MORPHOLOGICAL AND IMMUNOHISTOCHEMICAL STUDY OF THE NON- AMPULLATED PART OF THE DUCTUS DEFERENS OF THE CAMEL (CAMELUS DROMEDARIUS)

Assiut Vet. Med. J. Vol. 52 No. 111 October 2006

Dept. of Anatomy and Histology. Fac. of Vet. Med., Assiut University, Egypt. MORPHOLOGICAL AND IMMUNOHISTOCHEMICAL STUDY OF THE NON- AMPULLATED PART OF THE

DUCTUS DEFERENS OF THE CAMEL

(CAMELUS DROMEDARIUS)

(With One Table and 16 Figures)

By A.M. SALEH (Received at 30/8/2006)

دراسات مورفولوجية مناعية هستوکيميائية على الجزء الغير انبوری

للقناة الاسرية في الجمل وحيد السنام

عبد المهيمن مصطفی صالح اجريت دراسة مورفولوجية عيانيا ومجهريا بواسطة الميکورسکوب الضوئي والالکتروني الماسح وکذلک دراسة مناعية هستوکيميائية للجزء غير الانبوری للقناة الأسرية في الجمال وحيدة السنام السليمة ظاهريا وناضجه جنسيا. ينقسم الجزء الغير انبوري إلى ثلاثة مناطق تشريحية منطقة دانية شديدة التعرج، ومنطقة وسطی بسيطية التعرج، ومنطقة بعيدة غير

ص فحة

متعرجة. تغطى القناة الأسهرية بظهارة عمودية مطبقة کاذبة ومهدبة وترتکز على اساسية رقيقة. وتحمل الخلايا الظهارية اهدابا طويلة متفرعة في المنطقة الدانية بالمقارنة مع اهداب طويلة غير متفرعة في المنطقة الوسطى وخملات قصيرة في المنطقة السفلية. أما الغلالة العضلية فهي تتکون من طبقتين: داخلية دائرية وخارجية طولية. تم استخدام

9.5-DBH PGP وChAT لدراسة التوزيخ

التفاعلات المناعية الهستوکيميائية المضادة کل من العصبي العام والسمبثاوي والجارسمبثاوي للقناة الأسهرية. اظهرت هذه الدراسة أن الأعصاب الداخلية الموجودة في القناة الأسهرية تکون ليفة عصبية في النسيج تحت الطلائی واخرى في النسيج العضلى.

SUMMARY

Gross, light & scanning electron microscopic and immunohistochemical morphology of the non-ampullated part of the ductus deferens of sexually mature dromedary camels were studied with special reference to the changes and modifications occurring throughout its entire length. This part was subdivided into three anatomical regional segments; proximal tortuous (scrotal), middle less tortuous (inguinal) and distal straight (abdominal) segments. It characterized by simple and low folded mucosa. It lined by pseudostratified ciliated columnar epithelium with a thin lamina propria-submusosa. By scanning electron microscope,

Assiut Vet. Med. J. Vol. 52 No. 111 October 2006

stereocilia of epithelial cells were tall and branched in the proximal segment tall, non-branched stereocilia in the middle segment and thick short microvilli in the distal segment. The muscular coat arranged in two layers; the inner layer was circular, while the outer layer was longitudinal. The intramural nerve fibers of the ductus deferens form plexuses in subepithelial tissue and in the muscular coat in addition larger nerve bundles in the serosa. Immunohistochemical reaction to protein gene product-9.5 (PGP-9.5) was used to demonstrate the general innervation pattern. The adrenergic and the cholinergic innervation were demonstrated using by immunohistochemical reactions to DPH and CHAT. These structural variations along the length of the vas deferens suggest that it performs functions other than just as a passageway for spermatozoa.

Key words: Ductus deferens, camel, immunohistochemistry, scanning electron

microscope, mesoductus deferens.

