JOURNAL OF MORPHOLOGY 273:11991213 (2012)

Seasonal Spermatogenic Cycle and Morphology of Germ Cells in the Viviparous Lizard Mabuya brachypoda (Squamata, Scincidae)
Arlette Herna ndez-Franyutti1 and Mari Carmen Uribe2*
n Acade mica de Ciencias Biolo gicas, Universidad Jua rez Auto noma Laboratorio: Acuicultura Tropical, Divisio xico de Tabasco, 86039 Villahermosa, Tabasco, Me 2 a de la Reproduccio n Animal, Departamento de Biolog a Comparada, Facultad Laboratorio: Biolog noma de Me xico, 04510 Me xico D.F. de Ciencias, Universidad Nacional Auto ABSTRACT We describe seasonal variations of the histology of the seminiferous tubules and efferent ducts of the tropical, viviparous skink, Mabuya brachypoda, throughout the year. The specimens were collected monthly, in Nacajuca, Tabasco state, Mexico. The results revealed strong annual variations in testicular volume, stages of the germ cells, and diameter and height of the epithelia of seminiferous tubules and efferent ducts. Recrudescence was detected from November to December, when initial mitotic activity of spermatogonia in the seminiferous tubules were observed, coinciding with the decrease of temperature, photoperiod and rainy season. From January to February, early spermatogenesis continued and early primary and secondary spermatocytes were developing within the seminiferous epithelium. From March through April, numerous spermatids in metamorphosis were observed. Spermiogenesis was completed from May through July, which coincided with an increase in temperature, photoperiod, and rainfall. Regression occurred from August through September when testicular volume and spermatogenic activity decreased. During this time, the seminiferous epithelium decreased in thickness, and germ cell recruitment ceased, only Sertoli cells and spermatogonia were present in the epithelium. Throughout testicular regression spermatocytes and spermatids disappeared and the presence of cellular debris, and scattered spermatozoa were observed in the lumen. The regressed testes presented the total suspension of spermatogenesis. During October, the seminiferous tubules contained only spermatogonia and Sertoli cells, and the size of the lumen was reduced, giving the appearance that it was occluded. In concert with testis development, the efferent ducts were packed with spermatozoa from May through August. The epididymis was devoid of spermatozoa by September. M. brachypoda exhibited a prenuptial pattern, in which spermatogenesis preceded the mating season. The seasonal cycle variations of spermatogenesis in M. brachypoda are the result of a single extended spermiation event, which is characteristic of reptilian species. J. Morphol. 273:11991213, 2012. 2012 Wiley Periodicals, Inc. KEY WORDS: spermatogenesis; testicular efferent ducts; viviparous lizard; Mabuya cycle;
1

spermatogenesis, is as important as the female reproductive cycle in determining the reproductive pattern of a species (Trauth, 1979). Spermatogenesis is an essential process in the biology of reproduction and results in production of haploid spermatozoa from diploid spermatogonial cells. The study of the spermatogenesis requires the recognition and identication of germ cell morphology during their differentiation and their occurrence throughout the seasonal cycle. The differentiation of germ cells involves specic cellular events: mitosis of spermatogonia, meiosis of spermatocytes, and spermiogenesis of spermatids. This process is followed by spermiation, the release of sperm from the seminiferous epithelium (Bakst et al., 2007; Hess and de Franca, 2008). In reptilian sauropsids, as amniotic species, spermatogenesis has special evolutionary interest among vertebrates because germ cell development represents a strategy between the anamniotic clades (shes and amphibians) and the other amniotic taxa (birds and mammals; Gribbins et al., 2003, 2007, 2008, 2009; Rheubert et al., 2009a,2009b). In the testes of birds and mammals, germ cells are sequentially situated along the seminiferous tubules in a continuous spatial development of spermatogenesis that results in waves of sperm release from specic portions of the tubules during the reproductive cycle (Leblond and Clermont, 1952; Clermont, 1972; Lin
Additional Supporting Information may be found in the online version of this article. *Correspondence to: Mari Carmen Uribe, Departamento de Bioloa Comparada, Facultad de Ciencias. Circuito Exterior, Ciudad g noma de Me xico, InsurUniversitaria, Universidad Nacional Auto n Coyoaca n, 04510 Me xico, D.F. Me xico. gentes Sur 3000, Delegacio E-mail: uribearanzabal@yahoo.com.mx Received 22 January 2012; Revised 4 April 2012; Accepted 13 April 2012 Published online 13 July 2012 in Wiley Online Library (wileyonlinelibrary.com) DOI: 10.1002/jmor.20050

INTRODUCTION The identication and denition of the seasonal changes of the male reproductive cycle, e.g., the
2012 WILEY PERIODICALS, INC.

1200

NDEZ-FRANYUTTI AND M.C. URIBE A. HERNA

and Jones, 1990; Franca and Godinho, 2003; Hess and de Franca, 2008). In contrast, reptilian spermatogenesis develops a single population of spermatozoa, which is released during a single spermiation event (Mayhew and Wright, 1970; Guillette and ndez de la Casas-Andreu, 1987; Guillette and Me n-Santa Cruz et al., 1994; GribCruz, 1993; Villagra bins et al., 2003, 2007, 2008, 2009; Ferreira et al., 2009; Rheubert et al., 2009a,2009b). Consequently, the analysis of morphological and physiological characteristics of spermatogenesis in reptiles represents a main aspect in the understanding of the evolution of male germ cell development in vertebrates. Species of the genus Mabuya are tropical grass skinks that are distributed in regions of America, Asia, and Africa. These species present remarkable reproductive patterns having several unique specializations among lizards: a) the genus Mabuya includes oviparous and viviparous species in Asia and Africa. However, all the species of America are viviparous (Fitch, 1970; Vitt and Blackburn, 1983, 1991); b) the females may initiate gestation at a small body size and subsequently grow during gestation (Vitt and Blackburn, 1983); c) viviparous species have a long period of gestation (912 months; Vitt and Blackburn, 1983, 1991); d) the ovaries develop different types of eggs, some species ovulate macrolecithal eggs (containing abundant yolk; Vitt and Blackburn, 1983) while other species ovulate microlecithal eggs (containing little yolk; Webb, 1958; Vitt and Blackburn, 1983, 1991; Flemming, 1994; Hernandez-Franyutti et al., 2005); e) therefore, there are different levels of lecithotrophy (when most nutrients for the embryo are provided by the yolk), or matrotrophy (when most nutrients for the embryo are transferred from maternal tissues during gestation; Vitt and Blackburn, 1983, 1991; Blackburn, 2000, 2005; Uribe et al., 2006). Male reproductive activity of Mabuya species has been described as a continuous process by gross analysis of the testes (weight and volume), in M. striata (Simbotwe, 1980) and M. mabouya (Somma and Brooks, 1976; Ramirez-Pinilla et al., 2002); in other species, spermatogenesis was dened as a seasonal cycle, as in M. heathi (Vitt and Blackburn, 1983), M. capensis (Flemming, 1994). In these species, the peak of spermatogenesis occurred during spring and early summer, which is characteristic of the prenuptial pattern (Saint Girons, 1982). Histological changes in the seminiferous tubules of Mabuya were mentioned only in the study of M. capensis by Flemming (1994), but he did not present histological images. Mabuya brachypoda is a viviparous lizard distributed from the southeast of Mexico to Costa Rica in Central America (Webb, 1958). Some reproductive aspects have been studied in adult females of this species. The ovarian cycle and oogenesis of M. brandez-Franyutti chypoda were documented by Herna et al. (2005), who identied a seasonal process durJournal of Morphology

