Scanning electron microscopic study of hedgehog uteri

Scanning electron microscopic study of hedgehog uteri

TISSUE & CELL 1984 16 (2) 195-202 @ 1984 Longman Group Ltd D. LESCOAT”, M. SABOUREAUt, SCANNING ELECTRON OF HEDGEHOG UTERI Key words: Hedgehog, J...

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TISSUE & CELL 1984 16 (2) 195-202 @ 1984 Longman Group Ltd

D. LESCOAT”,

M. SABOUREAUt,

SCANNING ELECTRON OF HEDGEHOG UTERI Key words:

Hedgehog,

J. SEGALEN”

and Y. CHAMBON*

MICROSCOPIC

endometrium,

microvilli.

ciliated

cells, scanning

STUDY ultrastructure

ABSTRACT. Ultrastructural studies of hedgehog uteri (Erinaceus europaeus L.) have been made using animals in anestrus, in estrus and in estrus after sojourn of a week with a male. In estrus and anestrus the uterine epithelium is homogeneous, regularly interrupted by orifices of glands. It is composed of mlcrovillous cells only. Microvilli decrease m number and length in anestrus. A new type of cell, a ciliated cell, appears after copulation. Probable correlation of ultrastructural aspects of endometrium with hormonal situation is discussed.

Introduction Most recent research on ultrastructural characteristics of mammalian uterus has been focused upon a select group of laboratory and farm animals. The results may not be representative of events occurring in other mammals and especially in the insectivora which are generally conceded to be the most primitive order of living eutherian mammals. Studies of primitive species may be valuable for comparison with common experimental species and perhaps in evaluating the relative phylogenetic significance of morphological characters. In some insectivora, particular small uterine epithelial pits, sometimes real embryonic implantation chambers have been described (Van der Horst and Gillman, 1942; Feremutsch, 1948) and they will appear just after fertilization in the hedgehog (Hubrecht, 1888), Among the insectivora which have been the subject of investigation (Van der Horst and Gillman, 1942; Feremutsch, 1948; King et al., 1978; Prasad et al., 1979), little

research has been devoted to the uterus of the female hedgehog, a hibernating mammal (Hubrecht, 1888, 1889; Deanesly, 1934; Girodetab, 1967; Walin eta!., 1968; Balboni, 1969). To our knowledge, no research has been published using the scanning electron microscope (SEM) to investigate hedgehog uterine ultrastructure. The SEM approach permits rapid visualization of larger surfaces of tissues and the samples can be rotated in the specimen chamber, considerably facilitating observation. The present account reports SEM ultrastructural aspects of hedgehog endometrium under different conditions, at various times of the year, and is designed as a contribution to the comparative morphology of the genital tract. Materials and Methods Animals The female adult hedgehogs, Erinacew europaeus L., used in this study were caught near the ChizC Forest (Department Deux S&vres, Western Central France: latitude 46”09’N; longitude O”‘24’W). They were maintained outdoors under natural climatic conditions with free access to food throughout the year. They were grouped as follows: group 1, three animals in a waking state were killed in April; group 2, four animals in a waking state were sacrificed in April, after

‘Laboratoire d’Histologie et d’Embryologie. U.E.R. Medicales et Pharmaceutiques, UniversitC de Rennes; 2. avenue du Professeur L. Bernard, 35043 Rennes. France. tCentre d’Etudes Biologiques des Animaux Sauvages, C.N.R.S.. Villiers-en-Bois, 79360 Beauvoir-sur-Niort. France. Received 14 March 1983. Revised 5 March 1984. 195

LESCOAT,

196

having been placed in the presence of a male hedgehog for 1 week; group 3, four animals in a hibernating state were killed in December.

The uteri resemble those described in group 1 control hedgehogs but some differences have appeared. The luminal surface is similarly homogeneous, flat and without mucosa folds (Fig. 7). Glands are as numerous as in group 1

Group 1 The uterine lumen generally forms a wide slit (Fig. 1). The luminal surface of the endomet-

estrus hedgehog

uteri.

lumen (arrow) forms a wide mesometrial

Fig. 2. Luminal

surface is homogeneous

Fig. 3. Illustrating Fig. 4. Glands

depth of glands.

