeletal support for the eminentia olfactoria. A variable-sized eminentia olfactoria is present in most anurans, reaching large proportions in some ground-dwelling taxa, such as Bufo, but is best developed in distinctly fossorial species (Jurgens, 1971). In most anurans, the eminentia is supported by the cartilaginous nasal floor, with a minor contribution by the medial margin of the vomer. Examples of this condition are found in several species of Bufo (Baldauf, 1955, 1958; Michael, 1961; personal observation) and in Acris crepitans (Maglia et al. 2007b), among others. In Spea, however, the thickened portion of the nasal floor that contributes to the skeletal support for the eminentia olfactoria is extensively ossified, and the vomer constitutes an important component of the skeletal support (i.e. the entire lateral margin of the skeletal support is formed by the upturned medial margin of the vomer).

The adult skeleton of anurans, as well as the ontogenetic and structural changes that tadpoles undergo during development and metamorphosis have received considerable attention in recent years (e.g. de Jongh, 1968; Trueb, 1970; Jurgens, 1971; Roček, 1981; de Sá, 1988; Wiens, 1989; Rodriguez Talavera, 1990; de Sá & Trueb, 1991; Trueb & Hanken, 1992; Haas, 1996, 1999; Pugener & Maglia, 1997; Wild, 1997; Hall & Larsen, 1998; Maglia & Pugener, 1998; Sheil, 1999; Trueb et al. 2000; Sheil & Alamillo, 2005; and Banbury & Maglia, 2006, to name a few). Of these studies, however, only a handful included a description of the skeletal anatomy of the adult nasal capsules (e.g., Trueb, 1970; Jurgens, 1971; Roček, 1981) or their development (e.g. de Jongh, 1968; Roček, 1981; Haas, 1996; Hall & Larsen, 1998), likely because the extremely small size of the olfactory region of larvae and adults makes it difficult to examine. However, considering that large portions of the anuran phylogeny are still unresolved or highly controversial (see Ford & Cannatella, 1993; Hay et al. 1995; Haas, 2003; Frost et al. 2006), it is essential that investigators search for novel data sources (Jenner, 2001; Haas, 2003). Characters derived from the olfactory region of anurans have never been used in phylogenetic studies, although their informative value was recognized long ago (Jurgens, 1971), and is demonstrated herein. It is hoped that this work will serve as a ‘roadmap’ for future investigation of the skeletal morphology and larval development of the nasal capsules of anurans, and that future phylogenetic analyses of anurans based on morphology may include characters from the olfactory region.

Acknowledgments

We thank Jessica Mueller for her help with an early version of this manuscript and John Simmons and Linda Trueb at the KU Natural History Museum for allowing us to examine specimens under their care. We also thank Alex Haas and John Reiss for helpful discussions and Linda Trueb and an anonymous reviewer for critical comments and suggestions on the manuscript. Charles Huber provided invaluable assistance in the generation of the 3D reconstruction. Funding for this project was provided by NSF grant DBI-0445752.

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