On the of the Rabbit.

By

Ch. Togari. Form the Anatomical Laboratory, Aichi Medical College.

8 Figures (Plate XVIII-XIX).

One of the most interesting subjects in studying the is the changes occuring in the cells of the Graafian follicle in the time of the development of the corpus luteum after its rupture. As to this point, many divergence of opinions have been published, and still there is no comformity among the authors. Therefore, it is my purpose to investigate more thoroughly on the rabbit on the same question in addition to my previously published paper " On the Origin of the Corpus Luteum of the Mouse." As described in my previous work, the investigators thought that these cells are in different stages of development of the connective tissue cells, and their theories could be summarized as follows :— 1. The theca interna cells of the mature follicle, not maximal differentiated, retain yet their nature of reverting into the original connective tissue cells again, and can form the connective tissue reticul um among the lutein cells with mitotic divisions. This opinion was main- tained am-ong the authors of the granulosa origin theory of the corpus luteum such as, Sobotta in the mouse, the rabbit and the guinea-pig Meyer in man Iok a and Adac hi in the rabbit. Cohn explained that these cells can, furthermore, form the capillary blood vessels. 2. The theca interna cells can never be reverted into the fibroblasts, but invade among the granulosa lutein cells without any changes. This contention is maintained by Solomon and Gat e n by in man, the pig and the duck-billed platypus. 3. When the theca interna cells are multiplied by mitotic divisions after ovulation, and converted into the lutein cells with their gradual hypertrophy, contrary to the granulosa degeneration, then the theca interna origin theory will be asserted. This theory was proved by Buehler in man Clark also Doering in the pig; Jankowski in 338 Ch. Togari, man, the cow, the sheep, the pig and the guinea-pig ; Hegar in man ; Mats u y a m a in the rat ; and H i r o s e in man and the rabbit. 4. Although the theca interna cells can never be reverted into the connective tissue, yet they are utilized as the theca lutein cells to form the corpus luteum together with the granulosa lutein cells. This opinion is entertained by the authors of the double origin theory such as Van der Stricht in the bat ; Ho norê in the rabbit ; Voelker in the Sperm- ophilus citellus ; Loeb in the guinea-pig ; Corner in the sow ; Ts uk a- guchi intherabbit; Rabl, Seitz, Cohn, Marcotty, Schroeder and others in man. 5. The theca interna cells reach their maximal differentiation about the time of rupture and can not be reverted into the fibroblasts during the period of the corpus luteum formation, and they fulfill their function as such to degenerate sooner or later after rupture, suffering the pressure of the neighbouring tissue, and the corpus luteum is derived from the granulosa layer only. My former report on the mouse agrees with this view and a part of the theca interna cells of the human corpus lutem described by Meyer takes the same fate. The prosperity and decay of the granulosa layer during the corpus luteum formation is also the subject of repeated discussion, as theca interna cells mentioned above. The authors who insisted upon the epithelial or double origin theory noticed that the most of the granulosa cells remain after rupture and are converted into lutein cells only by their hypertrophy, as observed by Sobbota in the mouse ; H on orê also Co h n in the rabbit ; and many others in man : or both by their hyper- trophy and hyperplasy as noted by So bo tta in the rabbit and the guinea- pig ; Marshall in the sheep O'Donoghue in the marsupitalia ; Strak- os c h, Wall a r t, Schroeder and others in man. On the contrary, among, the authors who insist upon the theca interna origin theory, some have noticed that the granulosa layer of the follicles before rupture is composed. of a single layer of cells or has totally disappeared ; and others have remar- ked that the majority of it may flow out by rupture with the liquor folliculi, and that the remainder soon degenerates. Moreover, as to the retrogression of the corpus luteum, Sobotta has declared that it degenerates chiefly by the fatty metamorphosis of the lutein cells in the rabbit, contrary to the mouse in which it may exist throughout the whole life never undergoing a change. Rabl als supported the Sobota's views. Contrary to their descriptions, I have remarked that the corpus luteum of the mouse falls into retrogression by the pressure of the abjacent growing corpora lutea or follicles, or by the central cavity On the Corpus Lute= of the Rabbit. 339 formation of the , suffering the majority of the lutein cells in fatty degeneration and a part of them being converted into pigment cells. What may be, then, the fate of the corpus luteum of the rabbit ? Would it form the corpus fibrosum, by further contraction of the remaining connective tissue reticulum, after the disappearance of the entire mass of lutein cells, or the by its further hyaline degeneretion? In order to ascertain these questions, it is necessary to have an intensive investigation in comparative studies on a large variety of animals. And it is of some importance to compare the corpora lutea of the rabbit to those of the mouse. There is a very interesting contrast in the composition of ovaries of the rabbit and the mouse. These two animals belong to the same order but the ovary of the former is mainly composed of interstitial gland cells with some few corpora lutea and follicles whereas the latter is constructed chiefly with corpora lutea and follicles. Since the interstitial gland cells resemble very closely to the lutein cells, to avoid the possible mistake as some seem to have made of one for the other, it is of vital importance to differentiate them clearly by careful studies on their development and retrogression ; although the former is, of couse, derived from the theca interna cells of the atretic follicles. Literature as to the origin of the corpus luteum of the rabbit since Sobotta. The reports as to the origin of the corpus luteum of the rabbit are not of small number, but I want to survey them since Sobotta ; hence, the works before him are described in his copies, and they preserve historical interest only. So bo tta (1897) noticed that the lutein cells of the corpus luteum of the rabbit are derived from the granulosa cells of the mature follicle as the mouse ; the theca interna cells, rich in cytoplasm, penetrate into the lutein layer after they are reverted into the spindle-shaped connective tissue cells, and form the interstitial connective tissue reticulum. Honoré (1899)published the same results as Sobot ta' s on the origin ofthe lutein cells, but as to the fateof the theca interna cells of the Graa- fian follicles, his view was different from Sobob a's, namely, a few of the theca interna cells take part in the formation of the connective tissue reticulum among the granulosa lutein cells through the cell division, the majority of them remaining unchanged 6 to 11 days after copulation, and they are easily distinguished from the granulosa lutein cells. Cohn (1903) persisted also in the granulosa origin theory of the 340 Ch. Togari, corpus luteum, but as to the fate of the theca interna cells his view was quite different from the other authors, that these cells penetrate among the granulosa cells in the process of the corpus luteum formation, and change into the relatively wide capillaries communicated with the blood vessels of the theca interna. Ioka (1917) recognized that the corpus luteum is the epithelial organ, and the theca interna cells are reverted into the connective tissue reticulum among the lutein cells as So bo tt a' s. This process is fulfilled, according to So bot ta within 32 hours after copulation. However, he found a few theca interna cells without any changes even until 7 days after copulation. Bo r ell (1919) distinguished the granulosa cells from the theca interna cells by the vital staining as Iok a did, and proved that the lutein cells come from the former. Contrary to many works cited above, Hiros e (1920)mentioned that the corpus luteum is formed by the theca interna cells by their growth and multiplication as Mat s uy a ma (1919) noticed in the rat, and the majority of the granulosa cells are carried away at the time of ovulation, and after a short while the remainder of the granulosa disappear by degeneration. In the next year, he repeated the theca interna origin theory in the artificial corpus luteum of the rabbit produced by his method of injecting the human placental emulsion. At the thirtieth session of the Japanese Association of Anatomist at K- yoto Imperial University, Ts uk aguc hi (1922) interprets his preparations to indicate quite clearly that the lutein granules of the corpus luteum are derived from the plastosomes, and the granulosa lutein cells are distin- guished from the theca lutein cells cytologically. At the next session at the Tokyo Imperial University, he (1923) explained also with his prepara- tions that the lutein granules, corresponding to the secretory granules of the gland, are converted into colourless, transparent substance, and then absorbed as secretions. In the same year, Adac hi (1923) noticed that the lutein cells are derived from the granulosa cells by their hyperplasy kInd hypertrophy contrary to the theca interna cells of reverting into the spindle-shaped cells, both in the physiologic and artificial corpus luteum of the rabbit, the latter of which was produced by the Hiro se's method. As above described, some insist on that the theca interna cells in the rabbit may be converted into the so-called Seit z's theca lutein cells, and others, that they may be reverted into the fibroblasts to form the connec- tive tissue reticulum among the lutein cells, and that the various time On the Corpus Luteum of the Rabbit. 341 required to revert are recorded. In some extreme cases, they were noticed to change further into the blood vessels among the lutein cells. Besides these different opinions as to the changes of the theca interna cells occuring during the period of the corpus luteum formation, there are also the descriptions that deny the granulosa origin theory altogether.

