[ 423 ]

THE DEVELOPMENT OF THE RABBIT VAGINA

By EDNA B. CARRDepartment of Embryology, University College, London

Although it is agreed that the upper part of the rabbit's vagina develops from theMullerian canals, three conflicting views exist concerning the composition of thelower part. Langenbacher (1882) and Mihalkovics (1885) considered that the lowerpart, like the upper, is composed of Mullerian tissue. Later Baxter (1933) concludedthat this lower part is formed from the epithelium of the persistent bases of theWolffian ducts. Finally, Jost (1943a, b, 1947) considered that it is derived from tissueof the urogenital sinus, the Wolffian ducts completely disappearing. In view of thesediscrepant results, it seemed useful to undertake a new description of the develop-ment of the rabbit vagina, based on a full series of specimens prepared for histo-logical analysis.

MATERIALS AND METHOD

The foetuses used in this work were grouped according to the age since mating, andnot according to size, which is an unreliable index of age (Waterman, 1942; Jost,1947). Foetuses were obtained at fifteen stages, mostly at daily intervals between19 and 32 days post-coitum. At least two specimens of each age were examined.They were placed in a warm saline bath, the abdominal wall and pelvic girdle wereopened, Bouin's fixative was injected into the bladder and rectum, and the wholefoetus was then immersed in Bouin. This method always ensured satisfactoryfixation. The reproductive organs were dissected out, embedded in paraffin,sectioned at 10,t and stained in Weigert's haematoxylin and Orange G withErythrosin. This method of staining was found to emphasize the distinction betweenthe Wolffian and sinus cells. Graphical reconstructions were made of the reproduc-tive ducts using one specimen of each of the fifteen stages. The other specimen wasused to check the results obtained from each reconstruction.

RESULTS

(a) Identification of tissues

In the early stages the Wolffian cells appear as a columnar epithelium, the cytoplasmof which stains particularly with Orange G, while the nuclei stain lightly withWeigert's haematoxylin. In the later stages the' epithelial character is lost but thestaining properties remain the same. Because of its more closely packed nuclei, theMullerian tissue stains much more darkly with haematoxylin than does the Wolffiantissue. The distinction between the two epithelia is shown particularly in PI. 1,figs. 1 and 2. The urogenital sinus tissue consists of loosely packed irregular cells,the cytoplasm staining with both Orange G and Erythrosin, giving the cells a pinkercolour than that of the Wolffian tissue.

Anatomy 87 28

424 Edna B. Carr(b) Description of stages

It should be noted that throughout the description, posteriorly is synonymouswith caudally and anteriorly with cephalically.

19- and l9j-day embryo. At these stages the Mullerian canals open to the peri-toneal cavity by the ostia. Each Mullerian canal consists of a thick darkly stainingepithelium round a small lumen. As one proceeds caudally the canals approacheach other, becoming interposed between the two Wolffian canals, and joining eachother for a short distance to form the rudiments of the Mullerian vagina (Text-fig. 1).

Text-fig. 1. Dorsal reconstruction from 19k-day female embryo. x 1O0. w, Wolffian tissue;m, Mullerian tissue; s., sinus tissue projecting from the dorsal wall ofthe sinus; u.g.s., urogenitalsinus proper; p., projection from the Mullerianvagina. (These abbreviations apply to all figures.)

At 19 days the lumina of the two canals are still distinct though their walls are fusedmedially. At 191 days a complete fusion of the lumina occurs for a short distance.In both stages the two canals diverge posteriorly, each tapering away to nothing onthe wall of the Wolfflan canal of its own side, and ending before the sinus is reached.The Wolffian ducts are large with a thinner and more lightly staining epithelium

than that of the Mullerian canals. They are continuous from the mesonephros to theurogenital sinus. The lumina are large and considerably swollen just anterior totheir communication with the sinus. They open into two projections from the dorsalwall of the urogenital sinus which in turn open into the urogenital sinus proper.

21-day embryo. The Mullerian ducts have joined more extensively to form thevagina, and posteriorly the Wolffian canals become apposed to its lateral walls. Thedivergent posterior ends of the Mullerian canals are still present as in the previousstages. The solid base of the Mullerian -vagina divides into right and left arms adjacent

The development of the rabbit vagina 425to the Wolffian bulbs where these open into the two projections from the dorsalwall of the sinus.