ing which microlecithal eggs (0.91.3 mm in diameter) developed. The microlecithal eggs are considered the smallest described thus far in reptiles. The eggs attained maximum diameters during June-July. Macroscopic examination of the oviducts of M. brachypoda, collected between late June to late July by Webb (1958), showed embryos in the uterus. Histological analysis of the uterus of M. brachypoda during the annual cycle by Uribe et al. (2006), showed the presence of embryos between August and May. There are no previous descriptions of male reproductive system of M. brachypoda. Therefore, the goals of this study are to characterize the morphology of the germ cells during spermatogenesis and determine the changes of the seminiferous epithelia and efferent ducts in M. brachypoda during the annual cycle and to relate these characteristics to the long gestation period of the female reproductive cycle. MATERIAL AND METHODS
Male M. brachypoda (snout-vent length 5978 mm; N 5 36, 3 xico (18810 specimens per month), from Nacajuca, Tabasco, Me N, 9381 W), were collected during 2004. Meteorological data, mean monthly maximum and minimum temperatures and pre xico, were obtained from the Comision cipitation in Tabasco, Me xico (2004) (Supporting Information Nacional del Agua, Me Graph 1). The specimens were euthanized using ether and decapitation. A mid ventral incision opened the abdominal cavity. Maximum and minimum testicular diameters were measured. Testicular volume was subsequently calculated using the equation for the volume of an ellipsoid (V 5 4/3p (1/2L) (1/2W)2). Testes, and efferent ducts were removed and xed in alcoholic Bouins (815 h). After xation, the tissues were washed in 70% alcohol, dehydrated in a series of graded alcohols (80%, 96% and 100%), cleared in xylene, and embedded in parafn. Tissues were serially sectioned at 6 lm in longitudinal planes and stained with hematoxylin-eosin (Aguilar-Morales et al., 1996). The morphological changes of the seminiferous epithelium during the annual cycle were divided into eight phases (I to VIII) according to the classication described by Mayhew and Wright (1970) based on the morphological changes of the seminiferous epithelium in lizards of the genus Uma. This classication focuses on the type of germ cells bordering the tubular lumen during the annual cycle. The VIII phases are characterized by: I, mitosis of spermatogonia and no lumen; II, primary spermatocytes at luminal margin; III, secondary spermatocytes at luminal margin; IV, undifferentiated spermatids at luminal margin; V, spermatids in spermiogenesis at luminal margin; VI, spermatozoa in the lumen; VII, early regression with cellular debris in the lumen; VIII, complete regression, no mitosis of spermatogonia and reduced lumen. The progressive changes that occurred during the spermiogenesis were divided into seven steps according to Gribbins et al. (2009). The terms used for the two types of efferent ducts were ductuli efferentes and ductus epididymis as described by Sever (2010). Selected morphometrics such as: diameter and epithelial height of the seminiferous tubules, ductuli efferentes, and ductus epididymis were measured (N 5 25 per specimen) using a Carl Zeiss Axion Vision Imaging System. Digital photomicrographs were taken using an Olympus digital camera model C5050Z coupled to an Olympus CX31 microscope.

RESULTS The testes of M. brachypoda are ellipsoid structures (Fig. 1A,B), in the abdominal cavity and con-

SPERMATOGENESIS OF VIVIPAROUS LIZARD Mabuya brachypoda

1201

Fig. 1. Testes and epididymis of Mabuya brachypoda during different periods of spermatogenesis. A. Testes and epididymis during phase II of the seminiferous epithelium cycle. The seminiferous tubules of the testes are reduced. The ductus of the epididymis does not contain spermatozoa. B. Testes and epididymis during phase VI of the seminiferous epithelium cycle. The seminiferous tubules are larger compared to those seen in phase II in the Fig. 1A. The ductus epididymis encloses abundant spermatozoa. C. The seminiferous epithelium during phase II contains few germinal cells and there are no spermatozoa in the lumen of the seminiferous tubules. D. The seminiferous epithelium during phase VI contains abundant germinal cells and there are late spermatids at the apical end, adjacent to the lumen of the seminiferous tubules. Epididymis (E), late spermatids (1St), lumen (*), seminiferous epithelium (sep), spermatozoa (Sz), testes (T). Bars 5 A, B: 500lm, C, D: 100lm.

nected to the dorsal wall of the body by the mesorchium. Adjacent to the testis is the epididymis as a convoluted duct (Fig. 1A,B). The efferent ducts consisted of two segments. The proximal segment of the epididymis is closest to the testis and is formed by the ductuli efferentes. The distal segment is the ductus epididymis, which connects to the ductus deferens. The testes of M. brachypoda

present annual variations in testicular volume, diameter and epithelial height of the seminiferous tubules and diameter and epithelial height of the ductuli efferentes, and ductus epididymis (see supporting Information, Table 1). The reproductive cycle of M. brachypoda is highly seasonal. The phases of the spermatogenic cycle in the testis, and the presence of spermatoJournal of Morphology

