Ultrastructure and histochemistry of the male reproductive system of the genus Callinectes Stimpson, 1860 (Brachyura: Portunidae).

We described the ultrastructure and histochemistry of the reproductive system of five Callinectes species, and evaluate the seasonal variation in weight of the reproductive system and hepatopancreas by comparing annual changes of somatic indices. The somatic indices changed little throughout the year. In Callinectes, spermatogenesis occurs inside the lobular testes and, within each lobule, the cells are at the same developmental stage. Spermatogenesis and spermiogenesis follow the same development pattern in all Callinectes studied. Mature spermatozoa are released into the seminiferous ducts through the collecting ducts. Cells of the vas deferens are secretory as evidenced by rough endoplasmic reticulum, Golgi complex, and secretory vesicles that produce the seminal fluid. The anterior vas deferens shows two portions: proximal and distal. In proximal portion (AVDp), spermatozoa are clustered and embedded in an electron-dense, basophilic glycoproteinaceous secretion Type I. In the distal portion (AVDd), the spermatophore wall is formed by incorporation of a less electron-dense glycoproteinaceous secretion Type II. The secretion Type I change to an acid polysaccharide-rich matrix that separates the spermatophores from each other. The median vas deferens (MVD) stores the spermatophores and produces the granular glycoproteinaceous seminal fluid. The posterior vas deferens (PVD) has few spermatophores. Its epithelium has many mitochondria and the PVD seminal fluid changes into a liquid and homogeneous glycoprotein. Many outpocketings in the PVD and MVD help to increase the fluid production. Overall, the reproductive pattern of Callinectes is similar to other species that produce sperm plugs. The secretions of AVD, MVD, and PVD are responsible for the polymerization that forms the solid, waxy plug in the seminal receptacle. The traits identified here are common to all Portunidae species studied so far.

Ultrastructure of spermatozoa of spider crabs, family Mithracidae (Crustacea, Decapoda, Brachyura): Integrative analyses based on morphological and molecular data.

Recent studies based on morphological and molecular data provide a new perspective concerning taxonomic aspects of the brachyuran family Mithracidae. These studies proposed a series of nominal changes and indicated that the family is actually represented by a different number and representatives of genera than previously thought. Here, we provide a comparative description of the ultrastructure of spermatozoa and spermatophores of some species of Mithracidae in a phylogenetic context. The ultrastructure of the spermatozoa and spermatophore was observed by scanning and transmission electron microscopy. The most informative morphological characters analysed were thickness of the operculum, shape of the perforatorial chamber and shape and thickness of the inner acrosomal zone. As a framework, we used a topology based on a phylogenetic analysis using mitochondrial data obtained here and from previous studies. Our results indicate that closely related species share a series of morphological characteristics of the spermatozoa. A thick operculum, for example, is a feature observed in species of the genera Amphithrax, Teleophrys, and Omalacantha in contrast to the slender operculum observed in Mithraculus and Mithrax. Amphithrax and Teleophrys have a rhomboid perforatorial chamber, while Mithraculus, Mithrax, and Omalacantha show a wider, deltoid morphology. Furthermore, our results are in agreement with recently proposed taxonomic changes including the separation of the genera Mithrax (previously Damithrax), Amphithrax (previously Mithrax) and Mithraculus, and the synonymy of Mithrax caribbaeus with Mithrax hispidus. Overall, the spermiotaxonomy of these species of Mithracidae represent a novel set of data that corroborates the most recent taxonomic revision of the family and can be used in future taxonomic and phylogenetic studies within this family.

Prevalence, Abundance, and Intensity of Implanted Spermatophores in the Leech Haementeria officinalis (Glossiphoniidae: Hirudinida) from Guanajuato, Mexico.

