Functional Morphology and Comparative Anatomy of Appendicular Musculature in Cuban Anolis Lizards with Different Locomotor Habits Author(s): Wataru Anzai, Ayano Omura, Antonio Cadiz Diaz, Masakado Kawata and Hideki Endo Source: Zoological Science, 31(7):454-463. 2014. Published By: Zoological Society of Japan DOI: http://dx.doi.org/10.2108/zs130062 URL: http://www.bioone.org/doi/full/10.2108/zs130062

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© 2014 Zoological Society of Japan

ZOOLOGICAL SCIENCE 31: 454–463 (2014)

Functional Morphology and Comparative Anatomy of Appendicular Musculature in Cuban Anolis Lizards with Different Locomotor Habits Wataru Anzai1,2*, Ayano Omura1,3, Antonio Cadiz Diaz4,5, Masakado Kawata4, and Hideki Endo1 1

2

The University Museum, The University of Tokyo, Tokyo 113-0033, Japan Department of Biological Sciences, Graduate School of Science, The University of Tokyo, Tokyo 113-0033, Japan 3 Department of Global Agricultural Sciences, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo 113-8657, Japan 4 Department of Ecology and Evolutionary Biology, Graduate School of Life Sciences, Tohoku University, Sendai 980-8578, Japan 5 Faculty of Biology, The University of Havana, Havana 10400, Cuba

We examined the diversity of the musculoskeletal morphology in the limbs of Anolis lizards with different habitats and identified variations in functional and morphological adaptations to different ecologies or behaviors. Dissection and isolation of 40 muscles from the fore- and hindlimbs of five species of Anolis were performed, and the muscle mass and length of the moment arm were compared after body size effects were removed. Ecologically and behaviorally characteristic morphological differences were observed in several muscles. Well-developed hindlimb extensors were observed in ground-dwelling species, A. sagrei and A. bremeri, and were considered advantageous for running, whereas adept climber species possessed expanded femoral retractors for weight-bearing during climbing. Moreover, morphological variations were observed among arboreal species. Wider excursions of the forelimb joint characterized A. porcatus, presumably enabling branch-to-branch locomotion, while A. equestris and A. angusticeps possessed highly developed adductor muscles for grasping thick branches or twigs. These findings suggest divergent evolution of musculoskeletal characteristic in the limbs within the genus Anolis, with correlations observed among morphological traits, locomotor performance, and habitat uses. Key words:

adaptation, Anolis, appendicular musculature, habitat use, locomotion

INTRODUCTION In tetrapods, limb morphology closely reflects locomotor performance and ecological characteristics. Many studies have focused on the relationships between morphology, ecology, and behavior. Anolis is one of the largest genera of lizards, or the paraphyletic Lacertilia, with over 400 species (Poe, 2004). These lizards have radiated adaptively in the Caribbean Islands and show convergent evolution in form and habitat. Different morphologies and behaviors have evolved in this genus, reflecting adaptation to differing environments (Losos et al., 1998). Habitat specialists recognized as six “ecomorph” types based on the habitat use (TrunkGround, Trunk-Crown, Crown-Giant, Grass-Bush, Twig, Trunk) have evolved repeatedly (Williams, 1972, 1983). Multiple species or ecomorphs co-occurred sympatrically within a relatively small geographic area across the Greater * Corresponding author. Tel. : +81-3-5841-2824; Fax : +81-3-5841-8451; E-mail: [email protected] doi:10.2108/zs130062

Antilles (Losos, 2009; Schettino et al., 2010). Hence, anoles have become model organisms for the study of adaptive radiation and correlations between ecology, locomotion, and morphology. Some studies have reported correlations between the limb length and substrate uses such as perches of different diameters, or behaviors, such as jump frequency, in Anolis lizards (Pounds, 1988; Losos, 1990a; Irschick and Losos, 1998; Toro et al., 2004). However, despite the fact that differences in musculoskeletal characteristics serve as a basis for diversification of locomotion and behavior, quantitative data are lacking on the force production mechanisms and appendicular musculoskeletal morphology in this genus (Curtin et al., 2005; Vanhooydonck et al., 2006b). According to one study comparing two types of gekkotan lizards (Zaaf et al., 1999), some differences in musculoskeletal traits have been observed between terrestrial and arboreal species. A ground-dweller gekkotan (Eublepharis macularius) has well-developed knee and ankle extensor muscles, whereas a climbing gekkotan (Gekko gecko) has expanded humerus and femur retractors. These morphological characteristics may reflect adaptations to different loco-

