Print version ISSN 0716-9868
Rev. chil. anat. vol.15 n.1 Temuco 1997
REGIONAL HISTOLOGY OF THE DUCTUS EPIDIDYMIDIS IN THE DOG
(Canis familiaris, L.)
ESTUDIO HISTOLOGICO REGIONAL EN EL EPIDIDIMO DEL PERRO
(Canis familiaris, L.
* Schimming, B. C
** Vicentini, C. A.
* Orsi, A. M.
** Franceschini-Vicentini, I. B.
** Abreu-Rays, M. A.
* Department of Anatomy, Biosciences Institute of Botucatu, UNESP, São Paulo, Brazil
** Department of Biological Sciences, Faculty of Sciences of Bauru, UNESP, São Paulo, Brazil
This paper was supported by CAPES Grant.
SUMMARY: The epididymis of the dog, based on morphological differences, reveals five distinct and continuous zone in the duct. It is observed zone I and zone II, both of them localized into the head. The third zone, or zone III, comprises the distal head and all the body. The fourth and fifth zones (zones IV and V), are restricted to the tail, localized into the proximal and distal Cauda epididymidis, respectively. Each zone can be readly distinguished on the basis of its own duct morphological characteristics. This distinction concerns mainly to the overall duct histology, and also to cytological appearence and epithelial lining constitution.
KEY WORDS: 1. Epididymidis; 2. Dog; 3. Anatomy; 4. Regional histology.
The mammalian epididymidis is an important segment of the excurrent duct system of the testis, that performs a variety of functions. HAMILTON (1975) reported that epididymis is a dynamic structure of the seminal pathway, and its complex morphophysiology does not agree with the concept that epididymis would have merely a passive influence on sperm physiology. This view have support on several studies (see ROBAIRE & HERMO, 1988, for review).
It is evident that when passing through the epididymis, spermatozoa undergo morphological, physiological and maturational changes until they reach the tail of epididymis (Cauda epididymidis), where they are stored (BEDFORD, 1966, 1975; ORGEBIN-CRIST, 1967).
Histochemical and morphological studies in dog epididymidis have been published in the literature, with emphasis in protein secretion (NICANDER & MALMQVIST, 1977), localization and activity of hydrolases (SINOWATZ et al., 1979a), and of glycosidases (SINOWATZ et al., 1979b). Regional distribution of steroids in dog epididymidis has been carried too (BOUJARD et al., 1982). KAWAKAMI et al. (1991) reported changes in the epithelial lining of the epididymis, when the dog became sexually mature. An immunocytochemical study reveals the expression patterns of several cytokeratins, vimentin and desmin in the epididymis of the dog (WAKUI et al., 1994).
Despite such studies, stressing the importance of the morphofunctional knowledge of the dog epididymidis, little information is available on the regional histology of the duct (ORSI, 1983). Thus, the purpose of this study is to describe morphologically the regional histology of the ductus epididymidis in the dog, to provide a detailed structural baseline for specific studies of functional zonation in the epididymis of the dog, or, to comparate to other mammals.
MATERIAL AND METHOD
Ten adult, mongrel, sexually mature dogs (Canis familiaris, L.), after necropsy, had the epididymes, rapidly, dissected and isolated from the testes. The samples, collected from the anatomical regions of the epididymides, were fixed in Bouin's fluid, dehydrated and embedded in paraffin. Cross sections, 7 µm thickness, were stained with hematoxylin and eosin (H/E), and Mallory's and Masson's trichromes (ROMEIS, 1928; BEHMER et al., 1976). The tissues were observed in a Olympus BH-2 microscope.
The rounded tubules, obtained from several sections of different epididymal regions, were measured for height of epithelium, diameter of lumen and thickness of muscle coat. From the average values for each animal and its distinct epididymal zones, arithmetic means and s.d., for each histological epididymal zone, were calculated.
The epididymis in the dog, reveals five distinct and continuous histological zones in the duct. It is observed zone I or initial segment and zone II, both of them localized into the head. The zone III comprises the distal head and all the epididymal body. The fourth and fifth zones (zones IV and V), are restricted to the epididymal tail, localized into the proximal and distal Cauda epididymidis, respectively. The last region (V), is continuous to the vas deferens (Fig. 1).
