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International Journal of Morphology

versión On-line ISSN 0717-9502

Int. J. Morphol. v.22 n.2 Temuco  2004

http://dx.doi.org/10.4067/S0717-95022004000200001 

 

Int. J. Morphol., 22(2):109-112, 2004.

 

MORPHOMETRIC ANALYSIS OF THE DEVELOPMENT OF THE OVARIAN FOLLICLES IN PINEALECTOMIZED RATS

ANÁLISIS MORFORMÉTRICO DEL DESARROLLO DE LOS FOLÍCULOS OVÁRICOS EN RATAS PINEALECTOMIZADAS

 

*Katharine Raquel Pereira dos Santos; **Álvaro Aguiar Coelho Teixeira;*Eulâmpio José da Silva Neto; **Valéria Wanderley Teixeira & *Franklin Magliano da Cunha

* Departament of Morphology ­ Center of Sciences of the Health, Universidade Federal da Paraíba, João Pessoa ­ PB, Brazil.
** Department of Morphology and Animal Physiology, Universidade Federal Rural de Pernambuco, Recife ­ PE, Brazil.

Dirección para correspondencia:


SUMMARY: The present study had the objective of analyzing morphometrically the occupied area by the ovarian follicles in ovaries of pinealectomized female rats. To do so, 24 albino female rats (Rattus norvegicus albinus) were used, which were equally divided in the following groups: Group I ­ sham-pinealectomized rats and kept in the light/dark cycle (12/12 hours), for three months; Group II ­ pinealectomized rats and kept in the light/dark cycle (12/12 hours), for three months; Group III ­ pinealectomized rats and kept in the absence of light for three months; Group IV ­ sham-pinealectomized rats and kept in the absence of rats for three months. The results showed that the groups I and IV presented, respectively, average of 19,00 and 25,00, while groups II and III presented the highest average, 28,25 and 27,25, respectively, indicating, this way, a bigger occupied area by the follicles. It was observed a small amount of corpus luteum in the pinealectomized groups. So, we conclude that the pinealectomy can exert, in a certain period of time, some influence in the gonadal function, what might induce the formation of an anovulatory clinical patology and consequently the development of policystic ovaries.

KEY WORDS: 1. Pinealectomy; 2. Ovaries; 3. Morphometric; 4. Estrus.


RESUMEN: El estudio tuvo como objetivo analizar morfométricamente el área ocupada por los folículos ováricos en ovarios de ratas pinealectomizadas. Para ello, 24 ratas albinas (Rattus norvegicus albinus), fueron divididas igualmente en los siguientes grupos: Grupo I - Ratas falsamente pinealectomizadas y mantenidas en el ciclo claro/oscuro (12/12 horas), por tres meses; Grupo II - ratas pinealectomizadas y mantenidas en el ciclo claro/oscuro (12/12 horas), por tres meses; Grupo III - ratas pinealectomizadas y mantenidas en la ausencia de luz durante tres meses; Grupo IV ­ ratas falsamente pinealectomizadas, y mantenidas en la ausencia de luz por tres meses. Los resultados mostraron que los grupos I y IV presentaban, respectivamente, medias de 19,00 y 25,00, mientras que los grupos II y III presentaron las mayores medias, 28,25 y 27,25, respectivamente, indicando mayor área ocupada por los folículos. Fue observada una pequeña cantidad de cuerpos lúteos en los grupos pinealectomizados. De esta manera, concluimos que la pinealectomía puede ejercer, con el tiempo, alguna influencia en la función gonadal, lo que podrá inducir la formación de un cuadro clínico anovulatorio, y consecuentemente, el desarrollo de ovarios policísticos.

PALABRA CLAVE: 1. Pinealectomía; 2. Ovarios; 3. Morfometría; 4. Estro.