INTRODUCTION

The ductus deferens is a tubular structure connecting the tail of the ductus epididymis with the pelvic urethra (Popovic et al., 1973). It has been reported that it is a major storage site for sperms (Turner 1995). It probably plays important role in providing sperm with a suitable environment for their maturation, viability and protection during their passage through its terminal portion (Bergerson et al., 1994; Andonian and Hermo, 1999). The anatomical studies on the non-ampullated part of the ductus deferens have been conducted by several authors in different domestic animals. Schummer et al. (1979) in the domestic animals and Skidmore (2002) in the camel conducted the gross anatomical observation. Trautman and fiebiger (1957), Goswami et al. (1994) in camel, Wrobel and Dellmann (1993) in the domestic animals conducted the microscopic observation. The innervation had been studied histochemsitry and immunohistochemsirty by Duminika (1983) in the dog, Kujat et al. (1993) in bulls, Kaleczyc et al (1997) in boar and Mirabella et al. (2003) in water buffaloe. Studies on the camel ductus deferens are meager and focus mainly on some histological structures. There exist, so far, no reports on the scanning electron microscopy and the innervation of the camel ductus deferens. Therefore, the present study was undertaken to give more information on the morphology of the non-ampullated part of the ductus deferens in the dromedary camel grossly, histologically, immunohistochemically and by the scanning microscopy.

20

Assiut Vet. Med. J. Vol. 52 No. 111 October 2006

MATERIALS and METHODS The present study was carried out on the ductus deferens of 30 sexually mature and apparently healthy one humped camels (Camelus dromedarius) collected from Bany Ady slaughter house (Assiut - Egypt). Specimens were collected from the deferent segments of the ductus deferens and processed for light and scanning electron microscopy. Sections, 5-7um thick, were stained with H&E, Periodic acid Schiff (PAS) and Alcian blue (AB) techniques. Some morphometric aspects were performed using Leica Q 500 MC Image analyzer. For Scanning electron microscopy, specimens were immersed in 5% glutaraldehyde, post-fixed in 1% Osmium tetroxide, dehydrated in alcohol followed by amyl acetate and critical point dryed using liquid co2 and mounted on specimen stubs, sputter-coated with gold and examined in a JEOL 5400 LV scanning electron microscope.

Sample from the different segments of the ductus deferens were immunohistochemically treated for visualization of PGP-9.5-, DBH- and CHAT-positive nerves by light microscopy according to methods described fully by Kujat et al. (1993) and Saleh (2002).

RESULTS

The non-ampullated part of the ductus deferens of dromedary camel was relatively long and measured about 57.7 cm in length. It was enclosed in a narrow serosal fold, the mesoductus deferens. It was divided into three segments according to its position; scrotal, inguinal and abdominal. The scrotal one ran highly tortuous on dorsomedial aspect of the testis, measured about 25.5 cm in length (Table 1, Fig. 1, 2). The inguinal segment ran less tortuous in the inguinal canal medial to the vascular part of the spermatic cord, formed the longest segment of the ductus deferens (24.7 cm in length) (Table 1, Fig. 1). The abdominal segment, gained the abdominal cavity at the internal inguinal ring, ran dorsocaudally, entered the genital fold and continued as ampulla ductus deferens. It measured 16.5 cm in length and 2.6 mm in diameter (Table 1)

The wall of the non-ampullated part of the ductus deferens consisted of three circumferentially arranged layers around a centeral lumen: tunica mucosa, tunica muscularis and tunica serosa (Fig. 2, 3, 4). The latter was continuous with the mesoductus deferens and contained blood vessels, nerves and variable amount of adipose tissue. The lumen of the ductus deferens was narrow at the initial part, dilated distal-wards and frequently contained masses of sperms.

21

Assiut Vet. Med. J. Vol. 52 No. 111 October 2006

The tunica mucosa of the ductus deferens of the dromedary camels showed low longitudinal folds, lined with ciliated pseudostratified columnar epithelium. Four types of cells; principle, basal, apical and granular were recognized (Fig. 5).

The principle columnar cells predominated the lining epithelium of the ductus deferens. It characterized by acidohphilic foamy cytoplasm and rounded or oval basally situated nuclei. The basal cells were laid next to the basement membrane and characterized by slightly basophilic cytoplasm and ovoid or round nuclei. The apical cells are few in number and characterized by small deeply stained apical located nuclei. Granular cells were found among the principle columnar cells in the scrotal segment, extending from the basement membrane to the luminal surface. They presented wide apical portion, fine basiophilic granules in cytoplasm and round deeply stained nuclei (Fig. 5).