(G) with secretory

Fig. 5. Apical cell surfaces

AND CHAMBON

Group 2

Observations

Fig. 1, Uterine slit. X30.

SEGALEN

rium is relatively homogeneous; it is always flat, without any fold (Fig. 2). The surface is regularly interrupted by orifices of gland pits. Glands are numerous (139+7 glands/mm*) and well developed (Fig. 3); only some show signs of secretory activity (Fig. 4). In every sample, the luminal epithelium is composed of only one population of cells: microvillous cells with polygonal, flat or moderately convex apical surfaces of equal sizes (Fig. 5). Flat apices are well delineated by microvilli. Cell apices are sparsely or densely covered with microvilli irregular in length but often long and of dense appearance (Fig. 6). The appearance of uterine epithelium is similar in meso- and antimesometrial sides.

Methods Tissue was obtained by a mid-line incision performed under fluothan anaesthesia. Fixation was initiated in situ by perfusion via a carotid artery with 2.5% glutaraldehyde in O.lM phosphate buffer (pH=7.2). Endometrial tissues obtained from the meso- and antimesometrial sides were immediately immersed in the same fixative overnight. They were dehydrated in ethanol, then in acetone and dried in a Polaron E3000 critical point drying apparatus. The samples were coated with gold and examined in a Jeol JSM S35 scanning electron microscope.

Figs. 1-6. Control

SABOUREAU.

(Mtantimesometrial

with regularly

distributed

(AM) directed

glands (G). x300.

x2100. material.

x2800.

(A) are of similar

Fig. 6. Cell apices (A) are covered

area. x2800.

with long densely packed microvilli.

x8500.

Figs. 7-12. Uteri of estrus hedgehogs

after 1 week with a male.

Fig. 7. Luminal

mucosa folds, showing gland orifaces (G).

surface,

flat, without

Fig. 8. Spermatozoa

(arrow)

Fig. 9. Spermatoza

penetrating

Fig. 10. Lummal

epithelium

lying on epithelial

Fig. 12. Microvilli

the gland openings

shows microvillous

Fig. 1 I. Cell apices are polygonal are irregular

surface.

and flattened. in length.

x8500.

x 1400.

(arrow).

x850.

(M) and ciliated x2800.

(C) cells. x5600.

x40.

ULTRASTRUCTURE

OF HEDGEHOG

UTERI

(143&11 glands/mm2), are regularly distributed and are apparently not secreting. Uteri of two animals contain spermatozoa providing evidence of copulation. They are lying on the epithelial surface or, often, penetrating the gland openings (Figs. 8, 9). The luminal epithelium here shows two kinds of cell:microvillous cells and ciliated cells (Fig. 10). Microvillous cells are much more numerous. Their apices do not differ from each other in size. They are polygonal and flat or moderately convex (Fig. 1 I). Flat apices are well delineated by a border of microvilli. Microvilli are irregular in length and distribution. They are often long and abundant (Figs. 11, 12). Ciliated cells are distributed in small groups or singly among microvillous cells; they constitute 15% of the total number of cells but their distribution is very irregular, varying from 1 to 50% of the total number of cells from one place to another. Some microvilli are also present on ciliated cells (Fig. 10). SEM ultrastructural appearance is the same in mesoand antimesometrial sides of uteri. Group 3 The uterine lumen also forms a slit but is narrower than in the two preceding groups and uteri show some atrophy (Fig. 13). The luminal epithelial surface is homogeneous. It is very flat without any folding of mucosa (Fig. 14). Glands are less numerous than in groups 1 and 2 (91+7 glands/mm2). Only a few show sign of secretory activity (Figs. 15, 16). Their openings are regularly distributed. Uterine epithelium shows only microvillous cells. Cell apices of equal size protrude very slightly into the lumen or are flat (Figs. 17, 18). They are well delineated by a fringe of microvilli giving them a polygonal appearance. Microvilli numbers are different from one cell to another (Figs. 17, 18). Their distribution is also irregular on individual cells where some microvilli have drawn nearer to each other giving the apex a particular appearance (Fig. 17). They are, overall, much shorter and less abundant than in groups 1 and 2 (Fig. 18). Epithelial surfaces show identical characteristics in meso- and antimesometrial endometrium. Discussion SEM has been successfully rize the surface appearance

