RESEARCH ARTICLE

Morphology and Histology of the Tongue and Oral Chamberof Eublepharis macularius (Squamata: Gekkonidae), with SpecialReference to the Foretongue and its Role in Fluid Uptakeand Transport

Heather A. Jamniczky • Anthony P. Russell •

Megan K. Johnson • Stephane J. Montuelle •

Vincent L. Bels

Received: 16 October 2009 / Accepted: 22 October 2009

� Springer Science+Business Media, LLC 2009

Abstract Detailed descriptions of tongue morphology of

members of Squamata that refer to functional implications

other than food processing are rare. Herein we focus on the

morphology of the dorsal epithelium and internal structure

of the tongue of the Leopard Gecko, Eublepharis macu-

larius, emphasizing the foretongue and its relation to fluid

uptake. We employ both scanning electron microscopy and

serial histology to examine the morphology of the entire

tongue, its component regions, and its situation in the oral

chamber. We recognize five distinct morphological regions

of the dorsal tongue surface, each of which is distinctive

both morphologically and histologically. The foretongue

bears papillae quite different in structure and spacing from

those of all other tongue regions, and these non-glandular

structures are involved in gathering and transporting fluid

from the environment. Fluid unloaded from the foretongue

in the region of the vomeronasal sinus is channeled through

the network of cuboidal papillae and directed towards a

pair of compartments lateral to the tongue in which fluid

pools during a drinking bout. This allows the dorsal surface

of the mid- and hind-tongue, which are involved in food

processing and manipulation, to be largely segregated from

the pathway of fluid flow. We relate our findings to

descriptions of the tongue of other taxa, and propose

functional hypotheses for the observed morphology. This

study provides new anatomical information upon which

future studies of the functional morphology of the buccal

apparatus in the Gekkota can be based.

Keywords Tongue � Epithelium � Histology �Eublepharis macularius � Gekkonidae � Drinking

Introduction

The morphology of the tongue of members of Squamata is

known from a relatively large number of studies (e.g. Seiler

1891, 1892; de Rooij 1915; Camp 1923; Gnanamuthu

1937; Iwasaki and Miyata 1985; Iwasaki et al. 1985;

Schwenk 1985, 1986, 2000; Rabinowitz and Tandler 1986;

Smith 1986, 1988; Iwasaki 1990, 2002; Delheusy et al.

1994; Toubeau and Bels 1994; Herrel et al. 2005). Inves-

tigations of the gross anatomy of the squamate tongue,

supplemented by light and electron microscopy, across a

broad spectrum of subordinate taxa, have demonstrated

considerable morphological variation in the structure of the

dorsal lingual epithelium. Much attention has been paid to

describing the features of the tongue surface, and the

phylogenetic implications of these features have been

extensively discussed (e.g. Schwenk 1985; Smith 1988).

Several authors have also offered functional explanations

for the morphological diversity evident in squamate tongue

epithelia (e.g. Beisser et al. 1998, 2004; Schwenk 2000;

Iwasaki 2002; Herrel et al. 2005), mostly in relation to

adaptations for feeding in a terrestrial environment.

H. A. Jamniczky (&) � A. P. Russell � M. K. Johnson

Vertebrate Morphology and Palaeontology Research Group,

Department of Biological Sciences, University of Calgary,

2500 University Drive NW, Calgary, AB T2N 1N4, Canada

e-mail: hajamnic@ucalgary.ca

Present Address:H. A. Jamniczky

Department of Cell Biology and Anatomy, Faculty of Medicine,

University of Calgary, 3330 Hospital Drive NW, Calgary,

AB T2N 4N1, Canada

S. J. Montuelle � V. L. Bels

Museum National d’Histoire Naturelle, USM 302, 75005 Paris,

France

123

Evol Biol

DOI 10.1007/s11692-009-9072-9

As part of a larger project (Bels et al. unpublished)

investigating the biomechanics of oral processing in the

Leopard Gecko (Eublepharis macularius), we undertook a

scanning electron and light microscopical investigation of

the morphology of the tongue of E. macularius. We here

document its morphology in the context of drinking

behavior, in order to correlate the features observed with

their function in fluid collection and transport.