Fertilization through hypodermic implantation of spermatophores has been recorded in at least 4 groups of leeches: Glossiphoniidae, Piscicolidae, Ozobranchidae, and Erpobdelliformes. In Piscicola respirans (Piscicolidae), vector tissue responsible for sperm transfer from a specialized region of the body to the ovaries has led to the non-random attachment of spermatophores on the body surface of the recipient leech. It has been suggested that in glossiphoniid leeches, spermatophores are implanted in any part of the body surface of the recipient leech without a clear pattern or preference for region. In order to determine if the donor leech implants its spermatophores in a specific area of the conspecific recipient's body, we surveyed 81 specimens of Haementeria officinalis (Clitellata: Glossiphoniidae) from a wild population in Guanajuato, Mexico, and recorded the distribution of the spermatophores over the recipient's body surface. We describe for the first time a spermatophore of H. officinalis using scanning electron and light microscopy. Spermatophores were found attached dorsally between somites XVII and XXI 59.57% of the time, and the rest were found in other parts of the body, including on the ventral surface. The non-specific attachment for spermatophores does not support the presence of specialized tissue responsible for sperm transfer and instead attributes the placement of implantation to mechanical characteristics of the copulation process.

Ultrastructure of spermatogenesis and spermatozoa in agoutis during sexual development.

The aim was to study the ultrastructure of testicular parenchyma and define the morphological ultrastructure of spermatozoa of agoutis kept in captivity. Segments of testes from eight agouti males at prepubescence, prepuberty, pubescence and sexual maturity were fixed in glutaraldehyde. Laboratory procedures were performed for transmission electron microscopy. Spermatogonial cells of Type A - pale, Type A - dark, intermediate and Type B were found. Spermatocytes in the pachytene phase were abundant among primary spermatocytes. From the prepubertal phase, Sertoli cells exhibited invaginations in the nuclear membrane and lipid inclusions in the cytoplasm due to their phagocytic function. Leydig cells displayed higher metabolic activity during puberty as evidenced by the presence of lipid droplets. Spermatozoa were fully formed morphologically at prepuberty. The centriolar complex had partially degenerated and featured a centriolar space as in rodents. Sperm heads were tapered, without prominence of the acrosome or evidence of the perforatorium, differing from cavies, rats and mice. This is the first study to describe the ultrastructure of agouti spermatozoa. This research may assist as a basis for future work related to fertility and other biotechnologies applied to reproductive biology in agoutis.

Vitamin B12-induced spermatogenesis recovery in cimetidine-treated rats: effect on the spermatogonia number and sperm concentration.

The H2-receptor antagonist cimetidine is an antiulcer drug also used for the treatment of cancer due to its antiangiogenic effect. However, this drug has caused structural changes in the seminiferous tubules. Vitamin B12 has been used as a therapeutic agent for the treatment of male infertility. The supplementation of rats with vitamin B12 during cimetidine treatment has recovered the damaged seminiferous tubules, but how this vitamin restores the seminiferous epithelium has not been clarified. In this study, we evaluated whether vitamin B12 improves the number of spermatogonia, spermatocytes, and sperm concentration in cimetidine-treated rats. Adult male rats were treated for 50 days as follows: cimetidine group received 100 mg kg-1 b.w. of cimetidine, cimetidine-B12 group received cimetidine and 3 µg of vitamin B12-hydroxocobalamin, B12 group received only 3 µg of vitamin, and control group received saline. Sperm concentration was calculated and historesin-embedded testes sections were used for the quantitative analyses of spermatogonia (A; In/B) and spermatocytes. TUNEL method and PCNA immunofluorescence were performed. Cimetidine caused a significant reduction in sperm concentration. TUNEL-positive spermatogonia and spermatocytes were correlated to a significant reduction in the number of these cells. In cimetidine-B12 group, sperm concentration was higher than cimetidine group and a significant increase in the number of spermatogonia (stages II-VI) was correlated to a high incidence of PCNA-immunolabeled spermatogonia and spermatocytes. The results show that the supplementation of rats with vitamin B12 during cimetidine treatment increases sperm concentration and exerts a potential effect in the recovery of spermatogonia and spermatocytes.