used to characteof the lumen of

199

the adult hedgehog uterus, and, to our knowledge, this is the first report upon the uterine ultrastructure of this animal. The present paper shows, firstly, that SEM uterine appearances are different in hedgehogs active in April (in estrus) and in hedgehogs hibernating in December (in anestrus) and, secondly, that morphological changes occur after copulation. In estrus animals, the uterine cavity is wider and the number of glands/mm2 is significantly higher than in anestrus (P
ULTRASTRUCTURE

OF HEDGEHOG UTERI

were made on uteri of hedgehog, Erinaceus europaeus L. The morphological differences occurring in uterine epithelia were studied in hedgehogs awake in April (in estrus), in others awake in April (in estrus) but which have sojourned with a male for a week and in hedgehogs hibernating in December (in anestrus). In estrus and anestrus, uteri showed only microvillous cells with flat or slightly protruding apices covered with long microvilli in estrus and by very short and less numerous microvilli in anestrus. The number of glands is significantly lower in anestrus. After copulation, luminal epithelium resembled that of estrus and gland numbers were comparable. It was composed of abundant microvillous cells with long microvilii and of some ciliated cells. To our knowledge, ciliated cells had never been described in uteri of hedgehogs. Correlation of ultrastructural characteristics of endometrium with the hormonal situation was discussed.

Epithelium is composed of microvillous cells identical to those seen in estrus and of a new type of cell, a ciliated cell, absent from uteri of estrus unmated or anestrus animals. Ciliated cells are a normal component of uterine epithelium in certain mammals such 1962; Motta and as rabbits (Larsen, Andrews, 1976; Segalen et al., 1982) and humans (Dallenbach-Hellweg, 1971; More and Masterton, 1976) but their existence has never been mentioned in reports concerning rats (Tachi et al., 1970), mouse (Nilsson, 1958) or sheeps (Flechon et al., 1978). To our knowledge, the presence of ciliated cells has never been reported in hedgehog uterus. In mammals where they are present, it has been shown that their appearance or disappearance is dependent on the hormonal situation, and particularly on stimulatory or inhibitory effects due to variations in plasma progesterone and estradiol concentrations. Here, their presence only in the uteri of hedgehogs which have mated surely indicate that the hormonal state has changed after copulation. It may indicate that fertilization, which can only be regarded as a possibility because there are no indications of true pregnancy and because of the existence of sterile cycles in hedgehogs (Deanesly, 1934) has really occurred.

Acknowledgements

The authors thank M. T. Lavault and D. Quernee for technical assistance, R. Primault for photographical assistance and S. Lorre for typing the manuscript.

Summary Scanning electron microscopic observations

Figs. 1318. Anestrus hedgehogs uteri Fig. 13. Uterine lumen (arrow) forms a narrow slit. x30. Fig. 14. Epithelial surface is flattened and includes glands (G). x300 Fig. 15. Cell apices are well delineated by a fringe of microvilli. x2800. Fig. 16. Gland showing little secretory activity. ~5600 Fig. 17. Some cell apices (arrow) are marked by characteristic X5600.

Fig. 18. Microvilli are short and irregularly distributed. 14

x8500.

arrangement

of microvilli.