22-day embryo. The Mullerian canals are joined more extensively and are separatedno longer at their extreme posterior tips into right and left parts as in the previousstages (Text-fig. 2). The Mullerian vagina terminates in a dorsal sheet of tissueinserted between the Wolffian canals and bounded posteriorly by sinus tissue. Thelarge expansion of the posterior part of the Wolffian canals almost obscures theMullerian vagina when viewed from the dorsal side. The Wolffian canals still openinto the urogenital sinus.

Text-fig. 2. Dorsal reconstruction from Text-fig. 3. Dorsal reconstruction from22-day female embryo. x 50. 24-day female embryo. x 50.

23-day embryo. The Mullerian vagina is like that of the previous stage but pos-teriorly the solid tip has become separated by the urogenital sinus into dorsal andventral arms. The dorsal arm is embedded in the sinus tissue for some distance. Theanterior parts of the Wolffian canals have completely degenerated while posteriorlytheir openings into the urogenital sinus are blocked by loosely packed sinus cells.Thus the two projections from the dorsal wall of the sinus fuse to form a solid corein which the ends of the Mullerian canals terminate.

24-day embryo. The Mullerian vagina has increased in size while the Wolffiancanals have undergone further degeneration. The projection from the dorsal wall ofthe sinus ((s.) in Text-fig. 3) has grown anteriorly and forms a considerable structurejoining the Mullerian and Wolffian canals to the urogenital sinus.

25-day embryo. Further growth of the Mullerian vagina and of the projectionsfrom the dorsal wall of the urogenital sinus is accompanied by further degenerationof the Wolffian canals. Dorso-lateral projections (p. in Text-fig. 4) have grown out

28-2

426 Edna B. Carrfrom the Mullerian vagina to meet the Wolffian ducts. Posteriorly from here anextensive fusion of the Wolffian canals to the lateral walls of the Mullerian vaginahas occurred; although in the most posterior part they are separated from theMullerian by the sinus tissue.

26-day embryo. The Mullerian projections are more extensive while there is furtherdegeneration of Wolffian tissue at this age compared with that of the 25-day embryo.The left Wolffian duct still remains swollen at its base, where there is a smalllumen, whereas the right is reduced to a solid rod of cells.

-P.~~~~~~~~~~~~~~wu tP ~~~~~~~~~~~~~~~~~~~~~ 2Text-fig. 4. Dorsal reconstruction from Text-fig. 5. Dorsal reconstruction from

25-day female embryo. x 50. 27k-day female embryo. x 33.

27-day embryo. The Wolffian canals have undergone such a degree of degenerationthat nowhere is there a lumen present. The right Wolifian duct has fragmented.Both ducts at this stage have been reduced to a single cell in thickness in manyparts, and only by careful tracing from section to section is it possible to follow them.Posteriorly they are joined to the vagina by the two lateral projections of Mulleriantissue, and they become embedded in the wall of the vagina meeting the anteriorlyprojecting sinus tissue.At this stage the sinus tissue is not only embedded in the solid tip of the Mullerian

vagina, dividing it into dorsal and ventral arms, as in the previous stages, but alsoprojects anteriorly into the lumen, forming a solid core of sinus tissue. Hence theposterior end of the vagina is composed of this solid core of tissue, which extendsa short way along the sides while the Mullerian tissue forms the dorsal and ventralwalls.

272-day embryo. The Wolffian tissue is in a similar degenerate condition to that ofthe previous stage (Text-fig. 5). Posteriorly the right Wolffian duct passes to theinside of the Mullerian epithelium and emerges again to join the sinus tissue.

The development of the rabbit vaginaText-fig. 6 is the reconstruction of the dorsal view showing also the internal distribu-tion of tissues. The internal Wolffian and sinus tissue is seen here in relation to theother tissues.The degenerate condition of the Wolffian tissue and its incorporation beneath the

Mullerian tissue on the right side is seen in PI. 1, fig. 3. This section also shows theprojection of the Mullerian tissue which has extended from the vagina to incorporatethe Wolffian ducts.