Ultrastructure of spermatogenesis in Spix's yellow-toothed cavy (Galea spixii).

This was a pioneer study of the spermatogenic process from the onset of puberty in Spix's yellow-toothed cavies (SYC, Galea spixii) bred in captivity. The study aimed to characterize fine structure of spermatogenesis. Twelve testes from pubertal and post-pubertal SYC males were studied using transmission electron microscopy. Spermatogenesis can be divided into three phases: proliferation, meiosis, and spermiogenesis. In proliferation phase, three types of spermatogonia were identified and characterized as A(dark), A(pale), and B. In the second phase, spermatocytes (2n) undergo meiotic divisions that generate spermatids (n); the process begins in spermatocytes in the preleptotene stage when they increase their nuclear size, differentiating into spermatocytes in the leptotene stage when cell division is initiated. In addition, we found chromatin condensation, and formation of a structure composed of proteins that formed a central shaft and two lateral bars associated with pairing of homologous chromosomes. During spermiogenesis, the following main events occurred: condensation of nuclear chromatin, formation of acrosome with perfuratorium, elimination of residual cytoplasm, and development of the flagellum. The sperm head is different from that of other rodents. The endoplasmic reticulum and the Golgi complex are the two main organelles demonstrated during this process. These organelles collaborate through synthesis of proteins and hormones for the development of germ cells during spermatogenesis in SYC.

Spermatogenesis is seasonal in the large hairy armadillo, Chaetophractus villosus (Dasypodidae, Xenarthra, Mammalia).

Very little is known about the distinct reproductive biology of armadillos. Very few studies have investigated armadillo spermatogenesis, with data available only for Euphractus sexcinctus and Dasypus novemcinctus. In the present study, we analysed male germ cell differentiation in the large hairy armadillo Chaetophractus villosus throughout the year, describing a cycle of the seminiferous epithelium made of eight different stages. Evaluation of the testis/body mass ratio, analysis of the architecture of the seminiferous epithelium and the frequency of defective seminiferous tubules allowed identification of a temporal interruption of spermatogenesis during the period between mid-May to July (mid-end autumn) in correlation with very low testosterone levels. Overall, these results suggest that spermatogenesis is seasonal in C. villosus.

The effect of oviductal deleted in malignant brain tumor 1 over porcine sperm is mediated by a signal transduction pathway that involves pro-AKAP4 phosphorylation.

The interaction between sperm and oviduct results in the selection of sperm with certain qualities. Porcine oviductal deleted in malignant brain tumor 1, DMBT1 (previously called sperm-binding glycoprotein, SBG), has been proposed to be implicated in sperm selection through acrosome alteration and suppression of motility of a subpopulation of sperm that have begun capacitation prematurely. It produces in vitro acrosome alteration and decrease of motility of boar sperm, concomitant with tyrosine phosphorylation of a 97 kDa sperm protein (p97). We hypothesized that the phosphorylation of p97 may be a link between DMBT1 sensing by a subpopulation of boar sperm and its biological effect. In this work, p97 was identified by mass spectrometry and immunoprecipitation as a porcine homologue of AKAP4. Pro-AKAP4 was localized by immunofluorescence and subcellular fractionation to the periacrosomal membranes and was shown to be tyrosine phosphorylated by DMBT1 regardless of the presence of calcium or bicarbonate, and of cAMP analogs, protein kinase A inhibitors, or a protein kinase C inductor. A processed ∼80 kDa form of AKAP4 was also detected at the tail of boar sperm, which was not tyrosine phosphorylated by DMBT1 under the conditions tested. Immunohistochemistry of testis showed presence of AKAP4 in boar sperm precursor cells. The evidence presented here supports the involvement of AKAP4 in the formation of the fibrous sheath on boar precursor sperm cells and implicates the phosphorylation of pro-AKAP4 as an early step in the signal transduction pathway gated by DMBT1 that leads to sperm selection through acrosome alteration.