Comparative anatomy of limbs in Anolis

motor styles and habitat uses (Zaaf et al., 1999). However, no comparison of closely-related squamate taxa has been performed. In Anolis, for example, few studies have focused on the relationships between appendicular musculoskeletal traits and behaviors or habitat uses among species or ecomorphs (Vanhooydonck et al., 2006a, b; Herrel et al., 2008; Legreneur et al., 2012). Furthermore, the specific function of each muscle remains to be elucidated. Due to their morphological and ecological variety, the Anolis lizards are good models for investigating the effects of locomotor style and habitat use on musculoskeletal systems. In the present report, we explore differences in musculoskeletal morphology in the limbs of five Anolis species of different ecomorphs, recognized based on different locomotor styles, body sizes, body shapes, and habitat use patterns (Collette, 1961; Schluter et al., 1997; Beuttell and Losos, 1999; Butler et al., 2000). Anolis sagrei and Anolis bremeri are of the trunk–ground type that tends to be found on the ground or broad tree trunks (Schettino, 1999). These species have relatively long hindlimbs and high frequency of rapid running (Losos, 1990b). A trunk–crown anole, Anolis porcatus, is a wide-ranging arboreal species whose habit ranges from tree trunks to the narrow twigs near the canopy top (Schettino, 1999). These three species are mediumsized (see Table 1). In contrast, Anolis equestris is a remarkably large anole classified as crown–giant ecomorph (Losos, 2009). Individuals of this large species spend the greater part of their time on the high branches of tree crowns, and generally do not move particularly rapidly (Schettino, 1999). Anolis angusticeps is considered a twig type, characterized by its small, slender body and short limbs (Losos, 2009). This species often moves steadily and slowly on narrow surfaces (Schettino, 1999). The goal of this study is to examine the relationships among limb morphology, habitat uses, and locomotion styles in these five species by investigating and comparing musculoskeletal traits in analyses of power and the lever arm. MATERIALS AND METHODS Specimens Between September 2010 and September 2011, we captured a total of 21 adult Anolis lizards by hand or noose in Cuba (Table 1). To eliminate the effects of sexual dimorphism from the analysis, only adult male individuals were used, except for A. equestris, for which no male samples were found. Animals were anesthetized and fixed in 100% ethanol, and then were stored in 70% ethanol. Because dry muscles are easily frayed and difficult to isolate, the samples were preserved in 30% ethanol overnight before dissecTable 1. Anolis lizards specimens examined in this study. N indicates the number of individuals examined, SVL is the length from the snout to vent, mass is the weight measured just before dissection. The mean value and standard deviation of each measurement are indicated for each species. Ecomorph classification follows Losos (2009). species

ecomorph

n

SVL (mm)

mass (g)

A. sagrei A. bremeri A. porcatus A. equestris A. angusticeps

Trunk-Ground Trunk-Ground Trunk-Crown Crown-Giant Twig

5 4 4 3 5

55.5 ± 3.38 60.6 ± 4.69 67.7 ± 6.24 143.3 ± 8.01 40.5 ± 1.84

3.601 ± 0.55 5.329 ± 1.05 5.89 ± 1.91 62.95 ± 14.7 0.723 ± 0.096

455

tion. The fore- and hindlimbs of the specimens were dissected and each muscle was isolated under a microscope (S240; OLYMPUS, Tokyo, Japan). Eighteen forelimb muscles that are particularly important for locomotive joint motion and 22 muscles from hindlimb were chosen for measurements (Table 2) (Jenkins and Goslow, 1983; Zaaf et al., 1999; Gans et al., 2008; Herrel et al., 2008). Measurements Two parameters were measured and compared: muscle mass and length of the moment arm. Muscle force is often indicated by physiological cross-sectional area (PCSA). However, muscle force should correlate strongly with muscle mass. PCSA is proportional to the product of the muscle mass and the cosine of the pennation angle, and is inversely proportional to the product of the density of muscle and the fiber length. Hence, muscle mass were able to use for fully reliable index of muscular force (Fujiwara et al., 2011). Each isolated muscle was blotted dry and weighed (± 0.01 mg) using a SHIMADZU balance (AUW-220D; Kyoto, Japan). Moment arm length was defined as the distance from the center of rotation in each joint to the point of muscle insertion, thus representing the maximum possible moment arms (An et al., 1984; Fujiwara et al., 2011). Given the trade-off between torque and excursion, muscles with longer moment arms exert larger torque, and muscles with shorter moment arms produce grater excursion. Lengths were measured using calipers (± 0.05 mm). Statistical analyses To remove the body size effect among different growth stages and/or species, all measurements were corrected for size using linear regression (ln). For the muscle mass, residuals from the regression of the ln-transformed values against the ln-transformed total body mass were used. For the moment arm length, residual of the regression from the ln-transformed values against the ln-length of the limb bone on which the muscle inserts were used. All characters were tested for significance of differences among species by analysis of variance (ANOVA) with post hoc Tukey-Kramer multiple comparisons, using a level of significance of 0.05.