Water is a vital metabolic resource and it is anticipated

that the tongue, in those taxa that use this structure in the

drinking process, will be configured in such a way as to

promote effective and efficient transfer of water from the

environment to the oral chamber. Sherbrooke et al. (2007)

demonstrate this for the jaw angle region of the rain-har-

vesting Phrynosoma cornutum, implicating tongue, jaw,

and hypobranchial movements in the transfer of water from

the integument to the oral cavity. Furthermore, the oral

chamber acts as a conduit for this water between the tongue

and the glottis, and its structure is likely to be reflective of

the passage of water through it. In lizards such as gekko-

tans in which food prehension is jaw- rather than tongue-

based (Delheusy and Bels 1999; Schwenk 2000; Bels 2003;

Reilly and McBrayer 2007) we hypothesize that the tongue

is regionally specialized to accommodate both fluid and

food transport and processing.

To test this prediction, we investigate the structure of the

tongue in the Leopard gecko to determine whether func-

tional subdivision of the tongue, in relation to drinking and

food processing, is evident. We compare our results to

those that exist for other members of Gekkota (Gekko

gecko [pers. obs.], Gekko japonicus [Iwasaki 1990], Gon-

atodes antillensis, and Coleonyx variegatus [Schwenk

1985]), and suggest future research directions aimed at

furthering our understanding of the form and function of

the gekkotan and squamate buccal apparatus.

Methods

Three adult female (SVL 131–136 mm), one juvenile

female (SVL 70 mm), and one hatchling (SVL 53 mm)

female Eublepharis macularius were sacrificed by oral or

intracardiac administration of T-61 Euthanasia Solution

(Intervet Canada), according to a protocol approved by the

University of Calgary Animal Care Committee (protocol

#BI2005-22), following Canada Council on Animal Care

guidelines. Preliminary observations revealed that the

tongues of freshly sacrificed adult individuals were pro-

fusely coated with mucus, which remains in place after

fixation. To eliminate this problem in our experimental

sample so as to enable scanning electron microscopic

(SEM) visualization of tongue structure, we pretreated the

tongue as follows: the lower jaw and associated tongue

were excised from each individual immediately following

sacrifice, affixed with fine entomological pins to a paraffin

wax-lined, small, sealable plastic container and flooded

with pharmacological decongestant nasal spray (active

ingredient 0.1% xylometazoline hydrochloride). This was

allowed to sit on the tongue surface for several minutes,

and was then washed off by irrigating the tongue vigor-

ously with distilled water from a plastic wash bottle. Fol-

lowing several rounds of aqueous irrigation, the tongues

were fixed by immersion in 10% neutral buffered formalin

for 24 h, and stored in 70% ethanol.

Macroscopic structure of the tongue of E. macularius

was examined via macrophotography, photomicrography,

and SEM. For SEM observations, two adult tongues were

examined whole and one was sliced longitudinally prior to

examination. These were dehydrated using a graded etha-

nol series from 70% to 100%, critical point dried in a

Seevac CO2 Critical Point Dryer, mounted on � inch Zeiss

aluminum SEM mounts using colloidal silver paste, and

sputter coated with gold using a Hummer II Sputter Coater.

They were examined in the standard high vacuum mode of

a Philips/FEI ESEM XL30, and images were taken at

magnifications of 1259 and 5009 for each of the different

regions of the tongue surface. The tongue of the juvenile

female was photographed in situ after fixation and removal

of the lower jaw from the remainder of the head. Its tip was

subsequently removed and subjected to SEM.

Microscopic structure of the tongue of E. macularius

was visualized using serial histology. Adult tongues were

sliced sagittally, and each half was embedded separately in

paraffin wax following standard technique (Humason

1979) and oriented either for orthogonal or longitudinal

sectioning. Serial sections were cut at 10 lm and stained

with Masson’s Trichrome (Witten and Hall 2003, substi-

tuting Gill’s Haematoxylin for Mayer’s Haematoxylin).

Selected sections were photographed using a Nikon DS-L1

digital camera mounted on a Nikon 50i compound micro-

scope, integrated using a Nikon DS-5M camera control

unit. Images shown were not subjected to any post-capture

processing.

The tongue of the hatchling individual was examined in

situ by sectioning the entire head in the transverse plane.

This enabled the establishment of both fine structure and

the placement of the tongue in relation to other oral fea-

tures. Staining procedures were identical to those for the

sectioned adult tongues (see above).

Measurements of tongue structures were taken from the

histological longitudinal and cross sections of the adult

tongues. Slides were examined using the above-mentioned

microscope system. Measurements were taken directly

from the camera control unit using the measurement tool

calibrated to the appropriate magnification. Measurements

of papilla height, width, and spacing were taken from both

Evol Biol

123

longitudinal sections and cross sections of the tongues for

each of the five structural regions identified. Since the

absolute size of the structures may be influenced by the size

of the animal, the measurements were converted into two

ratios: height:width and spacing:width.