Histology and morphometry of the testes of adult domestic cats (Felis catus).

Testicles of 30 mongrel cats were analyzed histologically and morphometrically, divided into three groups: G1 (1-2 years old), G2 (over 2 and up to 4 years old) and G3 (over 4 and up to 6 years old). After orchiectomy and histopathology, the morphometric parameters studied were: thickness of the tunica albuginea (72 µm) and seminiferous epithelium (77.19 µm), perimeter (53.81; 90.57 µm), (54.80; 101.07 µm); area (174.23; 494.55 µm(2)), (176.68; 629.70 µm(2)); maximum diameter (14.94; 28.02 µm), (14.76; 31.66 µm); minimum diameter (13.25; 21.92 µm), (13.30; 24.52 µm); and shape factor (index for regularity of the format) (1.36; 1.36), (1.39; 1.35) of the nucleus and cytoplasm of spermatogonia and Leydig cells, respectively. The results can be used for comparative studies and contribute knowledge concerning the height of the seminiferous epithelium, thickness of the tunica albuginea and size of spermatogonia and Leydig cells.

Spermatophoric reaction reappraised: novel insights into the functioning of the loliginid spermatophore based on Doryteuthis plei (Mollusca: Cephalopoda).

During copulation, spermatophores produced by male coleoid cephalopods undergo the spermatophoric reaction, a complex process of evagination that culminates in the attachment of the spermatangium (everted spermatophore containing the sperm mass) on the female's body. To better understand this complicated phenomenon, the present study investigated the functional morphology of the spermatophore of the squid Doryteuthis plei applying in vitro analysis of the reaction, as well as light and electron microscopy investigation of spermatangia obtained either in vitro, or naturally attached on females. Hitherto unnoticed functional features of the loliginid spermatophore require a reappraisal of some important processes involved in the spermatophoric reaction. The most striking findings concern the attachment mechanism, which is not carried out solely by cement adhesive material, as previously believed, but rather by an autonomous, complex process performed by multiple structures during the spermatophoric reaction. During evagination, the ejaculatory apparatus provides anchorage on the targeted tissue, presumably due to the minute stellate particles present in the exposed spiral filament. Consequently, the ejaculatory apparatus maintains the attachment of the tip of the evaginating spermatophore until the cement body is extruded. Subsequently, the cement body passes through a complex structural rearrangement, which leads to the injection of both its viscid contents and pointed oral region onto the targeted tissue. The inner membrane at the oral region of the cement body contains numerous stellate particles attached at its inner side; eversion of this membrane exposes these sharp structures, which presumably adhere to the tissue and augment attachment. Several naturally attached spermatangia were found with their bases implanted at the deposition sites, and the possible mechanisms of perforation are discussed based on present evidence. The function of the complex squid spermatophore and its spermatophoric reaction is revisited in light of these findings. J. Morphol. 2012. © 2011 Wiley Periodicals, Inc.

Stages and duration of the cycle of the seminiferous epithelium in oncilla (Leopardus tigrinus, Schreber, 1775).

Six adult Leopardus tigrinus (oncilla) were studied to characterize stages of the seminiferous epithelium cycle and its relative frequency and duration, as well as morphometric parameters of the testes. Testicular fragments were obtained (incisional biopsy), embedded (glycol methacrylate), and histologic sections examined with light microscopy. The cycle of the seminiferous epithelium was categorized into eight stages (based on the tubular morphology method). The duration of one seminiferous epithelium cycle was 9.19 d, and approximately 41.37 d were required for development of sperm from spermatogonia. On average, diameter of the seminiferous tubules was 228.29 µm, epithelium height was 78.86 µm, and there were 16.99 m of testicular tubules per gram of testis. Body weight averaged 2.589 kg, of which 0.06 and 0.04% were attributed to the testis and seminiferous tubules, respectively. In conclusion, there were eight distinct stages in the seminiferous epithelium, the length of the seminiferous epithelium cycle was close to that in domestic cats and cougars, and testicular and somatic indexes were similar to those of other carnivores of similar size.