RESULTS Anatomy The appendicular musculature is illustrated in Fig. 1. The function, origin, and insertion data listed in Table 1 are principally based on A. sagrei. The nomenclature of Gans et al. (2008) is used. Additional references including Herrel et al. (2008), Jenkins and Goslow (1983), Oldham and Smith (1975), and Snyder (1954) were also consulted for the descriptions of the limb muscle. Muscle homology was conserved among all five species on a gross morphological level. Muscle mass The results of the comparison of normalized muscle mass are presented in Figs. 2 and 3. Statistically significant differences, presumably characterizing variations in ecology and behavior among Anolis species, were observed in some muscles. The ground-dweller anoles A. sagrei and A. bremeri were both equipped with notably larger knee extensors (M. iliotibialis, M. ambiens, M. femorotibialis externus, and M. femorotibialis internus) and ankle extensors (M. gastrocnemius and M. peroneus longus) than the other three species (Fig. 3). In contrast, the femoral retractor M. caudifemoralis longus was larger in the other three species, especially A. equestris and A. angusticeps (Fig. 3). These two species also possessed well-developed adductor muscles in the elbow joint, M. flexor carpi radialis or M. flexor carpi ulnaris

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Table 2. Function, origin, and insertion of muscles of limbs in anoles. Nomenclature of Gans et al. (2008) is used. Elbow adductor and abductor muscles were examined using the methods of determined in Fujiwara and Hutchinson (2012). *Deeper structures, not illustrated in Fig. 1. Function

Muscles

Abbr.

Origin

Insertion

forelimb humeral adductor M. pectoralis profundus

pecp

M. coracobrachialis longus

Interclavicular bone and ventral midline of the sternum cobra Posterior side of coracoid

Ventral side of deltopectoral crest of the humerus Medial epicondyle of humerus

M. scapulodeltoideus

dels

External side of deltopectoral crest

M. subcoracoscapularis; coracoid portion*

subc

Lateral surface of the suprascapula and the scapula Entire internal face of the coracoid

M. clavodeltoideus M. supracoracoideus

delc Interventral side of clavicle supco Ventrolateral aspect of the coracoid plate

External side of deltopectoral crest Lateral tuberosity of humerus

M. latissimus dorsi

ld

M. pectalis superficialis M. subcoracoscapularis; scapular portion*

pecs subs

Tendon to dorsal-proximal aspect of humerus Ventral side of deltopectoral crest Lesser tubercle

M. triceps complex

tri

Dorsal-lateral side of deltopectoral crest, Olecranon process of the ulna medial surface of the humeral shaft, tendon from the lateral surface of scapular, and scapulohumeral ligament.

M. biceps brachii M. brachialis anticus

bic bra

Tendon of ventral coracoid Deltopectral crest and ventral sides of the humeral shaft

Tendon of dorsal-proximal radius Tendon of dorsal-proximal radius

M. flexor carpi radialis

fcr

Entire medial aspect of radius shaft

M. flexor carpi ulnaris

fcu

Lateral aspect of the humeral entepicondyle Dorsal surface of entepicondyle

M. extensor carpi radialis M. ectensor carpi ulnaris

ecr ecu

Ectepicondyle of the humerus Ventral aspect of the ectepicondyle

Dorsal aspect of radius Ulnare and fifth metacarpal

M. extensor digitorum longus

edl

Short tendon from lateral side of ectepicondyle

Dorsal aspect of 2 and 3 metacarpals

M. flexor digitorum longus

fdl

Medial part of entepicondyle of the humerus

Tendon whichi splits to insert on the distal pharanges

humeral abductor

Lesser tubercle

humeral protractor

humeral retractor Aporoneurosis to the dorsal midline of the body Xiphisternal rib Medial aspect of the suprascapular certilage and posterior portion of scapula

elbow extensor

elbow flexor

elbow adductor

Eendon to fifth metacarpal

elbow abductor

wrist extensor

wrist flexor

Continued.

(Fig. 2). For knee flexors, the trend differed among muscles. Two muscles (M. flexor tibialis internus lateralis and M. flexor tibialis internus medialis) originating on the medial side of the ilioischiadic ligament were emphasized in A.

sagrei and A. bremeri, whereas two other muscles (M. flexor tibialis internus posterior and M. iliofibularis) from the lateral side of the ilioischiadic ligament were expanded in A. porcatus (Fig. 3).