The relatively small body size of E. macularius makes it

unsuitable for vascular study using conventional vascular

infiltration techniques. Instead, a specimen of Gekko gecko,

injected with red Microfil (FlowTech Inc, Carver, MA) for

another study (Russell et al. 1988), was examined com-

paratively in relation to vascular structure, and is figured

herein. Complete details on preparation of this specimen

are available elsewhere (Russell et al. 1988).

Three adult male specimens of E. macularius were

filmed drinking in lateral, dorsal, and frontal views using

an NAC HSV-500 C3 high-speed video system at 250 Hz.

Lizards were filmed under two conditions: (i) water

delivered on the substrate in front of the head (sometimes

labeled with barium) and (ii) water drops sprayed on a

sheet of glass arranged vertically in front of them. Lizards

were deprived of water for 1 week before filming, but were

fed crickets and mealworms ad libitum during this period.

During filming, lizards were placed in a small plexiglas

corridor and water was provided using a pipette. Record-

ings were transmitted to a computer for analysis of tongue

movement and deformation during each drinking cycle.

Quantitative analysis of the mechanics of drinking will be

presented elsewhere (Bels et al. in preparation).

Results

Anatomy

Schwenk (1985: Fig. 1) divided the lepidosaur tongue into

three regions: the tip, which consists of the bifurcated

anterior portion and a small area immediately posterior to

the bifurcated portion; and the remainder of the tongue,

which was subdivided into fore- and hindtongue by a line

placed halfway between the tip and the posterior end of the

tongue. Herein, we divide the tongue on the basis of ana-

tomical features rather than proportional dimensions, and

demonstrate that there are five distinctive morphological

regions on the dorsum of the tongue of E. macularius

(Fig. 1), four of which exhibit morphologically distinct

types of papillae on the dorsal surface. Papillae in each

region are differentiable on the basis of height, width, and

the density of their occurrence (Table 1). We designate

these regions (Fig. 1b) as follows: (1) extreme tongue tip

(no papillae); (2) tongue edges (feather-like papillae); (3)

foretongue (cuboidal papillae); (4) midtongue (leaf-like

papillae); and (5) hindtongue (two subregions, containing

long fungiform (5a) and short fungiform (5b) papillae

respectively). Each region is differently involved in oral

transport and is discussed separately below. Figure 1b

maps these regions and provides orientation and location

(Fig. 1a) information relating to each of the subsequent

figures.

The extreme tongue tip (Plate 1A–C) is bifid, covered

by keratinized squamous epithelium, and bears no papillae

(Plate 1A, B). Its thickened epidermal covering is smooth

and projects ventrally (Plate 1A) for a considerable dis-

tance before forming a sharp junction with the pleated

ventral surface of the more posterior region, that constitutes

Fig. 1 a Scanning electron micrograph of the dorsal surface of the

tongue of Eublepharis macularius. Scale bar = 2 mm, magnification

179. HIND, MID, and FORE refer to tongue divisions described by

Schwenk (1985); white boxes indicate sampled tongue regions shown

in subsequent SEM images. b Outline of tongue shown in Fig. 1a.

Numbered regions refer to tongue divisions described herein.

Abbreviations: gl glottis, pf papilla-free zone at base of tongue

Table 1 Relative dimensions of papillary structures on the dorsal

lingual epithelium of Eublepharis macularius. The extreme tongue tip

is not included, as papillae are absent from this region. Values given

are ratios, to remove the effects of differences in absolute size of the

specimens examined. Original measurements were recorded in lm

Papilla type/location Height:Width Spacing:Width

Feather-like/tongue edge 2.374 ± 0.18 0.228 ± 0.04

Cuboidal/foretongue 1.711 ± 0.09 0.195 ± 0.02

Leaf-like/midtongue 4.874 ± 0.30 0.337 ± 0.03

Long fungiform/hindtongue 2.293 ± 0.16 0.261 ± 0.05

Short fungiform/hindtongue 1.823 ± 0.19 0.281 ± 0.08

Evol Biol

123

the extensible, protrusible portion of the part of the tongue

that lies free in the oral cavity. This thickened epithelium

wraps around the distal extremity of the tongue and extends

for a very short distance on its dorsal surface (Plate 1B)

before forming a sharp junction with the cuboidal papillate

region (see below). The extreme tongue tip is underlain by

two extensions of the hyoglossus muscle, one on either side

of, and close to, the midline. The ventral part of the tongue

tip (Plate 1A) bears a midline sulcus that interdigitates

with a process rising from the floor of the oral chamber

(Plate 1C). This tongue-in-groove association between the

tongue tip and oral floor provides the guiding and trans-

lation mechanism across which the tongue tip slides during

protrusion when the mouth is minimally open during

drinking, resulting in a pivoting and inversion of the

anterior part of the tongue during fluid collection (descri-

bed further below).

