themselves around the ends of Sertoli's columns, a phenomenon which was formerly regarded as representing a copulation of the two elements, although it was clearly understood that no real fusion or interchange of chromatin occurred, but that the close relations of the two were for the purpose of furnishing nourishment to the developing spermatosomes. The whole forms a spermatoblast of von Ebner. Since the spermatids lining the lumen are changed into spermatozoa, and the process is repeated in the cells of the deeper layers as they come to the surface, the result is that the entire column is finally used up. The compensatory elements are supplied by the proliferation of the adjacent spermatogonia. The resulting products again divide, and thus build up an entirely new generation of spermatogenic cells. Hand in hand with these progressive phenomena occurs an extensive destruction of the cells taking part in spermatogenesis. This is shown by the presence of so-called karyolytic figures in the cells, which later suffer complete demolition.

These developmental changes are represented in the preced

Fig. 285.-Section of convoluted tubule from rat's testicle (after von Ebner, 88). The pyramidal structures are the sustentacular cells, together with spermatids and spermatosomes. Between these are spermatogenic cells, some of which are in process of mitotic division. Below, on the basement membrane and concealing the spermatogonia, are black points representing fat-globules, a characteristic of the rat's testicle. Fixation with Flemming's fluid.

ing schematic figure (Fig. 284), and may in part be observed in figure 285.

In mammalia it has been possible to trace the development of the spermatids into the spermatosomes. These phenomena have been studied and described by numerous writers, and although many conflicting views have been expressed, the essential steps of this process seem quite clearly established. The account here given is based in part on the recent observations of v. Lenhossék and the observations of Benda. Before considering the method of development of the spermatosomes from the spermatids, a few words concerning the structure of the latter may be useful. The sharply outlined spermatid possesses a slightly granular protoplasm and a round or slightly oval nucleus with a delicate chromatic network. In the protoplasm there is found a sharply defined globule, known as the sphere or sphere substance, which lies near the nucleus and

presents throughout a nearly homogeneous structure. This substance is first noticed in the spermatocytes, disappears during the cell-divisions resulting in the spermatids, and reappears in the latter. In the protoplasm of the spermatid, lying near the nucleus, there is further found a small globular body, the chromatoid accessory nucleus of Benda, smaller than the sphere and staining very deeply in Heidenhain's hematoxylin. A true centrosome may also be

found in the spermatid.

The nucleus of the spermatid develops into the head of the spermatosome, during which change the originally spheric nucleus. becomes somewhat flattened and at the same time assumes a denser structure and moves toward that portion of the spermatid pointing away from the lumen of the seminiferous tubule. Accompanying these changes in the nucleus, marked changes are observed in the shape and structure of the sphere, which marks the position of the future anterior end of the head of the spermatosome, and applies itself to the nucleus on the side pointing away from the lumen of the tubule. In this position it differentiates into an outer clear homogeneous zone and a central portion which stains more deeply and to which v. Lenhossék has given the name akrosome. From these structures are developed the head-cap and the lance of the spermatosomes, which differ in shape and relative size in the spermatosomes of the different vertebrates. Recent investigation seems to establish quite clearly that the axial thread of the tail is developed from the centrosome (from the larger, if two are present), which is situated at some distance from the nucleus. Soon after the beginning of the development of the axial thread the centrosome wanders to the posterior part of the future head of the spermatosome (the pole of the nucleus opposite the head-cap) and becomes firmly attached to the nuclear membrane in this position (observations made on the rat by v. Lenhossék, and on the salamander by Meves). The middle piece and the undulating membrane, it would appear, are differentiated from the protoplasm, although the question of the mode of their development is still open to discussion. The chromatoid body assumes a position near the axial thread at its junction with the cell membrane; its fate has not, however, been fully determined.

According to Hermann (97), the end-piece in the selachia is derived from the centrosome, the ring-shaped body from the invaginated half of the intermediate body of the spermatid formed during the last spermatocytic division, and the axial thread from filaments of the proximal half of the central spindle. The lance, according to him, represents a modified portion of the nuclear membrane of the

spermatid.

For further particulars regarding spermatogenesis see the investigations of v. la Valette St. George, 67-87; v. Brunn, 84; Biondi, Benda, Meves, and v. Lenhossék.

TECHNIC.

278. The ovaries of the smaller animals are better adapted to study than those of the human being, since the former are more easily fixed.

279. The germinal epithelium and its relations to the egg-tubes of Pflüger are best studied in the ovaries of young or newly born animals— cats, for instance, being especially well adapted to this purpose.

280. Normal human ovaries are usually not easily obtainable. Human ovaries very often show pathologic changes, and in middle life frequently contain but few follicles.

281. Fresh ova may be easily procured from the ovaries of sheep, pig, or cow in the slaughter-houses. On their surfaces are prominent transparent areas the larger follicles. If a needle be inserted into one of these follicles and the liquor folliculi be caught upon a slide, the ovum may as a rule be found, together with its corona radiata. That part of the preparation containing the ovum should be covered with a cover-glass under the edges of which strips of cardboard are laid. If no such strips are employed, the zona pellucida of the ovum is likely to burst in the field of vision, giving rise to a funnel-shaped tear. These tears have often been pictured and described as preformed canals (micropyles).

282. The best fixing fluid for ovarian tissue is Flemming's or Hermann's (vid. T. 17, 18), either of which may be used for small ovaries or pieces of large ovaries; safranin is then used for staining. Good results are also obtained with corrosive sublimate (staining with hematoxylin according to M. Heidenhain), and also with picric acid (staining with borax-carmin).