1202

NDEZ-FRANYUTTI AND M.C. URIBE A. HERNA

zoa in the ductus epididymis dened the seasonal changes (Fig. 1C,D). Between November and February, the size of the testis, its volume, diameter and the height of epithelium of the seminiferous tubules, as well as the diameter and epithelial height of the efferent ducts, in early spermatogenesis were minimal (Fig. 1A,C) but they were maximal between May and July, i.e., during late spermatogenesis (Fig. 1B,D; Table 1, supporting Information). In early spermatogenesis, the seminiferous epithelium of the testicular tubules contains few germinal cells and there are no spermatozoa in the lumina of the seminiferous tubules (Fig. 1C). The ducts of the epididymis do not contain spermatozoa. In late spermatogenesis, the seminiferous epithelia of the testicular tubules contain abundant germinal cells and there are spermatids at the apical end, adjacent to the lumen of the seminiferous tubules (Fig. 1D), and the ductus epididymis contain abundant spermatozoa. Testicular Components The wall of the testis is surrounded externally by the tunica albuginea, formed by vascularized connective tissue with abundant collagenous bers. The testes contain the seminiferous tubules lined by seminiferous epithelia. These epithelia consist of Sertoli cells and germ cells. Sertoli cells form a permanent epithelium and are distributed around the circumference of the seminiferous tubules (Fig. 2A). Sertoli cells have a basal nucleus, are frequently triangular in shape with one or two nucleoli. The seminiferous epithelium is limited by a basement membrane (Fig. 2A) that separates the germinal compartment inside the seminiferous tubules, from the interstitial compartment outside the seminiferous tubules. The interstitial compartment is formed by connective tissue composed of Leydig cells, broblasts, collagen bers, myo-epithelial cells, nerve bers and lymphatic and blood vessels. Morphology of Germ Cells During the Spermatogenesis Spermatogonia. Spermatogonia (Fig. 2BE) are located at the base of the seminiferous epithelium, bordering the basement membrane. During the annual cycle they proliferate by mitosis, producing a large population of spermatogonia or initiating meiosis and becoming primary spermatocytes. There are two types of spermatogonia: spermatogonia A (Fig. 2B), and spermatogonia B (Fig. 2CE). Spermatogonia A have an average diameter of 21lm; are ovoid in shape, and their nuclei are spherical and contain ne granular heterochromatin with one or two dense and round nucleoli. The spermatogonia B are slightly larger;
Journal of Morphology

have an average diameter of 23.3 lm and are more spherical than the spermatogonia A. Their nucleus and cytoplasm are similar to those seen in the spermatogonia A. During mitosis the bers of the spindle were easily observed (Fig. 2E). Primary spermatocytes. Primary spermatocytes (Fig. 2FK) are spherical cells with a round nucleus. The nucleus undergoes morphological changes during the progression of meiosis I. According to the morphology of the chromosomes, several steps of meiotic prophase I were seen: a) during the leptotene stage (Fig. 2F), the average cell diameter is smaller than that of the spermatogonia, attaining a 20.2 lm average diameter. The chromatin initiates condensation and the chromosomes appear as thin, densely stained threads. b) During zygotene (Fig. 2G), the average cell diameter increases to 23.2 lm and condensation of the chromosomes advances. Next, the chromosomes become more lamentous as the homologous chromosomes pair form. c) During the pachytene stage (Fig. 2H), the spermatocytes have an average diameter of 25.6 lm. The paired chromosomes are even thicker and shorter than those seen in zygotene and maintain their intense stained afnity. These spermatocytes were the most abundant among the primary spermatocytes. d) During the diplotene stage (Fig. 2I), the spermatocytes are at the maximum diameter for primary spermatocytes, they attained an average diameter of 26.8 lm. The paired and dense chromosomes become gradually separated but remain united in several sites, irregularly distributed all along the chromosomes, forming chiasmata. These cells were the rarest of primary spermatocytes. The primary spermatocytes divide forming the spindle and the chromosomes arranged into metaphase I (Fig. 2J), continue through anaphase I (Fig. 2K), and telophase I, and complete cytokinesis to produce the secondary spermatocytes. Secondary spermatocytes. Secondary spermatocytes (Fig. 2L) are the result of the rst meiotic division. Since secondary spermatocytes are rarely observed it is assumed that rapidly enter the second division of meiosis. Secondary spermatocytes are spherical in shape and smaller in average diameter (18.9 lm) compared to the primary spermatocytes. The nucleus is spherical and the chromatin consists of ne laments, indicating the continuation of meiosis. After the second meiotic division, the secondary spermatocytes form step 1 spermatids. Spermatids. Spermatids (Fig. 2MS), in their early stages (Fig. 2M), are spherical cells and smaller, compared to secondary spermatocytes, with an average diameter of 14.8 lm. The spermatids go through metamorphosis during spermiogenesis. The morphological changes of the spermatid included formation of the acrosomal system, chromatin condensation, elongation of the nucleus and

SPERMATOGENESIS OF VIVIPAROUS LIZARD Mabuya brachypoda

1203

Fig. 2. Sertoli cell nuclei and germ cells during different stages of spermatogenesis in the seminiferous tubules of Mabuya brachypoda. A. Sertoli cell nuclei in the base of a seminiferous tubule. B. Spermatogonium A is ovoidal in shape. C. Spermatogonium B is round in shape. D. Spermatogonium B round in shape and has two dense nucleoli. E. Spermatogonium B during metaphase and a Sertoli cell nucleus. F. Leptotene spermatocyte. G. Zygotene spermatocyte. H. Pachytene spermatocyte. I. Diplotene spermatocyte. J. Primary spermatocyte in metaphase I of meiosis. K. Primary spermatocyte in anaphase I of meiosis. L. Secondary spermatocytes. M. Spermatid in step 1 of spermiogenesis. N. Spermatid in step 2 of spermiogenesis. O. Spermatid in step 3 of spermiogenesis. P. Spermatids in step 4 of spermiogenesis. Q. Spermatids in step 5 of spermiogenesis. R. Spermatids in step 6 of spermiogenesis. S. Spermatids in step 7 of spermiogenesis. T. Spermatozoa in the lumen of the seminiferous tubule. Diplotene spermatocyte (DSc), Leptotene spermatocyte (LSc), Pachytene spermatocyte (PSc), Primary spermatocyte in metaphase I (MSc), Primary spermatocyte in anaphase I (ASc), Secondary spermatocytes (SSc). Sertoli cell nuclei (Se), Spermatid in spermiogenesis; step 1 (1St), step 2 (2St), step 3 (3St), step 4 (4St), step 5 (5St), step 6 (6St), step 7 (7St), spermatogonium A (SgA), spermatogonium B (SgB), Spermatozoa (Sz), Zygotene spermatocyte (ZSc), Bars 5 5lm.

formation of the agellum. These changes occur gradually through seven steps according to Gribbins et al. (2009). During the rst step (Fig. 2M), the spermatid contained a spherical nucleus with haploid and ne granular chromatin; steps two

(Fig. 2N), and three (Figs. 2O). Spermatids are characterized by the formation of the acrosomal system and the start of chromatin condensation; steps four (Fig. 2P), ve (Fig. 2Q), and six (Fig. 2R), spermatids are characterized by elongaJournal of Morphology

