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Revista chilena de anatomía

versão impressa ISSN 0716-9868

Rev. chil. anat. v.20 n.2 Temuco  2002

http://dx.doi.org/10.4067/S0716-98682002000200005 

Rev. Chil. Anat., 20(2):145-149, 2002.

MORPHOLOGIC ASPECTS OF THE ENDOMETRIUM, IN THE ESTRUS
PHASE, OF PINEALECTOMIZED RATS

ASPECTOS MORFOLÓGICOS DEL ENDOMETRIO, EN LA FASE DE ESTRO,
DE RATAS PINEALECTOMIZADAS
 
*Álvaro Aguiar Coelho Teixeira; **Manuel de Jesus Simões; *Joaquim Evêncio Neto & *Valéria Wanderley Teixeira

TEIXEIRA, C. A. A.; SIMÕES, M. J.; EVÊNCIO NETO, J. & WANDERLEY-TEIXEIRA, V. Morphologic aspects of the endometrium, in the estrus phase, of pinealectomized rats. Rev. Chil. Anat., 20(2):145-149, 2002.

SUMMARY: In this paper, it was analyzed morphologically the endometrium of pinealectomized rats, treated or not with melatonin, besides luteinizing hormones levels (LH), stimulating follicle (FSH) and prolactin (PRL). The experimental groups were: I ­ normal rats, during three months; II ­ normal rats, during six months; III ­ pinelalectomized rats evaluated after three months; IV ­ pinealectomized rats treated with melatonin for three months; V ­ pinealectomized rats evaluated after six months; VI ­ pinealectomized rats for three months, and after treated with melatonin for more three months. In the animals from groups III and V, the estrous ciclicity was modified, because beyond the increase in the incidence of the estrus phase, some animals developed a persistent estrus condition. In the other groups the estrous cycle was normal. The hormonal dosage showed that there was an increase in the LH levels in groups III and V, and a small reduction in group IV. There weren't changes between groups VI and II. The levels of FSH and PRL showed themselves reduced in all animals from the pinealectomized group. However, these results didn't show statistically meaningful differences. In groups III, V and VI the endometrium presented a normal aspect except by the presence of several endometrial glands, in most cases, dilated. In the animals that developed persistent estrus, it was observed a very fibrous stroma and rare endometrial glands, some of them presenting non-keratinazed pavement stratified epithelium. The animals in group IV showed a similar endometrium to the control groups. It was concluded that the pinealectomy can draw to the condition of persistent estrus, not producing great variation in the serumal levels of the gonadotrophic hormones, and inducing more effective changes in the glandular epithelium.

KEY WORDS: 1. Melatonin; 2. Endometrium; 3. Pineal; 4. Estrus.

INTRODUCTION

The pinealectomy and its effects on the photoperiodic sensibility have been examined in several mammals species, especially in rodents (Goldman & Darrow, 1983). These effects vary differently in relation to the species. In Turkish and European hamsters (Mesocricetus randti and Cricetus cricetus, respectively), for example, the pinealectomy allied to long photoperiods induces a testicle regression (Carter et al. 1982; Mason-Pévet et al., 1987). But, with the Syrian hamster, the pinealectomy prevents gonadal regression produced by the short photoperiod (Hoffman, 1979 and Reiter, 1980). In rats, the pinealectomy is associated to an increase of the gonadal function (Kinson & Peat, 1971). However, it has been found little available information about morphologic changes in reproductive organs of female pinealectomized rats, mainly in what concerns to the morphology of the endometrium.

Thus, the present study had the objective of collecting basic information about the morphology of the endometrium, in the estrum phase, of adult female rats submitted to pinealectomy, treated or not with melatonin, as well as evaluating the levels of luteinizing hormones (LH), stimulating follicle (FSH) and the prolactin (PRL).

MATERIAL AND METHOD

It was utilized 30 albino rats (Rattus norvegicus albinus), adults, from the EPM-1 Wistar lineage, proceeding from the Biotério Central da Universidade Federal de São Paulo ­ Escola Paulista de Medicina (UNIFESP ­ EPM), in which they were divided at random, in six groups, each one constituted of five female rats:

Group I ­ normal rats, kept in the clear/dark cycle (12/12 hours), during three months, that worked as control:

Group II ­ normal rats, kept in the clear/dark cycle (12/12 hours), during six months, that also worked as control;

Group III ­ rats submitted to the pinealectomy, kept in the clear/dark cycle (12/12 hours), and evaluated after three months;

Group IV - rats submitted to the pinealectomy, kept in the clear/dark cycle (12/12 hours), and treated with melatonin during three months;

Group V - rats submitted to the pinealectomy, kept in the clear/dark cycle (12/12 hours), and evaluated after six months;

Group VI - rats submitted to the pinealectomy, kept in the clear/dark cycle (12/12 hours), during three months and after treated with melatonin for three months more.

