In this article we will discuss about:- 1. Meaning of Gastrulation 2. Basic Mechanism in Gastrulation 3. Methods 4. Morphogenetic Movement 5. Different Chordates 6. Analysis of Mechanism.
Contents:
- Meaning of Gastrulation
- Basic Mechanism in Gastrulation
- Methods used to Study Gastrulation
- Morphogenetic Movement of Cells in Gastrulation
- Gastrulation in Different Chordates
- Analysis of Gastrulation Mechanism
1. Meaning of Gastrulation:
The blastula passes into the stage called gastrula by the process—Gastrulation. This process is extremely important in the ontogenetic process of an animal, because the blue-print of the future organisation is laid down during this phase.
During this crucial and dynamic process major presumptive organ-forming areas of the blastula become reorganised in a fashion that allows their ready transformation into the fundamental body plan of a species. Gastrulation is essentially a process of migration of cells from one place to the other in the embryo. Besides movement of cells, considerable nuclear differentiation also takes place.
In almost all animals it results in:
(i) The establishment and differentiation of three primary germinal layers—ectoderm, mesoderm and endoderm,
(ii) The establishment of nuclear differentiation and
(iii) The beginning of the control of genetic factors over development.
2. Basic Mechanism in Gastrulation:
The process of gastrulation involves following three cellular activities, cell-movement, cell-contact and cell-division. All these mechanisms are carried in a nicely co-ordinated and integrated way.
Number of factors are believed to be responsible for this coordination, but it has not been possible to pin point the final answer. It is undeniable that this process is controlled largely by intrinsic factors which are correlated with the external as well as internal conditions.
3. Methods used to Study Gastrulation:
The correct observation of incidences during gastrulation was started from the findings of W. Vogt in 1923. Vogt used vital dyes (Janus green and Neutral red) to mark the cells in an early gastrula and noted that cells during gastrulation actually migrate from one place to the other.
The vital dye technique of Vogt resulted into the application of several other methods:
(i) Visible differences in the cytoplasmic particles were used as natural marker,
(ii) Taging of the cells with carbon particles and
(iii) Taging of the cells with radioactive substances.
4. Morphogenetic Movement of Cells in Gastrulation:
During gastrulation, cells from one region of embryo move to another to take up their future fateful position. Two terms, emboly and epiboly which are quite opposite in their meanings, are generally applied to explain the process of movement.
Emboly means the throwing in or insertion of cells and epiboly signifies the extending upon. The movement of cells establishes a particular form and involved in organ formation in embryo—so this movement is designated as the morphogenetic movement. Fig. 5.15 shows the movement of cells in gastrulation.
Fundamentally, the morphogenetic movement is similar but the details of the process vary greatly.
Following types of cells movement occur:
Epiboly:
It involves the extension along the anteroposterior axis and peripheral divergence.
Emboly:
The inward movement of cells is classified into different types depending on the behaviour of migrating cells.
These are:
(i) Invagination:
It denotes the infolding of a layer of cells to form a cavity encircled by infolded cells. Generally in the gastrulation of Amphioxus and frog, the wall of the blastoderm is pushed inside the blastocoel. This creates a new cavity called the archentecon which communicates with the exterior by a blastopore.
This process of inpushing goes on and the inpushed layer forms the walls of the cavity. The archenteron (or primitive gut) completely obliterates the blastocoel.
(ii) Involution:
It implies the inward lotation of cells as seen in the gastrulation of amphibian and avian eggs. From one end near the edge of the blastoderm, the cells begin to move inwards to form the inner lining of the blastoderm.
(iii) Convergence:
It means the movement of cells to a particular region of the gastrula. In amphibian egg, the migration of cells to the external edge of the blastoporal lip is designated as convergence. The same phenomenon of convergence of cells is seen in the formation of primitive streak in chick embryo.
(iv) Divergence:
This phenomenon is opposite to convergence, when involuted cells diverge to take up their future positions inside the gastrula.
(v) Infiltration:
During this process, cells of the blastoderm infiltrate near the bottom of the blastocoel to form a second layer as seen in the gastrulation of chick.
(vi) Delamination:
This is a process of separation of a group of cells from others to form discrete cellular masses.
(vii) Extension:
The elongation of presumptive areas after they have moved inside the embryo is called the extension.
(viii) Cell proliferation:
It means the increase in the number of cells during gastrulation.
(ix) Concrescence:
It is similar to convergence. The cells from two sides migrate anteriorly along one axis, but in convergence the cells from two sides unite together and then move anteriorly.