The scanning electron microscopy revealed the presence of three different pictures in the luminal surface of the non-ampullated part of the ductus deferens. In the scrotal segment, it showed some non-ciliated cauliflower-shaped cells among the dense ciliated epithelium (Fig. 7). The ciliated cells were dominating and studded with dense tall (6.6 um) branched stereocilia (Fig. 8). The non-ciliated cells were few and presented micro-secrotory granules (Fig. 9). In the inguinal segment, the epithelial surface showed only ciliated cells. The stereocilia were dense non-branched and relatively shorter (4.3 um) than that of the scrotal segment (Table 1, Fig. 10). In the abdominal segment, the surface of the epithelium showed short (2.5 um) and thick microvilli (Table 1, Fig. 11).

PAS staining revealed mild to moderate reaction in the epithelial cells. The principle columnar cells contained large number of PAS positive granules in the apical zone and few ones in the basal portion. The secretory blebs and stereocilia showed intense PAS reaction. The basal cells showed weakly PAS reaction in the form of few fine positive granules (Fig. 6). With Alcian blue stain, the lining epithelium of the ductus deferens showed very weakly or no reaction.

The epithelium height reached its maximum value in the scrotal segment and decreased gradually toward the abdominal segment. It measured about 25.9, 20.21and 16.21um in the scrotal, inguinal and abdominal segments respectively (Table 1).

The lamina propria-submucosa composed of dense connective tissue with collagenous fibers. Some smooth muscle fibers derived from the tunica muscularis were scattered in the outermost part of the lamina propria. Lymphocytes and numerous blood vessels were also

22

Assiut Vet. Med. J. Vol. 52 No. 111 October 2006

demonstrated (Fig. 4). The thickness of this layer generally increased toward the termination of the abdominal segment. It measured 48.31um in the scrotal segment and reached about 70.31 um in the abdominal segment (Table 1).

The tunica muscularis consisted of muscular bundles arranged in two layers. The inner layer was circular, while the outer layer was longitudinal (Fig. 2, 3 and 4). It formed the thickest layer of wall of the ductus deferens. It measured about 575.42 um, 602.6 um 656.2 um in the scrotal, inguinal and abdominal segments respectively (Table 1).

The tunica serosa consisted of an outer mesothelial epithelium covering sub-serosal connective tissue layer. The latter layer contained numerous small-sized arteries and veins as well as venules, arteriolies, smooth muscle bundles and fibers, lymph vessels, nerve fibers and continued with the mesoductus deferens (Fig 2, 3 and 4.). At its outer part, the collagenous fibers appeared denser forming band-like structure containing numerous longitudinally oriented smooth muscle bundles extending to the mesoductus deferens.

The intramural innervation was generally studied by using PGP 9.5- IR. In the ductus deferens, the larger nerve bundles are situated in the tunica periductular tissue and mesoductus deferens and send finer bundles and solitary fibers to the muscular layer, subeepithelial and the vascular plexuses. These fibers formed two plexuses; one was situated in the muscular layer and the other in the subepithelial connective tissue (Fig. 12).

CHAT-IR and DBH-IR were used to differentiate the types of the nerve fibers. All nerve bundles that reach to the ductus deferens were contained a certain number of ChAT-positive fibers. Inside the ductular wall, the ChAT-positive nerves concentrated mainly in the subepithelial tissue and only very few fibers demonstrated in the muscular coat. The vascular wall was devoid from the cholinergic nerve fibers. DBH-IR allowed to visualization of virtually all fibers in the muscular coat and serosal layer and some nerve fibers in the subepithelial tissue and the vascular plexus. CHAT-IR is specific for the demonstration of cholinergic nerves. DBH-IR visualized the fine nerve fibers in the wall of the ductus deferens better than the general nerve marker (Fig. 14, 15, 16). Table 1: Showing the measurements of the diferent segments of the ductus deferens.

Length in cm Epithelium height in um Sterocilia height in um Lamina propria thickness in um Tunica muscularis thickness in um

Testicular 25.5+2.8 25.91.3 6.6 0.78 65.31 +2.57 675.42 = 8.6

Inguinal | 24.7 +2.4

20.21 +0.9 4.3+0.63 85.23 3.42 702.6 + 10.3

Abdominal

16.5 + 1.9 16.21 +0.67 2.5 +0.3 90.31 +3.95 756.2 + 9.4

23

Assiut Vet. Med. J. Vol. 52 No. 111 October 2006

.

