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vol.15 número2ASPECTOS MORFOLOGICOS E HISTOQUIMICOS DEL EPITELIO LUMINAL DEL OVIDUCTO DE LA PATA SILVESTRE (Cairina moschata, LINNAEUS, 1785) DURANTE LOS PERIODOS DE POSTURA Y NO POSTURAEFECTOS DEL ETANOL SOBRE LA EPIDERMIS DE FETOS DE RATA. ESTUDIO MORFOLOGICO Y MORFOMETRICO índice de autoresíndice de materiabúsqueda de artículos
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Revista chilena de anatomía

versión impresa ISSN 0716-9868

Rev. chil. anat. v.15 n.2 Temuco  1997

http://dx.doi.org/10.4067/S0716-98681997000200008 

ULTRASTRUCTURAL STUDY OF THE COAGULATING GLAND EPITHELIUM
OF THE RAT (Rattus norvegicus) AFTER VASECTOMY

ESTUDIO ULTRAESTRUCTURAL DEL EPITELIO DE LA GLANDULA DE
COAGULACION DE RATA (Rattus norvegicus) DESPUES DE LA VASECTOMIA

 

*
**
**
***
S. R. Melo
W. Mello Jr.
P. J. Garcia
V. H. A. Cagnon

 

* Department of Morphophysiological Science - State University of Maringá, Paraná - Brasil
** Department of Anatomy, Biosciences Institute, State University of São Paulo, Botucatu - Brasil
*** Department of Anatomy, Biology Institute, State University of Campinas, São Paulo - Brasil

SUMMARY: Morphological alterations of the coagulating gland of vasectomized rats were observed using light and transmission electron microscopy. The results demonstrated atrophy of the height of the secretory epithelium. Ultrastructural observations showed atrophy of the rough endoplasmic reticulum cisternae.

KEY WORDS: 1. Coagulating gland; 2. Rat; 3. Ultrastructure; 4. Vasectomy

  INTRODUCTION

Among the different procedures for the control of fertility in humans, vasectomy is becoming more popular, especially due to the simplicity of the surgical method (CASTRO,1988). The effects of vasectomy on the reproductive organs in laboratory animals have been investigated by several authors (ALEXANDER,1973; NEAVES,1975 and BARRAT & COHEN,1987). Nevertheless, few studies are found about the coagulating gland after vasectomy. According to SJOSTRAND (1965) the coagulating gland is observed in animals such as rabbit, rat, hamster, guineapig and monkey, but it is absent in man. Its secretion, together with that of the seminal gland, coagule and become a copulatory blockade on the female vagina. This blockade is related to the efficacy of fertilization for it prevents the sperm from leaving the female reproductive tract after coitus (CARBALLADA & ESPONDA, 1992). It has been demonstrated that a homologous relationship exists between the middle lobe of the human prostate and the coagulating gland of the mouse (PRICE, 1963). In spite of vasectomy being used as a contraceptive method, doubts and divergences still persist concerning the techniques employed and their consequences. Thus, the purpose of this study was to verify the possible morphological alterations in the secretory epithelium of the coagulating gland in rats after vasectomy.

MATERIAL AND METHOD

Thirty-two male albine rats (Rattus norvegicus) aging two months and with average weight 250 grams were used. Animals received solid diet (Purina) and water both "ad libitum". The rats were equally divided in two groups: vasectomized and simulated. All the animals were subject to surgery, which was carried out through abdominal midsagital incision under asseptic conditions. On the animals from the vasectomized group both deferens ducts (left and right) were clamped in two places 0.5 cm apart. Both were then doubly sectioned, resulting in a free segment of duct that was discarded. For sutures catgut lines 4-0 for peritoneum and muscle and 3-0 for skin were used. On the rats from the simulated group the procedure was essentially the same, except that the ducts were neither sectioned nor clamped. After periods of 60, 120, 180 and 360 days after surgery, four animals of each group were anesthetized with ethylic ether and subject to perfusion with glutaraldehyde 2.5%. The coagulating gland was removed and split into two portions. The first was subject to histological routine for observation under light microscope, where paraffin sections (7mm) of this tissue were processed and stained with hematoxilineosin. Morphometry was carried out through graded lens (10mm/100) coupled to Olympus microscope and 100x objective.

The parameters used for the measures were the lamina propria and the luminal surface of the epithelium.The second portion was immersed in glutaraldehyde 2.5% in phosphate buffer (pH 7.2) and processed according to the conventional technique for transmission electron microscopy. The ultra-fine sections, were stained with uranile acetate (WATSON,1958) and lead citrate (REYNOLDS,1963). Electronmicrographs were obtained with electronic microscope Philips EM 301.

