Reproduction in the male honey possum (Tarsipes rostratus: Marsupialia): the epididymis

The epididymis of the adult honey possum, Tarsipes rostratus, is enclosed by a heavily pigmented tunica vaginalis and lies with the testis in a prominent prepenile scrotum. It is connected to the testis by a single ductus efferentis and is lined by approximately equal numbers of cuboidal ciliated and principal cells. It is unusual for marsupials in having no well-defined compartments or fibrous septae and in having extensive convolutions of the duct only at the caudal flexure. Three principal functional zones (initial, middle, and terminal segments) were identified in the epididymis, based on epithelial type and ultrastructural evidence of sperm maturation. Luminal diameter increases progressively throughout the tract, and epithelial height variations (from about 2 to 20 microns) are greatest in the terminal segment. The epithelium itself is remarkably low (maximum of 21.6 microns) compared with that seen in the epididymis of other mammals. The thickness of the peritubular smooth muscle coat increases close to the junction of the epididymis and ductus deferens. Sperm concentrations were estimated from counts of sperm nuclei and thus can be no more than approximations. The figures are consistent, however, with a rapid increase in concentration in the initial segment, indicating extensive fluid resorption. Sperm concentrations appear to peak in the distal zone of the terminal segment, although sampling problems and wide variations in count make such a conclusion only tentative. Principal and basal cells are the predominant cell types in the epididymal epithelium. Basal cells are most abundant in the initial and distal middle segment. Principal cells show structural evidence of active exchange with the luminal contents and have abundant apical stereocilia, the structure of which depends on the epididymal zone. Other cell types occur less commonly in the epithelium. Lipid-rich and phagocytic principal cells are restricted to the middle and distal zones of the middle segment, respectively. Clear cells, restricted to the terminal segment, and halo cells were found in very low numbers. As in some other marsupials, principal cells (possibly specialized for this function) selectively remove cytoplasmic droplets and probably other cellular debris from the luminal contents. In Tarsipes, however, this process is not very efficient, and many discarded droplets pass through to the terminal segment where they form large masses of debris associated with aggregates of degenerating spermatozoa.     

Morphological changes in the efferent ducts during the main phases of the reproductive cycle of birds

The changes that take place in the efferent ducts during the major phases of the reproductive cycle of birds were studied morphologically using standard histological, morphometric, and ultrastructural methods in prepuberal, sexually mature and sexually active, and sexually mature but sexually inactive domestic fowl (Gallus domesticus), drake (Anas platyrhynchos), and guinea fowl (Numida meleagris). Profound structural and dimensional changes occurred in both segments (proximal and distal) of the efferent ducts and, in particular, in the nonciliated (Type I) cell of the proximal duct of sexually mature but inactive birds. The subapical tubulovacuolar system was markedly atrophic in nonciliated (Types I and II) cells and the numerous round dense globules of Type I cells that normally occurred in sexually active birds were replaced by fewer and more pleomorphic bodies containing lipofuscin granules in sexually resting birds. Lipid droplets, few and extremely large in inactive drakes but numerous and smaller in size in guinea fowls and domestic fowls, occurred in the Type I cell at both infra- and supranuclear levels of resting but not in prepuberal or sexually active birds. Ciliated cells in both segments of the ducts exhibited fewer and less profound phase-dependent changes ultrastructurally. Generally, the Type I cells of the proximal efferent duct appeared to be more sensitive to androgen deprivation than the Type II cell of the distal efferent duct or ciliated cells in both ducts. These morphologically phase-dependent features of the efferent ducts of birds may be used, together with or independent of testicular changes, in the determination of the status of the testis and epididymis of a male bird with regard to the reproductive cycle, especially in seasonally breeding species.     

