Non-invasive assessment of hormonal fluctuations during pregnancy in guanacos (Lama guanicoe) and its application in a wild population.

Obtaining endocrinological profiles using non-invasive methodologies by the measurement of hormone fecal metabolites is a widely used method to monitor ovarian activity and pregnancy in wild species. These tools allow the obtention of physiological information without causing capture-related stress on the individuals. In this research, we aimed to 1) biologically validate a non-invasive method to assess fecal progestagens and estrogens fluctuations during gestation in guanacos (Lama guanicoe) and 2) apply this technique to assess pregnancy in a wild free-ranging population. Fecal samples were collected through the gestation period (~12 months) of female guanacos in a 6.5-ha paddock. An increase in fecal metabolites of both hormones was detected. Progestagens increased gradually, in contrast to estrogens, which remained at basal values for most of the gestation period and peaked only a few days before calving. To assess pregnancy in wild free-ranging animals, fecal samples were collected from a population of La Payunia provincial reserve (Mendoza, Argentina) during the beginning of gestation and at the end of gestation. Through the first months of possible gestation, pregnant females represented between 40 and 80% of the population; at the end of gestation, only 20-40% of the females had confirmed pregnancies. Our results demonstrated that the polyclonal antisera and sexual hormone metabolite assays used here detect variations in the metabolites excreted through feces in guanacos and provide the possibility of non-invasive hormone monitoring of female reproductive status. Also, the findings in wild conditions suggest that natural abortions could have occurred during the first months of gestation. Although some abortions may be natural, the harsh environmental conditions that challenge the support of such a long gestational process may be another relevant factor to consider. The results obtained here enhance our understanding of the reproductive physiology of one of the most emblematic ungulates in South America.

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

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

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.