contents are extruded (fig. 5, b). The striæ in the membrane are believed to be minute pores, and are supposed, while the ovum is yet within the Graafian follicle, to permit the passage of granules of nutrient material into the interior of the ovum. After the ovum is discharged from the follicle, the spermatozoa may perhaps find their way into the ovum through these pores. According to Retzius the protoplasm of the ovum is united with the follicle-cells by fibres which pass through the pores of the zona.

Immediately surrounding the zona radiata, as the ovum lies within the mature Graafian follicle, is a thin stratum of granular substance, probably deposited upon the exterior of the ovum by the innermost cells of the discus proligerus, which immediately encircle the ovum within the follicle. When the Graafian follicle bursts and the ovum is set free, this granular material appears to imbibe water, and, as is specially noticeable in the ovum of the rabbit. swells up into a clear gelatinous envelope, which has been termed, from a possible homology with the white of the bird's egg, the albumen. But in the mammal this structure has not the nutritive importance to the embryo which is possessed by the corresponding formation in the bird, and it disappears during the passage of the ovum down the Fallopian tube.

The substance of the ovum within the tunica radiata is known as the vitellus or yolk (fig. 6, vi). It is a soft semi-fluid substance, composed mainly of protoplasm, which is filled with globules and granules (yolk-granules) of different

Fig. 5.-OVARIAN OVUM OF A MAMMIFER. (Allen Thomson.)

a, the entire ovum, viewed under pressure; the granular cells have been removed from the outer surface, the germinal vesicle is seen in the yolk substance within; b, the external coat or zona burst by increased pressure, the yolk protoplasm and the germinal vesicle having escaped from within; c, germinal vesicle more freed from the yolk substance. In all of them the macula is seen.

Fig. 6.-OVUM OF THE CAT; HIGHLY MAGNIFIED. SEMI-DIAGRAMMATIC. (E. A. S.)

zp, zona pellucida, showing radiated structure; vi, vitellus, round which a delicate membrane is seen; gr, germinal vesicle; gs, germinal spot.

sizes, but all small, and possessing a high index of refraction. Examined in the fresh condition, the protoplasm between the granules looks perfectly clear and structureless, but after treatment with suitable reagents, it may be seen to consist of a fine reticulum, which is especially fine and close near the periphery of the ovum, and also around the germinal vesicle, at which places the yolk granules are in less amount than elsewhere. The substances which occur within an ovum other than the nucleus and protoplasm, may, as in cells generally, be collectively designated deutoplasm"; they are regarded as furnishing a supply of nutrient matter to the protoplasm during the earlier stages of development.

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Embedded in the protoplasmic vitellus, usually eccentrically, is a large spherical

nucleus, which was termed by its discoverer, Purkinje, the germinal vesicle. This, which is about th inch in diameter, has all the characters of the nucleus of a cell: It consists of a nuclear membrane enclosing a clear material or matrix, embedded within which may be seen strands of karyoplasm, enclosing one or more well-marked nucleoli (fig. 6, gv). Frequently there is but one nucleolus, which is then large and prominent, and has received the name of germinal spot (macula germinativa, Wagner, 1835).

There is some doubt whether, before fertilization, there is another membrane (vitelline membrane) enclosing the vitellus within the zona radiata. The evidence of the presence of such a membrane is by no means clear, although its existence has been maintained by very competent observers (v. Beneden, Balfour).

The mammalian ovum (that of monotremes alone excepted) differs from that of other vertebrates in the relatively small amount of nutritive material (yolk granules, deutoplasm) which is embedded in its protoplasm. In fishes, amphibia, reptiles, and especially in birds, the amount of such nutritive material is vastly greater than that of the protoplasm itself, so that the very existence of the latter is obscured in most parts of the ovum, and it is only in the immediate neighbourhood of the germinal vesicle that the protoplasm can be distinctly

Fig. 7.-DIAGRAM OF A HOLOBLASTIC (ALECITHAL) OVUM (A) AND OF A MEROBLASTIC (TELOLECITHAL) OVUM (B). (E. A. S.)