The entire epididymal duct has a similar histological appearance, showing a pseudostratified epithelial lining, with stereocilia, covered by a myo-connective tissue sheath. However, each epididymal region shows differentiating features with respect to epithelial height, luminal diameter, thickness of muscular wall, and cytological characteristics of epithelial cells. Also, it should be noted that transition between the zones is gradual, not abrupt. Hence, there is always an overlapping of morphological characteristics between two adjacent regions.
Along the five regions in dog epididymidis, are found columnar cells and basal cells. Two different types of columnar cells could be distinguished: the predominant principal (P) cells, and a less numerous cell type named apical (A) cells. The P cells are observed in all the five zones, of the epididymal duct. They are tall cells, which extend from the basal lamina to the lumen. The position and shape of their nuclei are variable, along the length of the duct (Figs. 2-8). The basal (B) cells lie next to the basal lamina, and do not reach the tubular lumen. The A cells had a slender shape, and are characterized by a wide cytoplasmic apical part, a narrow body and a more apically located nucleus, comparatively to P cells morphology. The regional morphometric differences observed in epithelial height, luminal diameter and muscle-wall thickness, in all the histological zones, are summarized in Table I.
The zone I or initial segment , is characterized by an irregular and tall epithelium (45.60 ± 4.43 µm), which is the greatest value of the epididymal epithelial height observed. The epithelium contained all the cell types noticed. The tall columnar P cells have round to spherical nuclei with one or two distinct nucleoli, and usually the nuclei are localized in the lower half of the cells. The cell boundaries are indistinct, and a large number of long stereocilia projects from the apical cell surfaces to the lumen. B cells are seen in great number with ovoid nuclei and evident nucleoli. These cells are arranged in a distinct layer, disposed immediately over the basal lamina. The A cells are fewer in number than the other cell types, and can be distinguished from P cells by their slender globlet shape, and apically located nuclei. The irregular lumen of the ductus have a diameter of 125.85 ± 7.89 µm, and was devoid of spermatozoa. The duct is surrounded by a smooth muscle layer of about 11.05 ± 0.86 µm thick (Fig. 2).
The epithelial height of zone II is somewhat lower than those of zone I (41.85 ± 3.73 µm), but the luminal diameter (135.5 ± 10.03 µm) is little different than earlier zone. The P cells average 40 µm in height and their nuclei are ovoid, extremely dark with a conspicuous nucleoli. The adluminal stereocilia are long. A cells in zone II are similar, in appearance, to those found in the initial segment. The B cells are characterized by presence of nuclei of different shapes.The epididymal lumen in zone II appears regular, and contains rather immature spermatozoa in scarce number, and also cellular remnants. The muscular coat, surrounding the duct, almost does not vary in thickness (11.6 ± 0.91 µm).
Some features distinguished the epithelial lining of zone III. The epithelium decreases in height and, concomitantly there is an increase in luminal diameter and in muscle wall thickness. This zone was characterized by irregular epithelium (40.8 ± 1.72 µm), which accounted for the stellate appearance of the lumen.The epithelium contained the same three cell types, similar to other zones. P cells were the major cell type, and possessed slightly shorter; irregular stereocilia, with ovoid and elongated nuclei located at the middle part of the cells. A cells, with round to ovoid nuclei, are scarcely found, and B cells are similar to those of preceeding zones. The luminal diameter has average of 146.12 ± 8.08 µm, and the muscle coat is thicker than earlier zones (14.15 ± 1.79 µm) (Figs. 3 - 4).
The luminal and tubular diameters of the epididymal duct in zone IV are greater than those of zone III, while the epithelium in zone IV is lower avering 35 µm in height. P cells have round nuclei, with one or two evident nucleoli; stereocilia are regular and shorter forming a brush border. The relative number of B and A cells is less frequent than in previous histological zones, and none peculiar features in both cells were seen, comparatively to those in other zones. The tubular lumen ranges between 195 and 220 µm in diameter and contains few packed spermatozoa. The muscle coat surrounding the epididymal duct in zone IV (Figs. 5-6), is slightly thicker (16.25 ±1.78 µm), than in zone III.
