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Carnegie Stage 1 Introduction

Stage 1 is the unicellular embryo that contains unique genetic material and is an individually specific cell that has the potential to develop into all of the subsequent stages of a human being. It is the beginning of embryonic life and ontogenetic development that starts when an oocyte, arrested in metaphase of meiosis II, is penetrated by a sperm. This is the first event of fertilization. The embryo has a postfertilization age of approximately one day, is between 0.1 to 0.15 mm in diameter and weighs approximately 0.004 mg.

Fertilization is a series of events that begins when a sperm makes contact with an oocyte and ends with the intermingling of paternal (male) and maternal (female) chromosomes on the spindle at metaphase of the first mitotic division of the single cell. The events of fertilization require just over 24 hrs. to complete and normally take place in the ampulla of the uterine tube.

Stage 1 is divided into three substages; a, b and c. Stage 1a is referred to as the primordial embryo since all the genetic material necessary for the new individual, plus some redundant chromosomes, is now within a single plasmalemma (cell membrane). From the perspective of the female gamete it has also been named the penetrated oocyte. The fertilizing sperm has passed through the zona (capsula) pellucida and its plasmalemma has fused with that of the oocyte. Penetration activates the embryo into resuming its arrested meiosis II and after anaphase it enters telophase with the expulsion of the redundant chromosomes as a second polar body. This marks the beginning of Stage 1b in which the single-cell is referred to as the pronuclear embryo. From the perspective of the female gamete it has also been named the ootid because its female component is haploid like a spermatid. However, in the pronuclear embryo there are two separate haploid components: one maternal, or female, pronucleus and one paternal, or male, pronucleus. The pronuclei move toward each other and eventually compress their envelops where they lie adjacent near the center of the cell. Stage 1c is the last phase of fertilization and exists for a relatively short period. The pronuclear envelopes disappear and the parental chromosomes that were contained in separate pronuclei come together in a process called syngamy thereby establishing the genome of the embryo. The one-cell Stage 1c embryo is named the syngamic embryo or zygote. The chromosomes assume positions on the rapidly formed first mitotic spindle in preparation for cleavage.

The Carnegie collection contains no stage 1 specimens. The database for this stage is therefore limited to previously published images of specimens that best show the progression of development. Only images of those specimens that the authors considered healthy were selected for inclusion in the database. Grateful appreciation is expressed to all of the authors and their publishers for their generosity in permitting the use of their images in the database.

A measuring bar is present in most of the light micrographs (LM). The length of the measuring bar is based on a diameter of the one cell (including the zona pellucida) of 175 µm before fixation. The diameter of the cell, without the zona pellucida, is approximately 100 µm before fixation. The embryo undergoes shrinkage with fixation and embedding. Since the cytoplasm is more affected than the zona pellucida the subzonal (perivitelline) space becomes accentuated in fixed specimens. The original magnification is given in the legend of most scanning (SEM) and transmission (TEM) electron micrographs.

IN VITRO STAGE 1a SPECIMENS (click here to view figures)

In the primordial embryo the plasma membranes of the fertilizing gametes become confluent (Fig. 9). Non-fertilizing sperm may be present in the zona pellucida or in the subzonal space. The cytoplasm of the embryo protrudes where the fertilizing sperm enters. The protrusion is referred to as the fertilization cone (Fig. 3). It is devoid of microvilli and has numerous microfilaments forming a conspicuous band beneath the cell membrane. It later retracts supposedly drawing the sperm deep into the cytoplasm.

Once inside the cytoplasm the original envelope of the sperm vesiculates and is gradually dismantled. The head of the fertilizing sperm becomes decondensed and swells. Pronuclei are not yet present but a wave of granularity moves throughout the cytoplasm in 2 to 10 circular rotations. Some of the rotations are clockwise and some are counter clockwise. The movement lasts for twenty or more minutes and is referred to as the cortical reaction. The reaction involves the exocytosis of the contents of cortical granules. The membrane of the granules fuses with the overlying cell membrane then burst open releasing their contents into the subzonal space by a process similar to cell secretion (Figs. 18-21). Cortical granule release occurs three hours after sperm / oocyte fusion. Once the cortical granule contents are released into the subzonal space they interact with the inner part of the zona pellucida thus preventing further entrance of sperm into the oocyte and inhibiting polyspermia. The interaction is called the zonal reaction. (Fig. 22)

The second miotic division is completed at this time with the release of the second polar body into the subzonal space. Since the cytoplasm contains the genetic material from both the mother and father it is now technically an embryo (Figs. 1, 2, 8, 12-17).

IN VITRO STAGE 1b SPECMENS (click here to view figures)

The unique feature of pronuclear embryos is the presence of pronuclei which appear within 11 hours of in vitro insemination. The pronuclei each measure approximately 30 µm in diameter before fixation. The paternal (male) and maternal (female) pronuclei usually form simultaneously (Figs. 16, 17). The paternal pronucleus is usually larger and forms near the site where the sperm entered (fertilization cone); the maternal pronucleus forms near the site where the second polar body is extruded.

After their formation the pronuclei are located a distance from each other but subsequently each moves toward the center of the cell (Figs. 23, 24). By 15 hours after in vitro insemination they lie close to one another (Figs. 27-29). They remain closely associated until the onset of syngamy (See below). As they approach one another, adjacent areas of each pronuclear envelope appear to flatten out. At the same time nucleoli move from random locations within each pronucleus to line up in the flattened pronuclear envelope regions (Figs. 27-30). Nucleoli vary in size and number from as few as one to as many as nine. The envelope of the maternal pronucleus often begins to break down before the paternal one. The envelopes of the pronuclei do not fuse or interlock.

A straight line can be drawn from the polar bodies through the cytoplasm that divides the cell into equal parts. This line is called the polar axis (or anterior-posterior polar axis) and roughly indicates the location of the first cleavage plane. The pronuclei rotate into the polar axis and align along it (Figs. 25, 26). This is considered the correct position for syngamy and the first cleavage.

IN VITRO STAGE 1c SPECIMENS (click here to view figures)

The stage 1c embryo is called the syngamic embryo or zygote and is the last phase of fertilization. It is sometimes called a founder cell because it gives rise to embryonic stem cells. Stage 1c occurs about 20 hours after in vitro insemination but is elusive since it exists for only a brief period. The pronuclei quickly disappear when their envelopes break down (Figs. 46, 47, 50). Groups of chromosomes that were in the pronuclei assume positions in pairs on the rapidly formed first cleavage spindle thereby bringing about syngamy and the establishment of the integrated genome of the embryo. The chromosomes lie in an agranular zone in the central cytoplasm. Since a nuclear envelope does not form, a nucleus is not present at this substage.


Source: The Virtual Human Embryo.