Ultrastructure of the epididymis of the tammar,Macropus eugenii,and its relationship to sperm maturation

Summary The ductus epididymidis of the tammar is lined by an epithelium composed of principal, mitochondria-rich, apical and basal cells, and intraepithelial leucocytes. The epithelium is structurally differentiated into 6 zones referred to as the initial segment, middle segment (3 subdivisions) and terminal segment (2 subdivisions). The occurrence of the initial, middle and terminal segments corresponds quite closely to the anatomical differentiation of the epididymis into a head, body and tail.The initial segment epithelium in the tammar is lower and has shorter and more slender stereocilia than in other mammals which have been described. Otherwise, the structure of the epithelium has similar characteristics in the tammar to that described in other mammals.Spermatozoa begin to develop the capacity for motility within the initial segment, but only show structural signs of maturation in the middle segment. The sperm head rotates through 90 degrees in the proximal subdivision of the middle segment. The cytoplasmic droplet is detached and spermatozoa develop the capacity for motility in the middle subdivision of the middle segment. The cytoplasmic droplets are phagocytosed by the epididymal epithelium of the middle segment. Sperm storage appears to be the main function of the terminal segment.     

The lipofuscin in neuroglial cells of the optic nerve of Formosan rock-monkey (Macaca cyclopis)

The ultrastructure of lipofuscin granules in neuroglial cells of the optic nerve of the Formosan Rock-Monkey was investigated by electron microscopy. In the cytoplasm of astroglial cells, numerous irregular lipofuscin granules were characterized by the presence of large lipid droplets, small electron-dense pigment granules, and some lamellar structures. The lipofuscin granules of the oligodendroglial cells were composed largely of dense, coarse pigment granules, multilinear structures, and a few small lipid droplets. The lipofuscin granules in microglial cells were characterized by numerous lipid droplets in various sizes, small electron-dense pigment granules, and prominent lamellar structures. It was reported that the lipofuscin granules are wear-and-tear materials and products from the cells in lower functional activity. However, our observations suggest that the presence of lipofuscin granules in the neuroglial cells of the optic nerve is likely a characteristic product of active phagocytosis.     

Cytochemical and ultrastructural characteristics of the lining epithelium of the distal part of the epididymis in hamsters (Mesocricetus auratus)

In the terminal segment of the hamster epididymidis there was some evidence of micro-merocrine protein secretion a the level of the principal cells and clear evidence of granular secretion in the light cells, presumable of glycoproteins. The PAS and protein cytochemistry reactivities observed in both these cells, of the ductus epithelial lining, but especially in the light cells, are suggestive of mucopolysaccharides and protein complexes synthesis and secretion. This secretion is carried out to the epididymal epithelium from the lumen and luminal content. A complex of small vacuoles and vesicles appeared to form from the Golgi complex is showed in the principal cells. It was suggested that this complex may represented merocrine secretory vacuoles and vesicles in these cells. Dense granules, at the TEM level, are observed in all the cytoplasm of the light cells, with correspondence to similar PAS-positive granules observed in these cells, at the light microscope level. These granules, at the TEM level, are actually secreted to the epididymal duct lumen, by the apical cytoplasms of the light cells. Signs of absorption were suggested to the principal and light columnar cells through the ultrastructural observations of micropinocytosis, apical multivesicular bodies or great membrane-bounded vacuoles in the adluminal cytoplasms.     

Morphological features of the epididymal epithelium of gerbil, Meriones unguiculatus

Principal cells of the ducts epididymis of the Mongolian gerbil showed ultrastructural characteristics of lining epithelium cells close related to processes of protein secretion, and transcytosis occurring between adjacent principal cells which were mainly verified in the initial segment. Principal cells also presented roles of fluid phase and adsorptive endocytoses, as well as autophagic and heterophagic lysosomal activities mainly observed in the caput epididymis. Columnar (principal) cells of the corpus epididymidis presented great number of variable vesicles and vacuoles distributed in all the cytoplasmic levels occurring a progressive coalescence pattern among them, which help to guarantee formation of cytoplasmic channels for fluid phase transport between the tubular lumen and epididymal interstitium. Clear cells were presented in the initial segment and predominately in the cauda epididymis epithelium of the gerbil and showed marked ultrastructural characteristics of endocytosis activities occurrence, perhaps directly related to the turnover of fluid phase of spermatozoa stored into the lumen of the distal tail. Other epididymal epithelium cells were verified and described such as basal, halo, apical and dark cells, but they did not presented special ultrastructural features.     