Material and method. There is usualy no ovulation to be found, although the heat period appears directly after parturition in the rabbit. Hence, some of the materials were taken in such rabbits as they have been copulated in their majority within a few days following parturition, which were accurately dated ; with these materials, both the retrogression of the former corpus luteum graviditatis and the development of the corpus luteum are studied. And others, obtained at various durations following parturition without any stimuli, or after copulation performed in the heat periods, were utilized for the study of the corpus luteum degenera- tion by the former, and of the corpus luteum formation by the latter. 48 ovaries of the rabbits obtained in such manner are shown in the following table. 342 Ch. Togari, These rabbits were killed by striking on the neck or by chloroform narcosis. One side of the ovaries was cast into the Carnoy's mixture, embedded in celloidin, made serial sections of 10-15 M. in thickness, and stained with Best's carmine solution ; the other side of them was placed into Zenker's, Or t's or Flemming's solution, embedded in paraffin or celloidin-paraffin, made sections of 5-10 M. in thickness and stained with H a n s e n's hematoxylin-eosin, Heide n h a i n's iron hematoxylin, W ei- gert's elastic fiber stain and modified Tr a i na's connective tissue stain. The former preparations offered the study of the general sturucture of the ovary and the distribution of glycogen in the ovaries (The results of the latter will be published later). The latter was utilized for the inves- tigation of the minute structures of the follicle or the corpus luteum.

Observations. Mature follicle.