Mice spermatogonial stem cells transplantation induces macrophage migration into the seminiferous epithelium and lipid body formation: high-resolution light microscopy and ultrastructural studies.

Transplantation of spermatogonial stem cells (SSCs), the male germline stem cells, in experimental animal models has been successfully used to study mechanisms involved in SSC self-renewal and to restore fertility. However, there are still many challenges associated with understanding the recipient immune response for SSCs use in clinical therapies. Here, we have undertaken a detailed structural study of macrophages elicited by SSCs transplantation in mice using both high-resolution light microscopy (HRLM) and transmission electron microscopy (TEM). We demonstrate that SSCs transplantation elicits a rapid and potent recruitment of macrophages into the seminiferous epithelium (SE). Infiltrating macrophages were derived from differentiation of peritubular monocyte-like cells into typical activated macrophages, which actively migrate through the SE, accumulate in the tubule lumen, and direct phagocytosis of differentiating germ cells and spermatozoa. Quantitative TEM analyses revealed increased formation of lipid bodies (LBs), organelles recognized as intracellular platforms for synthesis of inflammatory mediators and key markers of macrophage activation, within both infiltrating macrophages and Sertoli cells. LBs significantly increased in number and size in parallel to the augmented macrophage migration during different times post-transplantation. Our findings suggest that LBs may be involved with immunomodulatory mechanisms regulating the seminiferous tubule niche after SSC transplantation.

Male reproductive apparatus and spermatophore morphology of the hermit crabs Pagurus brevidactylus and P. criniticornis (Anomura, Paguridae).

Spermatozoa of most crustacean species are nonmotile and are packed into spermatophores. In Decapoda, spermatophores are highly variable in morphology and can be useful in the solving of taxonomic and systematic questions, especially among the Anomura. In this study, the morphology and morphometry of the spermatophores of the western Atlantic hermit crabs Pagurus brevidactylus and P. criniticornis are described. The abdomen of fresh male specimens was dissected to expose the reproductive system and to extract the spermatophores, which were analyzed by stereoscopic, light, and scanning electron microscopy. The vas deferens can be divided macroscopically in three regions, all of them containing spermatophores. Tripartite spermatophores are composed of an elongated cylindrical main ampulla, a triangular accessory ampulla, a narrow cylindrical peduncle, and a round pedestal. Dimensions of the spermatophore components are positively correlated to the size of the crab. Morphological patterns observed in this study resemble those of other pagurid hermit crabs investigated to date. The morphological character distribution confirms classifications based on adult morphology and molecular analysis.

Ultrastructure of spermatogenesis in the white-lined broad-nosed bat, Platyrrhinus lineatus (Chiroptera: Phyllostomidae).

Spermatogenesis, the remarkable process of morphological and biochemical transformation and cell division of diploid stem cells into haploid elongated spermatozoa, is one of the most complex cell differentiations found in animals. This differentiation process has attracted extensive studies, not only because the process involves many radical changes in the cell shape and biochemistry, but also because the phases and steps of differentiation have provided a better basis for analyzing the seminiferous epithelium cycle. Thus, this study aimed to characterize ultrastructurally the spermatogenesis process in the bat Platyrrhinus lineatus in order to provide a basis for determining the stages of spermatogenesis and to facilitate comparisons of the process between bat species and other vertebrates. Based on ultrastructural characteristics three main types of spermatogonia could be accurately identified: A(d), A(p) and B; the differentiation of spermatids was clearly divided into 12 steps (steps 1-3: Golgi phase, steps 4-5: cap phase, steps 6-9: acrosomal phase and steps 10-12: maturation phase). The ultrastructure of spermatozoa, Leydig cells and Sertoli cells was characterized; and some processes including nucleolar disorganization and the formation of synaptonemal complexes, acrosome and chromatoid body were discussed. Based on our results we may conclude that the spermatogenic process of P. lineatus follows the pattern of mammals with some specificity, as the process of formation of the acrosome and the presence of the perfuratorium. By other side, the simpler ultrastructure of its spermatozoon shows a pattern more closely related to the sperm cells of humans and other primates.