Opposite effect of lipofuscin granules and melanosomes from human retinal pigment epithelium of eye on photooxidation of cardiolipin

The influence of lipofuscin granules and melanosomes from human retinal pigment epithelium on the light-induced photooxidation of cardiolipin liposomes and the generation of superoxide radicals was studied. Lipofuscin granules were able to stimulate, while melanosomes inhibited, the cardiolipin photooxidation. The visible light irradiation of both melanosomes and lipofuscin granules generated superoxide radicals with mean rates of 1.5 nmole/min/10(7) and 38 nmole/min/10(7) granules, accordingly. However, melanosomes but not lipofuscin granules reacted readily with superoxide radicals. Moreover, the rate constant of degradation of superoxide radicals in the presence of melanosomes was about five orders of magnitude higher than the rate constant of its photogeneration. Therefore, we propose that melanosomes in retinal pigment epithelium cells have a photoprotective role whereas lipofuscin granules may stimulate photodestructive reactions.     

Histochemical localization of carbonic anhydrase in the testis and epididymis of the boar.

The testis and epididymis of sexually mature, fertile boars were studied for localization of carbonic anhydrase (CA) using a cobalt precipitation technique. In the testis, cytoplasmic CA was found in the Sertoli cells and in the capillaries surrounding the seminiferous tubules. The epididymal duct was divided into initial, middle and terminal segments, and regional differences in CA activity were observed. The cell membranes of the basal cells were stained in the initial and middle segments. Strong cytoplasmic CA staining was present only in the apical cells in the initial and middle segments. The basolateral cell membranes were stained in the principal cells of the terminal segment and the ductus deferens. As a rule the capillaries surrounding the epididymal duct were unstained. The enzyme, specifically localized in regions of the male genitalia acting as sperm reservoirs, might be related to the quiescence of the stored spermatozoa by influencing the acid-base status of the epididymal fluid.     

Immunocytochemical localization of antagglutinin in the boar epididymis

Summary Antagglutinin, a specific protein synthesized by the boar epididymis, was localized by the biotin-streptavidin method in all the principal cells of the caput and corpus epididymidis as well as in the lumen of this organ. Intracellular staining, which was first detected in the initial segment, appeared stronger in the distal caput and in the corpus but diminished and disappeared in the caudal epididymal cells. In all the principal cells, a consistent reaction product was localized in the large Golgi complex. Only slight and diffuse immunoreactive material was detected in the cytoplasm, except in the middle caput where the heterogeneous reactive granules appeared to be intracellular sites of degradation of this protein. In the lumen, the intensity of reaction increased from the caput to the cauda. Antagglutinin appeared strongly associated with the luminal surfaces, especially around and between the stereocilia. However, the spermatozoa also exhibited a distinct pattern of immunostaining. The results are discussed in relation to protein secretion in the epididymis and to the role of antagglutinin in the gamete-interaction process.     

Histochemical localization of carbonic anhydrase in the testis and epididymis of the rabbit.

The distribution of carbonic anhydrase (CA) was studied in the testis and epididymis of mature, male rabbits using a cobalt precipitation method. CA was found only in the endothelium of the capillaries in the testis. The epididymal duct was divided into initial, middle and terminal segments. Strong cytoplasmic CA was present in the apical cells in the initial and middle segments. Vacuoles with CA staining in the membranes were found in the principal cells in the middle segment. Intensely stained basal cells were present in the terminal segment. In the last part of the terminal segment and the first of the ductus deferens the basolateral cell membranes were also stained. The function of the enzyme is discussed especially in relation to acidification of the epididymal fluid and facilitation of CO2 diffusion.     

