The Mammary Glands of the AmazonianManatee, Trichechus inunguis

(Mammalia: Sirenia): MorphologicalCharacteristics and Microscopic

AnatomyFERNANDA ROSA RODRIGUES,1* VERA MARIA FERREIRA DA SILVA,1

AND JOS�E FERNANDO MARQUES BARCELLOS2

1Laborat�orio de Mam�ıferos Aqu�aticos (LMA), Instituto Nacional de Pesquisas da Amazonia(INPA), Manaus, Amazonas, Brasil

2Laborat�orio de Histologia, Departamento de Morfologia (ICB), Universidade Federal doAmazonas (UFAM), Manaus, Amazonas, Brasil

ABSTRACTThe mammaries from carcasses of two female Amazonian manatees

were examined. Trichechus inunguis possesses two axillary mammariesbeneath the pectoral fins, one on each side of the body. Each papillamammae has a small hole on its apex—the ostium papillare. The mam-maries are covered by a stratified squamous keratinized epithelium. Theepithelium of the mammary ducts became thinner more deeply in the tis-sue and varied from stratified to simple cuboidal. There was no evidenceof glandular activity or secretion into the ducts of the mammary glands.Anat Rec, 297:1532–1535, 2014. VC 2014 Wiley Periodicals, Inc.

Key words: Amazonian manatee; Trichechus inunguis;mammary glands; anatomy; histology

INTRODUCTION

The presence of mammaries in females and the posi-tion of the genital orifices are the two main external sex-ually dimorphic characteristics of sirenians (Marmontelet al., 1992). Although they are cutaneous glands andnot immediately linked to the female reproductive tract,the mammaries are directly influenced by sexualhormones according to the reproductive status of theindividual (Gartner and Hiatt, 2007).

We examined the mammaries of the carcasses of twofemale Trichechus inunguis (designed PB1 and PB3)from a group of captive Amazonian manatees at theInstituto Nacional de Pesquisas da Amazonia (INPA) inAmazonas, Brazil. Following biometric measurements,anatomical descriptions of the mammaries wereprepared before they were removed and preserved in a10% formalin buffer for additional histological study.

Tissue fragments were sampled, dehydrated in etha-nol, cleared in xylene, and embedded in paraffin (Ban-croft and Stevens, 1996). The paraffin blocks were thencut into histological sections from 3 to 5 mm thick using

a microtome. Tissues were stained by standard histo-chemical techniques using hematoxylin and eosin (HE)(Carazzi, 1911) and Gomori trichrome stains (Gomori,1950), which indicate the presence of connective tissue,and Shiff ’s Periodic Acid (McManus, 1946) to identifymucins.

Grant sponsor: Conselho Nacional de Desenvolvimento Cient�ıficoe Tecnol�ogico (CNPq); Grant number: 131252/00–3; Grant spon-sors: Minist�erio da Ciencia e Tecnologia (MCT), Instituto Nacio-nal de Pesquisas da Amazonia (INPA), Projetos de PesquisasInstitucionais (PPI), Programa Piloto para a Protec~ao das Flo-restas Tropicais do Brasil (PPG-7).

*Correspondence to: Fernanda Rosa Rodrigues, Laborat�oriode Mam�ıferos Aqu�aticos (LMA), Instituto Nacional de Pesquisasda Amazonia (INPA), Av. Andr�e Ara�ujo, 2936, Petr�opolis,Manaus, Amazonas, Brasil 69011-970.E-mail: nanda_rodr@yahoo.com.br

Received 17 March 2012; Accepted 21 April 2014.

DOI 10.1002/ar.22956Published online 12 June 2014 in Wiley Online Library(wileyonlinelibrary.com).

THE ANATOMICAL RECORD 297:1532–1535 (2014)

VVC 2014 WILEY PERIODICALS, INC.

RESULTS AND DISCUSSION

An individual female T. inunguis has two mammariesthat are located one on each side of the body in the axil-lary region near the ventral insertion of the pectoral fins(Fig. 1). Macroscopic analysis revealed that the papillamammae is a rounded and protruding extension of theepidermis with a small hole in the center of its apex,known as the ostium papillare. The papilla is gray andlighter in color than the surrounding skin. The presenceof milk in the mammaries of PB3 (the older of the twoindividuals examined) was not observed because it hadnever reproduced prior to this investigation (Rodrigueset al., 2008). Some basic information and the biometricdata acquired from the two individuals used in thisstudy are presented in Table 1.

The majority of aquatic mammals have just one pairof mammaries, although walruses and polar bears havetwo pairs, usually located in an abdominal position.Cetaceans have a pair of mammary glands located insideof the mammary slits, next to the genital slit. In sire-nians, the mammaries are axillary, protuberant, and donot retract below the surface of the skin (Caldwell andCaldwell, 1985; Nishiwaki and Marsh, 1985; Pabst et al.,1999).

Sirenians are generally slow-moving animals with alow metabolic rate (Gallivan and Best, 1980), unlikecetaceans, which are normally highly active and fast-moving, with mammaries located below the surface ofthe skin to help minimize water resistance during swim-

ming (Pabst et al., 1999). Perhaps such streamlining isnot so important for the sirenians, given that they arethe only aquatic mammals with axillary mammaries;but more likely, the morphology and location of sirenianmammaries are pleisomorphic (De Jong and Zweers,1980; Pabst et al., 1999), rather than an aquatic adapta-tion, as their closest living relatives, elephants, alsohave one pair of axillary mammaries (Miall and Green-wood, 1878; Laursen and Bekoff, 1978).