Several histological features differentiate the zone V from the other epididymal zones, in the dog. Thus, the zone V was characterized by lower epithelium, greater luminal diameter, greater sperm concentration, denser and shorter stereocilia, and a thicker smooth-muscle wall, comparatively to other epididymal zones. The epithelial lining measures 25.8 ± 1.21 µm. P cells are also the most common epithelial cell type found in this zone. Usually, their nuclei are round and situated at the middle part of the cell but, because of the shortness of the P cells, nuclei rest near the basement membrane. The stereocilia still are regular and shorter, forming a brush border. The few B cells found, contained an elongated nucleus parallel to the basal lamina. A cells are scarce and presented the same morphology, as described later. The luminal diameter is 596.37 ± 32.45 µm, and the lumen is densely crowded with spermatozoa. The luminal occurrence of many packets of spermatozoa is a typical feature of this zone. The muscle coat is thicker (25.3 ± 2.53 µm), relatively to the other zones.
Epididymal morphology at the light microscopic level has been extensively reported in most species including the rat (REID & CLELAND, 1957), guinea pig (HOFFER & GREENBERG, 1978), buffalo (ABDOU et al., 1985), bull (GOYAL, 1985), hamster (VICENTINI & ORSI, 1987), cat (VIOTTO et al., 1988), goat (GOYAL & WILLIAMS, 1991), and black isogenic mouse (ORSI et al., 1993).
The epididymis, in all the species studied, so far is known, present variabilities in structure and function along its length. However, the precise number of epididymal regions may be species-specific (NICANDER, 1958; HAMILTON, 1975; RAMOS & DYM, 1977; GOYAL, 1985), and/or epididymal histological zonation may vary within the same species depending upon the criteria of classification (GLOVER & NICANDER, 1971).
Based upon morphological features such as: epithelial height, luminal diameter, thickness of muscle coat, cell types and regional differences in their distribution, the epididymis in the dog could be divided here into five distinct and continuous zones as follow, zones I and II are localized into the head; zone III comprises the distal head and all the body, and zones IV and V are restricted to the tail, localized into the proximal and distal cauda epididymidis, respectively. This morphological pattern in the dog epididymidis, is basically similar to that described in the hamster (VICENTINI & ORSI, 1987).
The epididymal zone I, in dog, exhibit the greatest value of epithelial height, associated to scarce spermatozoa inside the lumen suggesting fast transit of spermatozoa, which were similarly reported for the hamster and cat epididymal initial segments (VICENTINI & ORSI, 1987; VIOTTO et al., 1988, respectively).
According to NICANDER & GLOVER (1973) and FLICKINGER et al. (1978) descriptions, the epididymal zone II could be incorporated in the epididymal middle segment of rodent. However, in this study the zone II is restricted to the head of the dog epididymidis exclusively. Thus, the middle segment of the dog epididymidis, or zone III, comprises the distal head and all the body. Previously, ORSI et al. (1985) have postulated a similar description for the middle segment of the pig epididymidis, on the basis GLOVER & NICANDER (1971), previous morphological pattern for epididymal histological zonation in some mammals. Also, zone III showed irregular luminal surface and usually stellate-shaped lumen, which reflect variabilities on the epithelial height along this zone, upon comparative base (GOYAL & WILLIAMS, 1991).
Concerning to the zones IV and V in the dog Cauda epididymidis, our observations agree to ORSI (1983), previous description, relatively to localization of both zones at the epididymal terminal segment, similar to findings reported in hamster epididymidis (VICENTINI & ORSI, 1987). The zone V comprises the distal tail, and is characterized by lower epithelial lining, greater luminal diameter, shorter stereocilia, packed spermatozoa inside the tubular lumen, and a thicker smooth muscle wall, comparatively to the other histological zones in the dog epididymidis. Some of these morphological features, here described, were previously reported to the distal cauda epididymidis of other mammals, with function of spermatozoa storage (NICANDER, 1957, 1958; HAMILTON, 1975; WAITES, 1980; GREENBERG & FORSSMANN, 1983; ORSI et al., 1985; ABDOU et al., 1985; ROBAIRE & HERMO, 1988; GOYAL & WILLIAMS, 1991; BRIZ et al.,1993). Furthermore, EKSTEDT et al. (1991) considered the cauda epididymidis as a place for long-term storage of mature spermatozoa, a function that also could occur in the dog distal zone of epididymis.