Histochemical studies of the rat epididymis after treatment with alpha-chlorohydrin (U-5897)

Histochemical studies of the rat epididymis after treatment with alp ha-chlorohydrin (U-5897) are presented. 14 sexually mature male rats received either daily subcutaneous injections of 50 mg U-5907/kg body weight or distilled water for 20 days. The animals were sacrificed the day following the last injection. U-5897 induced temporary sterility as demonstrated by blocked transport of spermatozoa, and spermatogenic cells eliminated from the spermatogenic epithelium which became blocked in the caudal part of the epididymis. This resulted in the distension of the segment of the distal part of the epididymal duct and to the thinning of the epithelium which lined the altered segment. Alkaline and acid phosphatases, nonspecific esterases, succinate and glucose-6-phosphate dehydrogenases and reduced nicotinamide-adenine dinucleotide tetrazolium reductase in the unchanged part of the epididymal duct were comparable to control rats whereas the altered part of the epididymis showed these activities to much weaker degrees or to be absent altogether.     

Morphology of the bovine epididymis

The epididymis of the bull was divided into six regions, and morphological differences between regions were studied. The epithelium of all regions contained four cell types: principal and basal epithelial cells, and intraepithelial lymphocytes and macrophages. The epithelium of regions II-V also contained a few apical cells. Principal cells of all regions possessed an endocytotic apparatus including stereocilia underlain by canaliculi, coated vesicles, and subapical vacuoles (up to 1 micron in diameter); however, large vacuoles with a flocculent content and multivesicular bodies (up to 5 microns in diameter) were most numerous in regions II, III, and IV. The unique features of principal cells of region I were the presence of well-developed Golgi bodies, few lipid droplets, and whorls of smooth endoplasmic reticulum in the supranuclear cytoplasm. Numerous mitochondria, distended cisternae of rough endoplasmic reticulum, and dense granules characterized the infranuclear cytoplasm of the principal cells of regions II-VI; however, these features were more developed in region V. Apical cells were characterized by the apical location of the nucleus, many mitochondria in the apical cytoplasm, and few microvilli at the luminal border. Basal cells with few cytoplasmic lipid droplets were present throughout the length of the epididymis but appeared more numerous in region V. Intraepithelial lymphocytes were present at all levels of the epithelium but were never seen in the lumen. Intraepithelial macrophages containing heterogeneous granules, eccentric nuclei, and pseudopods were invariably seen near the basal area of the epithelium in all regions. These observations are discussed in an effort to define the role of each cell type in the epididymal epithelium.     

Somatostatin-like immunoreactivity in non-pyramidal neurons of the human entorhinal region

Summary The distribution of somatostatin-immunoreactive cells and processes throughout the human entorhinal region and subjacent white matter was examined either by the unlabelled antibody-enzyme method or by the avidin-biotin method. The brain slices were obtained at autopsy with a short post-mortem delay. The majority of somatostatin immunoreactive nerve cells was found in the inner principal layer and subjacent white matter. In addition, individually scattered immunoreactive neurons were observed in both the outer principal layer and lamina dissecans. The immunoreactive perikarya varied in shape and ranged in size from 10 to 30 m. Without exception the neurons could be classified as belonging to the group of non-pyramidal neurons. Each neuron gave rise to a few thick dendrites and a thin axon with a beaded appearance. In the adult human brain, the pattern formed by lipofuscin granules deposited in the nerve cells can be considered characteristic for the type of the neuron. Therefore, immunoreactive perikarya were documented, destained of chromogen and restained to demonstrate lipofuscin pigment and basophilic substance. It became evident from these studies that the previously immunoreactive cells were characterized by a large rounded and eccentrically located nucleus, sparse basophilic substance and, in most cases, a lack of lipofuscin granules. A few of the immunoreactive cells were laden with coarse pigment granules. The findings permit classification of entorhinal somatostatin-immunoreactive neurons as either non-pigmented or pigment-laden non-pyramidal neurons.Dedicated to Prof. Dr. J. Lang, Würzburg, on the occasion of his 65th birthdayA portion of the results has been presented at the annual meeting of the European Neuroscience Association 1986 in Marseille, France (Friederich-Ecsy et al. 1986)     

Histological localization of hydrolases in the epididymis of the dog]

