JOURNAL OF MORPHOLOGY 00:000–000 (2007) Morphology of the Femoral Glands in the Lizard Ameiva ameiva (Teiidae) and Their Possible Role in Semiochemical Dispersion Beatriz A. Imparato,1 Marta M. Antoniazzi,1 Miguel T. Rodrigues,2 and Carlos Jared1,2* 1Laborato´rio de Biologia Celular, Instituto Butantan, CEP 05503-900, Sa˜o Paulo, Brazil 2Departamento de Zoologia, Instituto de Biocieˆncias, Universidade de Sa˜o Paulo, Sa˜o Paulo, Brazil ABSTRACT Many lizards have epidermal glands in munication are poorly known. Most morphological the cloacal or femoral region with semiochemical func- papers on this subject deal with European, Asian, tion related to sexual behavior and/or territorial demar- Australian, and African species (Cole, 1966a,b; cation. Externally, these glands are recognized as a row Maderson, 1968, 1970, 1985; Maderson and Chiu, of pores, opening individually in the center of a modified 1970, 1981; Chiu and Maderson, 1975; Schaefer, scale. In many species the pores are used as systematic characters. They form a glandular cord or, in some spe- 1901, 1902; Cohn, 1904; van Wyk et al., 1992; Duj- cies, a row of glandular beads below the dermis, and are sebayeva, 1998). More recently, the morphology of connected to the exterior through the ducts, which con- pre-cloacal glands in an amphisbaenian (Amphis- tinuously liberate a solid secretion. Dead cells, desqua- baena alba) was reported and their role in the nat- mated from the secretory epithelium, constitute the ural history of this species was analyzed (Anto- secretion, known as ‘‘a secretion plug.’’ The present work niazzi et al., 1993, 1994; Jared et al., 1999). focusses on the morphology of the femoral glands of the Femoral glands are the most common semio- teiid lizard Ameiva ameiva, correlating it to the way in chemical sources in lizards (Jullien and Renous- which the secretion is deposited in the environment. The Le´curu, 1973) although some glandular scales or results here obtained are compared to those available ‘‘generation’’ glands (Maderson, 1967, 1968) have for other lizards and amphisbaenians. We observed that the diameter of the glandular pores did not show signifi- been described in cordylids in other regions of the cant differences between males and females. The glands body (van Wyk et al., 1992). Femoral glands are comprise germinative and secretory cells, which pass localized in one or two rows on the ventral side of through at least three stages of differentiation, during each thigh. The first morphological studies on fem- which an accumulation of cytoplasmic granules, with a oral glands date from the end of XIX century and glycoprotein content, occurs. The cells eventually die beginning of XX century (A´ braha´m, 1930), and and desquamate from the secretory epithelium, forming concerned primarily European species. The subject a secretory plug mostly constituted by juxtaposed non- was ignored by morphologists until the 1960s, when fragmented secretory cells. Because of the arrangement papers by Gabe and Saint-Girons (1965) and Cole of the rosette-like scales surrounding the femoral pores, (1966a,b) were published. Uva (1967), Gasc et al. we suggest that when the animal is in a resting position, with its femoral regions touching the ground, these (1970), Chiu and Maderson (1975) and Chauhan scales may be involved in the breakage of their respec- (1986) then reported additional data on other species. tive plugs, depositing tiny portions on the substrate. In Although semiochemical communication is very this manner, it seems that the method for signal disper- important in lizard biology, studies focusing on sion in this species involves specifically adapted struc- these aspects are rare (Mason, 1992; Cooper, 1994; tures and does not simply involve the chance breakage Cooper et al., 1996). However, many experiments of the plug, as the gland secretes it. Signal dispersion have demonstrated that several species are able to must also be intimately associated with the animal’s recognize chemical signals secreted by femoral movement within its territory. J. Morphol. 00:000–000, glands (Cooper and Vitt, 1984; Alberts, 1989, 1993; 2007. Ó 2007 Wiley-Liss, Inc. Lopez et al., 1998, Martin and Lopez, 2000). KEY WORDS: squamata; lizards; teiidae; femoral pores; The localization of glands in both lizards and epidermal glands; Ameiva ameiva; pheromone amphisbaenians suggests that signals are pas- Among the Squamata, semiochemicals have an *Correspondence to: C. Jared, Instituto Butantan, Laborato´rio de important role in sexual behavior, defense, preda- Biologia Celular, Av. Vital Brasil 1500, CEP 05503-900, Sa˜o Paulo, tion, territorial marking, orientation, intraspecific Brazil. E-mail: [email protected] aggregation, and parental care (Halpern, 1992). Published online in Besides their importance in squamate biology, the Wiley InterScience (www.interscience.wiley.com) morphology of the structures related to semiocom- DOI: 10.1002/jmor.10473 Ó 2007 