The above terms are coined for the convenience of analysing the events in gastrulation. Recent observations have established that it is essentially a phenomenon of integration. It was, therefore, felt necessary to understand the whole process for a meaningful comprehension of individual event.
5. Gastrulation in Different Chordates:
i. Amphioxus:
The blastula of Amphioxus contains the potential endodermal cells at the vegetal pole, i.e., hypoblast which forms the floor of the blastula. The presumptive organ forming cells (i.e., notochordal, mesodermal, epidermal, etc.), form the epiblast.
The epiblast constitutes the roof of blastula. The blastocoel is large. The dorsal crescent (presumptive neural and notochordal cells) lies in the future dorsal lip region of the blastopore while the ventral crescent (mesodermal area) occupies the ventral lip.
With the onset of gastrulation, an increase of mitotic activity is observed in the dorsal and ventral crescent regions. With the activity of the different cells, the endodermal plate invaginates into the blastocoel. During this process of invagination, the dorsal portion moves at a faster rate to touch a point which marks the anterior end of the developing embryo.
The notochordal cells, occupying the mid-dorsal region of the blastopore, involute and occupy a mid-dorsal position in the developing archenteron. Then the ventral crescents gradually converge on either side of the notochordal cells. Thus the roof of the archenteron is composed of mesodermal and notochordal cells.
This process of eraboly is accompanied by epiboly when the ectodermal and neural cells extend along the antero-posterior direction. The extension of ectodermal cells and the proliferation, involution and infolding of presumptive endodermal, notochordal and mesodermal cells result in the formation of a double-layered embryo (Fig. 5.17). The external layer forms the ectoderm.
The internal layer has a dorsomedian area of notochordal cells with two bands of mesodermal cells. The rest of the inner layer is formed of endodermal cells. Rapid cell proliferation, accompanied by emboly and epiboly causes an anteroposterior elongation of the gastrula.
As the developing gastrula elongates in the anteroposterior direction, the ventral crescent is gradually shifted dorsalward along the inner lateral side of the blastoporal lip. As a result of convergence, the mesoderm comes to lie on the two sides of the notochordal material at the dorsal blastoporal lip.
At the end of gastrulation the blastopore becomes smaller and is closed by ectodermal overgrowth. A neuroenteric canal is formed between the archenteron and developing neural tube.
Mesoderm differentiation:
The transformation of the neural plate to form the neural tube is associated with the formative of a shallow groove on either dorsolateral walls of archenteron. The cells forming these two grooves are smaller than other cells. The grooves become deeper and their edges come together.
Such fusion results in the separation of a solid notochordal rod along the mid-dorsal line. These two lateral grooves become divided by transverse partitions into enterocoelic pouches which grow between the endoderm and ectoderm (Fig. 5.17).
The cavities of these pouches retain their connection with the archenteron at the beginning which become subsequently lost. As a consequence paired hollow blocks of mesodmal cells are formed. Formation of hollow mesodermal blocks is observed only in the first two pairs of somites.
The posterior entcrococlic pouches are pinched off as solid blocks of mesodermal cells within which coelomic cavities are formed anew. This process is observed upto fourteenth pairs of somites. In the rest of the posterior segments, the two halves of the original folds meet to form a solid band of cells extending up to the blastopore. The mesodermal somites differentiate from the lateral bands.
On the basis of origion the mesoderm is divided into:
(a) Gstral mesoderm and
(b) Pristomial mesoderm.
The gastral mesoderm develops from the enterocoelic pouches, while the peristomial mesoderm differentiates from the lateral bands.
The somites or segmental mesoderms gradually grow ventrally on either side until they meet in the midventral line below the alimentary canal. The mesodermal sheet becomes double-walled enclosing coelome within themselves. The lateral plate mesoderm becomes thus splitted into (i) somatic mesoderm in association with ectoderm and (ii) splanchnic mesoderm in association with the endoderm.
ii. Frog:
In late amphibian blastula, the presumptive organ forming areas are oriented around the blastocoelic cavity.
The hypoblast is situated at the vegetal pole, while the epiblast is located at the animal pole. In the epiblast the notochordal cells, neural plate and epidermal areas are situated along the anteroposterior axis of the blastula with the notochordal cells located at the most posterior position.
At the end of the cleavage all the blastomeres remain stationary and none of them have shiftecf from its original position. But at the onset of gastrulation a great mass migration started to occupy their definite position in the developing embryo. Gastrulation begins with the appearance of a small cleft-like invagination at one side and just above the grey crescent (Fig. 5.18).