Localization and activity of five hydrolases (alkaline phosphatase, adenosine triphosphatase, acid phosphatase, nonspecific esterase and leucylamino-peptidase) were evaluated histochemically in the epididymides of mature dogs. In the ductuli efferentes, cilia and apical parts of the epithelial cells displayed high activity of alkaline phosphatase and adenosine triphosphatase. Strong activity of acid phosphatase, nonspecific esterase and leucylamino-peptidase was present in the basal and supranuclear zones of the epithelium of the ductuli efferentes. Stereocilia of all three segments of the ductus epididymidis showed a high activity of alkaline phosphatase. Positive adenosine triphosphatase reaction was confined to the stereocilia of the initial segment. A complex pattern of acid phosphatase activity was observed in the middle segment. The subdivision of the middle segment in four subsegments was therefore suggested. In the epithelium of the initial segment only a few nonspecific esterase-positive cells were seen. The infranuclear and basal areas of the epithelium in the middle segment and the supranuclear zone of the terminal segment displayed distinct nonspecific esterase activity. The possible contribution of the hydrolases to the function of the epididymis is discussed.     

Histology,ultrastructure, and seasonal variations in the bulbourethral gland of the African straw-colored fruit bat Eidolon helvum

We studied the morphological characteristics and seasonal changes of the bulbourethral gland of Eidolon helvum in a typical African tropical environment. Forty-eight bulbourethral glands were examined using gross anatomical, histological, histochemical, and ultrastructural techniques during the early rainy, late rainy, and peak dry seasons. The pear-shaped bilateral bulbourethral glands were located extra-abdominally in the inguinal region. Trabeculae from the capsule divided the parenchyma into numerous lobules of tubuloalveolar glandular acini. The mucosa was covered by a simple columnar epithelium consisting up of principal secretory cells, columnar dense cells and basal cells, which were progressively pronounced during the dry season. The principal cells contained eosinophilic granules, which were PAS positive while the dense cells did not show affinity for the stains. The mean gross weights, acini diameters, and epithelial heights were greater during the rainy season than the dry season. Ultrastructural evaluation showed that the cytoplasm of the principal cells contained well-developed Golgi complexes, rough endoplasmic reticulum, mitochondria, and secretory vesicles of varying electron densities and sizes. The secretory vesicles were numerous during the early rainy season, decreased during the late rainy season and were scanty during the peak dry season. The simple columnar epithelium observed during the rainy season was replaced by an undefined stratified epithelium during the dry season, and this was associated with cellular degenerations and regenerations. In conclusion, E. helvum has a typical mammalian bulbourethral gland, with a unique cell type, the dense cell whose functions are not well-understood. The gland exhibits cyclical seasonal variation in structure and secretory activity; being active during the early rainy season (breeding season), and showing the lowest activity during the dry season (non-breeding season). Glandular epithelial cell renewal occurs during the dry season in preparation for the next breeding season.     

Morphological examination of epididymal epithelium in the mule (E. hinnus) in comparison with parental species (E. asinus and E. caballus).

Following previous studies about the ultrastructure of male genital tract in parental species, a comparative study of epididymis of one of the possible hybrids, the mule, has been undertaken. Apart from small differences, general features of epididymal epithelium in the mule are similar to those of parental species. However, extension of our studies from the donkey to the horse to the hybrid permits a deeper insight into the morphology of this tract of excurrent duct. In the meantime, it is possible to evidence some features, sometimes shared with other species if taken separately, which in the whole characterize the epididymis in Equidae: the presence in principal cells of intranuclear inclusions and peculiar small granules in the basal cytoplasmic edge; the organization of groups of cells, likely to be principal ones, in such a way as to constitute intraepithelial crypts; a cumbersome presence of lipofuscinic matter all along the epithelium. Another interesting observation in the presence in the mule epididymis of well recognizable macrophages. All these data are discussed in comparison with parental species and with other species described in literature. Beyond any other consideration, it can be outlined that the complex morphology of the epithelium lining ductus epididymis in the mule is unaffected by the absence of spermatozoa, which are normally the target of the manifold functions of the epithelium itself.     