202

LESCOAT,

SABOUREAU,

SEGALEN

AND CHAMBON

References Balboni, G. C. 1969. Observations on the ovary of the hedgehog (Erinaceuc europaeus L.) in physiological and experimental hibernation and awakening. Acta Amt., Suppl. 56, 60-71. Baumeister, T. 1913. Die entwickhmgsvorglnge am keime des Igels (Erinoce~u europaeus L.) van seinem Uebcrtritt m den uterus bis zur ausbildung des mesoderms. Z. Wis. Zoo/. 105, I-86. Dallenbach-Hellweg, C. 1971. Histopathology of the Endometrium, pp. 125-135. Springer, Berlin. Deanesly, R. 1934. The reproductive processes of certain mammals. Part VI. The reproductive cycle of the female hedgehog. Phil. Tram. R. Sot. Land., B, 223, 239-276. Feremutsch, K. 1948. Der praegravide genitaltrakt und dte praeimplantation. Rev. Suivse de Zool.. 55, 567-622. Flechon, J. E., Guillemot, M. and Wintenberger-Torres S. 1978. Etude ultrastructurale du blastocyste ct dc I’implantation chez la brebis. In L’impkmtation de l’oeuf(eds. F. Du Mesnil Du Buisson. A. Psychoyos and K. Thomas), pp. 107-120. Masson, Paris. Girod, C., Dubois, P. and Cure, M. 1967. Recherches sur lea correlations hypophyso-genitales cher la tcmcllc dc Herisson (Erinacercs europaetu L.). Annls Endocr., 28, 581-610. Hubrecht, A. A. W. 1888. Keimblltterbildung und placentation des Igels. Arm. Am.. 3, SIO-515. Hubrecht, A. A. W. 1889. Studies in mammalian embryology. Q. Jl. Microsc. Sci.. 30, 2X3-404. Kanagawa, H., Hafez, E. S. E., Pitchford, W. C., Baechler, C. A. and Barnhart. M. I. 1972. Surface patterns in the reproductive tracts of the rabbit observed by scanning electron microscopy, Amt. Rec., 174, 205-226. King, B. F., Enders, A. C. and Wimsatt, W. A. 1978. The annular hematoma of the shrew yolk-sac placenta. Am. J. Amt., 152, 45-58. Larsen, J. F. 1962. Electron microscopy of the uterine epithelium in the rabbit, J. Cell Biol., 14, 49-64. More, I. A. R. and Masterton, R. G. 1976. The role of estrogen in the control of ciliated cells in the human cndometrium. 1. Reprod. Fur., 47, 19-24. Motta, P. M. and Andrew, P. M. 1976. Scannmg electron microscopy of the endometrium during the secretory phase. J. Amt., 122, 315-322. Nilsson, 0. 1958. Ultrastructure of mouse uterine surface epithelium under different cstrogenic influences. J. Ulrrast. Res., 1, 375-396. Prasad, M. R. N., Mossman, H. W. and Scott, G. L. 1979. Morphogenesis of the fetal membranes of an American mole. Scabpus aquatics. Am J. Amt., 155, 31-68. Psychoyos, A. 1978. Mecanismes de I’ovoimplantation. In L’implantation de [‘oeuf(eds. F. Du Mesnil Du Butsson. A. Psychoyos, and K. Thomas), pp. 21-26. Masson, Paris. Saboureau, M. and Dutourne, B. 1981. The reproductive cycle in the male hedgehog (Erinaceur europaew L.): a study ot endocrine and exocrine testicular functions. Reprod. Nutr. DCvelop., 21, 109- 126. Segalen, J., Lescoat, D. and Chambon, Y. 1982. Ultrastructural aspects of uterine secretion during the establishment of pregnancy in the rabbit: role of the egg. J. Amt., 135, 281-289. Smith. A. F. and Wilson, I. B. 1974. Cell interaction at maternal embryonic interface during implantation in the mouse. Ceil Tim. Res., 152, 525-542. Tachi, S., Tachi, C. and Lindner, H. R. 1970. Ultrastructural features of blastocyst attachment and trophoblastic invasion in the rat. J. Reprod. FM., 21, 37-56. Van der Horst, C. J. and Gillman, J. 1942. Pre-implantation phenomena in the uterus of Elephantuhu. S. Afr. J. Med. Sci., 7, 47-71. Walin. T., Soivio, A. and Kristoffersson, R. 1968. Histological changes in the reproductive system of female hcdgehoga during the hibernation season. Ann. Zool. Fem.. 5, 227-229.