Comparative anatomy of limbs in Anolis Table 2.

457

Continued.

Function

Muscles

Abbr.

Origin

Insertion

hindlimb femur adductor M. puboischiofemoralis externus* pife M. adductor femoris add

Ventral aspect of the pubis and ischium Puboischiadic ligament

Internal trochanter of femur Ventral aspect of femur, reach to epicondyle

M. iliofemoralis*

ilfem

Ventral-anterior aspect of the ilium

Lateral side of proximal femur

M. puboischiofemoralis internus

pifi

Anterodorsal aspec of the pubic plate and internal wall of the ischium.

Tendon to anterior proximal side of femur

M. caudifemoralis longus

cfl

Transverse process of caudal vertebra

Stout tendon to the femoral trochanter

M. iliotibialis

it

Aponeuroses from the lateral surface of ilium

M. puboischiotibialis

pit

M. flexor tibialis externus

fte

Aponeuroses from the ischiopubic ligament. Lateral-posterior aspect of ilioischiadic ligament Medial-anterior aspect of ilioischiadic ligament Medial-anterior aspect of ilioischiadic ligament Lateral-posterior aspect of ilioischiadic ligament

femur abductor

femur protractor

femur retractor

knee extensor

M. ambiens M. femorotibialis externus M. femorotibialis internus

Lateral side of broad tendon to head of tibia, common with the knee extensors. amb Aponeuroses from the pubis and Medial side of broad tendon to head acetabulum. of tibia. fetiex Lateral aspect of femoral shaft, for about Lateral side of broad tendon to head three quarter of its length of tibia fetiin Medial side of femoral shaft Medial side of broad tendon to head of tibia

knee flexor

M. flexor tibialis internus medialis ftim M. flexor tibialis internus lateralis* ftil M. flexor tibialis internus posterior ftip

Tendon to medial side of tibia head Tendon to proximolateral surface of tibia Mesial aspect of the proximal end of the tibial shaft, proximal side of pit Tendon to proximolateral surface of tibia Medial side of tibia head, common with the M. puboischiotibialis, distal side of pit. Tendon onto proximolateral aspect of the tibia Tendon of proximal fibular shaft

M. pubotibialis

pt

Aponeuroses from the pubic tubercle

M. iliofibularis

ilfib

Posteroventral margin of the iliac blade

M. gastrocnemius

gas

Stout tendon to proximolateral margin of the phalanges

M. peroneus longus M. flexor digitorum longus

perl fdl_h

Tendon from knee joint meniscus, femur on the tibial side, dorsal tubercle on the femur on the fibular side Tendon from fibular side of the femur Lateral epicondyle of the femur

M. tibialis anterior

ta

M. peroneus brevis

perb

Anteromesial aspect of the head of the tibia Lateral aspect of the fibular shaft

M. extensor digitorum longus

edl_h Dormesial aspect of the femoral epicondyle

Ventromesial aspect of the proximal end of the first metatarsal Posterodorsal side of the fifth metatarsal Dorsal side of third metatarsal

ankle extensor

Ventral side of the fifth metatarsal Tendon to the proximal heads of astragalocalcaneum and digits.