Histological analysis showed the surface of the mam-maries to be a typical stratified squamous keratinizedepithelium (Fig. 2A,B). It was possible to identify threelayers in the epidermis: (1) stratum basale, (2) stratumspinosum, and (3) stratum corneum. The presence ofcells rich with the dark brown pigment melanin in thebasal layer was also notable. Melanin granules werepresent in most keratinocytes, always in a supranuclearposition to protect the cell nucleus from ultraviolet light,and declining in density toward the superficial stratumcorneum. The cells of the basal layer were more baso-philic, probably due to the high metabolic activity associ-ated with renewal of the epithelium. The nuclei of cellsin the stratum basale were rounded, becoming ellipticalor absent toward the superficial layers of the epithelium.The stratum spinosum had a greater number of celllayers and the outermost stratum corneum was intenselyeosinophilic. Nutrition and maintenance of the epithelialcells is by capillary diffusion from underlying connectivetissues. No blood vessels were present in the epithelium.

The dermis was composed of two layers: (1) papillaryand (2) reticular. The more superficial papillary layerwas composed of loose connective tissue and had finger-like projections that formed the papillae dermales,becoming more dense deeper in the dermis to form thereticular layer. Both layers of the dermis were richlyvascularized. Hair follicles (Fig. 2A,B) and sebaceousglands were found in the dermis. Sweat glands were notfound in any section studied. In pinnipeds, Tedman andBryden (1981) associated the presence of sebaceous andsweat glands with lubrication of the papilla to decreaseabrasion during suckling.

The glandular tissue was concentrated in the centerof the mammary. The epithelium of the mammary ductsbecame thinner more deeply in the tissue and variedfrom stratified to simple cubic. PB3 showed a system ofducts lined by a simple epithelium of high cuboidal cells.However, there was no evidence of any glandular activ-ity or secretion into the lumen of the ducts of the mam-mary glands. Mammary ducts covered by stratifiedepithelium (Fig. 2A) were observed in PB1, but the glan-dular tissue was still undeveloped.

Fig. 1. Mammary of T. inunguis (PB3) located in the axillary regionnear the ventral insertion of the pectoral fin. The specimen was placedin dorsal decubitus position.

TABLE 1. Total Linear Body Length, Age, andBiometric Measurements of the Mammaries of the

Two Individual T. inunguis Examined in this Study

Individual

Total linearbody length

(cm) Age

Papilladiameter

(mm)

Papillaheight(mm)

R L R L

PB1 82.5 Newborn 8.0 8.0 4.0 3.0PB3 201.0 Six Years 25.0 28.0 18.0 20.0

R, right; L, left.

MAMMARY GLANDS OF T. INUNGUIS 1533

In T. inunguis, the skin lining the mammary is histo-logically similar to that of other aquatic mammals (Pabstet al., 1999; Rommel and Lowenstine, 2001). In a previ-ous study concerning the healing process of tissue dam-aged by solar radiation, Camargo et al. (1996) identifiedthe presence of a stratum granulosum in epidermal tis-sue sampled from two T. inunguis individuals. This find-ing suggests that the layers of the epidermis may varyaccording to their location on an animal’s body, as thematerial used by these researchers was derived from theback region where the skin is considerably thicker thanthat covering the mammaries.

Mammary gland tissue in T. inunguis showed similarhistological features to those found in the Florida mana-tee, Trichechus manatus latirostris. Of the 39 adultfemales of the latter species that were studied, eight werelactating. The presence of an active gland with largenumbers of alveoli (lined by cuboidal epithelium) wasdemonstrated in samples of one specimen. Frequently,secretion could be observed within the lumen (Marmon-tel, 1988). Unfortunately, neither of the individuals exam-ined in the current study had active mammary glands. InDugong dugon (Marsh et al., 1984), the presence of milkwas verified by compression and cutting of the mammarytissue. From those animals suspected of having milk,samples were collected for histological analysis. The speci-mens of D. dugon were classified according to theirdegree of glandular activity, but no histological informa-tion about the mammaries was given.

Because mammary glands are influenced by sexual hor-mones, their histology varies according to age and thephysiological conditions of the organism. Therefore, infuture studies it would be interesting to describe suchchanges by examining additional T. inunguis specimens ofdifferent ages and reproductive status, such as individualsthat are pregnant, lactating, and of a more advanced age.

ACKNOWLEDGEMENTS

Authors are grateful to Maria de F�atima Barbosa, Creu-zimar B. dos Santos S. da Silva and the team of techni-cians at the Laborat�orio de Histopatologia da Faculdadede Medicina da Universidade Federal do Amazonas(UFAM). They are also grateful to Dr. Christina MaedaTakiya along with colleagues at the Laborat�orio de Pato-logia Celular da Universidade Federal do Rio de Janeiro(UFRJ) and Antonio Selso S. de Oliveira from the Uni-versidade Federal Fluminense (UFF). All of these peoplewere extremely helpful in their respective contributionsto this study, providing technical support and the infra-structure necessary to complete the histological analy-ses. Authors thank Thiago R. Rodrigues for his digitalartwork. They also appreciate the assistance of Dr.Jason Alan Mobley in translating this manuscript fromPortuguese to English.

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