Book - The Frog Its Reproduction and Development 6
Chapter 6 - Blastulation
Internally the 8 -cell stage of the frog shows the beginning of a cavity where the well-formed animal hemisphere cells have completed their 6-sided cell membrane. Once a blastomere is formed it tends to acquire a certain degree of rigidity and will maintain its spherical cell boundaries independently of its neighbors. Cortical and vitelline membrane pressure may flatten cells slightly against each other. When segmentation of the egg occurs, dividing it into smaller and smaller cellular units, there naturally follows the appearance of an internal cavity known as the segmentation cavity or blastocoel. The aggregate becomes about 20 per cent larger than the total volume of the cells. The earlier cleavages (described above) tend to be perpendicular to the egg surface. However, after the 32-cell stage there appear division planes more or less parallel to the egg surface, cutting off surface protoplasm from inner protoplasm and yolk. With increased cell rigidity, following each division, these surface cells push against each other until they are lifted up and away from the underlying cells. A crude simile would be to hold four or more marbles tightly together in the hand, and no matter how much pressure is exerted there will always be a space in the center of the group of marbles (i.e., cells). In the frog's egg the formation of complete cells is rather slow, and the blastocoel is small and not readily apparent until about the 32-cell stage.
The blastocoel is therefore a cavity, and this stage of development is known as the blastula, overlapping the stages of cleavage. The cavity is enlarged with each of the early cleavages and it is filled with an albuminous fluid, arising from the surrounding cells. Even though the frog's egg is telolecithal, cleavage has been described as holoblastic (opposed to meroblastic) and the presence of this cavity distinguishes it as a coeloblastula (as opposed to stereoblastula which has no cavity). Since the horizontal cleavages appear toward the animal hemisphere, this newly forming blastocoelic cavity will appear in an eccentric position above the level of the equator, and slightly toward
Animal hemispheres^ Segmentotion cavity (blastocoel)
~ Vegetal hemisphere
Segmentotion covity (blastocoel)
Hemisected late blaslulo
Blastulation in the frog. (Redrawn and modified after Huettner.)
Frog blastula: sagittal section.
Progressive stages in blastulation.
the gray crescent side of the cleaving egg. It will remain in this position, beneath the animal hemisphere, until it is later displaced by the development of other cavities. The size of the blastocoel increases with the formation of smaller and smaller surrounding cells. The late blastula of the frog, by virtue of this blastocoel, has a volume displacement about 20 per cent greater than that of the fertilized egg.
Following the 32-cell stage the egg loses all semblance of rhythmic or synchronized cleavage and there is developed a gradient of cleavage with the most active region being at the animal pole and the least active being at the vegetal pole. In addition to this there are some characteristic changes in the bias tula of the frog embryo. First, the smaller pigmented animal pole cells tend to spread their activity in a downward direction toward the vegetal pole, by a sort of contagion, so that there is an actual migration of pigment-covered cells toward the vegetal pole. This, and the horizontal cleavages, result in increasing the cell layers but thinning of the roof of the blastocoel. Second, the horizontal cleavages in the very small animal pole cells give the blastula a double or multi-layered roof. The single outer layer of cells contains most of the superficial pigment and is now recognized as the epidermal layer which will give rise to epithelium, either of the integument or lining the nervous system. The inner tiers of cells of the blastular roof are less pigmented and are known collectively as the nervous layer because they will give rise largely to the neuroblasts of the nervous system.
Either on the surface of the whole blastula or in sections of the blastula, one can determine the margins of the downward-moving pigmented coat and active animal hemisphere cells and the consequent slight thickening of the equatorial band. This margin is known as the marginal zone, marginal belt, or germ ring. Such a marginal region of activity, where yolk is most actively being transformed into cytoplasm, is found in Amphioxus and chick, and probably in all vertebrate embryos. It will have much to do with the subsequent formation of the lips of the future blastopore.
The marginal zone is approximately equatorial in the blastula stage, except for the slight reduction in pigmented cells at the side where the original gray crescent was located. Opposite to this gray crescent region the wall of the blastula appears to contain relatively more layers of cells and is therefore somewhat thicker. These changes occur in anticipation of the next process in embryonic development, namely gastrulation.
There have been numerous attempts to explain blastocoel formation and the movements anticipating gastrulation. These have been based largely on purely physical phenomena. An attempt has been made recently to summarize these concepts, as follows (Holtfreter, 1947: Jour. Morph., 80:42):
If present in large numbers, the cells at the periphery of the body tend to establish a semipermeable layer, while the cells of the interior become separated from each other by secretion fluid. Thus the aggregation may be transformed into the configuration of a blastula or, if the amount of internal fluid increases further, into an epithelial vesicle. The occurrence of cylindrical stretchings and of migrations of the peripheral cells into the interior of the vesicle has been interpreted as possibly being caused by a surface tension lowering action of the inner fluid. Even in the absence of a gradient of surface tension, however, isolated embryonic cells tend to elongate and to migrate in one direction, with their original proximal pole leading the way, and possibly invagination movements may be partly due to this inherent dynamic tendency of the individual cell. Other factors influencing shape and movements of the cells in aggregate, or in a normal embryo, appear to be cell-specific differences of adhesiveness arising in the course of differentiation and making for a sorting out, grouping and aggregation of the various ceil types into tissues and organs. With the appearance of fibrous and skeletal structures, of local differences in mechanical stress and hydrostatic pressure, of differential growth and a variety of metabolic processes, so many new factors are introduced that the principle of interfacial tension loses more and more of its original primacy as a morphogenetic agent.