ankle flexor

458

W. Anzai et al.

genus are known to be tree climbers. However, our data indicate tri that well-developed extensor musdels cles in the knee and ankle joints are present in ground-dweller A. ecu sagrei and A. bremeri whereas the fcu fdl bra femoral retractor muscle, M. fcr tri caudifemoralis longus, is larger in fcu the other three species (Figs. 2 pecs and 3). Both A. sagrei and A. bic cobra ecu bremeri run on the ground or delc broad surfaces (Schettino, 1999). supco Their large hindlimb extensors are ecr pecp bra pecs presumably advantageous for bic edl pecp powerful kicking off from ground and other surfaces and fast running (Reilly, 1995, 1998; Fieler and Jayne, 1998; Herrel et al., 2008). C D pifi Anolis porcatus, A. equestris, it and A. angusticeps are rarely amb fetiex found on the ground, and lead a pit (cut) more arboreal life (Schettino, add 1999; Losos, 2009). For climbing cfl in lizards, limb retractors provide ftim propulsive force and counteract fdl_h gas pt gravity (Zaaf et al., 1999). Wellftip fte ilfib developed femoral retractor musamb ftip edl_h cles enable these lizards to supperb port their bodies and occupy tree perl substrates. These muscles are gas fetiin also useful for scansorial locomoedl_h tion (Zaaf et al., 1999). Though ta femoral retractors also function in running in lizards (Fieler and Jayne, 1998; Reilly and Delancey, 1997; Russell and Bauer, 1992; Spezzano and Jayne, 2004), some studies demonstrated the adaptive role of great femoral retractor muscles for climbing Fig. 1. The appendicular musculature of Anolis. (A) Lateral side of the trunk and left forelimb of A. bremeri; (B) Ventral side of the trunk and left forelimb of A. sagrei; (C): Dorsal side of the left (Snyder, 1954; Zaaf et al., 1999). hindlimb of A. porcatus; (D) Ventral side of the left hindlimb of A. equestris. The most superficial The data in this study is consistent M. puboischiotibialis have been cut. For abbreviations of muscle names, see Table 2. with the latter hypothesis. Five species of Anolis show different adaptive patterns for terrestrial or Moment arm arboreal substrates in terms of appendicular musculoskeleThe results of the comparison of size-corrected moment tal traits. These results are consistent with previous study of arm length in the five species are illustrated in Figs. 4 and other Anolis (Herrel et al., 2008) and/or gekkotans (Zaaf et 5. Moment arms of statistically significant shorter length al., 1999), hence other lizard genera may show similar patterns. were observed in the shoulder (M. pectoralis profundus and M. pectoralis superficialis) and elbow (M. brachialis anticus Different adaptive strategies for arboreal habitat and M. biceps brachii) muscles of A. porcatus compared to Muscles with shorter moment arms were observed in at least on another species (Fig. 4). In contrast, hindlimb the shoulder and elbow joints of A. porcatus compared to muscles showed little difference between species, except for other species (Fig. 4). This species is regularly found on the M. caudifemoralis longus, for which relatively longer values full spectrum of surface diameters in arboreal substrates, were observed in A. equestris and A. angusticeps. such as the trunk, branch, leaves, or narrow twigs (Losos, DISCUSSION 2009). The greater flexibility of the forelimb may be effective Terrestrial or arboreal adaptation in Anolis lizards for propulsion and stabilization in complex arboreal situaAnolis lizards are categorized as arboreal animals in tions, where a wider excursion of the forelimb is beneficial many studies (e.g., Peterson, 1973), as some species in the (Foster and Higham, 2012). According to Higham et al.

A

ld

B

Comparative anatomy of limbs in Anolis

dels

0.2

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−0.6

−0.1

−0.2

0.1

1 0 −1 −2

−0.2

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0.0

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0.0 0.1 0.2 0.3

0.0 0.1 0.2 −0.2

bra

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A. bremeri

0.0

A. porcatus

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A. sagrei

A. equestris

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ecu

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fcu

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pecs

ld

tri

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subs

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−0.4

−0.4

−0.2

0.0

0.0

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0.2

cobra

−0.2

pecp

459

A. angusticeps

Fig. 2. Boxplots showing the normalized values for the forelimb muscles mass among five species of Anolis. The vertical axis indicated the residuals from ln-transformed muscle mass regressed against ln-total mass. Superscript letters (a, b, and c) indicate that significant differences exist between species with different letters by ANOVA and Tukey-Kramer test (P < 0.05). For muscle abbreviations, see Table 2.

(2001), wider-ranging arboreal trunk–crown species are more capable of swift movements and angled turning than ground-dweller trunk–ground or inactive twig Jamaican Anolis species. A shorter moment arm provides wider excursion angles in each joint, thereby facilitating limb movement (Peterson, 1973). This muscular trait suitable for branch-tobranch locomotion was found to be a characteristic of forelimb of A. porcatus in this study. By contrast, the other two arboreal species, A. equestris and A. angusticeps, were equipped with large mass of adductor muscles in the elbow joints (Fig. 2). Moreover, in measurements of the mass of the four knee extensors, the internal muscles (M. ambiens and M. femorotibialis internus) in A. equestris and A. angusticeps were found to be larger than those in A. porcatus, while the external muscles (M. iliotibialis and M. femorotibialis externus) were smaller in A.

equestris and A. angusticeps (Fig. 3). Both species tend to spend time moving slowly and hanging on branches or twigs, which are nearly equal (in diameter) to their body thickness (Losos, 2009). In lizards, grasping ability with their manus is correlated with the development of M. flexor digitorum longus (Abdala et al., 2009). However, it appears that the two species often clasp branch not only by manus but also by their entire limbs and body (personal observations). When they hold on to the branch with short limbs as clipping, it is thought that they require strong forces through adduction of the limbs. Another notable point is that similar musculoskeletal characteristics were observed in the limbs of A. equestris and A. angusticeps, despite a hundredfold difference in body mass. This suggests that these two species, which are similar in terms of locomotion style, require similar appendicular musculoskeletal traits to achieve this

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add

ilfem 0.4 0.2

Fig. 3.