Assiut Vet. Med. J. Vol. 52 No. 111 October 2006

Assiut Vet. Med. J. Vol. 52 No. 111 October 2006

em

Assiut Vet. Med. J. Vol. 52 No. 111 October 2006

LEGENDS

Fig. 1: The non ampullated part of the ductus deferens consisted of

scrotal (sd) inguinal (id) and abdominal (ad) segments. Testis (t), Vascular part of the spermatic cord (V), Tunica vaginalis

(tv). Fig. 2: The scrotal segment run in highly tortuous manner. The deferent

duct (Arrows), Tail of the epididymis (e) Fig. 3: Due to tortuous manner of the ductus deferens, most of sections

were cut obliquely. The lumen (L) packed with spermatozoa surrounding by epithelium (e), lamina propria and muscular coat (tm). The mesoductus deferens is large (m) and expended in the non-attached side of the ductus deferens. The collagenous fibers appeared denser containing numerous smooth muscle bundles in the outer layer of the tunica serosa

and mesoductus deferens (Arrow), the inner layer. H&E x10. Fig. 4: The wall of the non-ampullated part of the ductus deferens

consisted of of epithelium (E), Lamina propria-submucosa (Lp), tunica muscularis ccomposed of inner circular (IM) and outer longitudinal (EM) layers of the smooth muscle fibers, and

tunica serosa (TS). Fig. 5: The The lining epithelium of the ductus deferens shows four

types of cells: principle (C), basal (Arrow), apical (arrow with double head) and granular (Arrow head). (oil im

x1000. Fig 6: The epithelium of the ductus deferens presents PAS-positive

granules in the columnar cells (star) and some in the basal cells

(arrow). (oil immersion lens), x1000. Fig. 7: In the scrotal segment, the luminal surface of the presents two

types of cells; ciliated (Stars) and non-ciliated cells (Arrows).

SEM, x 2000. Fig. 8: The ciliated cells show dense tall and branched steroocilia.

(Arrows) SEM, x 10,000. Fig. 9: The surface of the nonciliated cells shows micro-secrotory

granules. Fig. 10: In the inguinal segment, the epithelial surface presents only

ciliated cells cover with dense tall stereocilia. SEM, x 10,000. Fig. 11: In the abdominal segment, the epithelial surface covered with

microvilli. SEM, x10,000.

27

Assiut Vet. Med. J. Vol. 52 No. 111 October 2006

Fig. 12: The intramural innervation of the ductus deferens composed of

larger nerve bundles (Thick arrows) finer bundles and solitary sībers (Thin arrows)in the periductular tissue, foming dense nerve plexus in the subeepithelial tissue (Arrow heads) and fine

one in the muscular layer. PGP 9.5-IR, x100. Fig. 13: The ChAT-positive nerve fibers (Arrows) concentrated mainly

in the subepithelial tissue in addition to few fibers in the

muscular coat. Fig. 14: DBH-IR shows dense nerve plexus of fine nerve fibers in the

inner (im) and outer (em) muscular layer, in addition to few

fine fibers in the subepithelial tissue (e). DBH-IR, X200. Fig. 15: The vascular wall of the arteries and veins contains DBH

positive fibers. artery(a), vein(v). DBH-IR, x200. Fig. 16: The periductlar tissue contains thick DBH-positive fibers

(arrow) and the large nerve bundle are strongly DBH-IR. The small artery contains also DBH-positive fibers. DBH-IR, X200.

DISCUSSION

In the present study, the non-ampullated part of the ductus deferens of the camel is relatively long (66.2 cm in length average). It divided into three different segments; very tortuous scrotal, slightly tortuous inguinal and straight abdominal. Skidmore (2000) in the same animal reported that the ductus deferens ran tortuous at its initial part and measured 45-50cm in length. In other animals, the ductus deferens ran flexuous in the initial part and then straight in its most part and measured 52.5 - 82, 30 - 32 and 17 - 18 cm in buffaloe, donkey and dog respectively (Pal and Bharadwaj 1985; Salem 2003). In rat, buffaloes and donkey only two different structural segments were mentioned; extra-abdominal and intraabdominal (Hamilton and Cooper, 1978; Pal and Bharadwaj, 1985 and Salem, 2003).