Materials were taken from a rabbit 3 hours after copulation which was performed in the heat period occuring 15 days after parturition. Large, well-preserved Graafian follicles on the surface of the overy are spherical in foam, protrude fairly from the surface of the ovaries and display an almost transparent aspect. By microscopical observation, a part of the wall of the follicle facing the surface of the ovary was seen covered with squamous germinal layer composed of no more than 10 layers of cells owing to the accumulation of a large quantity of the liquor folliculi. The wall of the Graafian follicle consists of four layers which, to enumerate from the outer side, are the , the theca interna, the basement membrane and the . The theca externa is composed of 4 or 5 layers of connective tissue cells. Elastic fibers and fibers do not take part in the formation of the theca externa. The inner side of this layer has a clear cut limit with the theca interna, but the outer side has two different limits ; namely, a part facing to the surface has no clear cut limit with the tunica albuginea, and there exists 5 or 6 layers of compressed connective tissue cells between the germinal epithelium and the basement mem- brane, and the other for the most part has a clear cut limit with the so-called interstitial gland. The cell body represents an elongated spindle shape with poor cytoplasm, compared with those of the polygonal cells of the theca interna. In its center an elliptic nucleus is found stained dark with hematoxylin. The mitotic division is never recongnized in On the Corpus Lute= of the Rabbit. 343 this period. The theca interna is not equally thick all over the follicle and is of 1-3 layers of polygonal cells. The part of the follicle touching the surface of the ovary is only a layer of compressed cells or this layer is lacking therefore, no clear cut limits are noticed among the theca interna, the theca externa and the tunica albuginea. Polygonal cells of this layer do not run exactly in the same direction as the theca externa, but show a concentrical arrangement. They are large and polygonal, their cytoplasm show a honey-combed appearance and their nuclei are spherical or elliptical, containing a few chromatin. These cells do not exhibit any appearance of undergoing a change. Besides these polygonal cells, there exist also the spindle-shaped cells in various places of the theca interna, as found in the case of the mouse. They are connected with those of the theca externa, ruii usually through this layer diagonally and reach directly outside of the basement mem- brane, where they form an almost continuous line parallel to this membrane. These cells agree with those of the theca externa in size, form and chromaticity of their nuclei, so that both are noticed as one and the same which were derived from the ovarian stroma. Moreover, we are able to recognize the intermediate forms between the spindle-shaped and the polygonal cells, which suggest that the latter would be formed of the more differentiated spindle-shaped cells. Thus the sheath of the mature follicle of,the rabbit closely resembles that of the mouse, consisting of so-called Koel lik er's three layers, i.e., Stratum fibrosum externurn, Stratum medium cellulare and Stratum fibrosum internum. The capil- lary network in the theca interna is fairely congested except a part where it touches to the tunica albuginea, i.e., the so-called W al deyer's 'stigma.' The endothelial cells of these capillary wall are able to be distinguished from the spindle-shape cells in this layer by size, form, chromaticity and location of their nuclei, although there is a close resemblance. The basement membrane exists inside of the theca interna, surround- ing closely the granulosa layer concentrically, and shows a clear line tinged black with hematoxylin-eosiu. It is also stained deeply with the connective tissue stain, but not with the elastic fiber stain. The granulosa layer shows a condition similar to the appearance of the theca interim and is composed of 3 or 4 layers of cells at the super- ficial part, and of more than 8 layers of cells in the profound part. The cells of this layer are arranged densely and radially, and the outermost cells are arranged very regularly adherent to the basement membrane , contrary to the irregularly arranged cells facing the antrum folliculi . 344 Ch. Togari, The cone-shaped or rounded projection of the granulosa layer, i.e., the discus proligerus, can never be seen in the mature follicle of the rabbit, contrary to the mouse, and the corona radiata surrounding the ovum spread out a few strands of cells radially to the granulosa layer. This is due to the fact that the liquor folliculi begins to accumlate at many places almost at the same time especially near the ovum in the growing follicle : accordingly, certain of the epithelial cells are torn apart so as to form many fluid-filled spaces showing the sponge-like appearance of the granulosa layer, and as these spaces are enlarged, it finally lacks the discus proligerus formation. The mature follicle shows, therefore, the so-called Mjassbj edoff's Graafsches Blaeschen des Typus11,' its antrum folliculi being passed through by cords of cells which suspend the ovum on their center (Fig. 1). The granulosa cells are small round or oval, their cytoplasm are a few and look granular, their small round nuclei at the center are stained deeper in contrast with those of the polygonal cells of the theca interna, and contain no nucleoli. Among the granulosa cells in this period, many cell divisions are seen everywhere, but the degeneration appears occasionally in a few cells in the part facing the antrum folliculi. The so-called C al l-E xn er bodies in the granulosa layer are found in a large number not only in the growing follicles, but also in the mature follicles in the rabbit. They show spherical cavities in various shapes of reticular structure surrounded by the granulosa cells as the corona radiata shows. These may be concerned with the liquor folliculi formation. The liquor folliculi is further richer in quantity than that of the mouse, and stained granular with hematoxylin-eosin in the fixed preparations, contrary to the case of the mouse, in which it is stained usually in a thread-like appearace. Development of the corpus luteum. 1. Freshly ruptured follicle. Several freshly ruptured follicles are seen in the ovaries 10 hours after copulation. They can easily be recognized as a speck, visible to the naked eye, at the points of rupture on the ovarian surface. They are smaller than the mature follicle, somewhat collapsed, almost spherical in form and protrude fairly from the surface of the ovary, owing to the partial discharge of the and the granulosa cells with ovum, and their walls consist of the same layers as before ovulation. The limits of every layer are, therefore, clearly verified with hematoxylin-eosin, contrary to that of the mouse (Fig. 2). On the Corpus Luteum of the Rabbit. 345 At the point of rupture, there is no tunica albuginea with the germinal epithelium nor are the theca interna and externa to be seen, and the orifice of the rupture is surrounded by the granulosa cells only. This orifice is wider than that of the mouse, and at its border some hemorrhage can usually be noted (Fig. 2). The theca externa is a little different from that of the mature follicle, and its arrangement is rather irregular, but there is no cell division nor degeneration in this layer. The theca interna is a layer most affected in its arrangement in a mechanical way. By the decrease of the internal pressure of the follicle through rupture and some contraction of its wall, the theca interna seems as if it may be thicker than at the previous period, and at several places, it is projected into the granulosa layer as waves. So that two different " parts are formed in this layer : the thicker and the thinner part as seen in the mouse (Fig. 2 and 3). In the thicker part, there exist extended capillaries and a few hemorrhages which increase in the part facing the surface of the ovary. Polygonal cells of this layer show a looser arrangement than those of the former period, and are weak in chromaticity of their nuclei. Some - of these nuclei indicate the appearance of degeneration, showing their chromatin gathered in a mass, or flowing out into cytoplasm from one side of the nuclei after they were dissolved, although more frequently they become unstainable. The spindle-shaped cells of this layer are also changed a little in their arrangement, i.e., they rum obliquely or trans- versally, reach to the outside of the basement thembrane, where they run along the latter which now shows a wave-like course. No mitotic divisions are seen among these cells. Polynuclear leucocytes are seen in this layer. The besement membrane is noticed as a wave-like course, although. in some places forms not a continuous line when stained with hema- toxylin-eosin. . The majority of the granulosa cells remain after rupture, however, a few is extruded with the ovum, and there is no follicle as the majority of them are flowed out, as maintained by the authors who insist on the theca interna origin theory, although such follicles are often noticed that a part of the outermost cells of this layer is separated from the adjacent basement membrane, or some innermost cells may lie free as an island in the antrum folliculi without any change (Fig. 2). This layer also becomes somewhat thicker, since it lies in a smaller cavity than before, and is composed of 3 or 4 layers about the point of rupture, and the more we advance to the deeper part, the more cell layers 346 Ch. Togari, increase, so that the deepest part is composed of more than ten layers of cells. This layer has also been compressed through the rupture of the follicle, and its outermost zone shows a wave-like line corresponding to that of the innermost zone of the theca interna (Fig. 2 & 3). This wave- like line is one of the most important points to distinguish the follicle before and after ovulation. The cells of this layer show generally small vesicular forms with round or oval nuclei rich in chromaticity and contain nucleoli. There is still no mitotic division among these cells. The liquor folliculi stained thread-like appearance is smaller in amount than in the mature follicle and contains usually a little hemorrhage. 2. Ruptured follicle 18 hours after copulation. At the point of rupture, there exists also a speck seen macroscopically as that of the former period. The microscopical studies show that the ruptured follicles of this period are a little different from the freshly ruptured ones, and possess usually large apertures containing a little hemorrhage. The condition of the theca externa is similar to that of the mature follicles, showing the concentric arrangement, although the former becomes thicker than the latter containing 7 or 8 cells layer. Among these cells, no mitotic divisions can yet be seen, although they are rich in chromaticity of their nuclei. At the innermost part of the theca externa, I have found a notable fact that the intermediate forms of the spindle- shaped and the polygonal cells exist here and there. This fact sbwos that the formation of the polygonal cells from the spindle-shaped may be continued up to these stages, and these transitional forms may not be the pressed polygonal cells. Polygonal cells made in these periods may remain long as the so-called S ei tz's 'theca lutein cells' in the fully grown corpus luteum, contrary to the interpretation of many investigators that the ploygoiial cells of the ruptured follicles may be reverted into the spindle-shaped cells to form the connective tissue reticulum. The theca interna has decreased a little in its thickness, lacking its superficial part, and consisting of its profound part of two different portions, the thicker and the thinner, mentioned before. The nuclei of the polygonal cells are, on the whole, weak in chromaticity and contain no nucleoli. Some of them are even caryolytic or pycnotic. As a notable change in the arrangement of the spindle-shaped cells, some penetrate into the granulosa layer through the broken points of the basement membrane. A part of the endothelial cells of the capillary blood vessels On the Corpus Luteum of the Rabbit. 347 penetrate also, following the above described connective tissue. Polynu- clear leucocytes found in the theca interna are smaller in amount than those of the former period. The basement membrane in its larger parts has been broken and has disappeared. Thecondition of the granulosa layer is similar to that of the former period. This layer is composed of 6 or 7 layers of cells in the superficial part and 17 or 18 in the profound. Granulosa cells are usually loosely arranged, yet their limits with respect to one another are not clear, and some of them contain enlarged nuclei with nucleoli. The spindle-shaped cells penetrated into this layer above described exist in its outer zone, especially at the places tcliuching the thicker part of the theca interna. Some polynuclear leucocytes are also seen here and there. The fluid in the central cavity of the ruptured follicle is stained reticular, containing blood corpuscles and granulosa cells. 3. Ruptured follicle 24 hours after copulation. Macroscopically the appearance of the ruptured follicles of this stage is similar to that of the former period, except that some of them has a plug or hernina-like bulging. By the microscopical studies, the theca externa shows a slight wave- like course, being composed of 4 or 5 cells layer of concentric arrange- ment. Mitotic divisions among these cells are seen occasionally. The theca interna at this period on a whole is rather thicker than that of the former period, and the discrepancy between the thicker and the thinner part becomes very slight (Fig. 4). The cytoplasm of the polygonal cells gives a honey-combed appearance, and the nucleus with no nucleoli shows often the appearance of degeneration such as caryolysis or pycnosis. On the contrary, the nuclei of the spindle-shaped cells of connective tissue and the endothelial cells of the capillaries are rich in chromatin, and invade among the granulosa cells with mitotic divisions. The engorgement of the capillary blood vessels of this layer is not vigorous. Polyn.uclear leucocytes can be seen here and there. The besement membrane can scarecely be seen with hematoxylin- eosin. The granulosa layer, generally speaking, is thinner at the adjacent part of the large aperture made by rupture, and is composed of only a few layers of cells, and is thicker in the profound part being composed of 14 or 15 layers of cells. In some cases, a part of the granulosa layer exists as a plug hanging on the point of rupture (Fig . 4). This may be 348 Ch. Togari, produced by the internal pressure at the time of ovulation, not by the swelling of the granulosa cells, which causes a bulging of contents through the outer pole of wall, at the point previously weakened by rupture. This plug is composed of normal granulosa cells, fibrin, blood corpuscles and degenerating granulosa cells. The nuclei of the granulosa cells are generally rich in chromatin containing nucleoli, and enlarged a little, although the cytoplasm show no clear outer limits as found in former periods. With this appearance of nuclei, we can easily verify the gran- ulosa cells from the polygonal cells of the theca interna. The con nec- tive tissue cells and the endothelial cells which invade here from the theca interna are, generally, in advancing from the periphery to the inner border of the granulosa layer arrange id radially, without quite reaching the central cavity. Among the granulose cells, polynuclear leucocytes are also seen here and there. The central cavity of the ruptured follicle shows irregular round or oval forms of the fibrin mesh- works in which are contained a small amount of normal granulosa cells, blood corpuscles and the degenerating granulosa cells. 4. Ruptured follicle 33 hours after copulation. The aperture made by rupture can be seen macroscopically in this stage. The microscopical observations reveal mitotic divisions in the theca externa as in the former period. The limit of the theca interna and the granulosa layer becomes less clear as the relation between the thicker and the thinner part of the former. The theca interna has decreased in thickness and at several points polygonal cells can no longer be recognized, as the granulosa layer touches directly with the theca externa. In the polygonal cells, two kinds of cells can be distinguished ; the one is intact and the other in degeneration. Polynuclear leucocytes are also seen in this layer. The spindle- shaped connective tissue cells and the endothelial cells of the theca interna are penetrated into the granulosa layer with notable mitotic divisions. These cells in the granulosa layer are radial in arrangement, and spread almost through the whole layer. The distinction between both kinds of cells above described is one of the most difficult matters. The granulosa layer shows a similar thickness to that of the former period. These cells are enlarge, some of them possess the clear limits of one another. Polynuclear leucocytes are seen here and there. A few blood corpuscles can be marked out in the fibrin containing central cavity. 5. Corpus luteum 48 hours after copulation. According to macroscopical observations, the young corpus luteum On the Corpus Luteum of the Rabbit. 349

