Anatomy and histology of the male reproductive tract and spermatogenesis fine structure in the lesser anteater (Tamandua tetradactyla, Myrmecophagidae, Xenarthra): morphological evidences of reproductive functions.

The anatomy and histology of the male genital tract of the lesser anteater were studied. Fine details of spermatozoa regarding their genesis and morphology were also studied in six adult specimens. The testes lie in the pelvic cavity. The deferent duct emerges from the epididymis and opens into the ejaculatory duct, which drains into the membranous urethra. Accessory glands (prostate, seminal vesicle and bulbourethral gland) are histologically similar to those described in other mammals. The short penis presents an urethral orifice, while the corpus spongiosum becomes thinner at the end indicating the absence of a histologically defined glans. The seminiferous epithelium shows: (1) Sertoli cells with deep nuclear indentations, (2) spermatogonia with crusty-like chromatin, (3) spermatocytes at different stages of maturation and (4) three morphologically distinct stages of spermatid differentiation according to nuclear shape, acrosome development and chromatin condensation. Sperm heads appear oval. The length of the spermatozoa averages 67.33 ± 1.60 µm. Two specimens with inactive spermatogenesis were azoospermic. Their testes and epididymis presented sizes smaller than those with active spermatogenesis. These studies together with others in anteaters may contribute to successful breeding in conservation programmes.

Female reproductive tract of the lesser anteater (Tamandua tetradactyla, myrmecophagidae, Xenarthra). Anatomy and histology.

The morphological and histological features of the unusual reproductive tract of the female lesser anteater, Tamandua tetradactyla (Myrmecophagidae, Xenarthra), are described for the first time. The present study aimed to establish the main similarities and differences between this species and other xenarthrans. The populations of this species are declining rapidly for a number of reasons and our study is relevant to diverse programs related to its conservation. Studies were carried out on five female genital tracts of adult specimens. Ovaries were ovoid, presenting a medulla completely surrounded by the cortex, differently from that described in other xenarthans. Like in Dasypus but different from all other armadillos studied, single oocyte follicles were observed and a simple the uterus. The uterovaginal canal connects the uterus with the urogenital sinus. The simple columnar epithelium of the uterovaginal canal ends abruptly at a septum which resembles a hymen, where the transitional epithelium of the urogenital sinus appears. This ancestral feature is shared with that of other armadillos, except Tolypeutes matacus, which has a true vagina. Characteristics of the reproductive tract and sperm morphology of other Xenarthra are comparatively discussed. These observations suggest that important reproductive features are shared between the family Myrmecophagidae and the genus Dasypus, a basal group in the phylogeny of Xenarthra.

Absence of penile erections during paradoxical sleep. Peculiar penile events during wakefulness and slow wave sleep in the armadillo.

The electroencephalogram (EEG) together with electromyogram (EMG) of the ischiocavernosus, bulbocavernosus and levator penis muscles were chronically monitored across behavioral states of the armadillo Chaetophractus villosus. This animal has a very long penis, which exhibits remarkable phenomena during wakefulness (W), slow wave sleep (SWS) and paradoxical sleep (PS). During W it remains retracted within a skin receptacle. During SWS penile protrusion can be observed together with very complex movements. Protrusion is a non erectile event during which the penis remains out of its receptacle but without rigidity. Penile erections are observed only during SWS. Contrasting with other mammals, no erections occur during PS. During this phase the penile muscles share the atonia of the body musculature characteristic of that phase. Some reflections on mechanisms of those penile events are presented.

Characterisation of glycoconjugate sugar residues in the vomeronasal organ of the armadillo Chaetophractus villosus (Mammalia, Xenarthra).

Conventional carbohydrate histochemistry and the binding patterns of 21 lectins were analysed to characterise the glycoconjugate content in the components of the vomeronasal organ of the armadillo Chaetophractus villosus. The mucomicrovillous complex of the sensory epithelium bound most of the lectins studied. No reaction was observed with Con A, PSA, S-Con A and SBA, and the sustentacular cells were-stained with UEA-I, DSL, LEL, STL and Con A. The vomeronasal receptor neurons were labelled with S-WGA, WGA, PNA, UEA-I, STL, Con A, S-Con A, ECL and RCA120. The basal cell layer reacted with S-WGA, WGA, LCA, UEA-I, DSL, LEL, STL, Con A, JAC and VVA. The nonsensory epithelium exhibited a differential staining in relation to the different components. The mucociliary complex stained with ECL, DBA, JAC, RCA120, STL, LCA, PHA-E, PHA-L, LEL, BSL-I and VVA. However, SJA and UEA-I stained the mucus complex lining a subpopulation of columnar cells. The cytoplasm and cell membranes of columnar cells was labelled with DBA, DSL and LCA. The apical region of these cells exhibited moderate reactivity with LEL and SJA. None of the lectins bound specifically to secretory granules of the nonsecretory cells. Basal cells of the nonsensory epithelium were labelled with DSL, LEL, LCA, BSL-I and STL. The vomeronasal glands showed a positive reaction with WGA, DSL, LEL, LCA, DBA, PNA, RCA120 and SBA. Subpopulations of acinar cells were observed with ECL, S-WGA, Con A, S-Con A and DBA. PNA and RCA120 stained the cells lining the glandular ducts. In comparison with previous results obtained in the olfactory mucosa of the same group of armadillos, the carbohydrate composition of the vomeronasal organ sensory epithelium differed from the olfactory sensory epithelium. This is probably related to the different nature of molecules involved in the perireceptor processes.