The foretongue (Plate 1D–G) exhibits cuboidal papillae

that are of uniform shape (Plate 1D) and are the most

densely packed of all types of papillae described. They are

approximately 1.7 times as high as wide (Table 1). The

surface of each papilla is covered by keratinized stratified

squamous epithelial cells (Plate 1E). No goblet clusters or

glandular epithelium are present. In this region of the

tongue the hyoglossus muscle is divided into four longi-

tudinal, finger-like processes on each side (Plate 1F) that

span from just lateral to the midline to the region of

juncture of the smooth ventral surface of the tongue and the

papillate tongue edge. The four, finger-like processes align

with a series of longitudinal pleats on the ventral surface of

the tongue (Plate 1A) in the region posterior to the

smoothly-clad extreme tongue tip.

Lying dorsal to the hyoglossus in the foretongue is a

mass of interwoven intrinsic tongue musculature and

Plate 1 (A) Tongue of Eublepharis macularius, tip visualized using

SEM, ventral view, scale bar 500 lm, magnification 789; (B) Tongue

of Eublepharis macularius, tip visualized using Masson’s Trichrome/

light microscopy, longitudinal section, scale bar 100 lm, magnifica-

tion 1009; (C) Tongue tip visualized using Masson’s Trichrome/light

microscopy, transverse section, scale bar 1 mm, magnification 109;

(D) Tongue of Eublepharis macularius, foretongue visualized using

SEM, scale bar 100 lm, magnification 5009, showing cuboid

papillae; (E) Tongue of Eublepharis macularius, foretongue visual-

ized using Masson’s Trichrome/light microscopy, anterior cross-

section, showing cuboid papillae and musculature, scale bar 100 lm,

magnification 1009; (F) Tongue of Eublepharis macularius,

foretongue visualized using Masson’s Trichrome/light microscopy,

anterior cross-section, scale bar 1 mm, magnification 409; (G)

Tongue of Eublepharis macularius, foretongue visualized using

macroscopic photograph of tongue in dorsal view showing tongue tip

engorged with blood; (H) Tongue of Gekko gecko, injected with

Microfil to visualize vasculature; (I) and (J) show a detailed view of

the vasculature of the fore- and hindtongue, respectively, of Gekkogecko. Abbreviations: cp capillary, ct connective tissue sheath,

ft foretongue, ht hindtongue, hy hyoglossus, in invagination, lc lat-

eral buccolingual chamber, lg longitudinalis, mt midtongue, pc pos-

terior buccolingual chamber, pl pleat, tb non-papillate tongue base,

tr transversalis, ve verticalis

Evol Biol

123

intervening connective tissue, organized into transverse,

vertical and longitudinal components. The longitudinalis

component is less distinct at the distalmost extremity of the

tongue and merges with the deeper verticalis component,

such that there is no real distinction between them

(Plate 1E). The intrinsic transversalis muscle strands are

highly interwoven with the verticalis strands, and the latter

extend from deep within the tongue, dorsal to the hyo-

glossus bundles (Plate 1F). They spread dorsolaterally to

intersect and merge with the transversalis strands that lie

deep to the cuboidal papillae (Plate 1F). The intrinsic

muscle strands connect with connective tissue sheaths

lying at the base of the papillae (Plate 1E). There are no

spaces within the tongue body in this region (Plate 1F).

The cuboidal papillae in this region are highly vascu-

larized (Plate 1E, H–J), with their core containing expan-

ded tufts of vessels that render the tongue tip red in life

(Plate 1G), visibly distinguishing it from more posterior

regions. The visible evidence of such vascularity is likely

due to the relatively thinner keratinized tips of these

papillae compared with tips of those situated more

posteriorly on the tongue (see below), as the degree of

vascularity itself is not greater in this region (Plate 1H–J),

although the terminal vascular tufts are more pronounced

(the intrinsic vasculature of the more posterior papillae as

more attenuated, in association with the longer and more

gracile form of these papillae—see below). These cuboidal

papillae receive extensions of the verticalis component of

the underlying intrinsic muscle feltwork (Plate 1E). Con-

siderably more such musculature intrudes into the lateral

papillae of this region of the tongue (see below). There is

no central skeletal core in the foretongue (Plate 1F).