in this stage protrudes a little from the surface of the ovary, and the orifice of rupture can not usually be seen. The theca externa become a continuous layer surrounding the whole circle owing to the closure of the aperture. Hence, this layer may be spoken of as the capsule of the corpus luteum. The polygonal cells of the theca interna, i.e., so-called Seitz 's 'theca lutein cells' are seen inside of the fibrous coat of the corpus luteum. Spindle-shaped connective tissue cells and the endothelial cells penetrate notably into the granulosa layer with cell divisions, and some of them have pass3d through this layer to reach the 'fibrin containing central cavity . The arrangement of these cells in the granulosa layer is not simply radial, but also oblique or transversal . The granulosa layer is composed of 3 or 4 layers of cells in its superficial part and of about 18 or 19 in its profound part . The granulosa cells, i.e., so-called P fa n e us tiel's granulosa lutein cells ' are further enlarged, obtaining clear limits with respect to one another . The cells of the epithelial islands of the central cavity are also enlarged without any appearance of degeneration. The central cavity has been diminished in size, and contains usually epithelial islands and a little hemorrhage . Some of these contain leucocytes only in their fibrin meshwork . Among the luecocytes wandering in the capsule , the granulosa lutein layer and the central cavity, there are eosinophils in abundance at this stage . A large hemorrhage is not seen usually in the freshly ruptured follicles , nor in the young corpus luteum . But at this stage, a corpus haemorr- hagicum is noticed larger than the other corpora lutea in the same stage , and holds thinner granulosa lutein layer almost of the same dimension at every part of it as the commencement of the usual atrophy of the mature profound follicles.

6. Corpus luteum 72 hours after copulation . The young corpora lutea in this stage are noticed as pink protuber- ances on the surface of the ovary . The theca lutein cells originated from the polygonal cells of the theca intern a are situated inside the capsule of the corpus luteum. Their nuclei resemble closely those of the granulosa lutein cells in their size and chromaticity , so that the difference between both cells is very difficult , but in general, the latter can be distinguished form the former with their nucleoli . The granulosa lutein layer is composed of 7 or 8 layers of cells in its superficial part and about 20 in its profound part . The radial arrange- ment of the granulosa lutein cells becomes notable by further penetration , of the connective tissue and the capillaries from the theca intern a. Gran- 350 Ch. Togari, ulosa lutein cells with clear outer limits are enlarged, especially a few of them becomes large enough to be distinguished easily from the other gran- ulosa lutein cells or theca lutein cells. Even mitotic divisions can be observed among these cells. The granulosa lutein cells are, therefore, formed by their hyperplasy as well as hypertrophy in the rabbit, contrary to the simple hypertrophy in the case of the mouse. The spindle-shaped cells which invade among these cells, run generally radially with many cell divisions and spread out side branches obliquely or transversely. Capillaries containing blood corpuscles are seen in this layer near the fibrous coat of the corpus luteum. Polynuclear leucocytes which were first found in the sheath after rupture and then in the granulosa layer, are seen no more. As to the significance of the polynuclear leucocytes, Sobotta noticed that they may participates in the formation of the central connective tissue nucleus in the corpus luteum of the rabbit, like the wandering cells found during the period of the corpus luteum formation of the mouse. I have previ- ously explained in complete agreement with Sobotta that the mononu- clear wandering cells of the mouse would have the same function, but I have now found that it is polynuclear leucocytes in the rabbit quite contrary to the mononuclear wandering cells of the mouse. Therefore, all these wandering cells found in the copus luteum formation, would belong to the leucocytes and are not concerned with the connective tissue formation directly. The fibrin containing central cavity is reduced in size, and there exist spindle-shaped or stellate connective tissue cells, but no bl000d corpuscles. 7. Corpus luteum 96 hours after copulation. Corpora lutea 4 days after copulation are found on the surface of the ovary as pink protuberances. They are surrounded by the capsule of lamellar cells which make a clear limit from the neighbouring tissue. The theca lutein cells are noticed chiefly in the peripheral parts of the corpus luteum as groups of polygonal cells smaller than the granulosa lutein cells. The granulosa lutein cells are enlarged more than those of the former period, showing many mitotic divisions (Fig. 6). These cell divisions occur chiefly during a period from 3 to 4 days after copulation, and these may be caused by the fact that the granulosa cells of the mature follicle in the rabbit are smaller in number than the granulosa lutein cells of the fully grown corpus luteum. In other words, it may be said that when the number of the granulosa cells of the mature On the Corpus Luteura of the Rabbit. 331 follicles is equal to that of the granulosa lutein cells of the grown corpus luteum, there may occur the hypertrophy only in the course of the corpus luteum formation but when the former is smaller than the latter, it may be necessary that not only hypertrophy but also hyper- plasy to occur. The spindle-shaped cells are radial in arrangement, but also oblique or transversal, and enclose several lutein cells in the peripheral part of the corpus luteum as if they formed a lobe. Capil- laries containing blood corpuscles spFead not only throughout the whole of the lutain layer in radially arranged, but also in the central connective tissue nucleus (plug). This nucleus, found instead of the central cavity, consists usually of fibrin remnants and stellate or spindle-shaped connec- tive tissue cells.