Ultrastructural characterisation of the olfactory mucosa of the armadillo Dasypus hybridus (Dasypodidae, Xenarthra).

The ultrastructure of the olfactory mucosa of the armadillo Dasypus hybridus was studied. A comparison with the olfactory mucosa of another armadillo (Chaetophractus villosus) was made. The olfactory mucosa of D. hybridus shows many features which are similar to those of other mammals. Interestingly, it differs from the olfactory mucosa of the armadillo C. villosus. A suggestion is made that these differences may be due to differences in the digging habits of these species. In Dasypus, the supporting cells (SCs) showed dense vacuoles, multivesicular bodies and lysosome-like bodies probably related with the endocytotic system. The SCs show a dense network of SER presumably associated with xenobiotic mechanisms. The olfactory receptor neurons exhibit lysosome-like bodies and multivesicular bodies in their perikarya. These organelles suggest the presence of an endocytotic system. Duct cells of Bowman's glands exhibit secretory activities. Bowman's glands are compound-branched tubulo-acinar mixed glands with merocrine secretory mechanisms.

Identification and localisation of glycoconjugates in the olfactory mucosa of the armadillo Chaetophractus villosus.

Conventional histochemistry and the binding patterns of 22 biotinylated lectins were examined for characterisation of glycoconjugates in the components of the olfactory mucosa of the armadillo Chaetophractus villosus. The mucous lining the olfactory epithelium showed binding sites for DSL, WGA, STL, LEL, PHA-E and JAC. Only the basilar processes of the supporting cells stained for Con-A and S-Con A. The olfactory receptor neurons stained with LEL, LCA, Con A, S-Con A, JAC and PNA. The layer of basal cells did not react with any of the lectins studied. Bowman's glands in the lamina propria showed subpopulations of acinar cells reacting with SBA, S-WGA, WGA, STL, Con A, PSA, PNA, SJA, VVA, JAC and S-Con A, but in our optical studies with lectins we were unable to differentiate between mucous and serous cells in the way that is possible on electron microscopy. The ducts of Bowman's glands were labelled with S-WGA, STL, LEL, PHA-E, BSL-I and JAC. This histochemical study on the glycoconjugates of the olfactory mucosa in the order Xenarthra provides a basis for further experimental investigations.

The vomeronasal organ of the South American armadillo Chaetophractus villosus (Xenarthra, Mammalia): anatomy, histology and ultrastructure.

The vomeronasal organ (VNO) is a chemoreceptive structure that has not been extensively studied in the Xenarthran order. Tissue samples from the VNO of the armadillo Chaetophractus villosus were prepared for light and electron microscopy. The VNO is located in the anterior part of the base of the nasal septum. It is tubular in shape, approximately 18 mm in length and opens in the rostral region of the nasal cavity and with a blind caudal end. Its lumen is lined by sensory (SE) and nonsensory (NSE) epithelium. The SE shows sensory, supporting and basal cells whereas the NSE contains ciliated and nonciliated secretory cells and basal cells. At the ultrastructural level, the sensory cells appear as bipolar neurons with conspicuous microvilli on their free surface. The supporting cells of the SE contain numerous membrane-bound vesicles in their apical regions. A peculiar feature not found in other mammals, is the presence of concentric whorls of RER cisterns frequently observed in their basal expansions. Infiltrating plasma cells can be detected in the SE basal region close to the dorsal junctional area. This region also exhibits an unusual type of basal cell, probably responsible for the generation of new vomeronasal receptor neurons. The ciliated NSE cells exhibit numerous ovoids or irregularly shaped membranous protrusions projecting from the plasma membrane of the cilia. As far as we know, this is the first study reporting the presence of this feature in ciliated NSE cells. The nonciliated cells are characterised by scarce large secretory granules and apical microvilli. The vomeronasal glands are compound-branched tubuloacinar glands with serous acinar cells. Four types of secretory granules are present. The ducts of these glands reach the lumen in the dorsolateral region between the NSE and SE. Hypolemmal nerve terminals were observed contacting secretory cells. Fenestrated and nonfenestrated capillaries constitute the vascular supply to these glands. Plasma cells, intimately associated with acinar cells, were frequently observed.

