Gastrulation is the biological process that rearranges a simple ball of cells into a complex, multi-layered body structure. During this morphogenetic phase, cells are sorted into the three primary germ layers that will give rise to all the tissues and organs of the adult organism. The zebrafish, Danio rerio, is an exceptional model for observing this dramatic reorganization because its embryos develop externally and remain optically transparent throughout these early stages. This transparency allows biologists to watch the coordinated movements of thousands of cells in real-time, providing insight into the fundamental mechanisms of vertebrate body formation. The entire process of gastrulation defines the blueprint for the entire body plan.
Preparing for Shape Change: The Blastula Stage
Before the large-scale movements of gastrulation can begin, the embryo must first complete the cleavage stage, resulting in a structure called the blastula. This early embryo is organized into three distinct cell populations perched atop a massive, uncleaved yolk cell. The innermost population is the Deep Cells (DCs), a dome of motile, loosely organized cells. These cells form the bulk of the tissue that will eventually differentiate into the three germ layers.
The Enveloping Layer (EVL) is a single-cell thick epithelial sheet surrounding the deep cells. The EVL acts as a protective outer covering, forming a barrier that helps maintain the internal environment of the developing embryo. The cells of the EVL are tightly connected to one another, which is necessary for the coordinated spreading movement that follows.
The third population is the Yolk Syncytial Layer (YSL), an extra-embryonic structure unique to teleost fish. The YSL forms when marginal blastomeres fuse with the underlying yolk cell, creating a multinucleated layer (syncytium). Lying directly at the interface between the embryonic cells and the yolk, the YSL is positioned to influence the subsequent morphogenetic movements. It is subdivided into an internal part (I-YSL), which underlies the deep cells, and an external part (E-YSL), which extends past the blastoderm margin.
Enveloping the Yolk: The Role of Epiboly
The first, most conspicuous movement of zebrafish gastrulation is epiboly. This process involves the entire blastoderm, including the deep cells and the EVL, spreading downward to completely cover the large yolk cell. Epiboly begins when the yolk cell itself undergoes a slight upward doming, which helps to push the overlying deep cells radially outward.
The mechanical force driving this extensive spreading is primarily generated by the extra-embryonic tissues, specifically the YSL and the EVL. The YSL, a syncytium rich in cytoskeletal elements like microtubules and actin, actively contracts and pulls the overlying blastoderm margin toward the vegetal pole. This syncytial layer acts as an internal purse-string, generating the tension required for the massive sheet of cells to advance.
The EVL, as the outermost epithelial layer, must expand its surface area to keep pace with the spreading front. This expansion is achieved by cell shape changes and the rearrangement of cells within the layer. Simultaneously, the deep cells undergo radial intercalation, where cells from the inner layers move outward to join the outer layers. This cellular rearrangement effectively thins the entire blastoderm, which is a necessary prerequisite for the tissue to expand and cover the yolk.
The entire process requires tight coordination, as the EVL, the deep cells, and the YSL must advance together, maintaining their physical connections as they move past the equator of the yolk cell. Epiboly continues until the blastoderm margin closes entirely at the vegetal pole, a process that fully internalizes the yolk mass and establishes the primary outer boundary of the embryo.
Internalizing Cells: Involution and Germ Layer Positioning
As epiboly reaches approximately the halfway point over the yolk cell, a second, important set of movements begins, resulting in the internalization and rearrangement of the future body tissues. The first step is involution, where cells at the blastoderm margin turn inward and migrate beneath the superficial layer. This process separates the deep cell mass into two distinct layers: the inner layer, called the hypoblast, and the outer layer that remains on the surface, called the epiblast.
The hypoblast is the precursor to the mesoderm and endoderm germ layers, while the epiblast will form the ectoderm. Following internalization, the hypoblast cells begin a directed, collective migration toward the animal pole, moving over the I-YSL. This movement is coupled with two simultaneous and intricately linked processes: convergence and extension.
Convergence is the movement of cells toward the dorsal midline of the embryo, narrowing the tissue along the mediolateral axis (side-to-side). This narrowing is coupled with extension, a lengthening of the tissue along the anterior-posterior axis (head-to-tail). This coordinated narrowing and lengthening is driven by cellular behaviors such as mediolateral intercalation, where cells interdigitate between their neighbors to form a longer, narrower file.
The Wnt/Planar Cell Polarity (PCP) signaling pathway is a primary regulator of this convergence and extension, orienting the cellular movements and helping to coordinate the collective migration. This three-dimensional dance positions the newly formed germ layers relative to one another and physically elongates the developing embryonic axis.
Establishing the Body Plan: From Layers to Axis
The culmination of gastrulation movements results in the establishment of the three primary germ layers and the rough sketch of the vertebrate body plan. The epiblast, having remained on the embryo’s exterior, forms the ectoderm. The ectoderm gives rise to the outermost coverings, like the epidermis of the skin, and the entire nervous system, including the brain and spinal cord.
The hypoblast, which involuted inward, differentiates into the mesoderm and the endoderm, positioning them beneath the ectoderm. The mesoderm, the middle layer, forms the structural and circulatory components of the body:
- Muscle
- Bone
- Cartilage
- Blood
- Kidneys
The endoderm, the most internal layer, is the precursor to the lining of the digestive tract and associated organs like the liver, pancreas, and respiratory system.
Gastrulation movements are also directly responsible for establishing the major axes of the body. Convergence and extension physically define the elongated anterior-posterior axis, dictating where the head and tail structures will form. Signaling gradients, such as those involving Nodal and Bone Morphogenetic Protein (BMP) signals, establish the dorsal-ventral axis. The completion of these movements transforms a spherical mass into a three-layered, elongated structure with clearly demarcated body axes.