The epithelial lining of all histological zones in the dog epididymidis contains, mainly, three cell types: principal (P) cells, basal (B) cells and apical (A) cells. These cells have been reported in rat (HAMILTON, 1975), man (LÜDERS, 1976), opossum (ORSI et al., 1981), bull (ORSI et al., 1984), buffalo (ABDOU et al., 1985) and cat (VIOTTO et al., 1988). These different cell types are distinguished between themselves, mainly, by position and morphology of the nuclei (REID & CLELAND, 1957), and their functions have been extensively studied (ROBAIRE & HERMO, 1988). Although, clear cells, present in some rodent epididymidis (ROBAIRE & HERMO, 1988), were not observed in dog epididymidis. Also, clear cells are absent in other species including the monkey (RAMOS & DYM, 1977), guinea pig (HOFFER & GREENBERG, 1978), bull (GOYAL, 1985), and goat (GOYAL & WILLIAMS, 1991).
At the epithelium of the zones I and II, Caput epididymidis of the dog, a continuous layer of B cells were observed next to the basement membrane of the ductus, similarly to PAL & BHARADWAJ, (1989) observation on the basal cell pattern in the Corpus epididymidis of the Indian buffalo. The true cellular function of B cells seems to be hitherto unknown (ROBAIRE & HERMO, 1988), however BRIZ et al. (1993), recently, have discussed that B cells could be the "stem cells" of the epididymal cell population.
In rodents, A cells appear in proximal regions of the epididymis, mainly in the initial segment (HAMILTON, 1975; VICENTINI & ORSI, 1987; ROBAIRE & HERMO, 1988). In cat, VIOTTO et al. (1988) showed A cells in all the histological zones of epididymal head and body. On the other hand, A cells were, apparently, absent in the dog epididymidis (ORSI, 1983; KAWAKAMI et al., 1991). In contrast to those findings, in our present study, it has been found A cells in all the histological zones of the dog epididymidis, although in the zone V (distal tail), they are not so striking, owing to the lower epithelial height. Some cytophysiological correlations between A and P cells have been made, at immunocytochemical and ultrastructural levels (see ROBAIRE & HERMO, 1988, for revision).
Fig. 1. Fully exposed epididymis of the dog. Note the anatomical regions: caput (cp), corpus (cr), and cauda (cd), and their histological zones (roman numerals I-V), and the vas deferens (d).
Fig. 2. Initial segment (zone I). Note the principal (p), apical (a) and basal (b) cells, and the stroma periductal (rose). H & E. x 200.
Fig. 3. Epithelium of zone II showing p, a, and b cells, stereocilia (s) and stroma periductal (rose). H & E. x 200.
Fig. 4. Stellate lumen (stars) of the ductal sections in the zone III. H & E. x 100.
Table I - Morphological comparisons between different zones of the epididymis in the dog
|Epithelial height (µm)||45.60 ± 4.43||41.85 ± 3.73||40.8 ± 1.72||37.45 ± 1.24||25.8 ± 1.21|
|Luminal diameter (µm)||125.85 ± 7.89||135.5 ± 10.03||146.12 ± 8.08||204.25 ± 7.21||596.37 ± 32.45|
|Muscle-wall thickness (µm)||11.05 ± 0.86||11.6 ± 0.91||14.15 ± 1.79||16.25 ± 1.78||25.3 ± 2.53|
1P, principal; B, basal; A, apical Data are expressed as mean ± SD
Fig. 6. Epithelium of zone IV showing p and b cells. Note the stereocilia (s) and myoepithelial cells (arrowheads). H & E. x 400.
Fig. 7. The lower epithelium of the zone V showing the greater luminal sperm concentration (*). H & E. x 40.
Fig. 8. Zone V. The p and b cells are indicated. H & E. x 1000.
RESUMEN: La histología regional del conducto del epidídimo en el perro comprende cinco regiones histológicamente distintas y continuas. Dos de las zonas se localizan dentro de la cabeza (zona I o segmento inicial y zona II), una en la cabeza distal y en el cuerpo (zona III) y dos en la cola del epidídimo (zonas IV y zona V), situadas en la Cauda epididymidis, respectivamente. Cada zona puede distinguirse fácilmente sobre la base de variaciones de las características morfológicas del conducto. Esta distinción tiene relación con la histología del conducto en conjunto y también con el aspecto citológico y constitución del epitelio de revestimiento.
PALABRAS CLAVE: 1. Epidídimo; 2. Perro; 3. Anatomía; 4. Histología regional.
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Dirección para correspondencia:
Prof. Bruno Cesar Schimming
Department of Anatomy
Biosciences Institute of Botucatu
State University of São Paulo
Botucatu - 18618-000 -
São Paulo BRAZIL
Aceptado : 07-03-1997