Text-fig. 6. As Text-fig. 5, but to show Text-fig. 7T. Dorsal reconstruction fromgeneral distribution of tissues. x 33. 28k-day female embryo. x 833

28j-day embryo. At this stage (Text-fig. 7) the bases of the Wolffian canals havebecome completely incorporated internally to the Mullerian vaginal epithelium(Text-fig. 8). On the right side, the Wolifian duct is no longer in contact with thesinus tissue. Further degeneration in the anterior region of the Woiffian canals hastaken place.

29-day embryo. A great reduction in the Wolffian tissue has taken place at thisage (Text-fig. 9). Anteriorly, the Wolifian canals are reduced to a small vestige onthe tip of the projections from the Mullerian vagina. Internally, the Wolifiancanals have become fragmented and lie below the Mullerian epithelium (Text-fig. 10).The anterior projections of the sinus tissue which were joined to the posterior

ends of the Wolifian tissue have become internal to the Mullerian epithelium.P1. 1, fig. 4, shows the great reduction in the Wolifian tissue on the right side andits position internally to the Mullerian epithelium.

80-day embryo. Only a very small vestige of the Wolffian tissue now remainsexternally on the tips of the projections from Mullerian vagina. Internally, theWolifian remnants are even further reduced compared with the previous stage.

31-day embryo. The Wolffian tissue has now completely degenerated externallyon the left side while on the right side only a remnant was found embedded in the

427

428 Edna B. Carr

-w.

-m.

Text-fig. 8. AsAText-fig. 7, but to showgeneral distribution of tissues. x 33.

Text-fig. 9. Dorsal reconstruction from29-day female embryo. x 50.

-w.

w.

~~S.(Internal coreof sinus tissue)

Text-fig. 10. As Text-fig. 9, but to show general distribution of tissues. x 50.

The development of the rabbit vagina 429wall of the projection of the Mullerian vagina. Internally, a small fragment wasfound beneath the epithelium of the vagina of the right side. The sinus tissueforms a solid base to the vagina joining it to the urogenital sinus.

32-day embryo. Only the right Wolffian canal was found internally to the Mullerianepithelium, as a small group of cells only 40z in length (Text-figs. 11 and 12). Thesinus tissue forming the base of the vagina is still not canalized.

Text-fig. 11. Dorsal reconstruction from 32-day female embryo. x 50.

Text-fig. 12. As Text-fig. 11 but to show general distribution of tissues. x 50.

Edna B. Carr

DISCUSSION

Histological differentiation between the epithelia of the Mullerian ducts, Wolffianducts and uro-genital sinus is very clear in these specimens. The description of moststages shows that the limits between the different types of epithelia do not alwayscorrespond with the obvious morphological limits. For instance, sinus epitheliumextends dorsally and anteriorly from the urogenital sinus to form the terminal partof the morphologically continuous vagina; and the Mullerian vagina forms temporarydiverticula which join the degenerating Wolffian canals to it in the later stages. Inview of the complete sequence of events in development described here the mosttenable explanation is that the histological appearance of the epithelium indicatesits origin, and that actual outgrowths of sinus or Mullkrian epithelium are responsiblefor the arrangement observed. The alternative assumption, that a transformationof one type of epithelium into another occurs, seems very unlikely in view of theabsence of transitional stages.The changing position of the borders between epithelial types and the lack of

consistent correspondence between morphological and histological boundaries isperhaps responsible for the discrepant opinions about the development of the vaginawhich have been held by earlier workers (Langenbacher, Mihalkovics & Baxter).The results obtained in the present work confirm the view held by Jost (1943 a, b, 1947)that the vagina of the full-term rabbit foetus is derived mainly from Mulleriantissue, with sinus tissue forming its posterior part. Carleton (1931) found corre-sponding evidence in the adult for its composite nature, in his study in epithelialphagocytosis. He found that the lower third of the vagina was distinctly differentfrom the upper two-thirds, being columnar and non-stratified. Correspondingly,when Jost (1942) injected testosterone propionate, and Courrier & Jost (1942)injected pregneninolone into pregnant rabbits, they found stimulation of onlythe lower part of the foetal vagina.