Spermatogonial morphology and kinetics during testis development in mice: a high-resolution light microscopy approach.

Despite the knowledge of spermatogonial biology in adult mice, spermatogonial development in immature animals has not been fully characterized. Thus, the aim of this study was to evaluate the ontogeny of the morphological development of the spermatogonial lineage in C57BL/6 mouse testis, using high-resolution light microscopy. Spermatogonial morphology, chronology, and absolute number were determined for different ages postpartum (pp). The morphology of spermatogonia in immature mice was similar to that of adult spermatogonia, although their nuclear diameter was slightly smaller. The A(1) spermatogonia were first observed on day 2 pp, and only 24 h later, differentiating type A(3) and A(4) spermatogonia were observed in the seminiferous cords. This result indicated a shortening of the spermatogonial phase for immature mice of about ∼2.5 days when compared with adult mice and suggests that gonocytes and/or A(1) spermatogonia could directly become A(4) spermatogonia, skipping the developmental sequence of type A spermatogonia. These A(4) spermatogonia are functional as they develop into type B spermatogonia by day 5 pp. At day 8 pp, while differentiation to spermatocytes begins, the A(und) spermatogonia reach their maximal numbers, which are maintained through adulthood. The various details of the spermatogonial behavior in immature normal mice described in this study can be used as a baseline for further studies under experimental or pathological conditions.

Nucleolar cycle and its correlation with chromatoid bodies in the Tilapia rendalli (Teleostei, Cichlidae) spermatogenesis.

The aim of this study was: (1) to monitor the nucleolar material distribution using cytological and cytochemical techniques and ultrastructural analysis; and (2) to compare the nucleolar material distribution with the formation of the chromatoid body (CB) in the germ epithelium of Tilapia rendalli. Nucleolar fragmentation occurred during the leptotene of prophase I and nucleolus reorganization occurred in the early spermatid nucleus. The area of the early spermatid nucleolus was significantly smaller than that of the spermatogonia nucleolus. Ultrastructural analysis showed an accumulation of nuages, which form the CB, before nucleolar fragmentation in the spermatogonia cytoplasm. The CB was observed in association with mitochondrial clusters in the cytoplasm of primary spermatocytes, as well as in those of initial and later spermatids. In conclusion, the nucleolus seems to be related to CB formation during spermatogenesis of T. rendalli, because at the moment of nucleolus fragmentation in the primary spermatocytes, the CB reaches its largest area and it is able to complete important functions during spermatogenesis. The reorganized nucleolus of the initial spermatids has a lower area due several factors, one of which is the probable migration of nucleolar fragments from the nucleus to the cytoplasm, therefore playing a role in CB formation.

Neonatal gonocyte differentiation in Mongolian gerbil Meriones unguiculatus involves asynchronous maturation of seminiferous cords and rapid formation of transitional cell stage.

This study describes the neonatal differentiation of the Mongolian gerbil gonocytes, focusing on the relationship between its relocation to the basement membrane, apoptosis and postrelocation changes and also the distribution of androgen receptors (AR). Testes of gerbils from 1 to 35 days of age (d) were examined by high resolution light microscopy and immunocytochemistry for proteins PCNA, VASA, and AR as well as by the TUNEL method. Gonocytes were quantified according to degree of relocation into nonrelocated, relocating and relocated. Most of them were found in the center of seminiferous cords at 1 d but a small number of relocating and relocated gonocytes were already visible in the first postnatal day. After relocation, gonocytes change phenotypically to a transitional stage designated herein prospermatogonia. Both gonocyte relocation and transformation into spermatogonial lineage occur asynchronously in the seminiferous cords, mainly after 7 d. Gonocyte proliferation began before but peak after their relocation to basement membrane at the prospermatogonia stage. Higher levels of gonocyte apoptosis were found at 7 d and 21 d. From this time onward gonocytes were not found. Gonocytes and prospermatogonia showed high amounts of AR in their cytoplasm contrary to spermatogonial subtypes, indicating a possible AR inactivation in these cells. In conclusion, the process of gonocyte relocation in the gerbil extends until the second postnatal week, leads to their rapid differentiation into prospermatogonia and occurs simultaneously with the loss of androgen sensitivity. Differently from other laboratory rodents, the events regarding gonocyte maturation in the gerbil last longer and occur asynchronously in seminiferous cords.