This cleft-like invagination is crescent- shaped and represents the dorsal lip of the blastopore. As gastrulation progresses the crescent-shaped cleft continues to expand to assume a semicircular appearance, then becomes horse-shoe-shaped and finally forms a ring. This ring represents the blastopore. The blastopore becomes the focal point for gastrulation activities.
Migration of cells inside the gastrula starts along the newly-formed dorsal lip of blastopore and this inward pushing is caused by the endodermal cells which are folded inward (Fig. 5.19) and forward towards the future anterior end of the embryo. The upper margin of the blastopore is called the dorsal lip of the blastopore and the lower edge is designated as the ventral lip of the blastopore.
As invagination expands within the blastocoel, the prechordal plate cells from the upper part of the dorsal side move inward. The new cavity thus produced is called the archenteron which communicates to the exterior by the blastopore. With the further advancement of invagination, the archenteron continues to expand by obliterating the blastocoel.
The inward moving cells form a new border beneath the outer cells. The roof of the archenteron consists of the involuted layer which includes the endoderm and mesoderm. Beyond this layer lies the ectodermal layer. The floor of the archenteron is made up of a layer of endodermal cells, the derivatives of the large yolk cells which were located in vegetal hemisphere of blastula.
When the inward movement of the cells is in progress through the dorsal lip, another type of movement occurs on the outer side. The pigmented cells of the animal hemisphere started to enclose the macromeres of vegetal hemisphere. After completing the enclosure, the outer cells reach up to the ventral lip of it.
A small mass of macromeres remains uncovered for a while and acts as a plug of the blastopore. It is called yolk plug. At this stage, embryo is made up of two distinct strata, each of which is composed of many layers of cells.
Differentiation of three primary germ-layers:
The blastula of frog is mono-layered which in course of gastrulation becomes converted into a triploblastic stage, i.e., three cell-layered. These three layers are designated as the primary germ-layers (embryonic ectoderm, embryonic mesoderm and embryonic endoderm). All the organs of the developing embryo develop from these three primary germ-layers.
(a) Ectoderm:
The pigmented cells of the animal pole, which spread to enclose the macromeres of the vegetal hemisphere become differentiated into ectoderm.
(b) Endoderm:
The dorsal and lateral sheets of cells which form the roof of the archenteron represent the endoderm as well as mesodermal material. Upon completion of gastrulation, the roof and sides of the archenteron become lined by a single layer of endodermal cells which have differentiated from the involuted several celled thick archenteron roof.
(c) Mesoderm:
As soon as the endodermal sheet becomes separated dorsally and laterally from the involuted cells, mesodermal sheet is being formed between the endoderm and ectoderm. The mesodermal sheet starts its differentiation anteriorly and then proceeds gradually backwards.
The mesodermal sheet is divided into two halves by a narrow band of median cells which develop into notochord. Laterally the mesodermal sheets grow downward and finally the right and left mesodermal sheets unite in the mid-ventral line to become a continuous mesodermal sheet.
The three layers thus formed are ectoderm, mesoderm and endoderm. It is the special feature in amphibian development that gastrulation results into the formation of mesoderm first and then the endoderm.
iii. Chick:
The blastoderm has a central area free from yolk which is called the area pellucida, while the germ-wall with the adhering yolk constitutes the area opaca. In course of development, the blastoderm becomes converted into a double-layered structure—the upper one is the epiblast and the lower layer is called the hypoblast.
The space between these two layers is called the blastocoel while the space below the hypoblast is the primordial archenteron.
The epiblast contains presumptive ectodermal and neural areas at the anterior portion while the posterior half comprises of presumptive notochordal and mesodermal cells. The hypoblast transforms into the endoderm and the epiblast is converted into ectoderm and mesoderm (Fig. 5.20).
At the initiation of gastrulation, the hypoblast cells from the posterior end start migrating towards the anterior end of the embryo along the median line. Immediately after the inauguration of the movement in the hypoblast, the cells of the epiblast overlying the migrating hypoblast move downward towards the hypoblast.
These involuted cells occupy a position between the epiblast and hypoblast and migrate to the lateral and anterior ends between the epiblast and hypoblast. Movement of cells in the blastoderm of chick during gastrulation has been studied by Spratt (1946) by carbon particle technique.
With the activities of the epiblast and hypoblast, the presumptive mesoderm cells from the posterior half of the epiblast move posteriorly and converge from the lateral sides towards the median line. These converging cells begin to accumulate at the posteromedian border of the area pellucida as a raphe-like thickened structure. This marks the appearance of the primitive streak (Fig. 5.21).
