Early embryo orientation by direction of the cardinal point in natural clutches of two ranidae species inhabiting geographically distant regions

The azimuth (the least angle with the north-south direction) of the first cleavage furrow and anteroposterior axes of neurula was measured on projections of photographs of natural clutches. The azimuth distribution of the craniocaudal axis of Rana dalmatina neurulae in clutches from southern Italy and the first cleavage furrows of R. arvalis embryos in clutches from central Russia proved identical. Both craniocaudal axes and first cleavage furrows were mostly oriented from west to east. The azimuth distribution of the craniocaudal axis of R. arvalis neurulae in clutches subjected to repeated cold shock proved to be random. The causes and mechanisms of predominant orientation of the embryos in natural clutches of frogs are discussed. We propose that magnetic sensitivity is acquired by cytoskeleton elements, most likely microtubules, during reorganization in the course of normal development or due to experimental influences.     

The Establishment of Embryonic Axial Properties in the Nemertean,Cerebratulus lacteus

Experiments were performed to determine whether the first cleavage division plays a role in setting up the dorsoventral axis in embryos of the equal-cleaving nemerteanCerebratulus lacteus.Fertilized eggs were compressed to change the orientation of the first cleavage spindle, and thus the plane of the first cleavage division. One cell of the resulting two-celled embryos was then injected with lineage tracer to determine whether the first cleavage plane always maintains its normal relationships to the median and frontal planes or whether new relationships (and thus, novel cell lineages) could be created. Many of these compressed embryos gave rise to normal-appearing pilidium larvae in which the first cleavage plane had taken on various oblique angular relationships relative to the plane of bilateral symmetry and the dorsoventral axis of the larva. These findings indicate that the first cleavage plane can be dissociated from its normal relationships to these axial properties. Thus, the first cleavage division is not causally involved in the establishment of the dorsoventral and bilateral axes. We argue that the dorsoventral axis is specified prior to the first cleavage division.     

Quantitative aspects of RNA synthesis in normal and lithium-treated embryos ofXenopus laevis

Summary The treatment ofXenopus early embryos with lithium chloride produces exogastruale — embryos which fail to gastrulate normally and in which the rates of cell division are reduced. In the present study estimations of incorporations of (5-³H) uridine and the specific activities of the 5-ribonucleotide precursor pools showed that exogastrulae have higher rates of RNA synthesis per cell than control neurulae. Sub-cellular fractionations showed that a greater proportion of labelled RNA was retained in the nuclei of exogastrulae than of neurulae, while neurulae showed a greater incorporation into polysomes.     

Regulation and the modification of axial properties in partial embryos of the nemertean, Cerebratulus lacteus

 Embryos acquire axial properties (e.g., the animal-vegetal, dorsoventral and bilateral axes) at various times over the course of their normal developmental programs. In the spiral-cleaving nemertean, Cerebratulus lacteus, lineage tracing studies have shown that the dorsoventral axis is set up prior to the first cleavage division; however, blastomeres isolated at the two-cell stage will regulate to form apparently perfect, miniature pilidium larvae. We have examined the nature of axial specification in this organism by determining whether partial embryos retain the original embryonic/larval axial properties of the intact embryo, or whether new axial relationships are generated as a consequence of the regulatory process. Single blastomeres in two-cell stage embryos were injected with lineage tracer, and were then bisected along the second cleavage plane at the four-cell stage. Thus, the relationship between the plane of the first cleavage division and various developmental axes could be followed throughout development in the ”half-embryos”. While some embryo fragments appear to retain their original animal-vegetal and dorsoventral axes, many fragments generate novel axial properties. These results indicate that axial properties set up and used during normal development in C. lacteus can be completely reorganized during the course of regulation. While certain embryonic axes, such as the animal-vegetal and dorsoventral axes, appear to be set up prior to first cleavage, these axes and associated cell fates are not irreversibly fixed until later stages of development in normal intact embryos. In C. lacteus, the process whereby these properties are ultimately determined is apparently controlled by complex sets of cell-cell interactions. Received: 11 October 1996 / Accepted: 21 February 1997     

Experimental studies on a mutant gene (cl) in the Mexican axoloti which affects cell membrane formation in embryos from cl-cl females

The gene cl exerts a maternal effect in the Mexican axolotl resulting in an abnormal cleavage pattern. The early cleavage furrows appear partially depigmented and never continue completely around the egg. Subsequent divisions display a similar pattern which results in the vegetal hemisphere remaining uncleaved; but some portions of the animal hemisphere continue to cleave normally. Gastrulation is very rarely initiated.Several cytological abnormalities including polyploidy, broken chromosomes and fusion of nuclei are observed in mutant embryos. These abnormalities are likely secondary effects resulting from the absence of cell boundaries in the uncleaved portions of the embryo and account for its limited development. Cytochalasin B treatment of normal fertilized eggs produces phenocopies of the most severely affected mutant embryos. This suggests that the cl gene may directly affect the synthesis and/or distribution of a cell surface component which enables daughter cell membranes to be assembled and to adhere to one another.Cells from mutant blastulae were able to differentiate pigmented epidermis and neural tube when grafted to normal recipient blastulae or neurulae. This suggests that the gene is lethal only to cells derived from the vegetal cytoplasm or cortex, but not lethal to cells inheriting animal cytoplasm from $\text{cl}\text{cl}$ females.     