A simple method for taking photographs of histological sections without using neither photographic camera nor microscope

A simple photographic method for histological sections is described. This is a contact method which does not require the use of photographic camera. It is very similar to the contact print method used routinely in photographic laboratories. It is performed by placing the slides with the histological section, coverslip downwards, over the emulsion of the negative film. Then, they are illuminated with a common photographic enlarger during a brief period of time. Thus, a negative contact print of the section is obtained. This technique yields photographs of high quality focus, sharply contrasted with great detail. These results may also be obtained with low-contrast stained sections. This method is most suitable for photography of large or medium-sized histological sections (macros). This is often the case of neuroanatomical and immunohistochemical studies. Our method, being simple and quick, is recommended for routine use in laboratories

A simple method for taking photographs of histological sections without using neither photographic camera nor microscope

A simple photographic method for histological sections is described. This is a contact method which does not require the use of photographic camera. It is very similar to the contact print method used routinely in photographic laboratories. It is performed by placing the slides with the histological section, coverslip downwards, over the emulsion of the negative film. Then, they are illuminated with a common photographic enlarger during a brief period of time. Thus, a negative contact print of the section is obtained. This technique yields photographs of high quality focus, sharply contrasted with great detail. These results may also be obtained with low-contrast stained sections. This method is most suitable for photography of large or medium-sized histological sections (macros). This is often the case of neuroanatomical and immunohistochemical studies. Our method, being simple and quick, is recommended for routine use in laboratories

Olfactory mucosa of the South American armadillo Chaetophractus villosus: an ultrastructural study.

The sense of olfaction in armadillos plays an important role, suggested by the great development of the nasal structures, olfactory bulbs, and related brain regions. The mammalian olfactory mucosa is a privileged site of neuronal death and regeneration during the whole life span. A detailed knowledge of its ultrastructure is convenient for gaining insight into the factors controlling those phenomena. We performed this work in species not previously studied in order to provide a firm basis for further research on those factors. No information is available on the histology and ultrastructure of the olfactory mucosa in the order Xenarthra to which armadillos belong. Samples from the endoturbinals of the armadillo Chaetophractus villosus were prepared for light and electron microscopic examination by the usual conventional means. The olfactory epithelium of Chaetophractus villosus shows the classical three types of cells: supporting cells, olfactory receptor neurons, and basal cells. The olfactory neurons and the basal cells were similar to that described in other species. Two different types of supporting cells are described. An outstanding characteristic of the supporting cells is the normal presence of abundant phagosomes, apical secretory granules, apocrine-like protrusions, and highly developed smooth endoplasmic reticulum. Apoptotic bodies are frequently found in the infranuclear cytoplasm of supporting cells. The ductular epithelium of Bowman's glands reveals secretory activity. The lamina propria shows mixed Bowman's glands. Great development of smooth endoplasmic reticulum is observed in the mucous acinar cells. Evidence for merocrine and apocrine mechanisms in the Bowman's glands is presented. The presence of apoptotic bodies and phagosomes in supporting cells suggests a participation in the cellular events induced by cell death and proliferation of the olfactory epithelium. The variety of characteristics exhibited by the supporting cells of the olfactory mucosa may contribute to a deeper understanding of their scarcely known functions.

The brain of the armadillo Dasypus hybridus. A general view of its most salient features

The most salient neuroanatomical features of the brain of the seven banded armadillo Dasypus hybridus are described. The microscopic characteristics were studied by serial transverse and sagittal paraffin sections, stained with Nissl and Klüver-Barrera technique. This analysis comprises the telencephalon, diencephalon and mesencephalon. The most outstanding features of this brain are: 1) Great development of rhinencephalic structures (olfactory bulbs, olfactory tubercles, anterior commissure and pyriform cortex). 2) The relative size of the induseum griseum strongly suggests that this animal would be useful for a variety of studies on this structure. 3) The high position of the rhinal fissure on the lateral hemispheric wall determines the smallness of neocortex. Therefore, this armadillo is also useful for decortication studies. 4) Absence of a distinct pineal organ. 5) Conspicuous subfornical and subcommissural organs. 6) Absence of a distinct intermediate lobe in the hypophysis

The brain of the armadillo Dasypus hybridus. A general view of its most salient features

The most salient neuroanatomical features of the brain of the seven banded armadillo Dasypus hybridus are described. The microscopic characteristics were studied by serial transverse and sagittal paraffin sections, stained with Nissl and Kl³ver-Barrera technique. This analysis comprises the telencephalon, diencephalon and mesencephalon. The most outstanding features of this brain are: 1) Great development of rhinencephalic structures (olfactory bulbs, olfactory tubercles, anterior commissure and pyriform cortex). 2) The relative size of the induseum griseum strongly suggests that this animal would be useful for a variety of studies on this structure. 3) The high position of the rhinal fissure on the lateral hemispheric wall determines the smallness of neocortex. Therefore, this armadillo is also useful for decortication studies. 4) Absence of a distinct pineal organ. 5) Conspicuous subfornical and subcommissural organs. 6) Absence of a distinct intermediate lobe in the hypophysis

A simple method for taking photographs of histological sections without using neither photographic camera nor microscope.