The migratory cells after coming to this region move inwards and migrate anteriorly and laterally. The migratory cells of the epiblast move downward and occupy the position between the epiblast and hypoblast. These cells then diverge anteriorly and laterally as a broad middle layer of mesodermal cells along the primitive streak.
Daring its forward movement, it approaches the presumptive notochordal areas. With the anterior movement the streak also starts to move backward. The primitive streak is fully formed at about 18-19 hours after incubation. Gradually the area pellucida changes from a round to pear-shaped appearance.
The primitive streak represents the posterior region of the developing embryo and the embryo proper develops anterior to it. It is also an area of cell proliferation arid rapid growth. The primitive streak becomes very conspicuous in early embryonic life. It consists of a groove (Primitive groove) which is flanked on both the sides by two ridges (Primitive ridges).
It terminates anteriorly in a primitive pit, and posteriorly in a primitive plate. Immediately anterior to the primitive pit (which represents the defunct neuroenteric canal) lies an elevation, Hensen’s node or Head process.
In this area the mesoderm becomes thickened and projects from the primitive streak. With the formation of the Hensen’s node, the primitive streak regresses posteriorly arid the major organ forming areas become well established! A groove appears on the outer surface of the head process and the two folds unite to form a tube.
The formation and closure of the groove continue posteriorly. The entire process may be compared with the action of a zipper. As die closure of the groove comes to the posterior end, the backward movement ceases leaving an opening at the posterior end.
In the gastrulation of chick, the mesoderm differentiates lastly from the epiblast by the process of involution, elongation, expansion and extension. The hypoblast gives rise to endoderm and the epiblast differentiates into ectoderm and mesoderm.
iv. Mammal:
In the mammalian blastula (Blastocyst) the formative area (germ disc) is restricted at one end. The germ disc is composed of epiblast and hypoblast. In the embryo of pig, the gastrulation activities are observed at two centres—the posterior end forms the primitive streak while the anterior end forms the Hensen’s node.
The behaviour of these portions is almost similar to that observed in the gastrulation of chick. The mesodermal cells from the primitive streak move between the epiblast and hypoblast and form two wing-like areas.
The mesodermal cells are divided into:
(a) Embryonic mesoderm confined to the germ disc and
(b) Extra embryonic mesoderm.
6. Analysis of Gastrulation Mechanism:
Lillie (1913) established that the surface layer of the egg at first remains plastic but in course of development it loses its plasticity and becomes rigid. Spemann (1918) found that up to gastrulation, when the eggs are cut into two halves, each will form a complete embryo.
But after gastrulation, each half gives rise to half embryo. He also noticed that the half containing blastopole forms a complete embryo. He came to the conclusion that blastopore plays an important role in gastrulation.
Later he and Mangold (1924) grafted blastopore of one to the gastrula of another and demonstrated that the grafted blastopore influences the host tissue to form embryonic axis. They termed the blastopore as “organiser” and the influence of organiser as “induction”. Spemann’s lead was soon followed by different workers and considerable information became available regarding the nature of organiser.
It may be summarised that induction involves three distinct events:
(i) Evocation,
(ii) Individuation and
(iii) Competence.
The first two, evocation and individuation are the properties of organiser and the competence is the feature of the tissues on which the organiser acts.
In 1943, Holtfreter, working on the gastrulation mechanism of amphibian eggs, demonstrated that superficial cells are united by an extracellular surface coat and the beginning of invagination is due to the expansion of certain cells. This expansion according to him is caused by the change of surface tension due to the high pH of blastocoelic fluid.
Though many workers have questioned the findings of Holtfreter, it remained true that initiation of invagination is the property of localized cells. It may be due to local difference of pH or due to differential adhesiveness of the cells.
The work done to explore the nature of involution and epiboly also explained that the entire process is due to the nature of participating cells. It was demonstrated that the cells which are more adhesive are less mobile and on the contrary more mobile cells are less adhesive. Once this was understood attempts were made to explain the mechanism of gastrulation in terms of cellular adhesibility and cellular mobility.
In 1955, Townes and Holtfreter examined the interaction of different cell layers in amphibian gastrulae and demonstrated:
(a) That endoderm cells are less adhesive than mesoderm,
(b) That outer ectoderm is less adhesive than inner ectoderm and
(c) Mesoderm is less adhesive than inner ectoderm but more adhesive than endoderm.
Basing on this contention Stainberg (1964) proposed that disposition of different layers in a gastrula depends upon the adhesive nature of the cells. Outer ectoderm being less adhesive stays outermost. Mesoderm being less adhesive than inner ectoderm but more adhesive than endoderm remains in between the two.