1204

NDEZ-FRANYUTTI AND M.C. URIBE A. HERNA

tion of the nucleus; and step seven (Fig. 2S), is characterized by easy visualization of the agellum protruding into the lumen which suggests the germ cell is near to be released into the lumen. Spermatozoa. Once spermatids complete spermiogenesis (Fig. 2T), they are released into the lumen as spermatozoa. Spermatozoa are elongated, and the agellum is long and thin. Mature spermatozoa are transported to the efferent ducts. Annual Changes During the Seminiferous Epithelium Cycle The presence/absence of specic germ cells altered the morphology of the seminiferous epithelium during the annual cycle of Mabuya brachypoda through eight phases (I-VIII). Spermatogenesis was synchronous in all the seminiferous tubules. Phase I. The testes were in this phase during November and December (Fig. 3A,B). The majority of the seminiferous tubules contained only spermatogonia and Sertoli cells. The volume of the testes was 8.7 6 1.2 mm3. The diameter of seminiferous tubules was 61.8 6 2.2 lm. The seminiferous epithelium height was 28.3 6 1.7 lm. The lumen is occluded in most of tubules, and only a few of them had a visible small lumen (Fig. 3A). At this phase, recrudescence of spermatogenesis was detected undergoing spermatogonia in mitosis (Fig. 3B). As a consequence of the mitotic activity of spermatogonia, the number of germ cells in the seminiferous epithelium appeared to increase. Phase II. The testes were in phase II during January and February at which time the volume of the testes was 8.9 6 3.1 mm3(Fig. 3C). The diameter of the seminiferous tubules was 107.2 6 4.9 lm, and the seminiferous epithelium height was 31.4 6 1.0 lm. Spermatogonia continued proliferating entering into meiosis and formed abundant primary spermatocytes that were located at the margin of the lumen. Phase III. The testes were in phase III during March at which time the volume of the testes was 12.9 6 3.4 mm3 (Fig. 3D). The diameter of the tubules increased to 182.6 6 5.0 lm. The seminiferous epithelium heights also increased to 61.3 6 1.6 lm. Primary spermatocytes were numerous. Meanwhile, secondary spermatocytes were occasionally seen at the apical end of the seminiferous epithelium. Primary spermatocytes in pachytene of prophase I of meiosis were the most abundant type of germ cells identied in this phase. Occasionally, spermatids were also located at the margin of the tubular lumen. Phase IV. The testes were in phase IV during April at which time the volume of the testes was 47.7 6 5.5 mm3 (Fig. 4AD). The diameter of the seminiferous tubules was 271.7 6 6.7 lm, and the height of the seminiferous epithelium was 94.5 6
Journal of Morphology

2.4 lm.The seminiferous epithelium contained several layers of spermatids situated along the margin of the lumen. Most of the spermatids showed the initial morphological changes that occur during spermiogenesis, including acrosomal development and the initial elongation of the nucleus, as evidence of early spermiogenesis. Phase V. The testes were in phase V during May at which time the volume of the testes was 75.9 6 8.5 mm3 (Fig. 5AD). The diameter of the seminiferous tubules attained 306.2 6 7.4 lm, slightly larger than those seen in the previous stages. The height of the seminiferous epithelium attained 112.6 6 2.6 lm. The germinal epithelium formed large columns in which spermatids were located along the lumen. The number of primary spermatocytes decreased; meanwhile, the spermatids were very abundant. The spermatids advanced into late spermiogenesis presenting further elongation of the spermatid nucleus. Phase VI. The testes were in phase VI during June and July at which time the volume of the testes was 89.2 6 10.5 mm3 (Fig. 6AD). The seminiferous tubules attained their maximum diameter (356.6 6 17.4 lm), similar to that seen in the height of the seminiferous epithelia 122.1 6 5.9 lm. Spermatogenesis was completed during this phase. Numerous spermatids lined the margins of the seminiferous epithelium and spermatozoa were free within the lumen of the tubules (Fig. 6C,D). Phase VII. The testes were in phase VII during August at which time the volume of the testes was 52.6 6 12.0 mm3 (Fig. 7A,C). The diameter of the seminiferous tubules was reduced to 312.5 6 16.8 lm. During early regression (Fig. 7A,B), the seminiferous epithelium decreased in thickness to 89.2 6 5.4 lm; but, there were smaller regions with only three or four germ cell layers, containing spermatogonia, and some spermatids in spermiogenesis. During late regression (Fig. 7C), most of spermatocytes and spermatids disappeared, only remaining spermatogonia and Sertoli cells in the seminiferous epithelium. Abundant cellular debris, and some scattered spermatozoa were seen in the lumen of the seminiferous tubules. Phase VIII. The testes were in phase VIII during September and October at which time the volume of the testes was 20.6 6 9.0 mm3 (Fig. 7D). The seminiferous tubules attained the minimum diameter (77.7 6 1.8 lm), as the height of the seminiferous epithelium (11.6 6 1.4 lm). The lumen of the tubules was mostly occluded. The seminiferous epithelium contained, exclusively, spermatogonia and Sertoli cells. No mitotic activity of the spermatogonia was seen. Efferent Ducts The efferent ducts of M. brachypoda (Fig. 8AC) are ductuli efferentes and ductus epididymis. Near

SPERMATOGENESIS OF VIVIPAROUS LIZARD Mabuya brachypoda

1205

Fig. 3. Mabuya brachypoda, seminiferous tubules during phases I, II and III of the cycle. A. The seminiferous epithelium during phase I contains only Sertoli cells and spermatogonia. The lumen is very small or it is occluded. B. Spermatogonia in metaphase of mitosis are seen, where the bers of the spindle are evident. C. Seminiferous tubules during phase II of the cycle show spermatogonia A and B at the base of the seminiferous epithelium. Primary spermatocytes are abundant at the center of the tubules. The lumen is lightly larger than in the previous phase. D. Seminiferous tubules during phase III of the cycle. Secondary spermatocytes are seen at the luminal margin of the seminiferous epithelium. The lumen is clearly larger than in the previous phase. Lumen (*), Sertoli cells (Se), spermatogonia A (SgA) spermatogonia B (SgB), spermatogonia in metaphase of mitosis (arrowheads), primary spermatocytes (1Sc). Bars 5 12lm.

the efferent duct is observed the sexual segment of the kidney. The ductuli efferentes (Fig. 8AD) were the smallest efferent ducts. They were lined by cuboidal epithelium that had large stereocilia bordering the lumen (Fig. 8D) that we need to verify their presence at the electron microscope level. These ducts had a similar size throughout the reproductive cycle (Table 1, supporting Information). Their

minimal and maximal diameters were 17.5 6 3.4 lm from January through February, and 24.6 6 2.9 lm from June through July. Their epithelial height varied from 8.1 6 2.1 lm to 10.5 6 3 lm respectively. Only during the stage VII, at the end of the spermiation and during the early regression of the reproductive cycle in the seminiferous tubules, the ductuli efferentes contained a few spermatozoa.
Journal of Morphology

1206

NDEZ-FRANYUTTI AND M.C. URIBE A. HERNA

Fig. 4. Mabuya brachypoda, seminiferous tubules during phase IV of the cycle. A, B. Spermatids before metamorphosis at the luminal margin of the seminiferous epithelium. Some of them present early metamorphosis. The lumen is seen. C, D. Abundant spermatids during early metamorphosis are seen at the luminal margin of the seminiferous epithelium. Spermatids (St), spermatids in early metamorphosis (eSt), lumen (*). Bars 5 A: 20lm. B: 10lm. C: 30lm. D: 20lm.