 

INTRODUCTION

The pineal gland has the function of photoreceptive in fish, amphibious and reptiles, and as a biological clock in birds (Reiter, 1989). In human beings, its function is not totally explained, on the contrary it works as an endocrine gland that produces melatonin, hormone that modulates the liberation of gonadotrophins (Reiter, 1986).

The melatonin is related to the neuroendocrine regulation in several mammal species (Reiter, 1991). When female rats are under continuous illumination or pinealectomized, the ovarian weight and the incidence of estrum increase (Tamarkin et al.,1985). On the other hand the dark or the melatonin exogen administration in pinealectomized animals or kept in continuous illumination has an opposite effect (Wurtman et al.,1964).

The secretion of melatonin is related to the intensity and time if light exposition. The more luminous stimulus, the less synthesis, due to the light exposition inhibits the sympathetic fibers that innerve the pineal (Fritz & Speroff, 1982). The pinealectomy also eliminates detectable levels of melatonin, breaking the daily secretion pace, causing in rodents permanently active gonads because of the lack of the sign that transmits information about the length of the day (Tamarkin et al., 1985).

On the face of it, the objective of this research was to observe the influence of the pinealectomy in the development of ovarian follicles in female rats, during the estrus phase.

MATERIAL AND METHOD

It was used 24 female albino rats (Rattus norvegicus albinus) proceeding from the Biotery of the Department of Animal Morphology and Physiology form the Universidade Federal Rural of Pernambuco, with 90 days old, weighing nearly 200g. The animals were divided in four groups, as described below:

1.Group I ­ sham-pinealectomized rats (light/dark cycle 12/12 hours, for three months);
2.Group II ­ pinealectomized rats (light/dark cycle 12/12 hours, for three months);
3.Group III ­ pinealectomized rats (absence of light for three months);
4. Group IV ­ sham-pinealectomized rats (absence of light for three months).

Pinealectomy: it was realized with the animals under anesthesia, with atropine diluted in 1ml of distilled water, being administered 0,2ml of the solution. Next, it was used 0,2ml of xylazine and ketamine intraperitoneally. After the trichtomy and asepsis of the dorsal area of the head, it was realized the opening of the skin and subcutaneous tissue until the exposition of the cranial vault, where the periosteum was moved away with a scalpel. Using a micromotor of low rotation, a round fragment was taken out (+/- 4mm of diameter), having as reference the lambda, and put in a 0.9% of physiologic solution. Then it was done the venous sinus ligature and with the help of nippers, the pineal was removed. After that, the osseous fragment was replaced and the skin sutured. The same procedure was realized with groups I and IV (sham-pinealectomized), without removing the pineal.

Absence of light: It was obtained using a 3m2 room, refreshed to 22 C and walled in so that to avoid light access, where the animals from groups III and IV were kept during all experiment.

Morphometric analysis: At the end of the experiment, the female rats that presented the estrum phase, were anesthetized, the ovary collected, cleavaged and fixed in Boüin, for 48hrs. After these procedures, the fragments were processed for the inclusion in paraplast and submitted to the H.E. coloration. So as to determine the percentage of the ovarian area, occupied by the follicles, the laminas were observed in an ocular of 10x, containing in the interior a 25 dots Weibel reticulation. It was counted the dots that fell upon the follicles, with lens of 10x. It was determined four areas in the ovary, being 100 dots per animal, coming down to 400 dots per group.

Statistical analysis: It was realized analysis of comparisons in the average of the follicles among the groups, through the variation analysis to a delineation entirely casual and using the test of Turkey with 95% of significance.

RESULTS

The results showed that groups I and IV presented, respectively, means of 19,00 and 25,00, while groups II and III presented the highest means, 28,25 and 27,25, respectively, indicating a bigger area occupied by the follicles (Graphic I). We observed a small quantity of corpus luteum in the pinealectomized groups (Fig. 1).

Graphic I. Percentagem of the area occupied by the ovarian follicles.


Fig. 1. Rat ovary group II. Notice small quantity of corpus luteum (Cl). H. E. ± 34,28X.