STATISTICAL ANALYSIS : The statistical study of the mean height of the secretory epithelium of the coagulating gland (mm) was carried out through variance analysis for the factorial scheme 2 x 4 (two groups and four periods) in totally randomized experiments. The technique was complemented with Tukey's multiple comparisons test. Statistical conclusions were based on a 5% level of significance.

RESULTS

Light Microscopy: The secretory epithelium of the coagulating gland of simulated rats is columnar simple. Epithelial cells show elongated nucleus in central position in the cytoplasm (Fig. 1A). Vasectomized animals show diminished height of the epithelium in all periods (Tablea I). Epithelial cells exhibit cubic shape, with round nucleus in basal position on the cytoplasm (Fig. 1B)

 

Table I.- Average height of the secretory epithelium of the coagulating gland in the groups studied (µm).


 
Period after surgery (day)
Groups

60

120

180

360

Simulated

16.01 b

16.33 b

15.84 a

16.02 b

Vasectomized

13.18 a

13.07 a

13.74 a

10.60 a

C.V.= 12.17% SMD (5%) = 2.55        

Means followed by the letter (a-b) different according to Tukey multiple range test.

Electron Microscopy: The columnar epithelial cells of the simulated group exhibit irregularlyshaped nucleus, with evident nucleoli, that occupies a central position on the cytoplasm. On the basal region the endoplasmic reticulum (GER) forms dilated cisternae, and on the apical region mitochondria, Golgi apparatus and surface microvilli are observed (Fig. 2A,B,C). On the vasectomized group the epithelial cells generally exhibit cubic form with round nucleus in basal position (Fig. 3A). On the basal region the GER are atrophic (Fig. 3A,C), and on the apical region mitochondria, Golgi apparatus and surface microvilli are also observed (Fig. 3C). Only on the period of 360 days it is verified the presence of dense bodies on the cytoplasm (Fig.3B).



Fig. 1 Photomicrographs of the coagulating gland epithelium of rats 360 days after surgery. Observe in A: simulated tissue with columnar epithelium, centrally positioned nucleus stained with H.E 1,500X; in B: vasectomized tissue with atrophic epithelium, cubic cells, round basal nucleus. HE 1.500 X.





Fig. 2 Electronmicro-graphs of the epithelium of the coagulating gland of simulated group at 60 (A), 120 (B) and 180 (C) days after surgery. In A: observe columnar epi-thelium with central nucleus (N), lumen (L), and basal membrane (arrow), magnification 5,200X; in B: basal region with dilated GER cisternae, basal membrane (arrow), collagen fibers (col), magni-fication 11.500X; and in C: apical region contai-ning mitochondria (M), Golgi apparatus (G) and microvilli on the luminal surface (arrow), magni-fication 16,800X.

 

DISCUSSION

The observed alterations on the coagulating gland of vasectomized animals were similar to those found in castrated and vasectomized animals with hormonal imbalance. LUNG & CUNHA (1981) showed that the morphogenesis of the coagulating gland and seminal gland of mice are extremely sensitive to alterations on the normal conditions of androgenic hormones. GUYTAN et al. (1986) verified that rats treated with estrogen showed decrease on the volume of the glandular epithelium and increase on the volume of fibrousmuscular stroma in the ventral prostate and seminal vesicle. KIPLESUND et al., (1988)examining rats a week after castration, verified a sharp decrease on the height of the coagulating gland epithelium, showing that the coagulating gland exhibits androgenic dependence. RAJALAKSHMI & RAMAKRISHNAM (1991) studied castrated monkeys, which showed a significant decrease on the weight of the semen. They administered androgens, restabishing testosterone levels, and demonstrating once more the importance of hormonal action on the acessory sexual glands. GERERHAAS et al., (1991) reported the hypertrophy of Leydig cells, by means of morphometric analysis, in rats subjected to bilateral vasectomy.

The ultrastructural alterations found in the present study were also similar to those of castrated rats. GETERHAAS et al., (1991) verified that the GER resembles dilated vesicles in the Leydig cells of vasectomized rats, while it can be described as flattened tubules with few vesicles in control animals. Alterations concerning mitochondria and Golgi apparatus were not found. The authors atributted the alterations to the increase on LH levels. DAHL & TVETER (1973) described cytoplasmic decrease and generalized atrophy of the GER cisternae in the seminal vesicle of castrated rats. On the coagulating gland in the same condition it was observed that the GER cisternae on the basal regions persisted in many cells. BRANDES et al., (1962) reported a sharp decrease on the GER cisternae of epithelial cells from the coagulating gland of castrated adult rats. TONER & BAILLIE (1966) observed, after castration, that the volume of epithelial cells of the seminal vesicle of mice was reduced, and that the GER and the Golgi apparatus became disorganized. Secretory granules disappeared, leaving empty vacuoles and an increase on the number of dense bodies. DAHL & KJAERHEIM (1973) described alterations on the organelles of epithelial cells from the ventral, dorsal and lateral lobes of the prostate after castration. The major changes were decrease of the Golgi apparatus and GER, presence of large autophagic vacuoles and lipid droplets on the basal cytoplasm and increase on the number of dense bodies. DAHL & TVETER (1973) described that upon castration, due to the androgenic stimulus supression, the alterations were evident on the seminal vesicle and coagulating gland of rats. Both organs exhibited GER atrophy, cytoplasm decrease, dense bodies formation and autophagic vacuoles