Histological and stereological evaluation of zebrafish (Danio rerio) spermatogenesis with an emphasis on spermatogonial generations.

The zebrafish has become an important vertebrate model for basic and biomedical research, including the research field of the biology of reproduction. However, very few morphological and stereological data are available regarding zebrafish testis structure and spermatogenesis. In this careful histomorphometric evaluation of the testis, we studied spermatogonial cells using molecular markers, determined the combined duration of meiotic and spermiogenic phases, and examined the formation of the Sertoli cell barrier (tight junctions). We found at least nine spermatogonial generations and propose a morphology-based nomenclature for spermatogonial generations that is compatible with the one used in higher vertebrates. The number of germ cells per cyst increased dramatically (1 to approximately 1360 cells) from undifferentiated spermatogonia type A to early spermatids. The combined duration of meiotic and spermiogenic phases is approximately 6 days, one of the shorter periods among the teleost fish investigated to date. The number of Sertoli cells per cyst increased 9-fold during the maturational cycle of spermatogenic cysts and stabilized in the meiotic phase at a ratio of approximately 100 early spermatids per Sertoli cell (Sertoli cell efficiency). Similarly to mammals, Sertoli cell proliferation ceased in the meiotic phase, coinciding with the formation of tight junctions between Sertoli cells. Hence, the events taking place during puberty in the germinal epithelium of mammals seem to recapitulate the "life history" of each individual spermatogenic cyst in zebrafish.

High-resolution light microscopic characterization of spermatogonia.

It is possible to distinguish the morphological features of the spermatogonial nuclei and nucleoli and to further identify their distinct generations using an appropriate method to fix whole testes via vascular perfusion with glutaraldehyde, postfixation by immersion in reduced osmium, embedding in araldite, and staining of semithin tissue sections. A well-trained individual can distinguish each of the spermatogonial types in rodents (A(undiferentiated), A(1), A(2), A(3), A(4), In, and B) using this tissue preparation technique based on their morphological details and without correlation with the stages of the epithelium cycle or other parameters. The possibility of distinguishing each spermatogonial type by their morphological characteristics allows a more accurate evaluation of their kinetics during the spermatogenic cycle. Moreover, the understanding of spermatogonial behavior is a means to elucidate the functional control of the spermatogenesis, which consequently allows the determination of their effects on the fertility of humans and other animals.

Ultrastructural findings in murine seminiferous tubules as a consequence of subchronic vanadium pentoxide inhalation.

Vanadium (V) is a transition metal emitted to the atmosphere during the combustion of fossil fuels. Its current status as an atmospheric pollutant increases the need for information about the effects that this element might have on the reproductive health of exposed populations. The present study investigated changes in testicular ultrastructure following inhalation exposure of male mice to V (as vanadium pentoxide). Tissue V level was constant during the 12-week time period. We observed necrosis of spermatogonium, spermatocytes and Sertoli cells, as well as pseudo-nuclear inclusion and disruption of cellular junctions. Our findings stressed the importance of the hemato-testicular barrier in supporting the function of Sertoli cells and suggest as a possible target of V, tight junction proteins. Further analysis is needed in order to identify the role that reactive oxidative species (ROS) might have on these cellular junctions, and if a specific protein is the target of its toxic effects. The relevance of this report concerns the impact that metal air pollution could have on male fertility in dense cities with vehicular traffic problems.