An invagination that courses from dorsolateral to ven-

trolateral along the length of the tongue is situated dorso-

laterally in the extreme foretongue region (Plate 1C) and

reorients to a ventrolateral position as the posterior part of the

foretongue is approached (Plate 1F). This invagination

demarcates boundary between the papillate dorsal portion of

the tongue and the ventral tongue surface. It is bounded on its

dorsal border by epithelium that is heavily keratinized

(Plate 1F). Examination of the foretongue in situ (Plate 1A)

reveals that it is dished dorsally and makes up the ventral part

of a shallow anterior buccolingual chamber. Further poste-

riorly a pair of lateral buccolingual chambers are present,

demarcated by the juxtaposition of the papillate tongue

margin with the medial palatal shelves of the maxillae in the

roof of the mouth (Plate 2A, B). The dorsal portion of the

mid-tongue (Fig. 1a), with its much taller papillae (see

below), lying ventral to the narial passages (Plate 2A, B), can

be employed to seal off the narial passages and isolate them

from the lateral buccolingual chambers. This junction of the

tongue with the maxillary shelves occurs at the position of

the lingual invagination and the dense connective tissue that

underlies it. More posteriorly still, in the glottal region

posterior to the narial passages (Plate 2B), a central posterior

buccolingual chamber (Plate 2C, D) lies dorsal to another

dished area of the tongue.

The remainder of the tongue, whose role in fluid transport

is less pronounced, also exhibits a variety of papilla types

and extensive vascularity. The tongue edges (Plate 2E, F),

with the exception of the extreme tongue tip and the fore-

tongue, exhibit long, slender, feather-like papillae with wide

bases and narrower tips (Plate 2E) that are not keratinized.

These papillae demarcate the junction between the papillate

dorsal surface and smooth ventral surface of the tongue. The

marginal papillae bear small, orthogonally-projecting

glandular goblet clusters (Plate 2F), discussed further

below, are invaded by capillaries (Plate 1J), contain both

muscle and connective tissue, and are relatively widely

spaced at their tips but more closely packed at their bases

(Plate 2E, F; Table 1). They are approximately 2.3 times as

high as wide (Table 1).

The midtongue (Plate 2G, H) exhibits irregular, tall

papillae with leaf-like, posteriorly-swept apical surfaces.

These are less densely packed than those of the foretongue,

but more densely packed than those of the tongue edge

(Table 1). The stalk of each papilla is covered by pro-

jecting glandular goblet clusters (Plate 2H), whereas the

apical surface is covered by strongly keratinized, stratified,

squamous epithelial cells (Plate 2H). Indeed, the exposed

basal stems of all papilla types, except the cuboidal

structures of the foretongue region, exhibit densely-packed

populations of goblet clusters that produce a thick mucus

that coats the tongue (Plate 2I). They are particularly

prolific in the mid- and hindtongue regions. Histological

sections reveal that these cells are largely devoid of con-

tents, and only the cell membranes stain, producing a

ghost-like effect (Plate 2H, L). The papillae of the mid-

tongue are relatively and absolutely taller than any of the

other types described (Table 1), vascularized (Plate 1J),

and contain both muscle and connective tissue (Plate 2H).

In the free region of the midtongue, the papillae are present

on the lateral surface of the tongue but do not extend more

ventrally than this. More posteriorly the papillae extend

further ventrolaterally (Plate 2A) and the lingual invagi-

nation assumes a more ventrolateral location, and is lined

by a thick, keratinized, simple columnar epithelium sur-

rounded by connective tissue.

The anterior region of the midtongue is free of the floor

of the mouth and houses four bundles of the hyoglossus

muscle, arrayed ventrally and ventrolaterally on either side

of the ventral midline of the tongue. The left and right

hyoglossus components are segregated by a strong, trian-

gular connective tissue core (Plate 2A). The ventral surface

of the tongue is girdled by strongly-developed connective

tissue and is pleated in a fashion similar to that seen in the

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foretongue (Plate 1A, F). At the posteriormost extent of

the midtongue, the tongue transitions from being free in the

floor of the mouth (Plate 2A) to being fixed in place. The

muscle bundles in this region are stacked vertically and

surround the anteriormost extent of the entoglossal process

of the hyoid that lies in a central, ventrally-located sheath

(Plate 2A). Save for the highly-organized longitudinalis

bundles dorsally, the intrinsic musculature of the tongue is

less prevalent in the midtongue than it is in the foretongue,

and connective tissue is proportionally more dominant.

Lymph spaces are prominent in the central core region of

the midtongue. Large nerves and blood vessels are scat-

tered throughout. The substance of the tongue deepens

from anterior to posterior in this region.