The ductus epididymis (Fig. 8AC,E) was distal to the ductuli efferentes, and was larger than the ductuli efferentes. The diameter of the ductus epididymis and the height of the epithelial cells changed during the reproductive cycle. Minimum lumen size was measured as 67.8 6 8.1 lm in January-February; maximum lumen was 226.3 6 15.2
Journal of Morphology

lm between June through August, during the phases V to VII when masses of spermatozoa were in the lumen. The ductus epididymis was lined by columnar epithelium (Fig. 8E), and it hypertrophies during June-July. The height of the epithelium varied from 9.1 6 1.2 lm from JanuaryFebruary to 33.0 6 3.0 lm during June-July. From

SPERMATOGENESIS OF VIVIPAROUS LIZARD Mabuya brachypoda

1207

Fig. 5. Mabuya brachypoda, seminiferous tubules during phase V of the cycle. AD. Spermatids during middle metamorphosis and late metamorphosis are seen at the luminal margin of the seminiferous epithelium. The seminiferous epithelium presents columns extended to the lumen. Spermatids in middle metamorphosis (mSt) and late metamorphosis (lSt), columns (co), lumen (*). Bars 5 A: 40lm. B: 10lm. C: 20lm. D: 10lm.

May through August, the cytoplasm of the epithelial cells contained abundant secretory granules (Fig. 8E). DISCUSSION The results of our study are in agreement with earlier studies of reptilian sauropsids by Trauth

(1979). Histological analysis of Mabuya brachypoda shows the precise time and duration of reproductive activity and reects upon the environmental factors that may affect the cyclical changes of the reproductive system. Mabuya brachypoda males exhibit a strong seasonal cycle of spermatogenesis. Spermatogenic recrudescence was detected from November to
Journal of Morphology

1208

NDEZ-FRANYUTTI AND M.C. URIBE A. HERNA

Fig. 6. Mabuya brachypoda, seminiferous tubules during phase VI of the cycle. A, B. Late spermatids are seen at the luminal margin of the seminiferous epithelium. The seminiferous epithelium presents columns extended to the lumen. C, D. Abundant spermatozoa during spermiation are observed in the lumen of the seminiferous tubules. Late spermatids (lSt), lumen (*), spermatozoa (Sz). Bars 5 10lm.

December, when mitotic activity of spermatogonia and development of a central lumina in the seminiferous tubules were observed, coinciding with the decrease of temperature, photoperiod, and the beginning of the rainy season. From January to February, primary and secondary spermatocytes developed. From March through April, numerous
Journal of Morphology

spermatids in various stages of spermiogenesis were apparent in the apical layers of the seminiferous tubules. Spermiogenesis was completed from June through July, when the increases in temperature, photoperiod and rainy season occurred. During these months, the seminiferous tubules contained abundant spermatozoa, and the maxi-

SPERMATOGENESIS OF VIVIPAROUS LIZARD Mabuya brachypoda

1209

Fig. 7. Mabuya brachypoda, seminiferous tubules during phases VII and VIII of the cycle. A, B. During early regression, in phase VII of the cycle, the reduction of the seminiferous epithelium is evident. Spermatogonia, spermatids and spermatozoa are seen. Few spermatozoa are seen in the lumen of the seminiferous tubule. C. During middle regression, in phase VII of the cycle, the seminiferous epithelium is reduced to spermatogonia and Sertoli cells and abundant cellular debris (cd) in the lumen of the tubule are seen. D. Seminiferous tubules of Mabuya brachypoda in late regression, in phase VIII of the cycle. During complete regression, only spermatogonia and Sertoli cells are seen in the seminiferous epithelium. The lumen is very small or it is occluded. Cellular debris (cd), few spermatozoa (arrow head), lumen (*), reduction of the seminiferous epithelium (thick arrows), Sertoli cells (Se), spermatids (St), spermatogonia (Sg), spermatozoa (Sz), A: Bar 5 20lm. B: Bar 5 10lm. Bar 5 12lm. Bar 5 12lm.

mal testicular volume was detected. Testicular regression occurred from August through September when testicular volume and spermatogenic activity decreased; the seminiferous tubules contained only a few spermatocytes, the spermatids in

metamorphosis had diminished and the seminiferous epithelium decreased in thickness, having only three or four cell layers composed of spermatogonia, occasionally some spermatocytes and some remaining spermatids and spermatozoa. ProgresJournal of Morphology

1210

NDEZ-FRANYUTTI AND M.C. URIBE A. HERNA

Fig. 8. Epididymis of Mabuya brachypoda. A, B. Epididymis, testis and sexual segment of the kidney, during early spermatogenesis, in phase II of the seminiferous epithelium cycle. The ductuli efferentes of the efferent ducts have not spermatozoa. Stroma of connective tissue surrounds the efferent ducts. C. Epididymis and sexual segment of the kidney, during late spermatogenesis in phase VI of the seminiferous epithelium cycle. The ductuli efferentes have not spermatozoa, and the ductus epididymis contains abundant spermatozoa. The testis is observed near the epididymis. D. Detail of Fig. 8C. The ductuli efferentes have a cuboidal epithelium with large stereocilia in the apical end. The stroma surrounds the ductuli efferentes. E. Detail of Fig. 8C. The ductus epididymis has a large columnar epithelium and contains abundant spermatozoa. The stroma (S) surrounds the ductus epididymis. Ductuli efferentes (dep), ductus epididymis (Dep), epididymis (E), sexual segment of the kidney (SK), spermatozoa (Sz), stereocilia (arrowhead), stroma (S), testis (T). Bars 5 A: 500lm. B: 100lm. C: 500lm. D: 10lm. E: 20lm.

sively, with the advance of the regression, the spermatocytes and spermatids disappeared and the presence of hyaline material, abundant cellular debris, and some scattered spermatozoa were seen
Journal of Morphology

in the lumen of the seminiferous tubules. The regressed testes dened the total suspension of spermatogenesis. During October, the seminiferous tubules contained only spermatogonia and Sertoli