0.2 0.4

b

b

0.0 −0.4 0.2 −0.2 −0.6 0.4

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b

edl_h

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ilfib 0.1 −0.1

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b

ta

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b

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perb a

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fdl_h

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perl

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gas a

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pt

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fetiex

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it 0.1

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pifi

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A. sagrei A. bremeri A. porcatus A. equestris A. angusticeps

Boxplots showing the normalized vales for hindlimb muscles mass in the five species studied. For muscle abbreviations, see Table 2.

locomotion, although they differ considerably in body size. Variety of function of knee flexor muscles Among the knee flexors, the two muscles, M. flexor tibialis internus lateralis and M. flexor tibialis internus medialis, originating from the medial side were expanded in A. sagrei and A. bremeri, while the two other muscles, M. flexor tibialis internus posterior and M. iliofibularis, arising from the lateral side were emphasized in A. porcatus (Fig. 3). When Dipsosaurus dorsalis, a quadrupedal iguanid lizard, runs at

higher speeds on a broad surface, the femur becomes remarkably adducted during the stance phase for raising the hip height and increasing the stride length (Fieler and Jayne, 1998; Russell and Bels, 2001). Moreover, the M. flexor tibialis internus lateralis and medialis are active during adduction and retraction of the femur in running Sceloporus clarkii, as measured by electromyogram (Reilly, 1995, 1998), whereas no correlation was found between M. iliofibularis and locomotor performance in Sceloporus woodi (Higham et al., 2011). Therefore, we postulate that the M. flexor tibi-

Comparative anatomy of limbs in Anolis

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delc

0.0 0.1 0.2 0.3

−0.6

−0.4

−0.2

−0.2 −0.4

b

dels

cobra a

0.2 0.4

a

a

0.0

0.2

pecp a

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Fig. 4. Boxplots showing the normalized values for the forelimb muscles moment arm among five species of Anolis. The vertical axis indicated the residuals from ln-transformed length of moment arm regressed against ln-length of the bones which on the muscle insert. Superscript letters (a, b, and c) indicate that significant differences exist between species with different letters by ANOVA and Tukey-Kramer test (P < 0.05). For muscle abbreviations, see Table 2.

alis internus lateralis and M. flexor tibialis internus medialis are required for faster running in locomotion of A. sagrei and A. bremeri through femoral adduction. In contrast, the strong development of M. flexor tibialis internus posterior and M. iliofibularis in A. porcatus are apparently effective for maintenance of the femoral position on a nearly horizontal plane and holding the center of body mass close to the substrate with climbing. These findings indicate that the muscles regarded as “knee flexors” in previous studies play varying roles in locomotor performance. More detailed investigation with a focus on the function of each muscle is required. For instance, electromyographic patterns of limb movement may be quantified, as done by Jenkins and Goslow, 1983; Reilly, 1983, 1995 in Anolis and other genera of Lacertilia. In conclusion, the single genus Anolis has evolved a rich diversity of appendicular musculoskeletal morphology and

adaptive strategy within the Cuban Islands. The present comparative analysis demonstrated a correlation between the myological characteristics evolution and locomotor styles and habitat uses in Anolis lizards. Hindlimb extensors, which are used for running, are better developed in terrestrial species, whereas arboreal species has acquired larger femoral retractors for climbing. Two different strategies were demonstrated in the forelimb of the arboreal species: wider excursion angles with shorter moment arms enabling quick movement in the forest crown, and better development expanding adductor muscles for hanging on branches. Differences in knee flexor muscles indicated diverse functions depending on the species. Two medial muscles used in adduction have developed in the ground-dwellers, and two lateral muscles involved in abduction have evolved in wellclimbers. Degrees in development of these muscles are cor-

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W. Anzai et al.