283. The treatment of the Fallopian tubes is the same as that of the intestine; in order to obtain cross-sections of a tube it is advisable to dissect away the peritoneum near its line of attachment and then distend the tube before fixing. It is instructive to dilate the tube by filling it with the fixing agent, thus causing many of the folds to disappear.

284. No special technic is necessary in fixing the uterus and vagina. The epithelium is, however, best isolated with one-third alcohol (vid. T. 128).

285. Seminal fluid to which normal salt solution has been added may be examined in a fresh condition. The effect upon the spermatozoa of a very dilute solution of potassium hydrate (1% or weaker) or of a very dilute acid (acetic acid) is worth noticing. The spermatozoa of salamandra maculosa show the different structural parts very clearly (lance, undulating membrane, marginal thread, etc.). In macerated preparations (very dilute chromic acid), or in those left for some time in a moist chamber, the fibrillar structure of the marginal and axial threads may be seen quite distinctly. The spermatozoa may also be examined in the form of dry preparations (treatment as for blood), stained, for instance, with safranin. Osmic acid, its mixtures, and osmic vapors are useful as fixing agents, certain structures being better brought out so than by employing the dry methods.

286. In examining the testicle (spermatogenesis) it is advisable to begin with the testis of the salamander, which does not show such complicated structures as do the testes of mammalia. Here also either Flemming's or Hermann's fluid may be used as a fixing agent, the latter being

followed by treatment with crude pyroligneous acid (vid. T. 18). For the salamander Hermann recommends a mixture composed of 1% platinum chlorid 15 c.c., 2% osmic acid 2 c.c., and glacial acetic acid 1 c.c., and for mammalia the same solution with double the amount of osmic acid. This fluid is allowed to act for some days, the specimen then being washed for twenty-four hours in running water and carried over into alcohols of ascending strengths. Paraffin sections are treated as follows: Place for from twenty-four to forty-eight hours in safranin (safranin 1 gm. is dissolved in 10 c.c. of absolute alcohol and diluted with 90 c.c. of anilin water; vid. T. 119). After decolorizing with pure or acidulated absolute alcohol the sections are placed for three or four hours in gentian-violet (saturated alcoholic solution of gentian-violet 5 c.c. and anilin water 100 C.C. c.), and are then placed for a few hours in iodo-iodid of potassium solution until they have become entirely black (iodin 1, iodid of potassium 2, water 300); finally, they are washed in absolute alcohol, until they become violet with a dash of brown. The various structures appear differently stained: for instance, the chromatin of the resting nucleus and of the dispirem, bluish-violet; the true nucleoli, red; while, on the other hand, in the aster and diaster stages the chromatin stains red.

It is of especial importance that small testicles should not be cut into pieces before fixing, as this causes the seminal tubules to swell up and show marked changes, even in regions at some distance from the cut (Hermann, 93, I).

The treatment of the remaining parts of the male reproductive organs requires no special technic.

VI. THE SKIN AND ITS APPENDAGES.

A. THE SKIN (CUTIS).

THE skin consists of two intimately connected structures—the one, of mesodermic origin, is the true skin, corium or dermis; the other, of ectodermic origin, is the epidermis or cuticle. The superficial layer of the corium is raised into ridges and papillæ which penetrate into the epidermis, the spaces between the papillæ being filled with epidermal elements. Thus, the lower surface of the epidermis is alternately indented and raised into a system of furrows and elevations corresponding to the molding of the corium.

In the epidermis two layers of cells may be observed-the stratum Malpighii, or stratum germinativum (Flemming), and the horny layer, or stratum corneum. According to the shape and characteristics of its cells, the stratum germinativum may also be divided into three layers-first, the deep or basal layer, consisting of columnar cells resting immediately upon the corium; second, the middle layer, consisting of polygonal cells arranged in several' strata, the number of the latter varying according to the region of the body; and third, the upper layer, or stratum granulosum, which is composed, at most, of two or three strata of gradually flattening cells characterized by their peculiar granular contents.

All these cell layers consist of prickle cells, and for this reason the stratum Malpighii is sometimes known as the stratum spinosum. When these cells are isolated by certain methods, their surfaces are seen to be provided with short, thread-like processes. In section the cells appear to be joined together by their processes. Since it has been proved that the processes of adjacent cells do not lie side by side, but meet and fuse, they must be regarded as belonging alike to both cells. Between the fused processes, which are known as intercellular bridges, there exists a system of channels which is in communication with the lymphatic system of the corium. The prickles just mentioned are variously regarded by different investigators; some considering them to be exclusively protoplasmic

Fig. 286.-Under surface of the epidermis, separated from the cutis by boiling. The sweat-glands may be traced for a considerable part of their length; X40: a, Sweatgland; b, longitudinal ridge; c, depression; d, cross-ridge.

processes of the cells, others regarding them as derived from the membranes of the cells composing the stratum Malpighii. Ranvier and others ascribe a fibrillar structure to the peripheral portion of the cellular protoplasm, and, according to them, these fibrillæ, surrounded by a small quantity of indifferent protoplasm, form the processes. Ranvier has also shown that such fibrillæ may extend from one cell around several others before reaching their ultimate destination in other cells at some distance. (Fig. 288.) The cells of the stratum granulosum contain peculiar deposits of a substance to which Waldeyer has given the name of keratohyalin. This substance occurs in the form of irregular bodies varying in size and imbedded in the protoplasm. The nuclei of such cells always