WILEY-LISS, INC. 2 B.A. IMPARATO ET AL. sively deposited in the environment during locomo- Histochemistry tion of the animals within their territory (Maderson, Historesin and paraffin sections were subjected to the follow- 1986; Jared et al., 1999). Among lizards possessing ing histochemical staining procedures (according to Bancroft femoral glands, signal dispersion involves the and Stevens, 1996): periodic acid-Schiff (PAS) and alcian blue breakage of small portions of the secretion plugs. pH 2.5, for identification of neutral and acidic mucosubstances, These remain on the substrate, releasing chemical respectively, and bromophenol blue, for identification of pro- signals for a period of time that must be related to teins. Sudan black B was used for identification of lipidic sub- stances. both the natural history and environment of a par- ticular species (Alberts, 1993). Ameiva ameiva is a teiid lizard with a broad geo- Photomicrography graphic distribution in Central and South Ameri- Photographs were obtained by using a Leica DMLB micro- cas, from Panama´ to South Brazil and North Ar- scope equipped with a Leica MPS60 photographic system. gentina (Vanzolini et al., 1980; Colli, 1991). The aim of the present paper was the study of the mor- phology and some aspects of the histochemistry of Scanning Electron Microscopy the femoral glands of the teiid Ameiva ameiva, A strip of skin from the femoral region containing 5–7 femo- and the comparison of the results with those ral pores was removed from the animals, fixed in Karnovsky so- obtained for other lizards and amphibaenians. We lution (Karnovsky, 1965) for 24 h, post-fixed in 1% osmium te- hope to contribute to the integrative knowledge of troxide, incubated in 1% tannin, and postfixed once more in 1% osmium tetroxide (Mizuhira and Futaesaku, 1974; Simionescu the natural history of lizards, especially in relation and Simionescu, 1976). The material was dried by the critical to chemical communication. Our data suggest that point method, coated with gold, and examined in a JEOL SM in Ameiva ameiva the differentiated scales con- 6100 and in a JEOL JSM 5300, operating between 15 and 25 kV. taining the femoral pores and the secretion plugs are structures adapted to the dispersion of chemi- Transmission Electron Microscopy cal signal on the substrate and seem to be associ- ated with the locomotion of these animals through- Fragments of the femoral glands (separated from the skin) of out their territory. about 1 mm3 were fixed in Karnovsky solution, pH 7.2 (Kar- novsky, 1965) for 24 h, post-fixed in 1% osmium tetroxide, con- trasted in 2% uranyl acetate, dehydrated in ethanol and embed- ded in epoxy resin. Ultrathin sections (60 lm) were obtained in MATERIALS AND METHODS a Sorvall MT 6000 ultramicrotome and were examined with a LEO 906E, operating at 80 kV. Animals Adult individuals of Ameiva ameiva Linnaeus 1758 were col- lected at Palmas (State of Tocantins, Brazil) and maintained for Morphometrical Study 2 years in the vivarium of the Laboratory of Cellular Biology of Adult male (N ¼ 14) and female (N ¼ 14) specimens of the Instituto Butantan. They were housed in terraria contain- Ameiva ameiva from the zoological collection of Museu de Zoo- ing humid earth at the bottom with dry branches above and logia da Universidade de Sa˜o Paulo, all from the same popula- pieces of plastic pipe for shelter. They were daily exposed to tion, were studied. The number and the diameter of the pores sunlight during the morning period. Food was supplied weekly in the femoral region of males and females were quantified and and was composed of a variation of live items (beetles - Tenebrio possible differences in both sexes were analyzed by Student molitor, crickets - Gryllus sp, cockroaches - Pycnocellus surina- test. The diameters of the pores of three regions in the pore row mensis), fruits, eggs, and minced beef. (the two extreme regions, named P1 and P3, and the middle For the morphological studies, six animals were sacrificed region, named P2) were measured. Because data were normally with a intraperitoneal overdose of sodic thiopental just after distributed and had homogeneous variances, they were ana- being collected from nature. Samples of the skin of the femoral lyzed by ANOVA test. region containing the femoral glands were removed and pro- cessed as described below. Voucher specimens were deposited in the collection of the RESULTS Museum of Zoology, University of Sa˜o Paulo (MZUSP). General Anatomy and Histology of the Femoral Glands Histology In Ameiva ameiva (Fig. 1), the ventral skin of the thighs, in both males and females, shows a sin- The skin containing the femoral glands was cut in strips of gle row of pores (Fig. 2), whose number varies 2 about 3 3 5mm using a razor blade and fixed in 4% parafor- from 17 to 23 in males and from 16 to 22 in maldehyde in PBS 0.1 M, pH 7.2 for 24 h. The material was dehydrated and embedded either in historesin (glycol methacry- females.