add

−0.2 −0.6 −1.0 0.0 −0.4

fdl_h

0.0

0.5

0.5 1.0

−0.4 0.2 0.4

−1.5

−0.6

−1.0

−0.5

−0.5

−0.2

0.0

perb

edl_h 0.0 −0.5

A. sagrei A. bremeri A. porcatus A. equestris A. angusticeps

−1.0

0.4 0.0 −0.4 −0.8

0.5 0.0

0.5

−0.2 −0.4

pt

0.4

0.2 0.0

perl

gas

ta

0.8

−0.4

−0.2 0.4

ftip

−0.2

0.0

0.0 −0.5 0.2

0.0 0.2 0.4 0.6

0.2 0.0

0.0 −0.4 −0.2

ftil

0.4

0.8

−0.4

ftim

0.5

1.0

b

fte

pit

0.2

0.2 −0.2

0.0

b

a

ilfib

−0.5

0.2

0.2 0.0

Knee ext. 0.4

cfl

−1.0

pifi

ilfem

−0.4 −0.2

−0.5

−0.4

0.0

−0.2

0.5

0.0

1.0

0.2

pife

Fig. 5. Boxplots showing the normalized values for the forelimb muscles moment arm among five species. Results for four knee extensors are illustrated collectively in Knee ext, as these muscles insert a common tendon. For other muscle abbreviations, see Table 2.

related with that in the proximity of their body center to the surface. These interpretations were based on biomechanical theory applicable to all lacertilian clades. Similar analyses should be conducted more widely in the species of Anolis and other lizards to clarify the relationships between musculoskeletal morphology, locomotor style, and habitat use in lizards. ACKNOWLEDGMENTS We are grateful to Lazaro Echenique-Diaz, Hirosh Akashi, and Hajime Wakasa for helping us collecting specimens. We also thank Shin-ichi Fujiwara, Daisuke Koyabu, Mugino Kubo, Yasuhisa Nakajima, Soichiro Kawabe for helpful advice. We also thank reviewers for comments that significantly improve the manuscript. Collection and exportation permits were provided by the Centro de Control y Gestión Ambiental (CICA) of the Agencia de Medio Ambiente de Cuba. This work was supported by MEXT/JSPS KAKENHI Grant Number 22405002, 23658253, 24370035 to HE

and 22405008 to MK.

REFERENCES Abdala V, Manzano AS, Tulli MJ, Herrel A (2009) The tendinous patterns in the palmar surface of the lizard manus: functional consequences for grasping ability. Anat Rec 292: 842–853 An KN, Takahashi K, Harrigan TP, Chao EY (1984) Determination of muscle orientations and moment arms. J Biomech Eng-T ASME 106: 280–282 Beuttell K, Losos JB (1999) Ecological morphology of Caribbean anoles. Herpetol Monogr 13: 1–25 Butler MA, Schoener TW, Losos JB (2000) The relationship between sexual size dimorphism and habitat use in Greater Antillean Anolis lizards. Evolution 54: 259–272 Curtin NA, Woledge RC, Aerts P (2005) Muscle directly meets the vast power demands in agile lizards. Proc R Soc B 272: 581– 584 Collette BB (1961) Correlations between ecology and morphology in

Comparative anatomy of limbs in Anolis anoline lizards from Havana, Cuba, and southern Florida. Bull Mus Comp Zool 125: 135–162 Fieler C, Jayne BC (1998) Effects of speed on the hindlimb kinematics of the lizard Dipsosaurus dorsalis. J Exp Biol 201: 609–622 Foster KL, Higham TE (2012) How forelimb and hindlimb function changes with incline and perch diameter in the green anole, Anolis carolinensis. J Exp Biol 215: 2288–2300 Fujiwara S, Hutchinson JR (2012) Elbow joint adductor moment arm as an indicator of forelimb posture in extinct quadrupedal tetrapods. Proc R Soc B 279: 2561–2570 Fujiwara S, Endo H, Hutchinson JR (2011) Topsy-turvy locomotion: biomechanical specializations of the elbow in suspended quadrupeds reflect inverted gravitational constraints. J Anat 219: 176–191 Gans C, Gaunt AS, Adler K (2008) The Skull and Appendicular Locomotor Apparatus of Lepidosauria. Biology of the Reptilia, Vol. 21, Morphology I. Society for the study of Amphibians and Reptiles, Salt Lake City Herrel A, Vanhooydonck B, Porck J, Irschick DJ (2008) Anatomical basis of differences in locomotor behavior in Anolis lizards: a comparison between two ecomorphs. Bull Mus Com Zool 159: 213–238 Higham TE, Davenport MS, Jayne BC (2001) Maneuvering in an arboreal habitat: the effects of turning angle on the locomotion of three sympatric ecomorphs of Anolis lizards. J Exp Biol 204: 4141–4155 Higham TE, Korchari PG, McBrayer LD (2011) How muscles define maximum running performance in lizards: an analysis using swing- and stance-phase muscles. J Exp Biol 214: 1685–1691 Irschick DJ, Losos JB (1998) A comparative analysis of the ecological significance of maximal locomotor performance in Caribbean Anolis lizards. Evolution 52: 219–226 Jenkins FA, Goslow GE (1983) The functional anatomy of the shoulder of the savannah monitor lizard (Varanus exanthematicus). J Morphol 175: 195–216 Legreneur P, Homberger DG, Bels V (2012) Assessment of the mass, length, center of mass, and principal moment of inertia of body segments in adult males of the brown anole (Anolis sagrei) and green, or carolina, anole (Anolis carolinensis). J Morphol 273: 765–775 Losos JB (1990a) Ecomorphology, performance capability, and scaling of West Indian Anolis lizards: an evolutionary analysis. Ecol Monogr 60: 369–388 Losos JB (1990b) The evolution of form and function: morphology and locomotor performance in West Indian Anolis lizards. Evolution 44: 1189–1203 Losos JB (2009) Ecology and Adaptive Radiation of Anoles: Lizards in an Evolutionary Tree. University of California Press, London Losos JB, Jackman TR, Larson A, Queiroz K, Schettino LR (1998) Contingency and determinism in replicated adaptive radiations of island lizards. Science 279: 2115–2118 Oldham JC, Smith HM (1975) Laboratory Anatomy of the Iguana. WC Brown Company Publishers, Dubuque Peterson JA (1973) Adaptation for arboreal locomotion in the shoulder region of lizards. Ph. D. Thesis, University of Chicago