Grown corpus luteum.

1. Corpus luteum 5-9 days after copulation.

Extremely protruded corpora lutea from the surface of the ovary have reached almost definite size and obtained almost definite structure. The corpus luteum is clearly marked off from the surrounding interstitial gland by the capsule derived from the theca externa of the mature follicle. The theca lutein cells in the small polygonal shape, exist not only among the granulosa lutein cells or between them and the capsule, but also among the spindle-shaped cells of the fibrous coat. These latter cells may be derived from the spindle-shaped cells in situ, not from the theca lutein cells penetrated into the capsule. The gra,nulosa lutein cells contain a nucleus, or rarely two nuclei, and they usually lack with mitotic division. Their cytoplasm are granular or partly honey-combed and stained red with eosin, although in most cases weaker than the interstitial cells. Among the lutein cells, many capillary blood vessels are seen with connective tissue cells, which converge into one another to surround the several lutein cells. The central connective tissue nucleus is formed chiefly with spindle-shaped cells.

2. Corpus luteum 10-26 days after copulation. The fully grown corpora lutea are almost twice as lage as the mature follicle and in general, extremely protrude from the surface of the ovary, without some of them being situated in the depth of the cortex. They are perfectly marked of from the surrounding interstitial gland by the capsule, being composed of 2-4 layers of connective tissue cells. 352 ( h. Togari, The theca lutein cells are polygonal in shape with small round or oval nuclei containing no nucleoli, and are found several cells grouped together between the granulosa lutein cells and the capsule ; in other words, the polygonal cells of the mature follicle partly remain as the theca lutein cells chiefly at the peripheral parts of the fully grown corpus luteum in the rabbit, whereas in the muse they are degenerated in the early period of the corpus luteum formation (Fig. 7). The preparation fixed in Flem ming's solution andastained with iron-hematoxylin shows that these cells are equal to the interstitial gland cells in their structure and staining. The granulosa lutein cells are large polygonal, their nuclei are large and spherical with one or several nucleoli, aud thir cytoplasm are in the granular or honey-combed appearance stained red with eosin. The cells of the granulosa plug on the surface of the ovary at the point of rupture, are also converted into lutein cells covered with the germinal epithelium, and show honey-combed appearance larger in size than the lutein cells of the other parts. The granulosa lutein cells somewhat resemble those of the interstitial gland, but the former are different not only by their genesis, but also by their size and osmophil granules in their cytoplasm from the latter. Hence, these two kinds of cells are distinguished clearly with preparations fixed in the Flemmin g's solution, and that stained with iron-hematoxylin. The fully grown corpus luteum sets in degeneration gradually as the granulosa cells of the ruptured follicle convert gradually into the lutein cells. Some of the lutein cells of the corpus luteum graviditatis 26 days after copulation are clearly in degeneration. In the cerpus lgteum ovula- tionis 25 days after copulation, the retrogression of the lutein cells is more remarkable than in the former, and they degenerate in groups here and there. The connective tissue cells from the spindle-shaped cells of the theca interna and externa, converge into one another to surround almost all lutein cells, contrary to the interstitial gland in which several cells are usually surrounded as a lobe. In the central part of the corpus luteum, the connective tissue cells assemble together to form a nucleus in various size and shape. This central nucleus exists without fail in the fully grown corpus luteum, contrary to the mouse, in which some are absorbed already in this period. These connective tissue cells have, for the most part, the spindle-shaped nuclei Nthich are easily distinguished from those of the lutein cells. There is no mitotic divisions to be seen among the connective tissue nucleus and reticulum. On the Corpus Luteum of the Rabbit. 353 The capillaries run with the connective tissue reticulum among the lutein cells and finally come into contact with one side of almost every lutein cell. Besides these capillaries, the small blood vessels are seen at the peripheral part of the lutein layer or in the connective tissue nucleus of the corpus luteum. Some of the capillaries at the central part of the corpus la' 3nm are extremely extended as if they were a central sinus.

Retrogression of the corpus luteum. For the study of the retrogression of the corpus luteum of the rabbit, the ovaries during the period from 1. to 55. day after parturition are made into serial sections. Because the situation of the corpora lutea is hardly noticed macroscopically in their higher degree of the retrogression, although they are seen as a projection on the surface of the ovary in its early periods. The lutein cells of the corpus luteum graviditatis begin to degenerate gradually even in its gestation period as spoken of before, and the figures in their retrogression become very notable shortly after parturition. Some of the corpus luteum graviditatis have wholely disappeared leaving no trace by the degeneration of the lutein cells, and by the absorption of the connective tissue and the blood vessels among them at 22 days after parturition, although some of them are still in degeneration showing a few lutein cells among the reduced connective tissue reticulum at 45 days after parturition, and some are converted into a kind of corpus fibrosum containing pigment cells derived from the partly remaining lutein cells at 55 days after parturition. Thus, the time required for the retrogression of the corpus luteum is very variable, and this may per- phaps dependent upon the property of the individuals, although the original size of the fully grown corpus luteum, the pressure of the adjacent tissue and the blood supply for the corpus luteum may have no little influence upon the retrogression of the corpus luteum. Generally speaking, the corpus luteum graviditatis may have, in short, disappeared leaving no trace or may be left as a corpus fibrosum some 30 days after parturition. The corpus luteum in retrogression has usually a clear limit against the adjacent tissue by its capsule as noticed by Cohn, although it may take a wave-like course, owing to the unequal reduction of the lutein ' cells (Fig. 8). This fibrous coat is also gradually reduced as the lutein cells are diminished in number, and consists usually of 1-3 layers of cells. In some cases, however, a part of this coat is totally absorbed. 354 Ch. Togari,