The pinealectomy was realized in previously anaesthetized animals with thiopentone sodium, in the dosage of 50mg/kg, by intra peritonial way, and after it was proceeded the technic preconized by Kuszak & Rodin (1997). The melatonin (Sigma Chemical Co., St. Louis, MO., USA) was managed in the water recipient in ambar glasses, in the dosage of 10µg/night/rat (0,4µg/ml), from 19h to 7h. During the day period it was made water restriction (Gibbs & Vriend, 1983; John, et al., 1992 and Trentini et al., 1992). At the end of the experiment, the animals that presented the estrus phase were guillotined in the morning, always at the same time (8:00-10:00hrs.), and the hormonal levels in the measured serum by radioimmunoassay (RIA), using the "NIAMDD-Rat" procedures, to the LH and FSH, and "NIAMD-Rat-Prolactin", to the PRL (Ronnekleiv et al., 1973 and Acuña-Castroviejo et al., 1993), having the values expressed in ng/ml. The average of the hormonal dosages were submitted to the Test of Turkey with 95% of significance. To the morphologic analisys it was taken two fragments from the medium third of each uterine cornu, fixed in the liquid of Boüin, later processed to the inclusion in "paraplast", and colored through hematoxilin-eosine (H.E.).

RESULTS

The animals from groups G IV and G VI presented normal estrous cycles, compared to the control groups. In the animals from groups G III and G V, the estrous ciclicity was changed, due to an increase in the estrus phase incidence, in more then 50% of the cycle (Graphic I), where some animals developed the condition of persistent estrus. The serumal levels of the LH, FSH and PRL hormones, of the animals from the experimental groups presented variations when compared to the respective control groups. However these results have not shown statistically meaningful differences among the animals from these groups (Table I). The morphologic analisys showed that animals from groups G III, G V and G VI the endometrium presented itself covered by cubic or cylindrical simple epithelium, with some leococits and rare mitosis figures. In the endometrial stroma, it was showed up two very distinct regions: one narrower and more cellular, located right under the covering epithelium, and the other, bigger, near the miometrium, with a large concentration of collagen fibers arranged randomly or concentrically around the endometrial glands (Fig. 1). The endometrial glands were in a large amount, mostly dilated presented, where in some, it was observed neutrophiles in their lumen (Fig. 2). In the animals from groups III and V, that developed persistent estrus, the endometrium presented some characteristics as the animals that were in the cycle, but the endometrial glands were scarce, and in some of them the epithelium presented areas of stratification (Figure 3). In the animals from group IV, the endometrium was similar to the control groups.


Graphic I. Frequency of the estrous cycle phase in the studied groups.



Fig. 1. Rat endometrium group III. Notice a great concentric arrangement of collagen fibers around the endometrial glands (arrow). H. E. ± 340 X.


Fig. 2. Rat endometrium group III. Notice endometrial glands, containing infiltered leococitary mainly neutrophiles (arrow). H. E. ± 340 X

.

Fig. 3. Pinealectomized rat endometrium (per-sistent estrus). Notice in the stroma, endometrial gland presenting stratified epithelium (arrow). H. E. ± 340 X

DISCUSSION

The pinealectomy is reported due to meaningfully increase the incidence of the estrus phase (Chu et al., 1964 and Gittes & Chu, 1965), what was observed in the pinealectomized rats without melatonin treatment, during three or six months. The persistent estrus condition developed in some rats is probably associated to high levels of estrogen in these animals. According to Chiórboli (1970), there is an increase in the production of androgens and estrogens in the ovary of the rats in persistent estrus. Besides, the removal of the pineal gland in rats is followed by the increase of circulatory estrogens (Benson & Orts, 1972). That indicates a regulatory activity of the melatonin in the maintenance of the estrous ciclicity in these animals.

The hormonal dosages showed a small increase of the LH in the pinealectomized groups without melatonin treatment, and a decrease of the FSH independent or not of the melatonin administration. However, these results can be considered normal, as they were not statistically meaningful, thus, they are according to the results obtained by Acuña-Castroviejo et al., where the levels of these gonadotrophins returned to normal values 20 days after the pinealectomy. About the prolactin, it was observed a decrease in its levels in all pinealectomized groups, treated or not with melatonin; it suggests that the pinealectomy must exert a negative feedback on the release of this hormone, because according to Relkin et al., 1972, the increase of the pineal activity takes to a decrease in the PIF production (Prolactin Inhibitory Factor) increasing the levels of prolactin in the plasma.