The lumen of the ductus deferens was wider in the scrotal and inguinal segment in comparison with the abdominal one; it frequently contained masses of sperms. These results are disagreed with the results of Stallcup and Griffon (1969) in bull, and Goswami et al. (1994), in camel. The ducts deferens was lined by pseudostratified columnar epithelium and studded with stereocilia as confirmed by scanning electron microscopy. These results are similar to that mentioned by Goswami et al. (1994) in camel, Wrobel and Dellman (1993) in the domestic animals, Schimming (2001) in tufted capchin monkey and

28

Assiut Vet. Med. J. Vol. 52 No. 111 October 2006

Salem (2003) in donkey. In agreement with Pal and Bharadwaj (1985) in buffaloes and Salem (2003) in donkey, the epithelial lining of extra abdominal part (scrotal and inguinal) of ducts deferens was higher than that of the abdominal one. These results disagree with that observed by and Goswami et al. (1994) in the same animal, Hoffer and Greenberg (1978) in guinea pigs, Kennedy and Heidger (1979) in rats

The present study demonstrated that the lining epithelium of the camel duct deferens consisted of four types of the cells; principle, basal apical, dark stained cells. While Goswami et al. (1994) in the same animal, Pal and Bharadwaj (1985) in buffaloes and Salem (2003) in donkey, mentioned that the lining epithelium composed mainly of principle and basal cells.

The principle cells were the major cell type; they possessed rounded or ovoid basally situated nuclei. By scanning electron microscope, the surface of these cells showed dense tall-branched stereocilia in the scrotal segment, dense less tall non-branched stereocilia in the inguninal segment and thick short microvilli in abdominal segment. The stereocilia were microvilli projections of the cel membrane increased greatly the surface area of the duct cells (Niemi 1965). The cytologic characteristics of principle cells, as stereocilia, apical blebblings, primary and secondary lysosomes, seem to indicate that these cells are mainly responsible for the absorptive function (Paniagua et al., 1981). These cells eliminated water from the lumen of the duct during and after ejaculation into the underlying vascular channel in the lamina propria in order to concentrate the sperms (Liebish, 1990; Andonian and Hermo, 1999).

The basal cells were oval or spherical with slightly basophilic cytoplasm and round nuclei. They were probably undifferentiated cells that are capable of differentiation into columnar cells (Paniagua et al., 1981; Nistal et al. 1992). It could be suggested that the principal, apical, and basal cells seem to be related to a cell cycle, in which the basal cells are the stem cells.

The apical cells were also described by Kennedy and Heidger (1979) in rat, Paniagua et al. (1981) & Nistal et al. (1992) in man and Schimming (2001) in tufted capuchin monkey. These cells might represent old or "exhausted" columnar secretory cells that have undergone mitochondrial hyperplasia and loss of organelles involved in glycoprotein synthesis (Nistal et al. 1992). It might be involved in the acidification of the seminal plasma or transport of electrolytes, hydrogen ions and water across the mucosa (Schimming, 2001).

29

Assiut Vet. Med. J. Vol. 52 No. 111 October 2006

The granular cells which found only in the scrotal segment, had wide apical portion, basiophilic granular cytoplasm and round der stained nuclei. By scanning electron microscope, appeared cauliflower in shape and presented secrotory granules and short microvili on their surface. They were also reported in others mammals as rat (Rodríguez & Bustos-Obregón, 1993), in the tufted capuchin monkey (Schimming, 2001). These cells appeared to be involved in microapocrine secretion, in addition they aid in phagocytosis of the sperm remnants or residual bodies (Hamilton and Cooper, 1978). Some authors considered these cells as: holocrine secretory cells (Martan et al., 1964), dead or dying columnar cells (Hoffer, 1976 and Paniagua et al., 1981).

The lining epithelium of the camel ductus deferens contained PAS-positive granules in the columnar and basal cell similar to that mentioned by Stallcup and Roussel (1968) in the bull. While, in buffaloes, the PAS-positive granules demonstrated only in the columnar cells (Pal and Bharadwaj 1985). With Alcian blue stain, the lining epithelium of the ductus deferens showed very weakly or no reaction. This indicated that the epithelium of the camel ductus deferens contained mainly neutral mucopolysaccharids.