As a closely resembled figure to that of the corpus luteum degeneration, there exist the interstitial gland cells in a certain territory perhaps origi- nated one and the same atretic follicle, are in degeneration, having faintly stained cytoplasm and pycnotic nuclei. The latter figure, however, usually lacks the capsule originated from the theca externa, and the cells in degeneration contact directly with neighbouring intact interstitial gland cells. In exceptional cases, the fibrous coat surrounding this figure is noticed, but even in this figure a transitional part wheie the degenerat- ing interstitial gland cells run into the intact may be found without fail by the careful researches with serial sections. Hence, the difference between both figures in retrogression can usually be made by the capsule, except the appearance of cells and the distribution of the connective tissue and the blood vessels among them. The degeneration of the lutein cells begins at places spotted over the whole corpus luteum, as seen in the mouse, not from the peripherial part nor from the central part. The lutein cells which compose the plug hanging on the surface of the corpus luteum degenerate, however, earlier than the other part of it. In the poriods of vigorous retrogression, groups of degenerating lutein cells with pycnotic nuclei were noticed here and there, which could never be seen in the mouse ; and at these places or around the blood vessels, many mast cells are found individually or grouped (Fig. 8). According to L e h n e r, there is much divergence of opinions as to the significance of the appearance of the mast cells. Among these, Cohen, Try b and others believed that " Die Mastzellen Zerfalls- oder Abbaupro- dukte aufnehmen and zu Granula verarbeiten." The mast cells in my case would have the function to take up the degenerated products of the corpus luteum as noticed by these authors, and they may have no func- tion of producing the connective tissue fibers as proposed by certain other authors. Because, there is, in reality, no increase of the connec- tive tissue in the retrogression of the corpus luteum of the rabbit, although many authors recognized its multiplication in the time of the retrogression of it in man and other mammals. In short, mast cells would have the phagocytio action as Lehner noticed and play an im- portant rOle in the absorption of the remnants of the degenerating corpora lutea. As the lutein cells contain much osmophil substance in their late stage, fatty degeneration may be the chief cause of the retrogression of the corpus luteum. The lutein cells which were large polygonal with spherical nuclei and granular or honey-combed cytoplasm deeply stained On the Corpus Luteum of the Rabbit. 355 with eosin, gradually decrease in size and fade in colour. Their further retrogression renders the nuclei of the lutein cells frequently hypochro- matic or caryolytic, and some pycnotic owing to the flowing out of the fluid of the nuclei, or gives them the appearance of the chromatin gathered at the nuclear membrane only or that of caryorrhexis. And in the more advanced, the nuclei disappear totally leaving vacuolated cytoplasm. As exceptional cases, the following figures may be observed in the degenerating corpus luteum : 1. A lutein cell contains two or more nuclei, through the fusion of adjacent lutein cells. 2. The lutein cells are almost entirely radially arranged, as seen in the corpus luteum formation, by the early absorption of the connective tissue among them. 3, Eosinophil masses exist here and there, which may be formed by the colloid degeneration of the lutein cells. But there can never be seen the hyaline degeneration or the calk deposit in the lutein cells. In the above described manner, the majority of the lutein cells disappear leaving no trace, but some of them are converted into pigment cells containing yellowish pigment in the cytoplasm. These pigment cells are usually seen among the honey-combed lutein cells or in the remained connective tissue reticulum. As to the origin of the pigment, the rabbit falls in with the same category of the mouse. The connective tissue reticulum and the central nucleus of the cor- pus luteum are gradually absorbed and disappear following the fatty degeneration of the lutein cells, and the growth of them can scarcely be met with. In some cases, however, they remain after the disappearance of the lutein cells, and form a kind of corpus fibrosum. The situation of these bodies is not definite, some existing at superficial and others at the profound part of the ovary. The corpus albicans made by the further contraction and hyaline degeneration of this corpus fibrosum can never be seen in my whole sections. The majority of the blood vessels are obliterated and absorbed gradually together with the connective tissue, and a part of them is reverted into those of the ovarian stroma. Their multiplication or hyaline degeneration can also never be observed as in the case of the mouse. Summary. Concerning the results in studying the corpus luteum of the rabbit, the following observations are to be particularly emphasized. A. Development of the corpus luteum. 1. The majority of the polygonal cells of the theca interna of the 356 Cli. Togari, mature follicle degenerate and disappear during the corpus luteum formation, but a part of them remains and forms the theca lutein cells of the fully grown corpus luteum. 2. The granulosa lutein cells are derived from the granulosa cells of the mature follicle after rupture, not only by their hypertrophy but also by their hyperplasy. The nuclei of the granulosa lutein cells contain usually nucleoli, which appear soon after ovulation, although they can not be seen in the mature follicle. 3. The connective tissue reticulum among the lutein cells and the central connective tissue nucleus are formed from the spindle-shaped cells of the theca interna and the theca externa by their multiplication' with cell divisions. 4. The blood vessels in the corpus luteum are derived from those of the theca interna, their larger stems (small arteries and veins) existing chiefly at its peripherial part, and the capillaries in the central part where they may sometimes extend extremely as a sinus. B. Retrogression of the corpus luteum. 1. The theca lutein cells in their retrogression do not revert to the former connective tissue cells, but are absorbed after their degeneration as the granulosa lutein cells. 2. The majority of the granulosa lutein cells disappear by their fatty degeneration, and a part of them after they were converted into the pigment cells. 3. The connective tissue reticulum and the central connective tissue nucleus are gradually absorbed and disappear following the degeneration of the lutein cells, but remain long in part to form a kind of the corpus fibrosum. Even this corpus fibrosum disappears finally, not forming the corpus albicans by the further contraction and hyaline degeneration of the former. 4. The capsule of the corpus luteum originated from the theca externa of the Graafian follicle, remains until the late stage of the retrogression of the corpus luteum, and may be utilized as the criterion of the distinction between the corpus luteum and the interstitial gland in their retrogression (C o h u). 5. The majority of the blood vessels of the lutein layer are oblite- rated and abosorbed gradually together with the connective tissue of the corpus luteum, and a part of them is reverted to the stroma vessels with- out hyaline degeneration. 6. By the retrogression of the corpus luteum graviditatis, a little colloid, but scarcely no calk, is noticed. On the Corpus Luteum of the Rabbit. 357