More than 95% reversal of left-right axis induced by right-sided hypodermic microinjection of activin into Xenopus neurula embryos

In recent years, genes that show left-right (L-R) asymmetric expression patterns have been identified one after another in vertebrate gastrula-neurula embryos. However, we still have little information about when the irreversible L-R specification is established in vertebrate embryos. In this report, we show that almost 100% of the embryos develop to be L-R-inverted larvae after microinjection of activin molecules into the right lateral hypodermic space of Xenopus neurula embryos. After right-side injection of 10-250 pg activin protein, both early neurulae just after gastrulation movement (stage 13-14) and late neurulae just before neural tube closure (stage 17-18) showed almost 100% reversal of the heart and gut L-R axes. At higher doses of activin, more than 90% of the L-R-inverted embryos showed L-R reversal of both heart and gut. The survival ratio of the right-injected 4-day embryos was 90% on average. In the left-injected embryos, the occurrence of L-R inversion was less than 2% as observed in normal untreated siblings (1.7%). When the same amount of activin (1-50 pg) was microinjected into both sides of neurula embryos, the incidence of L-R inversion was reduced to 58%. The injection of activin along the dorsal midline in the trunk region also randomized the visceral L-R axis. Injection of activin into the right side changed normal left-handed expression of Xnr-1 to right-handed or bilateral expression. In contrast, left-handed expression of Pitx2 was switched to the right side by right activin injection. This is the first report of a method that achieves complete inversion of the visceral L-R axis by treatment of embryos at the neurula stage. Activin not only acts on the neurulae to cancel the original L-R specification up to the late neurula stage, but also rebuilds a new L-R axis whose left side coincides with the injection side. It is suggested that the left and right halves of neurulae have equal potential for L-R differentiation.     

Early cleavage in Phoronis muelleri (Phoronida) displays spiral features

The view that early cleavage in Phoronida follows a radial pattern is widely accepted. However, data supporting this characterization are ambiguous. Studies have been repeatedly reporting variation between individual embryos, and the occurrence of embryos exhibiting oblique divisions or nonradial cell arrangements. Such embryos were often considered to represent variation within radial cleavage, or artificial appearances. Cleavage in Phoronis muelleri was previously characterized as “derived radial,” but also oblique spindles and cell elongations, and shifted cell arrangements were observed. We studied the early cleavage in P. muelleri applying 4D microscopy, fluorescent staining, and confocal laser scanning microscopy. To deal with the problem of variation we provide statistical evaluations of our data. These show that oblique divisions do not represent variational abnormalities. In fact, they reveal that most cells divide obliquely from the third cleavage onwards. What is more, in almost all cells the axis of the third cleavage is inclined dextrally. The fourth cleavage is even stronger sinistrally pronounced. Subsequently, the pattern of alternating cleavage orientation is largely restricted to animal and vegetal blastomeres. As a result of the obliqueness of divisions, four cells encircle the poles in most embryos. Cross furrows are occasionally present. We found no indications for radial cleavage in P. muelleri. In contrast, the observed cleavage displays several characters consistent with the pattern of spiral cleavage. A close relation of phoronid and spiralian cleavage is also suggested by molecular phylogenies, allying both groups in the Lophotrochozoa. We suggest our findings to represent morphological support for this lophotrochozoan/spiralian affinity of Phoronida.     

The cytoskeletal involvement in cellularization of theDrosophila melanogaster embryo

Summary The distribution and arrangement of cytoskeletal components in the early embryo ofDrosophila melanogaster were examined by thin-section electron microscopy to elucidate their involvement in the formation of the cellular blastoderm, a process called cellularization. During the final nuclear division in the cortex of the syncytial blastoderm bundles of astral microtubules were closely associated with the surface plasma membrane along the midline where a new gutter was initiated. Thus the new gutter together with the pre-formed ones compartmentalized the embryo surface to reflect underlying individual daughter nuclei. Subsequently such gutters became deeper by further invagination of the plasma membrane between adjacent nuclei to form so-called cleavage furrows. Nuclei simultaneously elongated in the direction perpendicular to the embryo surface and numerous microtubules from the centrosomes ran longitudinally between the nucleus and the cleavage furrow. Microtubules often appeared to be in close association with the nuclear envelope and the cleavage furrow membrane. The plasma membrane at the advancing tip of the furrow was always undercoated with an electron-dense layer, which could be shown to be mainly composed of 5–6 nm microfilaments. These microfilaments were decorated with H-meromyosin to be identified as actin filaments. As cleavage proceeded, each nucleus with its perikaryon became demarcated by the furrow membrane, which then extended laterally to constrict the cytoplasmic connection between each newly forming cell and the central yolk region. The cytoplasmic strand thus formed possessed a prominent circular bundle of microfilaments which were also decorated with H-meromyosin and bidirectionally arranged, similar in structure to the contractile ring in cytokinesis. These observations strongly suggest that both microtubules and actin filaments play a crucial role in cellularization ofDrosophila embryos.     