Only a small part of the latter is represented. The yolk or food material is represented in both by clear globules, which in B are seen vastly to preponderate, except in the immediate neighbourhood of the germinal vesicle.

recognized (fig. 7, B). It is here also that, after fertilization, the more active changes in the ovum occur, and it is this part alone in which in the bird and most other oviparous vertebrates the process of division or segmentation of the yolk and consequent formation of embryonic cells proceeds. Hence these ova are said to undergo a process of incomplete segmentation, only a part of the ovum appearing to undergo development, and they are accordingly termed meroblastic to distinguish them from those (like the mammalian ova) in which the yolk or nutritive material is everywhere in relatively small proportion to the protoplasm, the whole of which undergoes division after fertilization, and participates in the formation of the embryo (holoblastic ova). This small amount of nutritive material in the mammal is obviously related to the fact that the mammalian ovum early acquires an attachment to the maternal system from which it is then able directly to derive its nutriment, whereas the meroblastic ovum of oviparous vertebrata necessarily contains all the nutriment required by the developing bird, reptile, or fish, until it is sufficiently advanced in development to emerge from the egg and obtain food independently. Although, however, the mammalian ovum is holoblastic, it is none the less clear, from a comparison of the early stages of its development with that of the bird, that the ancestors of the mammalia must have had ova of the meroblastic type.

Balfour has further conveniently distinguished between those ova in which there is a great accumulation of nutritive or yolk material at one pole (telolecithal ova, as in the bird, reptile, and fish amongst vertebrates), those in which the accumulation of yolk is in the middle of the ovum (centrolecithal ova, as in arthropods), and those in which it is scattered pretty equally in small amount throughout the protoplasm without any very marked accumulation

1 Purkinje discovered the germinal vesicle in the bird's ovum in 1825; that of mammals was first noticed by Coste in 1833.

(alecithal ova, as in mammals, Amphioxus, echinoderms). It is clear that these conditions of arrangement of the proto- and deuto-plasm within the ovum are the main factors in determining variations in the process of segmentation.

Maturation of the ovum. Formation of polar globules.-Either before its escape from the Graafian follicle, or immediately after, the ovum undergoes a peculiar change, preparatory to, but nevertheless altogether independent of fertilization, which consists of a process of unequal cell-division or germination, and results in the extrusion from the vitellus of two minute spherical bodies (fig. 8), which have

HOURS AFTER IMPREGNATION.

(Bischoff.)

On the zona a, spermatozoa are seen, and others in the perivitelline space; b, the polar globules.

been termed the polar globules or directive corpuscles, from a supposition that their presence determines the pole at which the first segmentation will take place should the ovum become fertilized. It is, however, ncertain whether there is any constant relationship of this kind, but it is none the less clear that the extrusion

of the polar globules is an event of the highest importance for the due development of the ovum, since until this has happened the ovum appears to be incapable of complete fertilization and segmentation. What is actually extruded is a small part of the nucleus of the ovum, or, to speak more precisely, two small parts of its nucleus in succession, probably surrounded by a very thin investment of protoplasm. Prior to this extrusion, the germinal vesicle approaches the periphery of the vitellus, loses its distinctness of outline, and after passing through phases which are characteristic of a nucleus which is about to divide, does actually undergo a division into two, the one part being extruded into a space (perivitelline), which has become formed in consequence of the shrinking or contraction of the ovum, and the other part remaining in the vitellus, only, however, to repeat the process of division, and to form a second extruded globule. The remainder of the germinal vesicle, which is now termed the female pro-nucleus, leaves the periphery of the vitellus for a situation nearer to the centre, where, if fertilization should supervene, it awaits the advent of the male pro-nucleus, which is formed from a spermatozoon. After the two pronuclei have come together, a new and complete nucleus is formed by their conjugation.

The actual formation of polar globules has not hitherto been observed in the human ovum, although there is no doubt whatever that it takes place. In the rabbit various stages in the process have been traced by E. v. Beneden and Rein, and it has also been noticed in other mammals. But the details of the process have been made out most precisely (by Fol, Hertwig, and others) in the transparent ova of echinoderms, and more recently and minutely (by 3. v. Beneden, Carnoy, Boveri, Zacharias, and others) in Ascaris megalocephala, a thread-worm parasitic in the horse, in which all the changes can be followed in one and the same ovum. or the various phases fixed by means of reagents in different ova, and these may afterwards be stained and studied with the utmost minuteness. The successive changes in such ova are represented in figs. 9 and 10. The polar globules remain visible for a time in the perivitelline fluid, and are even seen, should the ovum become fertilized, during the early stages of segmentation, but they ultimately disappear and are not known to take any further part in the subsequent changes which the ovum undergoes.