The ductus deferens has been classically described as exhibiting a folded mucosa with portions of the epithelium and underlying lamina propria projection into the lumen Goswami et al. (1994) in camel, Wrobel and Dellmann (1993) in the domestic animals, and Salem (2003) in donkey. In the monkey the mucosal folds appeared in the middle and distal segments of duct. Kennedy and Heidger (1979) observed a folded mucosa only in the terminal segment of vas deferens of the rat. In the camel, the ductus defrens showed low or absence mucosal invaginations but not have uniform lumen.

The tunica muscularis of the camel ductus deferens consisted of many muscular bundles arranged into inner thick circular and outer thin longitudinal layers similar to that stated by Goswami et al. (1994). In the donkey, it consisted of inner interwoven and outer longitudinal layers of smooth muscle fibers (Salem, 2003). While in stallions, bulls, boars and carnivores, it consisted of intermingled circular, longitudinal and oblique smooth muscle layers (Trautman and Fiebiger 1957; Wrobel and Dellmann, 1993). In guina pigs and buffaloes, it consisted of inner and outer longitudinal and middle circular layers of smooth muscle fibers (Hoffer and Greenberg, 1978; Pal and baradwaj 1985). The muscular coat of the camel duct deferens increased in thickness distalwards. The increased thickness of the muscular coat in the abdominal segment, may

Assiut Vet. Med. J. Vol. 52 No. 111 October 2006

assisted in maintaining the speed of ejaculated sperms within this part as nearly as that of the initial part (Salem, 2003).

The serosal layer of the ductus deferens of the camel is characterized by presence of smooth muscle bundles distributed in its different parts. In its outer part, the collagenous fibers became denser and formed a band-like structure just under the mesothelium and containing some smooth muscle bundles. The muscle bundles might be derived from the outer layer of the muscular coat. Our results agree with that of other domestic animals reported by Trautman and Fiebiger (1957) Schummer et al. (1979) Pal and Bharadwaj (1985). In donkey, the band like structure in the outer layer of the subserosal layer contained continuous longitudinally oriented layer of the smooth muscle bundles decreased toward the mesoductus deferens but the other part the serosa not contained any muscular bundles (Salem, 2003). The presence of longitudinally oriented smooth muscle bundles within the serosa of the ductus deferens, probably enable it to accommodate itself with the shorting and lengthing action, exerted by contraction of muscular coat during ejaculation. The unique (band-like) arrangement of dense collagenous fibers in the outer layer of the subserosa may indicated that, this layer lay under tension and consequently need more support and great resistance to pulling force (Salem 2003).

The intramural nerve fibers were demonstrated immunohistochemically by using well-established general nerve markers PGP.5-IR (Thompson et al. 1983; Wang et al. 1990). It showed the distribution of the large and fine fibers in the wall of the ductus deferens and the components of the vascular plexuses. The intramural innervation of the camel ductus deferens composed of larger nerve bundles in the serosal and the periductalar tissue, in addition finer bundles and solitary fibers in the ductular wall formed thick subepithelial and fine intramuscular nerve plexuses. The specific innervations were demonstrated by using adrenergic nerve markers (DBH-IR) and cholinergic nerve marker (CHAT-IR) (kujat et al., 1993; Saleh, 2002). DBH-IR allowed to visualization of virtually all fibers in the muscular coat and serosal layer and many nerve fibers in the subepithelial tissue in addition to the vascular plexus and therefore represented postjunctional sympathetic fibers. As in the boar (Kaleczyc et al., 1993), the DBH positive fibers were dense in both the inner and outer muscular layer. Adiverging innervation pattern was observed in the dog and bull ductus deferens where the adrenergic innervation was occurred only in the inner muscular layer, while the other muscle layer contained few fibers

31

Assiut Vet. Med. J. Vol. 52 No. 111 October 2006

(Sjoestrand, 1965, and Kujat et al., 1993). In our result DBH-IR cleared the fine nerve fibers in the wall of the ductus deferens better than the general nerve marker. Similar result has been reported by Saleh (2002) in the innervation of the camel testis and epididymis.