γ-Tubulin,microtubule arrays,and quadripolarity during sporogenesis in the hepatic<Emphasis Type="Italic"> Aneura pinguis</Emphasis> (Metzgeriales)

This is the first report on the organization of a quadripolar microtubule system (QMS) in polyplastidic meiosis of a hepatic with polar organizers (POs). Unlike the monoplastidic sporocytes of mosses and hornworts, in which meiotic quadripolarity can be traced to plastid division and migration, sporocytes of Aneura pinguis are polyplastidic and tetrahedrally lobed before the QMS is organized. Whereas the QMS in mosses and hornworts is plastid-based, the QMS of A. pinguis is focused at four POs where gamma tubulin (-tubulin) is concentrated. An aster of microtubules emanates from each PO centered in the four cytoplasmic lobes and the opposing radial microtubules interact to form the QMS that envelops the nucleus. A functionally bipolar spindle is gradually formed as the four poles converge in pairs on either side of opposite cleavage furrows. The resulting spindle remains quadripolar. Although -tubulin is most concentrated in the deeply concave poles straddling cleavage furrows, it also extends into the spindle itself. Telophase groups of chromosomes curve around the polar cleavage furrows and a phragmoplast that originates in the interzonal region guides a cell plate that extends to the equatorial cleavage furrows. Discrete POs are reformed at opposite tips of the elongated dyad nuclei in prophase II and microtubules radiating from them give rise to the spindles of second meiosis. Spindles remain sharply focused and -tubulin extends into distal portions of the spindle. Interzonal phragmoplasts that expand to join with pre-established cleavage furrows mediate cytokinesis resulting in a tetrad of spores. Each young tetrad member has a radial microtubule system emanating from the nucleus.     

Calcium waves along the cleavage furrows in cleavage-stage Xenopus embryos and its inhibition by heparin

Calcium signaling is known to be associated with cytokinesis; however, the detailed spatio-temporal pattern of calcium dynamics has remained unclear. We have studied changes of intracellular free calcium in cleavage-stage Xenopus embryos using fluorescent calcium indicator dyes, mainly Calcium Green-1. Cleavage formation was followed by calcium transients that localized to cleavage furrows and propagated along the furrows as calcium waves. The calcium transients at the cleavage furrows were observed at each cleavage furrow at least until blastula stage. The velocity of the calcium waves at the first cleavage furrow was approximately 3 microns/s, which was much slower than that associated with fertilization/egg activation. These calcium waves traveled only along the cleavage furrows and not in the direction orthogonal to the furrows. These observations imply that there exists an intracellular calcium-releasing activity specifically associated with cleavage furrows. The calcium waves occurred in the absence of extracellular calcium and were inhibited in embryos injected with heparin an inositol 1,4,5-trisphosphate (InsP3) receptor antagonist. These results suggest that InsP3 receptor-mediated calcium mobilization plays an essential role in calcium wave formation at the cleavage furrows.     

THE ANATOMY AND THREE-DIMENSIONAL MORPHOLOGY OF ANNULARIA HOSKINSII SP. N.

Foliage attached to calamitean shoots is described from coal ball petrifactions of Middle and Late Pennsylvanian age. Leaves correspond to the compression-impression genus Annularia and thus represent the first attached members of this genus to be recognized as petrifactions. Individual leaves contain a single unbranched vascular bundle flanked by wide lateral laminar areas which occupy more than half the leaf cross sectional area. Stomata are confined to broad bands within concave furrows which parallel the vascular bundle on the abaxial leaf surface. Epidermal cells within these furrows are in rows aligned obliquely to the leaf axis, and the rows angle outward at a slight angle towards the leaf margin. Convolutions of the leaf-bearing axes result from nodal diaphragms which are oblique to the shoot axis. Whorled leaves apparently radiate outward in the plane of each obliquely positioned nodal diaphragm. This petrified material helps explain the apparent flattening of entire nodal diaphragms and leaf whorls within the same plane as seen in compression specimens. Annularia hoskinsii sp. n. is proposed, and the systematic position of structurally preserved Annularia foliage relative to the genus Dicalamophyllum is discussed.     