The fact that throughout the whole animal kingdom the extrusion of polar globules from the ovum as it becomes mature is almost universal, and that a similar process has also been observed to occur in plants indicates the great importance of the phenomenon. The significance will be further discussed after the process of fertilization of the ovum has been described.

1 The ovum may, however, receive a spermatozoon before the completion of the formation of polar globules.

Fig. 9.-FORMATION OF THE FIRST POLAR GLOBULE IN THE EGG OF ASCARIS MEGALOCEPHALA. (v. Gehuchten.)

A. The ovum with the germinal vesicle transformed into a spindle of (achromatic) fibrils: from the poles of the spindle other fibrils radiate into the protoplasm. At the equator of the spindle eight portions of chromatin are visible; cs, head of a spermatozoon which has previously entered the ovum, and is becoming transformed into the male pro-nucleus; m, gelatinous membrane of the ovum.

B. The chromatin particles are seen separated into two sets. The achromatic fibrils are not shown in this preparation. The ovum is considerably shrunken.

C. Half of the germinal vesicle is extruded into a perivitelline space, and along with a portion of protoplasm is becoming separated off from the ovum as a polar globule. The extruded half includes four of the chromatin particles; the other four remain in the ovum; m', membrane dividing the polar globule from the ovum.

A. The remainder of the germinal vesicle (after extrusion of the first giobule, g1) has again become transformed into a spindle of achromatic fibrils, with the four remaining chromatin particles at the equator of the spindle.

B. The spindle, now irregularly Y-shaped, is seen approaching the surface of the ovum; g', first polar globule; ns, male pronucleus which has become formed from a spermatozoon.

C. Extrusion of half of the germinal vesicle remainder.

D. Completion of the process; the second polar globule, g, is now separated from the ovum ; i contains two of the chromatin particles. The other two remain in what is left of the germinal vesicle, n which now forms the female pronucleus; ns, male pronucleus; g', first polar globule.

Fertilization.—The ovum, after its expulsion from the Graafian follicle is received upon the fimbriated end of the Fallopian tube. The fimbriæ are covered by a prolongation of the ciliated lining of the tube, and the action of the cilia serves to propel the minute ovum into and along the tube towards the uterus. In this passage it may, if impregnation have occurred, meet with the spermatozoa, one or more of which may penetrate the zona pellucida, and fertilize the ovum. It is possible in some instances for fertilization to occur on the fimbriated extremity of the tube, or in the body of the uterus, but it is probable that in most cases it happens in the tube itself.

It is probable that normally only a single spermatozoon enters the vitellus. If it should happen that two or more enter, normal development does not as a rule occur. Exceptions to this rule have, however, been recorded.

The changes in the ovum which accompany fertilization have, like those which result in the formation of the polar globules, been studied most satisfactorily in the transparent ova of echinoderms and in Ascaris. In the former (fig. 11) the spermatozoa

Fig. 11. FERTILIZATION OF THE OVUM OF AN ECHINODERM. (Selenka.)

8, spermatozoon; m.pr, male pronucleus; f.pr, female pronucleus.

1. Accession of a spermatozoon to the periphery of the vitellus; 2. Its penetration, and the radial disposition of the vitelline granules; 3. Transformation of the head of the spermatozoon into the male pronucleus; 4, 5. Blending of the male and female pronuclei.

may be seen to penetrate the gelatinous investment which here takes the place of a zona pellucida, and the head, of one only as a rule, to imbed itself in the periphery of the ovum, which becomes slightly protruded at the point of contact. According to v. Beneden's account, the spermatozoon always enters in Ascaris at a particular part of the ovum (polar disc), at which part there is an aperture in the vitelline membrane (micropyle). When once it has passed into the ovum, this aperture becomes closed, and the head of the spermatozoon rapidly increases in size, and acquires the appearance of a nucleus which, in contra-distinction to the remains of the germinal vesicle, or female pro-nucleus, is termed the male pro-nucleus. Soon it leaves the periphery, and passes towards the centre of the ovum in the direction of the female pro-nucleus. In its passage through the protoplasm it appears to exercise a peculiar attraction upon the granules in that substance, for these become arranged in its vicinity in radiating lines. The tail of the spermatozoon has in the meantime disappeared, whether by being cast off or by blending with the protoplasm of the ovum has not certainly been made out. As the male pro-nucleus approaches the female pro-nucleus, the latter moves somewhat to meet it, and presently the two pro-nuclei come into contact and together form a new nucleus, com