SPERMATOGENESIS OF VIVIPAROUS LIZARD Mabuya brachypoda

1211

cells, and the lumen became completely occluded. Similar reptilian spermatogenic seasonality was documented primarily, in temperate species, and it was indicated that spermiation occurred during the warmer months of the year (Licht, 1984; Aldridge et al., 1990; Gribbins et al., 2009). Meanwhile, spermatogenesis in some tropical species has been described both as a continuous (Licht, ndez-Gallegos et al., 2002; Ferreira 1984; Herna et al., 2009; Gribbins et al., 2009), or as seasonal; if seasonal it was triggered either by the rainy season or the photoperiod (Fitch, 1970; Licht, 1984; n-Santa Guillette and Casas-Andreu, 1987; Villagra rez-Bautista et al., 2006; Cruz et al., 1994; Ram Ferreira et al., 2009), a similar pattern to that described in temperate species. The seasonal morphological changes of the testis in M. brachypoda coincided with those reported for ovarian seasonality of this species (HernandezFranyutti et al., 2005). According to this report, females were in early vitellogenesis during May through July, and ovulation occurred during July through August. The concurrence of gametogenesis between males and females of M. brachypoda indicates synchrony in the reproductive cycles of both sexes. The synchrony of gametogenic cycles of males and females was seen in other species of the genus Mabuya, as M. heathi (Vitt and Blackburn, 1983), and M. capensis (Flemming, 1994). The histological analysis of testes and epididymis of M. brachypoda evidences that the males reached the maximum level of spermatogenic activity, before mating, corresponding to a prenuptial pattern, as it was described by Saint-Girons (1982) and Licht (1984). Other lizards were documented as having prenuptial spermatogenesis, including Barisia imbricata (Guillette and Casas-Andreu, 1987) and M. capensis (Flemming, 1994). Lizards, usually have a prenuptial pattern of reproduction, in contrast, multiples species of snakes as Thamnophis sirtalis parietalis (Krohmer et al., 1987), Opheodrys aestivus (Aldridge et al., 1990), and Seminatrix pygaea (Gribbins, 2010), have a postnuptial pattern, as the spermatozoa are produced after the completion of the mating season and stored in the male ducts until the following mating season. In the prenuptial pattern, both events, spermatogenesis and mating, are initiated by the same hormonal condition, called the associated pattern (Whittier and Crews, 1987). In the postnuptial pattern, spermatogenesis and mating are temporally separated, occurring at different times of the year. Separate hormonal patterns may exist, and this is known as dissociated pattern of reproduction (Whittier and Crews, 1987). The observation of the VIII phases of the annual spermatogenic cycle in the seminiferous epithelium of M. brachypoda, based in the classication used by Mayhew and Wright (1970), indicated the sea-

sonality of the testicular cycle. Spermatogenic activity in M. brachypoda, wherein all of the seminiferous tubules have similar stages of spermatogenesis, in a chronological uniformity, during the annual reproductive cycle, coincides with that of ndezseveral species of reptiles, such as lizards (Me n-Santa Cruz et al., de la Cruz et al., 1988; Villagra 1994; Ochotorena et al., 2005; Gribbins et al., 2007; Gribbins, 2010; Rheubert et al., 2009a,2009b), a turtle (Gribbins et al., 2003; Gribbins, 2010), alligators (Gribbins et al., 2006) and a snake (Gribbins et al., 2008; Gribbins, 2010). In all of these species, there is a synchronous development of a single population of germ cells during spermatogenesis followed by a single spermiation event. Gribbins (2010, 2011) compared the spermatogenesis in several reptilian sauropsids considering that the spermatogenic strategy in reptiles, releasing sperm in a single event, is reminiscent to that described in anamniotic taxa (shes and amphibians). The testis of shes (Grier and Uribe, 2009), and amphibians (Uribe et al., 1994; Ogielska and Bartmanska, 2009; Uribe, 2009), contains tubules or lobules where the germinal cells develop in a cyst or spermatocyst. The cysts are formed when spermatogonia are enclosed by Sertoli cells. Within the cysts, germ cells undergo the events of mitosis, meiosis and spermiogenesis as a single cohort. When spermiation occurs, the cysts open and release spermatozoa into the tubules or lobules, from where they travel to the deferent ducts. In contrast, in all amniotic taxa (sauropsids, birds and mammals), the testes contain seminiferous tubules that are lined by permanent, seminiferous epithelium formed by Sertoli cells and germ cells, and the germ cells are not within spermatocysts. As described in several species of reptilian sauropsids by Gribbins (2010, 2011), the testes of M. brachypoda have Sertoli cells and spermatogonia as a permanent seminiferous epithelium, throughout the year. During spermatogenesis, the germ cells commence meiosis while moving into the seminiferous epithelium from the base to the lumen and are subsequently released into the lumen at spermiation. During the reproductive season, in both birds and mammals, the seminiferous epithelium contains Sertoli cells associated with layers of germ cells. The germ cells are in different stages of spermatogenesis (Leblond and Clermont, 1952; Hess and Franca, 2008). Unlike reptilian sauropsids, which have synchronous spermatogenesis along the lengths of tubules, in birds and mammals there is a successive order of spermatogenic phases along the seminiferous epithelium which occurs in a wave of cyclic cell division and sperm development (Lin and Jones, 1990; Hess and Franca, 2008). Leblond and Clermont (1952) recognized in the rat the cellular associations that occur along the seminiferous epithelium during the spermatogenic phases. Later, the seminiferous epitheJournal of Morphology

1212

NDEZ-FRANYUTTI AND M.C. URIBE A. HERNA

lium cycle was studied in several species of mammals, such as mouse, hamster, gerbil, guinea pig, ram, monkey (Clermont, 1972), domestic cat (Franca and Godinho, 2003), grey squirrel (Tait and Johnson, 1982), and in birds as the Japanese quail (Lin and Jones, 1990; Jones and Lin, 1993). Jones and Lin (1993) concluded that in birds there is a well-dened cycle of the seminiferous epithelium, and spermatogenesis involves the activities of germ cells within and between successive generations to produce cellular associations of the seminiferous epithelium, which are essentially similar to those described in mammals. Therefore, as it was reviewed by Gribbins (2011), the functional strategy for germ cell development in reptilian sauropsids is synchronized within the seminiferous epithelium along the lengths of the tubules, and there is a single spermiation event. As a consequence, reptiles represent a dynamic of spermatogenesis between the anamniotes shes and amphibians, and the amniotes birds and mammals. Several studies described annual changes in the epididymis of reptilian sauropsids (Mayhew and Wright, 1970; Dufaure and Saint-Girons, 1984; Ravet et al., 1987; Mesure et al., 1991; Ferreira et al., 2009; Sever, 2010; Rheubert et al., 2010), coinciding with our observations in M. brachypoda. These studies correlate the cyclic changes in the testes with those seen in the epididymis and its secretory activity, in particular, in the ductus epididymis. Sever (2010) considers that, during the reproductive cycle, the functions of these ducts include reabsorption of luminal uid and active secretion. Rheubert et al. (2010) documented the presence of glycoproteins in the secretions of the ductus epididymis in the Gecko (Hemidactylus turcicus), and suggested that the function of the glycoproteins in the epididymal secretions perhaps produce an environment for sperm storage and/or maturation. Mesure et al. (1991), studying the epididymis of the lizard Lacerta vivipara, suggested that these secretions are discharged into the lumen of the ductus epididymis where they bind to the heads of the spermatozoa. Ferreira et al. (2009) and Sever (2010) consider the importance of analyzing the morphological changes of the epididymis and its secretions in the interpretation of its role in the maturation of the spermatozoa. The sexual segment of the kidney is a distal portion of the nephron of squamates, and is observed near the testis and epididymis. The sexual segment of the kidney is believed to provide seminal uid that mixes in the ureter with sperm (Sever et al., 2002; Sever and Hopkins, 2005). Thus, spermatogenesis, including the cell types and the dynamics of the seminiferous epithelium cycle and features of the duct system in reptiles, provide essential aspects for the understanding of the evolution of male reproduction in vertebrates.
Journal of Morphology