Comparison of the corpus luteum of the mouse and the rabbit. The results obtained from the comparison of the corpus luteum of the mouse, formerly published, and that of the rabbit, above described, are as follows : (This description is laid upon the foundation by the Sob° tta's report as to the corpus luteum formation of the rabbit). The corpus luteum and the mature follicle of the rabbit are larger than those of the mouse, and the former is several times as large as the latter, although the duration of the corpus luteum graviditatis existence between the two kinds of animals is of but little difference. The mature follicles of the rabbit are rich in liquor folliculi, but poor in the granulosa layer compared with that of the mouse. The so-called C all-E x ner bodies which are considered as they may serve for the liquor formation, are seen in the mature follicle of the rabbit, whereas they are almost none in the mouse. The is lacking in the rabbit showing the mature follicles of so-called Mj ass oj edo ff's 'Gr a af- sches Blaeschen des Typus II', contrary to the mouse which possesses `Girra a fsches Blaeschen des Typus I.' The polygonal cells are a little larger in amount than those of the mouse. The spontaneous ovulation of the rabbit is usually lacking, although the heat period comes following parturition ; contrary to the mouse in which it may usually occur without any external stimuli at any time during a period beginning at 23 and ending at 33 hours after parturition. The relative positions of all the layers of the follicle directly after ovulation in the rabbit is much clear than that of in the mouse. He- matoxylin-eosin would bring them out distinctly in case of the rabbit but it is not quite so in the mouse. This difference may be caused by the quantity of the liquor folliculi flowed out at the time of ovulation. The small primary breeding occurs usually at the time of rupture, but the large secondary seldom during the corpus luteum formation in the rabbit ; contrary to the mouse, in which the primary breeding takes place rather rarely, but the large secondary more frequently than the former. The congestion of the ovary during the corpus luteum forma- tion of the mouse is more remarkable than that of the rabbit. The majority of the polygonal cells of the theca interna of the ma- ture follicles degenerate and disappear during the corpus luteum forma- tion, but a part of them remain and are seen in the fully grown corpus luteum as the theca lutein cells, that is very different from the mouse, in which all the polygonal cells degenerate and disappear during the period of the corpus luteum formation, the lutein cells being derived from the 358 Ch. Togari, granulosa layer only. The granulosa lutein cells are multiplied with cell divisions during a period from 3 to 4 days after copulation in the rabbit. This phew., menon is lacking in the mouse and the lutein cells of the latter are formed only enlarged granulosa cells of the mature follicle. The connective tissue reticulum among the lutein cells are derived from the spindle-shaped cells of the theca externa and interna in both animals. In the rabbit these cells are distributed minutely from the early period of the corpus luteum formation and they are converged into one another and surround almost all the lutein cells in the fully grown corpus luteum. On the contrary, in the mouse they are not so minute at first, and then they gradually become minutely to surround the several , lutein cells so as to form a lobe in the fully grown corpus luteum. The wandering cells found during the period of the corpus lutenrn formation are polynuclear leucocytes in the rabbit, but mononuclear in the mouse, and the former is smaller in number than the latter. The blood vessels of the corpus luteum penetrating from the theca interna of the mature follicle, are the same in the arrangement to the connective tissue reticulum in both animals, and the rabbit possesses the small arteries and veins chiefly at the peripheral part of the corpus luteum, but the mouse capillaries only. The central connective tissue nucleus is commonly formed in the rabbit, but in the mouse, its formation is indefinite coincident with the liquor accumulation in the central part of the young corpus luteum. The orifice made by rupture is wide and remains opened several days long in the rabbit, but it is narrow and closes perfectly within a few hours after ovulation in the mouse. The corpus luteum of the rabbit in the early period of its formation shows usually the appearance of the offener Kelch ' owing to the long existence of the orifice of rupture, but in the mouse, it frequently shows a vesicle by the accumulation of the liquor in the central part of the corpus luteum. In the rabbit, some of the granulosa epithelium which evaginated from the orifice of rupture and hung on the surface of the ovary as a mushroom is converted into lutein layer. In the mouse, this pheno- menon, however, can never be seen. The corpus luteum graviditatis degenerates actively soon after parturition showing the groups of degenerating lutein cells with wander- ing mast cells here and there in the rabbit ; contrary to the mouse, in which it exists long after parturition, and then degenerates gradually On the Corpus Luteum of the Rabbit. 359 being pressed passively by the adjacent growing follicles or corpora lutea. Consequently the outer border of the corpus luteum in the rabbit is clearly marked off from the surrounding tissue to the late stage of its degeneration, quite opposite to the mouse. The central connective tissue nuclei in 'the corpora lutea exist long, some forming the corpora fibrosa and the other disappearing with the degeneration of the lutein cells in the rabbit. Contrary to this, in the mouse, even in the fully grown stage, some of them are absorbed and have disappeared already ; therefore, in the advanced stages of the corpus luteum degeneration they can never be seen. The yellowish pigment cells found in the degenerating corpora lutea are less frequent and smaller in amount in the rabbit than in the mouse. In the retrogression of the corpus luteum in the mouse, the central cavity formation in the ovary plays an important rOle, but this cavity formation is lacking in the ovary of the rabbit.