The hindtongue (Plate 2I–L) is differentiable into two

regions (Fig. 1b): in its central region (Fig. 1b, region 5A)

it exhibits long fungiform papillae (Plate 2I, J), whereas

more laterally the fungiform papillae are shorter (Fig. 1b,

region 5b; Plate 2K, L). The stalks of both sets of papillae

are covered in glandular goblet clusters. Apically they bear

keratinized stratified squamous epithelial cells (Plate 2J,

L). The lateral walls of both types of papillae are irregular,

and both types are more densely packed than are the

papillae of the tongue edge, but less densely so than those

of the midtongue (Table 1). Both long and short fungiform

papillae are wider at the base than are the papillae of the

fore- and midtongue and the tongue edges, and contain

more muscle tissue and more elaborate vasculature

(Plate 1H–J; Plate 2J, L). Long fungiform papillae exhibit

posteriorly-swept apical aspects (Plate 2I, J), whereas short

fungiform papillae occasionally exhibit branched apical

aspects (Plate 2K). The lingual invagination, shrouded

Plate 2 Tongue of Eublepharis macularius. (A) Midtongue visual-

ized using Masson’s Trichrome/light microscopy, cross-section, scale

bar 1 mm, magnification 109, showing lateral buccolingual cham-

bers; (B) Macroscopic view of roof of mouth, (macroscopic view of

tongue in lower right corner); (C) Hindtongue visualized using

Masson’s Trichrome/light microscopy, cross-section, scale bar 1 mm,

magnification 109, showing central posterior buccolingual chamber;

(D) Non-papillate region of hindtongue anterior to glottis visualized

using Masson’s Trichrome/light microscopy, cross-section, scale bar

1 mm, magnification 109; (E) Tongue edge, SEM, scale bar 100 lm,

magnification 5009; (F) Tongue edge, Masson’s Trichrome/light

microscopy, longitudinal section, scale bar 100 lm, magnification

1009; (G) Midtongue, SEM, scale bar 100 lm, magnification 5009,

showing papillae with posteriorly-swept tips and goblet clusters; (H)

Midtongue, Masson’s Trichrome/light microscopy, longitudinal

section, scale bar 100 lm, magnification 1009, showing papillae

and goblet cells; (I) Hindtongue region A, SEM, scale bar 100 lm,

magnification 4619, showing mucus on papillae; (J) Hindtongue

region A, Masson’s Trichrome/light microscopy, cross-section, scale

bar 100 lm, magnification 2009, showing vasculature and intrinsic

musculature at higher magnification; (K) Hindtongue region B, SEM,

scale bar 100 lm, magnification 4619, showing papillae; (L)

Masson’s Trichrome/light microscopy, cross-section, scale bar

100 lm, magnification 2009, showing goblet clusters. Abbreviations:

ac anterior buccolingual chamber, ch choana, cp capillary, dc dense

connective tissue, eh entoglossal process of hyoid, go goblet cluster,

hy hyoglossus, in invagination, lc lateral buccolingual chamber,

mc mucus, mx maxilla, np narial passage, pc posterior buccolingual

chamber, ps medial palatal shelf of maxilla

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123

by connective tissue, partially underlies the hindtongue

(Plate 2C).

Anteriorly, the hindtongue exhibits stout, vertically

stacked, paired subdivisions of the hyoglossus muscle

ventrolaterally, flanking, but at some distance from, the

central niche in which the lingual process of the hyoid

apparatus sits (Plate 2C). At the very base of the tongue

there is a central, papilla-free zone (Fig. 1, Plate 2D). The

hyoglossus muscle is much less discrete at this point, and

no subdivision into bundles is evident.

Behaviour

High-speed cinematography reveals that E. macularius

gathers fluid by cyclical protrusive-retractive motions of

the tongue, resulting in a fluid bolus being gathered by the

foretongue and eventually conveyed to the stomach by

head-tossing behavior and deglutition. In fluid gathering,

the extreme tongue tip acts as a fulcrum over which the

papillate foretongue pivots and inverts (Fig. 2a, b, d, e).

This pivot mechanism results in presentation of the dorsal

surface of the foretongue directly to the fluid as it rolls

out of the mouth (Fig. 2c, f, g). The foretongue is

greatly expanded on the substrate as it touches the water

(Fig. 2f, g), maximizing the surface area presented to the

water. Following several such cycles, emersion is achieved

by the head being raised. The protraction-retraction cycles

of the tongue continue briefly.