ACKNOWLEDGMENTS The authors thank Marcela E. Aguilar-Morales n for help in histological a-Alarco and Adriana Garc processing, Ana Isabel Bieler Antolin and Gabino De la Rosa Cruz for the kind technical assistance Antonio Herwith digital photography; and Jose ndez Go mez for the excellent digital preparation na of histological gures and Edgar Abraham Lozano ndez Vidal for the asMendoza and Ulises HernA sistance in the preparation of the Supporting Information Graph. LITERATURE CITED
o-Bello B, Salinas-Rosales P. 1996. Aguilar-Morales M, Coutin cnicas histolo gicas y citoqu micas. Las Manual general de te Prensas de Ciencias Facultad de Ciencias, Universidad noma de Me xico, Me xico, D.F. Me xico. p 106. Nacional Auto Aldridge RD, Greenhow J, Plummer MV. 1990. The male reproductive cycle of the rough green snake (Opheodrys aestivus). Amphibia-Reptilia 11:165172. Bakst MR, Akuffo V, Trel P, Brillard JP. 2007. Morphological and histochemical characterization of the seminiferous epithelial and Leydig cells of the turkey. Anim Reprod Sci 97:303 313. Blackburn DG. 2000. Classication of the reproductive patterns of amniotes. Herpetol Monogr 14:371377. Blackburn DG. 2005. Amniote Perspectives on the Evolution of Viviparity. In: Uribe MC and Grier HJ, editors. Viviparous Fishes. New Life Publications, Homestead, FL. pp 319412. Clermont Y. 1972. Kinetics of spermatogenesis in mammals: Seminiferous epithelium cycle and spermatogonial renewal. Physiol Rev 52:198236. n Nacional del Agua. Gobierno de Me xico. 2004. Me xico, Comisio D.F. Dufaure JP, Saint-Girons H. 1984. Histologie compare de lepididyme et de ses secretions chez les reptiles (lezards et serpents). Arch Anat Microsc 73:1526. Ferreira A, Silva DN, Van Sluys M, Dolder H. 2009. Seasonal changes in testicular and epididymal histology of the tropical lizard, Tropidurus itambere (Rodrigues, 1987), during its reproductive cycle. Braz J Biol 69:429435. Fitch HS. 1970. Reproductive cycles of lizards and snakes. Univ Kans Mus Nat Hist. Misc Publ 52:1247. Flemming AF. 1994. Male and female reproductive cycles of the viviparous lizard, Mabuya capensis (Sauria: Scincidae) from South Africa. J Herpetol 28:334341. Franca LR, Godinho CL. 2003. Testis morphometry, seminiferous epithelium cycle length, and daily sperm production in domestic cats (Felis catus). Biol Reprod 68:15541561. Gribbins KM. 2010. Temperate Reptilian Spermatogenesis: A nNew Amniotic Mode of Germ Cell Development. In: Herna ndez de la Cruz F, Me ndez Sa nchez JF, dez Gallegos O, Me n en Reptiles: Morfolog a, Ecolog a y Evoeditors. Reproduccio n, pp 137167. lucio Gribbins KM. 2011. Reptilian Spermatogenesis. A Histological and Ultrastructural Perspective. In: Cheng Y, editor. Spermatogenesis 1:3 Landes Bioscience, Austin, TX, pp 250269. Gribbins KM, Gist DH, Congdon J. 2003. Cytological evaluation of spermatogenesis in the Redeared Slider, Trachemys scripta. J Morphol 255:337346. Gribbins KM, Elsey RM, Gist DH. 2006. Cytological evaluation of the germ cell development strategy within the testis of the American Alligator,Alligator mississippiensis. Acta ZoolStockholm 87:5969. Gribbins KM, Mills EM, Sever DM. 2007. Ultrastructural examination of spermiogenesis within the testis of the ground skink, Scincella laterale (Squamata, Sauria, Scincidae). J Morphol 268:181192.

