Fertilization and the first cleavage mitosis in insects

Fertilization in animals is now considered to be of the "sea urchin type"; that is, haploid male and female pronuclei completely fuse shortly after sperm entry into the egg, followed by the formation of a mitotic spindle to allow cleavage mitoses to proceed. However, two other patterns of fertilization and early embryonic mitosis in some animal species are known: an Ascaris type and a gonomeric type. The gonomeric type of fertilization in insects and other arthropods is not well known and is quite different from the sea urchin and Ascaris types. In the present article, the author examines the peculiar gonomeric fertilization, using mainly the silkworm as an example.     

Fertilization potential and electrical properties of the Xenopus laevis egg

The membrane potential of Xenopus eggs was monitored continuously from prior to fertilization until early cleavage. A rapid decay of the initial potential of -33.1 +/- 8.1 (SD) mV (N = 14) upon impalement to a value of -19.3 +/- 4.2 (SD) mV (N = 68) suggested that insertion of the first electrode caused depolarization. Outward and inward rectification were observed when the resting potential was made more positive than about 5 mV or more negative than about -30 mV. Eggs were not activated by this level of current injection. Fertilization and activation evoked a membrane depolarization which was influenced by the external Cl- concentration, the nature of the halide species, and 4,4-diisothiocyanostilbene-2,2-disulfonic acid. Smaller transient depolarizations were associated with the initial stages of the fertilization potential but not with activation. Only when the fertilization potential was significantly diminished, as in high external Cl- or in the presence of Br- or I- solutions did polyspermy ensue. The input resistance of the unfertilized egg was 13.2 +/- 9.8 M omega (N = 26) and decreased about 200-fold at the peak of the fertilization potential to 0.077 +/- 0.020 M omega (N = 9). Ninety minutes after the onset of the fertilization potential and about 6 min after the start of furrow formation the membrane began a series of cleavage cycle-associated hyperpolarizations. These were unaffected by either the external Cl- concentration or other halide species. Reduction in amplitude of the fertilization potential had no apparent effect upon the normal elevation of the fertilization envelope or upon cleavage and later development. The fast electrical block to polyspermy appears to have a lower threshold in Xenopus compared with other species and is also effective at negative membrane potentials.     

Early egg contractions and patterned parasynchronous cleavage in a living insect egg

Summary Fifteen movements of egg ooplasm occur prior to the formation of the embryo of the beetle,Callosobruchus maculatus. These movements are associated with nuclear division taking place before formation of the embryo. Yolk globules exhibit a coordinated pattern of saltation and quiescence with these movements. The significance of this yolk activity for the movement of nuclei within the yolk mass is discussed. During the syncytial blastoderm stage the surface of the egg is infolded around the surface nuclei. Nuclear division during this period appears to produce localized stretching of the surface which in turn results in a yolk contraction. Division is parasynchronous and patterned with a wave of anaphase configurations spreading across the egg. The significance of such mitotic gradients to patterning of insect segments is discussed.     

Fertilization stimulates lipid peroxidation in the sea urchin egg

Arachidonic acid is rapidly taken-up by Strongylocentrotus purpuratus eggs and eventually incorporated into cellular lipids. During the first few minutes following fertilization the arachidonic acid that has not been incorporated into other lipid forms is oxidized to a hydroxy-fatty acid. In vivo, the time of arachidonic acid conversion coincides with the transient period of increased intracellular free calcium after fertilization. In vitro, this lipid peroxidizing activity has been shown to be initiated by micromolar calcium. Taken together with the presence of Ca2+-stimulated lipase, these results suggest that calcium regulates both the release of polyunsaturated fatty acids from cellular lipids and their subsequent oxidation. The physiological function of lipid hydroxides or hydroperoxides in sea urchin fertilization is unknown. A possibility is that they may be important in regulating the many membrane permeability changes occurring within minutes after fertilization.     

Delay of cleavage of the Arbacia egg by ultraviolet radiation

While our data do not permit us to state the exact locus or mode of action of ultraviolet radiation in the Arbacia egg, certain general conclusions may be reached. The amount of delay of cleavage of these eggs is determined by two principal factors: (1) The extent of an effect, resulting from photochemical action induced by ultraviolet radiation, which is reversible in a biological sense, the reversibility not being directly dependent upon the process of cell division. (2) The sensitivity of the cell division process to the effects of the ultraviolet-induced photochemical reaction. This factor varies with the stage of cell division, the cell being insensitive during a period corresponding to most of mitosis. It seems likely that these findings may apply to cell division in general, but, since the quantitative relationships observed must, in this case, reflect the integration of two semi-independent factors, the over-all picture may appear quite different for different kinds of cells.     

Fertilization triggers activation of Fyn kinase in the zebrafish egg

Fertilization results in the tyrosine phosphorylation of several egg proteins and studies have shown that tyrosine protein kinase activity is required for successful fertilization. The Fyn protein kinase has been detected in eggs of the sea urchin, frog and rat, although measurement of fertilization-induced changes in Fyn kinase activity have only been successful in the sea urchin system. The present study demonstrates the presence of Fyn kinase in the zebrafish egg and the stimulation of this enzyme at fertilization. Activation of Fyn was detected as early as 30 seconds post-fertilization and increased approximately six-fold by 2 minutes post-insemination. The activation of Fyn in the zebrafish egg required sperm and was not observed in spontaneously activated eggs.     