Conclusions. All the layers of the mature follicles of the rodent may take at least the following changes during the period of the corpus luteum formation and during its retrogression, considering the results obtained by the comparative study of the corpora lutea in the mouse and the rabbit. A. The sheath of the mature follicle consists of two layers ; the theca externa composed of spindle-shaped cells and the theca interna of polygonal cells and the spindle-shaped cells which are morphologically identical with those of the theca externa. The capillary blood vessels exist among these cells and form a delicate anastomosing plexus near the basement membrane. In the mature follicle just before rupture, almost all the blood vessels which lie at the part so-called Wald eyer 's ' Stigma ' are driven out to the deeper part of the follicle by pressure ; so that the breedings are very small or could never be seen by the rupture of the follicle. Some young corpora lutea with wide and long existing orifice of rupture, lack the vigorous accumulation of the blood, although some are in the state of the corpus haemorrhagicum, when the orifice closed early in the corpus luteum formation. 1. The theca externa surrounding the outermost part of the follicle is concentric in its arrangement. When the mature follicle is ruptured, its arrangement becomes more or less irregular, and then a part of this layer takes part in the formation of the connective tissue reticulum of the corpus luteum, and the other larger part remains to form a capsule of 360 Oh. Togari, the corpus luteum by recovering its concentric -arrangement. The changes which take place in the capsule during the period of the corpus luteum degeneration in these two animals may be classified as follows : namely, the one disappeares early by its absorption or is mixed with the stroma connective tissue, while the other exists long to its late stage showing a clear limit against the adjacent tissue. 2. The polygonal cells of the theca interna are formed by the enlarged spindle-shaped cells they degenerate and disappear during the period of the corpus luteum formation in a certain species, but in the other, some of them remain as the theca lutein cells near the capsule of the fully grown corpora lutea, although the majority of them degenerates soon after rupture. Hence, the existence of the theca lutein cells is quite different in different animals. 3. The spindle-shaped cells of the theca interna in the mature follicle show their arrangement as the sheath falls under nearly so-called three layers of. Koellik er. They penetrate into the granulosa layer radially with mitotic divisions during the early period of the corpus luteum formation, some of them reaching the central part where they form the central connective tissue nucleus, and the other larger parts of cells among the lutein cells converge into one another, so as to form the connective tissue reticulum. The connective tissue nucleus and the reticulum are not usually increased during the period of the corpus luteum degeneration, but they are absorbed and disappear together with the degeneration of the lutein cells, or some of them remain to form the corpus fibrosum. The latter never suffers further contraction and hya- line degeneration to form the corpus albicans. 4. The invasion of the capillaries of the theca interna into the granulosa layer before rupture can never be seen in the normal condi- tion. They penetrate into the granulosa layer following the spindle- shaped connective tiosue cells after rupture, and they lie also radially arranged at first and then give side branches to come to contact with one side of almost every lutein cells, some invading into the connective tissue nucleus. These blood vessels in the time of retrogression are obliterated and absorbed gradually. B. From the primitive follicles up to the mature follicles, these is always the basement membrane, consisting of delicate connective tissue fibers densely arranged. It can never be seen that it separate into delicate fibers to penetrate into the granulosa layer before rupture, although it disappears entirely soon after ovulation. C. The granulosa layer of the mature follicle shows a similar On the Corpus Luteuua of the Rabbit. 361 relation in its thickness to that of the theca interna, its outermost cells being arranged regularly. Directly after ovulation, the majority of the granulosa cells remain rather in a loose arrangement, especially their outermost cells becoming irregular, although a few of them are dis- charged with the liquor folliculi and ovum. The degenerative process can never be seen among the granulosa cells before or after ovulation. These cells are, afterwards, converted into lutein cells by their hypertrophy only or both by their hypertrophy and hyperplasy. The nuclei of the granulosa lutein cells contain usually nucleoli contrary to the granulosa cells of the follicle, and the cytoplasm are at first in granular appearance, but become gradually honey-combed by depositing lipoid. During the degeneration period of the corpus luteum, their nuclei show manifold figures of degeneration, and the honey-combed appearance of the cyto- plasm becomes further remarkable and at last large vacuolated or homogenous. Thus the majority of the lutein cells disappear by fatty met-amorphosis, but some of them are converted into pigment cells containing lipoid, and then the latter also disappear ultimately. Even in the corpus luteum graviditatis, in general, we can scarcely met with colloid and calk. D. The liquor folliculi may be produced by the secretion of the granulosa epithelium and the transudation from the blood vessels. The chief cause of the ovulation may be the rapid increase of the internal pressure by the accumulation of the liquor folliculi, not by the contrac- tion of the smooth muscle fibers or elastic fibers in the wall of the follicle, as it is advocated by several authors. Nor is the rupture of the follicle caused by the penetration of the capillaries in the theca interna into the granulosa layer, although their congestion may play an import- ant rOle for ovulation. The quantity of the liquor folliculi flow out with ovum by rupture and the liquor accumulation in the central cavity of the young corpora lutea are different in different animals. In conclusin, I wish to express my hearty thanks to Professor T. As a i for his guidance and useful advices.

Explanation of figures (Plate XVIII.XIX). Fig. 1. A part of the wallof the mature follicle, showing the Mj ass oj edoff 's so-called Graaf schesBlaeschen des TypusII' with its ovumin the secondmatura- tion division(3 hours after copulation). ZeissOk, K 2, Obj. 16 mm . Fig. 2. Freshly ruptured follicle, showing the orifice of rupture and the majority of remaininggranulosa cells (10 hours after copulation). Zeiss Ok, K8. Obj.16 mm. 362 Ch. Togari,

Fig. 3. Enlarged view of a part of the figure 2., showing the thicker and the thinner part of the theca interna. Zeiss Ok. K8. Obj. 8 mm. Fig. 4. Young corpus luteum, showing the granulosa plug at the point of rupture (24 hours after copulation). Zeiss Ok, K2, Obj. 16 mm. Fig. 5. A part of the young corpus luteum, showing the degeneration of the polygonal cells of the theca interna and the invasion of its capillary (33 hours after copulation). Zeiss Ok. K12, Obj. 8 mm. Fig. 6. A part of the young corpus luteum, showing the mitotic division of a granulosa lutein cell (96 hours after copulation). Zeiss Ok. K8, Obj. 16 mm. Fig. 7. A part of the fully grown corpus luteum, showing granulosa and theca lutein cells (10 days after copulation). Zeiss Ok. K12, Obj. 8 mm. Fig. 8. A part of the corpus luteum in degeneration, showing the clear outer limit by the capsule and the manifold degenerating figures of the lutein cells (5 days after parturition). Zeiss Ok. K12, Obj. 8 mm. Abbreviations. AF antrum folliculi point of rupture BM basement membrane GR membrana granulosa BV blood vessel H honey-combed lutein cells in C capsule of the corpus degeneration luteum M granulosa lutein cell in mito- CP capillary tic division CT connective tissue OR orifice of rupture G group of degenerating OV ovum lutein cells with pyc- TA tunica albuginea notic nuclei TE theca externs, GL granulosa lutein cells TI theca interna GP granulosa plug on the TL theca lutein cells

Bibliography. 1. A d ac hi, K., Vber die kuenstlichen Erzeugung von • Corpus luteum mittels Injektion der Plazentaremulsion (I. Mitteilung). Osaka Igakkai Zasshi. Bd. 22. 1923. 2. He tt, J., Das Corpus luteum der Dohle. Arch. f. Anat. u. Entw. Bd. 97. 1923. 3. -, Das Corpus luteum der Molches. Zeitschr. f. Anat. u. Entw. Bd. 68. 1923. 4. , Das Corpus luteum der Zauneideckse. Zeitschr. f. mikr.-anat. Forsch. Bd, 1. 1924. 5. Hiros e, T., On the artificial production of the corpus luteum in the rabbit. Nippon Fuginka Gakkai Zasshi. Vol. 16. 1921. 6. L e h n e r, J., Das Mastzellen-Problem und die Metachromasie-Frage. Ergebn. d. Anat. u. Entw. Bd. 25. 1924. 7. M atsu y a m a, R., On the formation of the corpus luteum. Jikken Igaku Zasshi. Vol. 3. 1919. 8. Mj ass oj e doff, S. W., Zur Frage iiber die Struktur des Eifollikels bei den Saeugetieren. Arch. f. mikr. Anat. u. Entw. Bd. 97. 1923. 9. Solomon s, B. & G a ten b y, B., Notes on the formation, structure and physiology of the corpus luteum of man, the pig and the duck-billed platypus. Journ. of Obst. a. Gyn. of British Empire. Vol. 31. 1924. 10. Toga r i, Ch. On the origin of the corpus luteum of the mouse. Aichi Journ. of Exper. Med. Vol. 1. 1923. 11. , On the retrogression of the corpus luteum of the mouse. Aichi Journ. of Exper. Med. Vol. 1. 1924. TAF.XVIII. FOLIAANATOMICAJAPONICA,BD.IV. OV

C.Togari. FOLIAANATOMICA JAPONICA, BD. IV. TAF. XIX.

C. Togari.