SPERMATOGENESIS OF VIVIPAROUS LIZARD Mabuya brachypoda

Gribbins KM, Rheubert JL, Siegel DS, Collier MH, Sever DM. 2008. Histological analysis of spermatogenesis and the germ cell development strategy within the testis of the male Western Cottonmouth Snake, Agkistrodon piscivorus leucostoma. Ann Anat 190:461476. Gribbins KM, Rheubert JL, Poldemann EH, Collier MH, Wilson B, Wolf K. 2009. Continuous spermatogenesis and the germ cell development strategy within the testis of the Jamaican Gray Anole,Anolis lineatopus. Theriogenology 72:484492. Grier HJ, Uribe MC. 2009. Male Reproductive System: Testis, Spermatogenesis and Testicular cycles. In: Jamieson B., editors. Reproductive Biology and Phylogeny of Fish (Agnatha and Bony Fishes). Science Publishers, Inc Eneld Chapter 4, pp 119142. Guillette LJ, Casas-Andreu G. 1987. The reproductive biology of the high elevation Mexican lizard Barisia imbricata. Herpetologica 43:2938. ndez de la Cruz FR. 1993. The reproductive Guillette LJ, Me cycle of the viviparous Mexican lizard Sceloporus torquatus. J Herpetol 27:168174. ndez-Frayutti A, Uribe MC, Guillete LJ. 2005. Oogenesis Herna in the viviparous matrotrophic lizard Mabuya brachypoda. J Morphol 265:152164. nHernandez-Gallegos H, Mendez-De La Cruz RF, Villagra Santa Cruz M, Andrews RM. 2002 Continuous spermatogenesis in the lizard Sceloporus bicanthalis (Sauria: Phrynosomatidae) from high elevation habitat of central Mexico. Herpetologica 58:41521. Hess RA, de Franca LR. 2008. Spermatogenesis and Cycle of the Seminiferous Epithelium. In: Yan Cheng C. editor. Molecular Mechanisms in Spermatogenesis. Landes Bioscience and Springer Business Media, Rio Grande, St. Austin, TX. USA. pp 115. Jones RC, Lin M. 1993. Spermatogenesis in birds. Oxford Rev Reprod B 15:233264. Krohmer RW, Grassman M, Crews D. 1987. Annual reproductive cycle in the male red-sided garter snake, Thamnophis sirtalis parietalis: Field and laboratory studies. Gen Comp Endocr 68:6475. Leblond CP, Clermont Y. 1952. Denitions of the stages of the cycle of the seminiferous epithelium in the rat. Ann NY Acad Sci 55:548573. Licht P. 1984. Reptiles. In: Lamming GE, editor. Marshalls Phyisiology of Reproduction. Reproductive Cycles of Vertebrates. Churchill Livingstone, Edinburgh, pp 206282. Lin M, Jones RC. 1990. Spatial arrangement of the stages of the cycle of the seminiferous epithelium in the Japanese quail, Coturnix coturnix japonica. J Reprod Fertil 90:361 367. Mayhew WW, Wright SJ. 1970. Seasonal changes in testicular histology of three species of the lizard Genus Uma. J Morphol 130:163186. ndez de la Cruz FR, Guillette LJ, Villagra n-Santa Cruz M, Me Casas-Andreu G. 1988. Reproductive and fat body cycles of the viviparous lizard Sceloporus mucronatus. J Herpetol 22:112. Mesure M, Chevalier M, Depeiges A, Faure J, Dufaure JP. 1991. Structure and ultrastructure of the epididymis of the viviparous lizard during the annual hormonal cycle: Changes of the epithelium related to secretory activity. J Morphol 210:133145. Ochotorena Sanz A, Uribe MC, Guillette LJ. 2005. Seasonal gametogenic cycles in a Cuban tropical lizard, Anolis porcatus. J Herpetol 39:443454. Ogielska M, Bartmanska J. 2009. Spermatogenesis and Male Reproductive System. Anura. In: Ogielska M, editor. Reproduction of Amphibians. Science Publishers, Inc. Eneld, Chapter 2, pp 3499. rez-Bautista A, Luja VH, Balderas-Valdivia C, Ort z-Pulido Ram R. 2006. Reproductive cycle of male and female spiny lizards,

1213

Sceloporus melanorhinus, in a tropical dry forest. Southwest Nat 51:157162. Ramirez-Pinilla MP, Serrano VH, Galeano JC. 2002. Annual reproductive activity of Mabuya mabouya (Squamata, Scincidae). J Herpetol 36:667677. Ravet V, Courty Y, Depeiges A, Dufaure JP. 1987. Changes in epididymal protein synthesis during the sexual cycle of the lizard, Lacerta vivipara. Biol Reprod 37:901907. Rheubert JL, McHugh HH, Collier MH, Sever DM, Gribbins KM. 2009a. Temporal germ cell development strategy during spermatogenesis within the testis of the Ground Skink, Scincella lateralis (Sauria: Scincidae). Theriogenology 72:5461. Rheubert JL, Poldemann EH, Eckstut ME, Collier MH, Sever DM, Gribbins KM. 2009b. Temporal germ cell development strategy during mixed spermatogenesis within the male Mediterranean Gecko, Hemidactylus turcicus (Reptilia: Gekkonidae). Copeia 4:791798. Rheubert JL, Sever DM, Geheber AD, Siegel DS. 2010. Proximal testicular ducts of the Mediterranean Gecko (Hemidactylus turcicus). Anat Rec 293:21762192. Saint-Girons H. 1982. Reproductive cycles of male snakes and their relationships with climate and female reproductive cycles. Herpetologica 38:516. Sever DM. 2010. Ultrastructure of the reproductive system of the Black Swamp snake (Seminatrix pygaea). VI. Anterior testicular ducts and their nomenclature. J Morphol 271:104 115. Sever DM, Stevens R, Ryan TJ Hamlett WC. 2002. Ultrastructure of the reproductive system of the black swamp snake (Seminatrix pygaea). III. Sexual segment of the male kidney. J Morphol 252:238254. Sever DM, Hopkins WA. 2005. Renal sexual segment of the Ground Skink, Scincella laterale (Reptilia, Squamata, Scincidae). J Morphol 266:4659. Simbotwe MP. 1980. Reproductive biology of the skinks Mabuya striata and Mabuya quinquetaeniata in Zambia. Herpetologica 36:99104. Somma CA, Brooks Gr. 1976. Reproduction in Anolis oculatus, Ameiva fuscata and Mabuya mabouya from Dominica. Copeia 2:249256. Tait AJ, Johnson E. 1982. Spermatogenesis in the grey squirrel (Sciurus carolinensis) and changes during sexual regression. J Reprod Fert 65:5358. Trauth SE. 1979. The testicular cycle and timing of reproduction in the Collared Lizard (Crotaphytus collaris) in Arkansas. Herpetologica 35:184192. Uribe MC. 2009. Spermatogenesis and Male Reproductive System. Urodela. In: Ogielska M, editor. Reproduction of Amphibians. Science Publishers, Inc Eneld, Chapter 3, p 100124. mez R os G, Brandon RA. 1994. Spermatogenesis Uribe MC, Go in the Urodele Ambystoma dumerilii. J Morphol 222:287299. ndez-Frayutti A, Guillete LJ Jr. 2006. InterUribe MC, Herna embryonic regions of the viviparous matrotrophic lizard Mabuya brachypoda. J Morphol 267:404414. n-Santa Cruz M, Me ndez-de la Cruz FR, Parra-Ga mez Villagra nico del lacertilio Sceloporus L. 1994. Ciclo espermatoge mucronatus (Reptilia: Phrynosomatidae). Rev Biol Trop 42:289296. Vitt LJ, Blackburn DG. 1983. Reproduction in the lizard Mabuya heathi (Scincidae): A commentary on viviparity in new world Mabuya. Can J Zool 61:27982806. Vitt LJ, Blackburn DG. 1991. Ecology and life history of the viviparous lizard Mabuya bistriata (Scincidae) in the Brazilian Amazon. Copeia 1991:916927. Webb RG. 1958. The status of the Mexican lizards of the genus Mabuya. Sci Bull Univ Kansas 17:13031313. Whittier JM, Crews D. 1987. Seasonal Reproduction: Pattern and Control. In: Norris DO, Jones RE, editors. Hormones and Reproduction in Fishes, Amphibians and Reptiles. Plenum Press, New York, pp 385409.

Journal of Morphology