Radiographic images of E. macularius drinking barium-

labelled water (Bels et al. unpublished) reveal that the

tongue tip is loaded with fluid, and that water is unloaded

from the tongue into the anterior buccolingual chamber in

the region of the vomeronasal sinus. In the following cycle

fluid is removed from this region and transported to the pair

of lateral buccolingual chambers (Plate 2C), at the same

time that a new bolus of fluid is being gathered by the

foretongue and transported to the anterior buccolingual

chamber. The original bolus of fluid moves laterally from

the anterior buccolingual chamber to the lateral buccolin-

gual chambers by traversing the tongue laterally in the

region of the junction of the foretongue and midtongue.

Fluid is thus kept segregated from the mucus-rich part of

the tongue (Plate 2I) and is ultimately guided to the val-

vular glottal area, posterior to the region of air flow, where

it enters a third, central chamber (Plate 2C, D) prior to

being swallowed, assisted by an upward tilting of the head.

The hindtongue moves very little in the protraction-

retraction cycles of the foretongue, and remains essentially

Fig. 2 Frames from high-speed cinematography of E. macularius during drinking in frontal (a–c) and dorsal (d–g) views. The foretongue pivots

over the extreme tongue tip and expands laterally as its dorsal surface is presented to the liquid. See text for further details

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123

fixed relative to the skull, with only the mid- and foretongue

being displaced appreciably. The hindtongue is involved in

fluid transportation only inasmuch as it forms the medial and

dorsal boundaries of the internal oral compartments.

Discussion

Our observations allow us to relate the macroscopic

structure of the tongue and oral chamber of E. macularius

to the process of fluid uptake. The tongue is generally

similar, both in macroscopic and histological morphology,

to the tongue of other lizards. Iwasaki (1990) described

three types of papillae on the tongue of Gekko japonicus:

dome-shaped, which correspond to the cuboidal papillae

described here in the foretongue region (Plate 1D–G);

fan-shaped, which correspond to the leaf-like papillae

described in the present study in the midtongue region

(Plate 2G, H); and scale-like, which correspond to the short

and long fungiform papillae observed in the present study

in the hindtongue region (Plate 2I–L).

The lack of papillae on the extreme tongue tip was also

noted by Delheusy et al. (1994) for Oplurus cuvieri.

Although they reported similar foretongue papillae for that

taxon, the remainder of the tongue of O. cuvieri differs

noticeably from that of E. macularius. Papillae present in

the fore, mid- and hindtongue of O.cuvieri are generally

cylindrical in shape, and their tips are far less elaborate

than the leaf-like and fungiform papillae observed herein.

We describe the presence of glandular goblet clusters on

the exposed stalks of papillae in the mid- and hindtongue

regions. Iwasaki (1990) noted a large number of cells

containing secretory granules on the tongue of G. japoni-

cus, and Delheusy et al. (1994) reported abundant mucus

secretions in O. cuvieri. Rabinowitz and Tandler (1986)

described the morphology of papillae on the tongue of

Anolis carolinensis and designated these as ‘plumose’, on

the basis of the presence of ‘plume cells’. Comparison of

these cells (Rabinowitz and Tandler 1986: Fig. 7) to the

goblet clusters observed in E. macularius (Plate 2E–L)

suggests a structural similarity, although there are mor-

phological features that seemingly preclude their homol-

ogy. The goblet clusters we describe are more cuboidal and

exhibit few cellular contents. They invest the stems of the

papillae and extend from the papillary base to the kera-

tinized tip, whereas the plume cells of Rabinowitz and

Tandler (1986) are restricted to the apical portion of the

papillae. It is possible that the clusters we describe repre-

sent a more elaborately expressed form of those observed

by Rabinowitz and Tandler (1986), but further study is

required to test this hypothesis.

Interestingly, we failed to establish the presence of taste

buds in E. macularius. Schwenk (1985) found evidence for

widespread presence of taste buds across Lepidosauria, but

among the Gekkota he noted their absence from the tongue

of Gonatodes antillensis, and their presence only on the

hindtongue of Coleonyx variegatus. Our results provide

further evidence that the Gekkota may largely lack taste

buds. Although the explanation for this phenomenon may

be phylogenetic, there may also be a functional or adaptive

significance to it.

Our findings allow us to demonstrate the morphological

basis for fluid transport in relation to the structure of the

tongue and the associated oral regions of E. macularius.

This functional complex consists of papillae of particular

shapes and orientations; the cross-sectional morphology of

the tongue in its various regions; the association of cross-

sectional morphology with the morphology of the roof of

the mouth and the narial passageways; the form of the

lateral regions of the buccal cavity and their segregation

from the dorsal region by maxillary flanges; the muscular

organization of the tongue facilitating the interaction of

these features; and the form and pattern of distribution of

the lingual papillae.