The light microscopy of the endometrium of groups III, V and VI, showed in the superficial epithelium, similar results to the ones described by Simões (1997) in rats in the physiologic estrus, suggesting that the pinealectomy exerts little influence on the endometrium covering epithelium. But, in the appropriate blade it was observed a great deal of concentration of collagen fibers, besides hyperplasic glands. According to Drobnik & Dabrowski (1996) the pinealectomy induces a collagen increase while the treatment with melatonin inhibits this effect. As it was mentioned, the pinealectomy increases the cycling estrogen in rats, what suggests that this hormone is the responsible for the hyperplasia of endometrial glands. Though, even after the oral reposition of melatonin for more three months (group IV), glands hyperplasia and the collagen were not altered. But the treatment right after the pinealectomy (group IV) was enough to avoid such modifications. Concerning to the changes in the glandular epithelium of rats in persistent estrus, it suggests that atypical metaplastic changes occur, in response to the continuing action of the estrogen and absence of melatonin, during a certain period of time, because according to Vrontakis et al. (1993), the biggest estrogenic effect in the uterus is the stimulus of DNA synthesis and the cellular proliferation.

This way, it was concluded that the melatonin contributes to the maintenance of estrous ciclicity in rats; the pinealectomy can take to the persistent estrus condition, however it does not produce great variation in the serumal levels of the gonadotrophic hormones (LH, FSH and prolactin), and induces more effective changes in the glandular epithelium; the oral reposition of melatonin after the pinealectomy prevents the changes in the glandular epithelium.

RESUMEN: En este trabajo evaluamos morfológicamente el endometrio de ratas pinealectomizadas, tratadas o no con melatonina, además fueron medidos los niveles de los hormonas luteinizante (LH), folículo estimulante (FSH) y prolactina (PRL). Los grupos experimentales fueron: I. Ratas normales tratadas con melatonina durante tres meses; II. Ratas normales, con seis meses de tramiento; III. Ratas pinealectomizadas y evaluadas después de tres meses; IV. Ratas pinealectomizadas y tratadas con melatonina por tres meses; V: Ratas pinealectomizadas y evaluadas después de seis meses de tratamiento; VI. Ratas pinealectomizadas después de transcurridos tres meses tratadas con melatonina por otros tres meses. En los animales de los grupos III y V, la ciclicidad estral fue alterada, pues además del aumento en la incidencia de la fase de estro, algunos animales desarrollaron la condición de estro permanente. En los demás grupos el ciclo estral fue normal. Las dosis hormonales mostraron que hubo un aumento en los niveles del LH en los grupos III y V, y una pequeña reducción en el grupo IV. No hubo alteración entre los grupos II y VI . Los niveles del FSH y PRL se mostraron reducidos en todos los animales de los grupos pinealectomizados. Estos resultados no muestran diferencias estadísticamente significativas. En los grupos III, V y VI el endometrio presentó aspecto normal, excepto por la presencia de innumerables glándulas endometriales, en su mayoría dilatadas. En los animales que desarrollaron estro permanente, se observó estroma bastante fibroso y glándulas endometriais escazas, algunas presentaron epitelio estratificado pavimentoso no queratinizado. Los animales del grupo IV, mostraron el endometrio semejante al de los grupos controles. Concluimos que la pinealectomía puede llevar a condición de estro permanente, no pruduciendo grandes variaciones en los niveles séricos de las hormonas gonadotróficas e induciendo modificaciones más efectivas en el epitelio glandular.

PALABRA CLAVE: 1. Melatonina; 2. Endometrio; 3. Pineal; 4. Estro.

REFERENCES

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Gittes, R. F. & Chu, E. W. Reversal of the effect of pinealectomy in female rats by multiple isogeneic pineal transplants. Endocrinology, 77:1061-7, 1965.         [ Links ]

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Dirección para correspondencia:
Prof. MSc. Álvaro Aguiar Coelho Teixeira.
Universidade Federal Rural de Pernambuco.
Departamento de Morfologia e Fisiologia Animal.
Área de Histologia
Rua Dom Manoel de Medeiros s/n ­ Dois Irmãos
CEP: 52.171-900
Recife-PE.
BRAZIL

e-mail: aact@bol.com.br

Recibido : 04-04-2002

Aceptado: 22-05-2002 


* Department of Morphology and Animal Physiology, Universidade Federal Rural de Pernambuco, Recife ­ PE, Brazil.

** Histology of Discipline, Universidade Federal de São Paulo ­ EPM, São Paulo, Brasil.