Structure of the inguinal gland in the mountain hare in various states of reproductive activity. I. The sebaceous glands

Histology and ultrastructure of sebaceous glands of the integumental inguinal gland of the hare were studied during the periods of low and high reproductive activity (in winter and in spring breeding season). It was established that the glands were functioning continually (cells in acini of all the animals used contained numerous secretory granules). The expansion of agranular endoplasmic reticulum and Golgi complex in secretory cells of sexually active animals, especially of males, shows an increased intensity of gland secretory process during the breeding season.     

A morphologic study of the ductus of the epididymis of Sus domesticus

The head, body, and tail regions of the epididymal duct (or caput, corpus, and cauda epididymis) in two healthy and sexually mature Sus domesticus males were examined by light microscopy and by scanning or transmission electron microscopy. The epididymal duct is lined with a pseudostratified epithelium with stereocilia and covered by a muscular-connective tissue sheath that is thickest in the tail region. Diameter of the epididymal duct and height of epididymal epithelium are maximal in the head region. Length of the sterocilia and spermatic density are higher in the head and body regions. Somatic cells are abundant in the tail region. The epididymal epithelium is made up of five cell types: basal cells, principal cells, clear cells, narrow cells, and basophilic cells. Abundant secretory units are observed in the supranuclear cytoplasm of columnar principal cells. Each mature secretory unit is constituted by electron-dense secretion granules covered by more than eight layers of cisternae of reticulum between which the mitochondria are intercalated. In the apical cytoplasm the isolated secretion granules become larger and less electron dense. The apical surface is covered by numerous sterocilia. Basal cells are pyramidal and less high than principal cells. The clear cells, arranged between the principal cells, are characterized by the presence of abundant vesicular elements and electron-lucid secretion granules, and by an apocrine secretory process. The narrow cells are characterized by their highly vacuolized cytoplasm. Intermediate cell typologies can be found among basal, principal, clear, and narrow cells, which could be four developmental stages of the same cell type. The basophilic cells are spheroidal and are found at different levels between the epithelial cells and in the connective tissue underlying the epithelium. © 1993 Wiley-Liss, Inc.     

Retinal pigment epithelium lipofuscin proteomics

Lipofuscin accumulates with age in the retinal pigment epithelium (RPE) in discrete granular organelles and may contribute to age-related macular degeneration. Because previous studies suggest that lipofuscin contains protein that may impact pathogenic mechanisms, we pursued proteomics analysis of lipofuscin. The composition of RPE lipofuscin and its mechanisms of pathogenesis are poorly understood in part because of the heterogeneity of isolated preparations. We purified RPE lipofuscin granules by treatment with proteinase K or SDS and showed by light, confocal, and transmission electron microscopy that the purified granules are free of extragranular material and associated membranes. Crude and purified lipofuscin preparations were quantitatively compared by (i) LC MS/MS proteomics analyses, (ii) immunoanalyses of oxidative protein modifications, (iii) amino acid analysis, (iv) HPLC of bisretinoids, and (v) assaying phototoxicity to RPE cells. From crude lipofuscin preparations 186 proteins were identified, many of which appeared to be modified. In contrast, very little protein ( approximately 2% (w/w) by amino acid analysis) and no identifiable protein were found in the purified granules, which retained full phototoxicity to cultured RPE cells. Our analyses showed that granules in purified and crude lipofuscin preparations exhibit no statistically significant differences in diameter or circularity or in the content of the bisretinoids A2E, isoA2E, and all-trans-retinal dimer-phosphatidylethanolamine. The finding that the purified granules contain minimal protein yet retain phototoxic activity suggests that RPE lipofuscin pathogenesis is largely independent of associated protein. The purified granules also exhibited oxidative protein modifications, including nitrotyrosine generated from reactive nitrogen oxide species and carboxyethylpyrrole and iso[4]levuglandin E(2) adducts generated from reactive lipid fragments. This finding is consistent with previous studies demonstrating RPE lipofuscin to be a potent generator of reactive oxygen species and supports the hypothesis that such species, including reactive fragments from lipids and retinoids, contribute to the mechanisms of RPE lipofuscin pathogenesis.