According to the statistical analysis, there was no meaningful difference between the means to the level of 5% of probability, through the test of Turkey (Table I).


Table 1. Means of the area occupied by the ovarian follicles/group.


GROUP

Means of ovarian follicles1

G I

19,00 a

G II

28,25 a

G III

27,25 a

G IV

25,00 a

C.V. (%)

19,73


1 Means followed by the same letter does not differ statistically between them by the test of Turkey (P < 0,05).

DISCUSSION

The morphometric analysis showed that groups II and III presented the highest average area occupied by the follicles, besides being very close, independently of the photoperiodic conditions they were submitted. We must mention yet the presence of a few luteining bodies in these groups. According to Soares et al. (1997; 1998) pinealectomized female rats presented a great deal of follicle concentration and a decrease of corpus luteum. On the other hand, Schlatt et al. (1993) mentioned the presence of ovarian follicles only in female pinealectomized hamsters and kept in short photo periods (8 hours light/16 hours dark) for 27 weeks. These results suggest that the melatonin can perform an important role in the regulation of the ovarian cycle in these animals, where its absence would take a decrease of the ovulation. In fact, Teixeira et al. (1997) observed that ovaries of pinealectomized rats, treated right after with melatonin, presented a decrease of the ovarian follicles and a great deal of luteining bodies. Thus, we concluded that the pinealectomy can exert, as time passes, some influence in the gonadal function, what can induce the formation of an anovulatory clinical patology and consequently the development of policystic ovaries.

REFERENCES

Fritz, M. A. & Speroff, L. The endocrinology of the menstrual cycle: the interaction of folliculogenesis and neuroendocrine mechanisms. Fertility and Sterility, 38, 509-29, 1982.         [ Links ]

Reiter, R. J. The pineal gland: an important link to the environment. Physiol. Sci., 1:202-5, 1986.         [ Links ]

Reiter, R. J. The pineal gland. Endocrinology, 1:240-53, 1989.         [ Links ]

Reiter, R. J. Pineal melatonin cell biology of its synthesis and of its physiological interactions. Society for Endocrinology, 2:151-80, 1991.         [ Links ]

Schlatt, S.; Niklowitz, P.; Hoffmann, K. & Nieschlag, E. Influence of short photoperiods on reproductive organs and estrous cycles of normal and pinealectomized female djungarian hamsters, Phodopus sungorus. Biol. Reprod. 49, 234-50, 1993.         [ Links ]

Soares-Jr, J. M.; Teixeira, A. A. C.; Evêncio-Neto, J.; Simões, M.J. & Baracat, E.C. Aspectos morfológicos do ovário de ratas pinealectomizadas. Anais da SOGESP. IV Jornada Paulista de Obstetrícia e Ginecologia, 20, 1997.         [ Links ]

Soares-Jr, J. M.; Simões, M. J.; Rodrigues de Lima, G. & Baracat, E.C. Aspectos morfológicos das alterações ovarianas recentes de ratas pinealectomizadas. Arquivos do Instituto Biológico. v. 65(Supl):101, 1998.         [ Links ]

Tamarkin, L.; Baird, C. J. & Almeida, O. F. X. Melatonin: A Coordinating Signal for Mammalian Reproduction? Science, 227:714-20, 1985.         [ Links ]

Teixeira, A. A. C.; Evêncio-Neto, J.; Simões, M. J.; Soares-Jr, J. M. & Baracat, E.C. - Aspectos morfológicos do ovário de ratas pinealectomizadas e tratadas com melatonina. Arquivos do Instituto Biológico, 64 (Supl): 65, 1997.         [ Links ]

Wurtman, R. J.; Axelrod, J.; Chu, E. W. & Fischer, J. E. - Mediation of some effects of illumination on the rat estrous cycle by sympathetic nervous system. Endocrinology, 75: 266-72, 1964.         [ Links ]

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

Received : 19-01-2004
Accepted: 05-03-2004

 

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