Free amino acids in the early cleavage stages of the rabbit egg

In the first half of gestation, mouse yolk sac contains levels of N-acetyl-β-hexosaminidase activity and a ratio of N-acetyl-β-hexosaminidase: β-glucuronidase much higher than those of other embryonic tissues or the decidua. The properties of the yolk sac enzyme are very similar to those in the other tissues, suggesting that the observed difference in activity is quantitative rather than qualitative. In blastocyst cultures, the vesicular growths derived from the inner cell mass possess patterns of N-acetyl-β-hexosaminidase activity similar to yolk sac in vivo, but different from trophoblast cells developing in the same cultures. It is therefore proposed that the vesicular growths contain at least some differentiated yolk sac cells.     

Fertilization potential and polyspermy prevention in the egg of the nemertean, Cerebratulus lacteus

We investigated the electrical properties of the egg of the nemertean worm Cerebratulus, and found evidence that an electrically-mediated polyspermy block operates for a period of about 1 hr after fertilization. At fertilization, in natural or artificial sea water, the membrane potential shifts from its resting level of about -66 mV to a peak of about +43 mV, and in most cases remains greater than 0 mV for more than 1 hr. The average potential during the first 30 min is +22 +/- 8 mV (SD, n = 12). When the external Na+ concentration is reduced from 486 to 51 mM (choline substituted) the fertilization potential amplitude is reduced; the average potential during the first 30 min is -27 +/- 21 mV (SD, n = 5). Eggs inseminated in 51 mM Na+ sea water become polyspermic, indicating that polyspermy prevention depends on an electrically-mediated mechanism. The electrical block is required for about 60 min, since transfer to 51 mM Na+ sea water during this period results in polyspermy. During the first hour following fertilization, the egg is also developing a permanent, nonelectrical block; the degree of polyspermy which results upon transfer to low Na+ sea water decreases progressively with time. The permanent block appears to be at the level of the egg plasma membrane or glycocalyx, since the egg envelope is not a barrier to sperm penetration, nor does its removal induce polyspermy. Electron micrographs show no obvious changes in the morphology of the extracellular layers, plasma membrane or cortex of the egg after fertilization.     

The cleavage pattern of the axolotl egg studied by cinematography and cell counting

Summary The temporal pattern of cleavage in the egg of the axolotl,Ambystoma mexicanum, was studied 1. by time-lapse microcinematography, and 2. by counting the total number of blastomeres dissociated at successive stages.Eggs were filmed from the one-cell stage till the early gastrula either (A) simultaneously from above and below with a double-camera assembly, or (B) from the side with a single camera.The animal blastomeres divide synchronously from the 2nd up to and including the 10th cleavage. The cycle length is roughly constant from the 3rd till the 10th cleavage. The cycle from the 2nd to the 3rd cleavage is slightly longer, while that from the 1st to the 2nd cleavage is about 20% longer. After the 10th cleavage the synchrony of divisions is lost owing to variable lengthening of cell cycles in individual blastomeres. Gastrulation starts around the onset of the 15th cleavage in the animal blastomeres.The analysis of films taken in side view reveals seven recurring cleavage waves, from the 5th till the 11th cleavage. Cells in the animal, equatorial and vegetative regions in sequence repeatedly pass through the three successive phases of the cleavage cycle—rounding-up, division, and relaxation—but with a shift in phase. The start of the 10th cleavage division of the slowest vegetative cells more or less coincides with that of the 11th division of the animal cells; from then on the cleavage waves become increasingly obscured.Morulae and blastulae were dissociated by placing them in 1/15 M phosphate buffer (pH 7.8) for the duration of 2–3 cleavage cycles and then removing the vitelline membrane. In this solution cell divisions continued without disturbance of the temporal cleavage pattern. The dissociated cells were fixed either just prior to the onset of the next cleavage (up to the 10th cleavage) or at those times when cleavageswould have been expected, had there been no lenthening of cleavage cycles (beyond the 10th cleavage). The total cell number was counted, dividing cells being scored as two.Prior to the 11th cleavage the total cell number increased exponentially. Beyond the 10th cleavage the rate of increase was considerably lower. At the time when gastrulation would have started if the egg had not been dissociated, the total cell counts were 13,000–15,000, whereas the number anticipated without lengthening of cleavage cycles would be of the order of 130,000 (2¹⁷).The application of Balfour's rule to amphibian eggs is criticized.     

Fertilization selection on egg and jelly-coat size in the sand dollar Dendraster excentricus

Organisms with external fertilization are often sperm limited, and in echinoids, larger eggs have a higher probability of fertilization than smaller eggs. This difference is thought to be a result of the more frequent sperm-egg collisions experienced by larger targets. Here we report how two components of egg target size, the egg cell and jelly coat, contributed to fertilization success in a selection experiment. We used a cross-sectional analysis of correlated characters to estimate the selection gradients on egg and jelly-coat size in five replicate male pairs of the sand dollar Dendraster excentricus. Results indicated that eggs with larger cells and jelly coats were preferentially fertilized under sperm limitation in the laboratory. The selection gradients were an average of 922% steeper for egg than for jelly-coat size. The standardized selection gradients for egg and jelly-coat size were similar. Our results suggest that fertilization selection can act on both egg-cell and jelly-coat size but that an increase in egg-cell volume is much more likely to increase fertilization success than an equal change in jelly-coat volume. The strengths of the selection gradients were inversely related to the correlation of egg traits across replicate egg clutches. This result suggests the importance of replication in studies of selection of correlated characters.