SUMMARY

1. The development of the rabbit vagina is described in detail from reconstruc-tions made from serial sections of foetuses of 19-32 days gestation.

2. The Mullerian ducts join to form the anterior part of the vagina. Two dorso-lateral diverticula were formed from Mullerian tissue at 25 days and incorporatedthe bases of the degenerating Wolffian canals. These diverticula were still present at32 days. It was found that the Wolffian ducts degenerate almost completely leavingonly a small group of cells embedded in the epithelium of the right Mulleriandiverticulum. A projection from the dorsal wall of the sinus elongates duringdevelopment and forms the posterior part of the vagina.

3. These results support the view of Jost that the vagina of the rabbit is formedfrom Mullerian and sinus tissue but provide no evidence in support of the view ofBaxter that it is formed from the Mullerian and Wolffian tissues or of Mihalkovicsand Langenbacher that it is formed from Mullerian tissue alone.

I am indebted to Prof. G. R. de Beer for his help in the initial stages, and sub-sequently to his successor Mr M. Abercrombie for all his valuable help and advice inthe completion and presentation of this work.

430

Journal of Anatomy, Vol. 87, Part 4

CARR-THE DEVELOPMENT OF THE RABBIT VAGINA

Plate I

The development of the rabbit vagina 431I especially wish to thank Mr H. Barker for his technical advice and skill, especially

in the cutting of serial sections; and Miss J. Hubbard for the photographs.

REFERENCESBAXTER, J. S. (1933). The development of the vagina of the rabbit. J. Anat., Lond., 67, 555-562.CARLETON, H. M. (1931). Studies on epithelial phagocytosis. I. Proc. Roy. Soc. B, 108, 1-10.COURRIER, R. & JOST, A. (1942). Intersexualitk foftale provoquee par la pregn6ninolone au cours

de la grossesse. C.R. Soc. Biol., Paris, 136, 395-396.JOST, A. (1942). Intersexualite experimentale chez l'embryon de lapin. Etude du vagin et de

l'utricule prostatique. Ann. Endocr., Paris, 3, 121-124.JOST, A. (1943a). Etude d'embryologie et de physiologic genitales chez le lapin. Ann. Endocr.,

Paris, 4, 221-238.JOST, A. (1943b). Sur le vagin de la lapine. C.R. Soc. Biol., Paris, 137, 392-393.JOST, A. (1947). Recherches sur la differentiation sexuelle de l'embryon de lapin. Arch. Anat. micr.

Morph. exp. 36, 151-200, 242-270, 271-315.LANGENBACHER, L. (1882). Beitrag zur Kenntniss der Wolff'schen und Muller'schen Gange bei

Saugern. Arch. mikr. Anat. 20, 92-108.MIHALKOVICS, VON G. (1885). Untersuchungen uber die Entwickelung des Harn- und Geschlechts-

apparates der Amnioten. Int. Mschr. Anat. Histol. 2, 41-62, 65-106, 284-306, 307-339,348-385, 387-433, 435-485.

WATERMAN, A. J. (1942). Variation in rabbit embryos. Anat. Rec. 84 (Abstract), 513-514.

EXPLANATION OF PLATE

The photomicrographs of sections are shown in anterior view so that the actual left side of thebody is the right side of the illustration.Fig. 1. Transverse section showing the comparative sizes and appearances of the Wolffian and

Mullerian canals in a 21-day foetus. x 115.Fig. 2. Transverse section to show the degenerating Wolifian ducts adhering to the lateral walls

of the Mullerian vagina in a 25-day foetus. x 180.Fig. 3. Transverse section showing the remains of the Wolifian tissue being incorporated in the

wall of the Mullerian vagina in a 27j-day foetus. On the right is shown the Mullerian projec-tion which joins the Wolffian duct to the vagina. x 50.

Fig. 4. Transverse section of the 29-day foetus. The Wolffian tissue appear as a small group ofcells in the wall of the projections from the Mullerian vagina. x 65.

Fig. 5. Transverse section of the base of the vagina and the urogenital sinus, just anterior to theirjunction. The tip of the Mullerian vagina is seen connected to the sinus region. x 65.