When placed into the context of its in situ location, the

foretongue is dished dorsally and makes up the ventral part

of an anterior buccolingual chamber (Plate 1C), lying

anterior to the non-olfactory portions of the narial pas-

sageways. The cuboidal papillate region of the foretongue

in this location lies medial to the opening of the nares into

the oral chamber (Plate 1C), thus segregating the water-

laden portion of the tongue from the olfactory area. The

spacing between the papillae is wide (Plate 1D, E), and,

upon tongue extension, creates capillary channels for fluid

pickup as the dorsal surface is stretched and expanded

(Fig. 2e, f, g) by being rolled out of the mouth (with the

extreme tip remaining within the confines of the buccal

cavity; Fig. 2c). During protraction, the tongue moves over

the mandibular symphysis and expands laterally as it does

so—a function attributable to the longitudinalis compo-

nents of the intrinsic tongue muscles that lie in bundles at

the base of the papillae (Plate 1E, F). The tines of the

foretongue remain in the buccal cavity, employing the cleft

in the tongue and the process that rises from the floor of the

mouth beneath it, as a fulcrum-and-anchor device. The

rounded, laterally spread dorsal surface of the foretongue is

extended and presented to the fluid (Fig. 2b, c). During

retraction fluid is held in the interpapillary pockets and no

bead of fluid is evident on the tongue surface during

retraction (Bels et al. unpublished). The curled tongue is

straightened as it is drawn into the mouth (Fig. 2). The

absence of mucus glands from the foretongue is compatible

with its fluid transportation function, allowing fluid to enter

deep into the interpapillary spaces. Contraction of the

transversalis muscles (Plate 1E) will narrow the foreton-

gue, reducing the spaces between the papillae, and the

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123

verticalis components (Plate 1E) will shorten the papillae.

Collectively these two muscle masses are likely responsi-

ble for the change of configuration of the foretongue during

retraction, resulting in compression of the interpapillary

spaces and the forcing of fluid onto the dorsal surface of the

tongue, enabling its collection in the anterior buccolingual

chamber (Plate 2A–D), ventral to the maxillary shelves and

lateral to the non-olfactory portions of the narial passage-

ways. The form of the tongue and the roof of the mouth are

configured such that fluid travels into the lateral chambers

(Plate 2A) and remains segregated from the narial pas-

sages. Interestingly, Herrel et al. (2005) report the presence

of papillae on the ventral surface of the foretongue of

Lacerta spp. (which they term ‘fungiform’) that are very

similar in shape to the cuboidal papillae described herein.

These species are known to use the ventral aspect of the

tongue to gather water when drinking.

The presence of muscle fibers within papillae, as noted

for A. carolinensis by Rabinowitz and Tandler (1986),

allows neurological control of the movements of these

structures. Such control would permit the lizard to alter-

nately open and close the spaces between papillae, and to

alter their dimensions. This would facilitate both collection

and retention of water in the foretongue region during

pickup, and release at the time of transfer, and would also

allow differential control of the amount of mucus present

on the tongue. We suggest, given the cloying nature of

mucus secretions, that papillae would be relatively

expanded in order to increase the amount of mucus present

on the tongue during feeding, and would be relatively

contracted in order to decrease the amount of mucus

present on the tongue during drinking. Further experi-

mentation is required to test this hypothesis.

Previous authors (e.g. Schwenk 1985; Smith 1988;

Beisser et al. 1998, 2004; Iwasaki 2002; Herrel et al. 2005)

have postulated that the presence of papillae on the tongue of

sauropsids is explained either phylogenetically, or by the

need for adaptation for food procurement and processing in a

terrestrial environment. Food prehension is jaw- rather than

tongue-based in Gekkotans (Delheusy and Bels 1999;

Schwenk 2000; Bels 2003; Reilly and McBrayer 2007),

leaving the tongue free for extra-oral functions. Herein we

present the first evidence of a fluid procurement and trans-

portation function for the papillate Gekkotan tongue, and

relate this to the overall form and configuration of the oral

chamber. Spectacle-wiping (Bustard 1963) by the tongue may

also involve the use of the non-glandular part of the tongue,

allowing for cleaning without the deposition of mucus on the

spectacle surface. Although not a feature of eublepharid ge-

kkotans, because they have eyelids, this behaviour would

likely have co-opted the non-glandular part of the tongue as an

exaptation in all gekkotans that have a spectacle (see Cogger

[1967: Plate 7] for an example of this behaviour).

Acknowledgments We thank K. Kardong for discussion, and

P. Wise for providing specimens. This research was supported in part

by an NSERC Discovery Grant to APR.

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