A simple photographic method for histological sections is described. This is a contact method which does not require the use of photographic camera. It is very similar to the contact print method used routinely in photographic laboratories. It is performed by placing the slides with the histological section, coverslip downwards, over the emulsion of the negative film. Then, they are illuminated with a common photographic enlarger during a brief period of time. Thus, a negative contact print of the section is obtained. This technique yields photographs of high quality focus, sharply contrasted with great detail. These results may also be obtained with low-contrast stained sections. This method is most suitable for photography of large or medium-sized histological sections (macros). This is often the case of neuroanatomical and immunohistochemical studies. Our method, being simple and quick, is recommended for routine use in laboratories.

The brain of the armadillo Dasypus hybridus. A general view of its most salient features.

The most salient neuroanatomical features of the brain of the seven banded armadillo Dasypus hybridus are described. The microscopic characteristics were studied by serial transverse and sagittal paraffin sections, stained with Nissl and Klüver-Barrera technique. This analysis comprises the telencephalon, diencephalon and mesencephalon. The most outstanding features of this brain are: 1) Great development of rhinencephalic structures (olfactory bulbs, olfactory tubercles, anterior commissure and pyriform cortex). 2) The relative size of the induseum griseum strongly suggests that this animal would be useful for a variety of studies on this structure. 3) The high position of the rhinal fissure on the lateral hemispheric wall determines the smallness of neocortex. Therefore, this armadillo is also useful for decortication studies. 4) Absence of a distinct pineal organ. 5) Conspicuous subfornical and subcommissural organs. 6) Absence of a distinct intermediate lobe in the hypophysis.

Standardization of fixation, processing and staining methods for the central nervous system of vertebrates.

This paper reports the standardization of methods used for processing and embedding various vertebrate brains of different size in paraffin. Other technical details developed for avoiding frequent difficulties arising during laboratory routine are also reported. Some modifications of the Nissl and Klüver-Barrera staining methods are proposed. These modifications include: 1) a Nissl stain solution with a rapid and efficient action with easier differentiation; 2) the use of a cheap microwave oven for the Klüver-Barrera stain. These procedures have the advantage of permitting Nissl and Klüver-Barrera staining of nervous tissue in about five and fifteen minutes respectively. The proposed procedures have been tested in brains obtained from fish, amphibians, reptiles and mammals of different body sizes. They are the result of our long experience in preparing slides for comparative studies. Serial sections of excellent quality were regularly obtained in all the specimens studied. These standardized methods, being simple and quick, are recommended for routine use in neurobiological laboratories.

New and peculiar cytoplasmic membranous bodies in the acinar cells of the harderian gland of the armadillo Chaetophractus villosus.

A new and peculiar morphological feature in acinar cells of the Harderian gland of the South American armadillo Chaetophractus villosus (Mammalia, Dasipodidae) is reported. The gland of adult males and females was studied at macroscopic, microscopic and electron microscopic levels. The gland is the largest structure in the bony orbit. It is located in its medial (nasal) and basal side. It shows a tubuloalveolar structure characterized by large alveoli with a single layer of columnar or cuboidal cells. Myoepithelial cells are located between the secretory ones and the basement membrane. The peculiar morphological feature consists of large intracellular membranous bodies located in the supranuclear cytoplasmic region. They are seen in every acinar cell of males and females. Their size is prominent being almost as large as the nucleus. Only one body is observed in each cell. The structure of the bodies displays an outstanding geometrical pattern which differs completely from other membranous structures described in other species.

New and peculiar cytoplasmic membranous bodies in the acinar cells of the harderian gland of the armadillo Chaetophractus villosus.

A new and peculiar morphological feature in acinar cells of the Harderian gland of the South American armadillo Chaetophractus villosus (Mammalia, Dasipodidae) is reported. The gland of adult males and females was studied at macroscopic, microscopic and electron microscopic levels. The gland is the largest structure in the bony orbit. It is located in its medial (nasal) and basal side. It shows a tubuloalveolar structure characterized by large alveoli with a single layer of columnar or cuboidal cells. Myoepithelial cells are located between the secretory ones and the basement membrane. The peculiar morphological feature consists of large intracellular membranous bodies located in the supranuclear cytoplasmic region. They are seen in every acinar cell of males and females. Their size is prominent being almost as large as the nucleus. Only one body is observed in each cell. The structure of the bodies displays an outstanding geometrical pattern which differs completely from other membranous structures described in other species.