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Poe S (2004) Phylogeny of anoles. Herpetol Monogr 18: 37–89 Pounds JA (1988) Ecomorphology, locomotion, and microhabitat structure: patterns in a tropical mainland Anolis community. Ecol Monogr 58: 299–320 Reilly SM (1995) Quantitative electromyography and muscle function of the hind limb during quadrupedal running in the lizard Sceloporus clarki. Zoology 98: 263–277 Reilly SM (1998) Sprawling locomotion in the lizard Sceloporus clarkii: speed modulation of motor patterns in a walking trot. Brain Behav Evol 52: 126–138 Reilly SM, Delancey M (1997) Sprawling locomotion in the lizard Sceloporus clarkii: the effects of speed on gait, hindlimb kinematics, and axial bending during walking. J Zool Lond 243: 417–433 Russell AP, Bauer AM (1992) The m.caudifemoralis longus and its relationship to caudal autotomy and locomotion in lizards (Reptilia: Sauria). J Zool Lond 227: 127–143 Russell AP, Bels V (2001) Biomechanics and kinematics of limbbased locomotion in lizards: review, synthesis and prospectus. Comp Biochem Phys A 131: 89–112 Schettino LR (1999) The Iguanid Lizards of Cuba. University Press of Florida, Gainesville Schettino LR, Losos JB, Hertz PE, Queiroz KD, Chamizo AR, Leal M, et al. (2010) The anoles of Soroa: aspects of their ecological relationships. Breviora 520: 1–22 Schluter D, Price T, Mooers AO, Ludwig D (1997) Likelihood of ancestor states in adaptive radiation. Evolution 51: 1699–1711 Snyder RC (1954) The anatomy and function of the pelvic girdle and hindlimb in lizard locomotion. Am J Anat 95: 1–45 Spezzano LC, Jayne BC (2004) The effects of surface diameter and incline on the hindlimb kinematics of an arboreal lizard (Anolis sagrei). J Exp Biol 207: 2115–2131 Toro E, Herrel A, Irschick D (2004) The evolution of jumping performance in Caribbean Anolis lizards: solutions to biomechanical trade-offs. Am Nat 163: 844–856 Vanhooydonck B, Aerts P, Irschick DJ, Herrel A (2006a) Power generation during locomotion in Anolis lizards: an ecomorphological approach. In “Ecology and Biomechanics: A Mechanical Approach to the Ecology of Animals and Plants” Ed by A Herrel, CRC Press, Boca Raton, pp 253–269 Vanhooydonck B, Herrel A, Damme RV, Irschick DJ (2006b) The quick and the fast: the evolution of acceleration capacity in Anolis lizards. Evolution 60: 2137–2147 Williams EE (1972) The origin of faunas. Evolution of lizard congeners in a complex island fauna: a trial analysis. Evol Biol 6: 47−89 Williams EE (1983) Ecomorphs, faunas, island size, and diverse end points in island radiations of Anolis. In “Lizard Ecology: Studies of Model Organism” Ed by RB Huey, ER Pianka, TW Schoener, Harvard University Press, Cambridge, pp 326–370 Zaaf A, Herrel A, Aerts P, Vree FD (1999) Morphology and morphometrics of the appendicular musculature in geckoes with different locomotor habits (Lepidosauria). Zoomorphology 119: 9–22 (Received March 27, 2013 / Accepted March 10, 2014)