The cholinergic innervation of the camel ductus detferens was demonstrated with CHAT-IR. All nerve bundles that reach to the ductus deferens were contained a certain number of cholinergic fibers. Inside the ductular wall, the cholinergic fibers concentrated mainly in the subepithelial tissue and showed very few fibers in the muscular coat. The vascular wall was devoid from the cholinergic nerve fibers. In the bull vas deferens, the cholinergic fibers formed well develop plexus in the subepithelial tissue and inner muscular wall and few fibers found in the outer muscular layer (Kujat et al. 1993). The adrenergic innervation has important role in the spermatozoa transport during emission of semen from the epididymis through the ductus deferens (Billups et al. 1990 and Kempinas et al. 1998). Restriction of the spermatozoa transport in the vas deferens has been observed after adminstration of sympathetic nerve blocking agents (Leidl, 1965). The adrenergic nerves appeared to stimulate the movement of the ductus deferens, while the cholinergic nerves seem to inhibit the movement of it (Shirai et al. 1973). In this study ChAT-IR and DBH-IR were not able to visual all the nerve that detected by general nerve marker. The non-demonstrated fibers may be peptidergic nerves and need further studies to clear the different types of the nerve fibers in the wall of the ductus deferens.

REFERENCES

Andonian, S. and Hermo, L. (1999): Cell and segment specific

localization of lysosomal and secretory proteins and endocytic receptors in epithelial cells of the cauda epididymidis and vas

deferens of the adult rat. J. Androl, 20:415-29. Bergerson, W.; Amserlgruber, W.; Sinowatz, F. and Bergerson,

M. (1994): Morphological evidence of sperm maturation in the ampulla ductus deferentis of the bull. Cell Tissue Res., 275:

537-41. Billups, K.L.; Tillman, S. and Chang, T.S. (1990): Ablation of the

inferior mesenteric plexus in the rat: alteration of sperm storage

in the epididymis and vas deferens. J Urol 143: 625-629. Duminika, V.A. (1983): Neural apparatus of the wall of the ductus

deferens in the dog. Arkh Anat Gistol Embriol. 84:16-23.

ITI.

Assiut Vet. Med. J. Vol. 52 No. 111 October 2006

Goswamm, S.K.; Sing, Y. and Nagpal, S.K. (1994): Histological and

histochemical studies in ductus deferens of camel: Non

glandular part. Indian J. Animal Sci., 64: 1328-1331. Hamilton, D.W. and Cooper, T.G. (1978): Gross histological variations

along the length of the rat vas deferens. Anat. Rec., 190: 795

810. Hoffer, A.P. (1976): The ultrastructure of the ductus deferens in man.

Biol. Reprod., 14: 425-43. Hoffer, A.P. and Greenberg, J. (1978): The structure of the epididymis,

efferent ductules and ductus deferens of the guinea pig: a light

microscope study. Anat. Rec., 190: 659-78. Kaleczyc, J.; Timmermans, J.P.; Majewski, M.; Lakomy, M. and

Scheuermann, D.W. (1997): Immunohistochemical characteristics of nerve fibres supplying the porcine vas deferens. A colocalisation study. Histochem Cell Biol. 107:

229-4. Kempinas, W.D.; Suarez, J.D.; Roberts, N.L.; Strader, L.F.; Ferrell, J.;

Goldman, J.M.; Narotsky, M.G.; Perreault, S.D.; Evenson, D.P.; Ricker, D.D. and Klinefelter, G.R. (1998): Fertility of rat epididymal sperm after chemically and surgically induced

sympathectomy. Biol Reprod 59: 897-904. Kennedy, S.W. and Heidger, JR. P.M. (1979): Fine structural studies of

the rat vas deferens. Anat. Rec., 194: 159-80. Kujat, R.; Rose, C. and Wrobel, K.H. (1993): The innervation of the

bovine ductus deferens: comparison of a modified acetyl cholinesterase-reaction with immunoreactivities of cholinacetytransferase and panneuronal markers.

Histochemistry 99: 231-239 Leidle (1965): New Ergebniss der Andrologie. Springer, Berlin

Heiderberg-New York. Martan, J.; Risley, P.C. and Hruban, Z. (1964): Holocrine secretory

cells of the human epididymis. Fert. Steril., 15: 180-7. Mirabella, N.; Squillacioti, C.; Varricchio, E.; Genovese, A. and

Paino, G. (2003): Innervation of vas deferens and accessory male genital glands in the water buffalo (Bubalus bubalis). Neurochemical characteristics and relationships to the

reproductive activity. Theriogenology. 59: 1999-2016. Niemi, M. (1965): The fine structure and histochemistry of the epithelial

cells of the rat vas deferens. Acta anat., 60: 207-19.