We conclude with this study that the bilateral vasectomy induces the atrophy of the secretory epithelium of the coagulating gland in rats; the morphological response of the epithelium is a decrease on the GER cisternae, specially on the basal region.







Fig.3 - Electron-micrographs of the epithelium of the coagulating gland of vasectomized group at 180(A), and 360(B and C) days after surgery. Observe in A: basal nucleus (N), atrophic GER cisternae on the basal regions, basal membrane (arrow), magnification 10,400X; in B: nucleus (N) and presence of dense bodies (cd), magnification 13,200X; in C: epithelial cell with basal nucleus (N) and atrophic GER cisternae on the basal region, basal membrane (arrow), mag-nification 13,200X

RESUMEN: Las alteraciones morfológicas de la glándula de coagulación de las ratas vasectomizadas, fueron observadas utilizando microscopía de luz y microscopía electrónica de transmisión. Los resultados demostraron disminución en la altura del epitelio secretor. Observaciones ultraestructurales mostraron atrofia de las cisternas del retículo endoplasmático granular.

PALABRAS CLAVE: 1. Glándula de coagulación; 2. Rata; 3. Ultra-estrutura; 4.Vasectomía.

REFERENCES

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BARRAT, C. L. R. & COHEN, J. Quantitation of sperm disposal and phagocytic cells in the tract of short and long term vasectomized mice. J. Reprod. Fertil., 81:377-84, 1987.         [ Links ]

BRANDES, D., GROTH, D. P. & GYORKEY, F. Occurrence of lysosome in the prostatic epithelium of castrate rats. Exp. Cell. Res., 28:61-8, 1962.         [ Links ]

CARBALLADA ,R., ESPONDA. P. Role of fluid from seminal vesicles and coagulating glands in sperm transport into the uterus and fertility in rats. J. Reprod. Fertil., 95:639-48, 1992.         [ Links ]

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DAHL, E., TVETER, K.J. The ultrastructure of the accessory sex organs of the male rat. Z.Zellforsch., 144:179-89, 1973.         [ Links ]

GETERHAAS, B.; BORNSTEIN, S. R. & JARRY, H. Morphological and hormonal changes following vasectomy in rats, suggesting a functional role for Leydigcell associated macrophages. Horm. Metab. Res., 23: 373-8, 1991.         [ Links ]

GUYTAN, F., BELLIDO, C., AGUILAR, R. LUCENA, M.C. Morphometric analysis of the rat ventral prostate and seminal vesicles during prepubertal development: effects of neonatal treatment with estrogen. Biol. Reprod., 35:219-25, 1986.         [ Links ]

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LUNG, B. & CUNHA, G. R. Development of seminal vesicles and coagulating glands in neonatal mice. I. The morphogenetic effects of various hormonal conditions. Anat. Rec., 199:73-88, 1981.         [ Links ]

NEAVES, W. B. Biological aspects of vasectomy. In: Handbook of Physiology Endocrinology. Washington: American Physiological Society. p.383-404, 1975.         [ Links ]

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RAJALAKSHMI, M. & RAMAKRISHNAM, P.R. Evaluation of the ability of a new long acting androgen ester to maintain accessory gland function in castrated Rhesus monkey. Contraception, 43:83 - 89, 1991.         [ Links ]

REYNOLDS , E. S. The use of lead citrate at high pH as an electron opaque stain in electron microscopy. J. Cell. Biol., 19:208, 1963.         [ Links ]

SJOSTRAND, N. O. The adrenergic innervation of the vas deferens and the accessory male genital gland. Acta. Physiol. Scand.,169-75, 1965.         [ Links ]

TONER, P. G. &BAILLIE, A. H. Biochemical, histochemical and ultrastructural changes in the mouse seminal vesicle after castration. J. Anat., 100:173-88, 1966.         [ Links ]

WATSON, M. L. Staining of tissue sections for electron microscopy with heavy metals. J. Biophys. Biochem. Cytol., 4:475, 1958.         [ Links ]

Dirección para correspondencia:
Prof. Silvana Regina de Melo
Department of Morphophysiological Sciences
State University of Maringá
Av. Colombo 5790
87020900
Maringá-Paraná
BRAZIL

Recibido : 17-05-1997
Aceptado: 21-10-1997

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