33

Assiut Vet. Med. J. Vol. 52 No. 111 October 2006

Nistal, M.; Santamaría, L. and Paniagua, R. (1992): The ampulla of the

ductus deferens in man: morphological and ultrastructural

aspects. J. Anat., 180: 97-104. Pal, C. and Bharadwaj, M.B.L. (1985): Anatomical studies on the vas

deferens of buffaloes (Bubalis) at different stages of maturity

I.Non-ampullated part. Indian Veterinary Journal 62: 668-674. Paniagua, R.; Regadera, J.; Nistal, M. and Abaurrea, M.A. (1981):

Histological, histochemical and ultrastructural variations along the length of the human vas deferens before and after puberty.

Acta anat., 111: 190-203. Popovic, N.A.; Mc Leod, D.G. and Borski, A.A. (1973): Ultrastructure of

the human vas deferens. Investig. Urol., 10:266-77. Rodríguez, A. and Bustos-Obregón, E. (1993): Histophysiological and

morphometric studies of the post-natal development of rat vas

deferens. Andrologia, 25:29-37 Saleh, A.M.M. (2002): Morphological studies on the innervation of the

spermatic cord, testis, and epididymis in the camel. Ph. D.

Thesis Assiut (Egypt). Salem, A.O. (2003): Morpological and morphometerical studies on the

non-ampullated part of the ductus deferens of the donkey

(Equus asinus) Assiut Vet. Med. J., 49: 18-42. Schimming, B.C. (2001): Morphological study of the vas deferens in the

tufted capuchin monkey Cebus apella Rev. Chil. Anat. Vol.

19. Schummer, A.; Nickel, R. and Sack, W.O. (1979): The viscera of the

domestic mammals. 2nd Ed. Verlag Paul Parey, Berlin.

Hamburg. Shirai, M.; Sasaki, K. and Rikimaru, A. (1973): A histochemical

investigation of the distribution of adrenergic and cholinergic nerves in the human male genital organs. Tohoku J. Exp. Med.

111: 281-291. Sjoestrand, N.O. (1965): The adrenergic innervation of the vas deferens

and accessory male genital glands. Acta Physiol Sc and

65: 1-82. Skidmore, L. (2000): Anatomy of the Camel Reproductive Tract. In

Recent Advances in Camelid Reproduction, L. Skidmore and G.P. Adams (Eds.), International Veterinary Information Service (www.ivis.org), Ithaca, New York, USA.

34

.

Assiut Vet. Med. J. Vol. 52 No. 111 October 2006

Stallcup, O.T. and Roussel, J.D. (1968): Histochemistry of epithelial

cells of the vas deferens of young Holstein bull. J. Dairy Sci.

18: 996. Stallcupand and Griffon (1969): Histochemical localization of some

enzymes in the ampulla of bull ductus deferens. J.

Reproduation and Fertility 18: 180-81. Thompson, R.J.; Doran, J.F; Jackson, P.; Dhillon, A.P. and Rode, J.

(1983) PGP 9.5 a new marker for vertebrate neurons and

neuroendocrine cells. Brain Res. 278: 224-228. Trautmann, A. and Fiebiger, J. (1957): The male genital organs. In

fundamentals of the histology of domestic animals. Translated and revised from 8" and 9th German Ed., (1949) By Habel, R. E. and Biberstein, E. L. Comstock publishing associates Itheca.

New York. Turner, T.T. (1995): On the epididymis and its role in the development

of the fertile ejaculate. J. Androl., 16: 292-8, 1995. Wang, L.; Hilliges, M.; Jernberg, T.; Wiegleb-Edstrom, D. and

Johansson, O. (1990): Protein gene product 9.5 immunoreactive nerve fibres and cells in human skin. Cell

Tissue Res. 261: 25-33. Wrobel, K.H. and Dellmann, H.D. (1993): Male reproductive system. In

textbook of Veterinary Histology. 4th. Ed. Edited by Dellmann, H.-D. P., Lea & Febiger. Philadelphia.

35