Axis determination in eggs of Xenopus laevis: A critical period before first cleavage,identified by the common effects of cold,pressure and ultraviolet irradiation

Exposure of eggs of Xenopus laevis to a temperature of 1.0°C for 4 min or a pressure of 8000 psi for 5 min in a critical period before first cleavage results in embryos exhibiting a reduction and loss of structures of the body axis. The deficiencies occur in a craniocaudal progression which is dose dependent. In the extreme, totally axis-deficient embryos with radial symmetry are formed. Maximum sensitivity to cold and pressure occurs at 0.6 of the time from fertilization to first cleavage and extends from approximately 0.4 to 0.8, the period between pronuclear contact and mitosis, and the approximate period of gray crescent formation. The effects of cold and pressure resemble those previously reported for uv irradiation in that (1) the types of axis-deficient embryos produced are morphologically indistinguishable; (2) sensitivity in all cases ends before 0.8; (3) cold and uv effects, although not those of pressure, can be prevented by cotreatment with D₂O; and (4) impaired eggs can be rescued by oblique orientation. We interpret these results as follows: during the 0.4–0.8 period the egg reorganizes its contents in a manner critical for subsequent development of the embryonic body axis. The reorganization process involves cytoskeletal elements, some of which are sensitive to cold, pressure, and uv, and protected by D₂O. Rescue by oblique orientation can be understood as the result of a gravity-driven reorganization of the egg's contents, supplanting the normal mechanochemical process impaired in treated eggs.     

Evolution of development in the sea star genus Patiriella: clade-specific alterations in cleavage

Examination of early development in five species of the Patiriella sea star species complex indicates that the ancestral-type radial holoblastic cleavage (Type I) is characteristic of P. regularis and P. exigua, whereas cleavage in species from the calcar clade followed multiple alternatives (Types II-IV) from holoblastic to meroblastic. Considering that invariant radial cleavage is thought to play a role in embryonic axis formation in echinoderms, we documented the details of blastomere formation in Patiriella sp. and followed development of the embryos. In Type II cleavage, the first and second cleavage planes appeared simultaneously at one pole of the embryo, dividing it directly into four equally sized blastomeres. In Type III cleavage, the first and second cleavage planes appeared simultaneously, followed promptly by the third cleavage plane, dividing the embryo directly into eight equally sized blastomeres. In Type IV cleavage, numerous furrows appeared simultaneously at one end of the embryo, dividing it into 32-40 equally sized blastomeres. Confocal sections revealed that embryos with cleavage Types II-IV were initially syncytial. The timing of karyokinesis in embryos with Types II and III cleavage was similar to that seen in clutch mates with Type I cleavage. Karyokinesis in embryos with Type IV cleavage, however, differed in timing compared with Type I clutch mates. Alteration in cleavage was not associated with polarized distribution of maternally provided nutrients. For each cleavage type, development was normal to the competent larval stage. Although variable blastomere configuration in the calcar clade may be linked to possession of a lecithotrophic development, other Patiriella species with this mode of development have typical cleavage. The presence of variable cleavage in all calcar clade species indicates that phylogenetic history has played a role in the distribution of this embryonic trait in Patiriella. The plasticity in early cleavage in these sea stars indicates that this aspect of early development is not constrained against change and that there are many ways to achieve multicellularity.     

The dorsoventral axis is specified prior to first cleavage in the direct developing sea urchin Heliocidaris erythrogramma

Previous fate mapping studies as well as the culture of isolated blastomeres have revealed that the dorsoventral axis is specified as early as the 2-cell stage in the embryos of the direct developing echinoid, Heliocidaris erythrogramma. Normally, the first cleavage plane includes the animal-vegetal axis and bisects the embryo between future dorsal and ventral halves. Experiments were performed to establish whether the dorsoventral axis is set up prior to the first cleavage division in H. erythrogramma. Eggs were elongated and fertilized in silicone tubes of a small diameter in order to orient the cleavage spindle and thus the first plane of cell division. Following first cleavage, one of the two resulting blastomeres was then microinjected with a fluorescent cell lineage tracer dye. Fate maps were made after culturing these embryos to larval stages. The results indicate that the first cleavage division can be made to occur at virtually any angle relative to the animal-vegetal and dorsoventral axes. Therefore, the dorsoventral axis is specified prior to first cleavage. We argue that this axis resides in the unfertilized oocyte rather than being set up as a consequence of fertilization.     

Density dependent growth in adult brown frogs Rana arvalis and Rana temporaria – A field experiment

In species with complex life cycles, density regulation can operate on any of the stages. In frogs there are almost no studies of density effects on the performance of adult frogs in the terrestrial habitat. We therefore studied the effect of summer density on the growth rate of adult frogs during four years. Four 30 by 30 m plots in a moist meadow were used. In early summer, when settled after post-breeding migration, frogs (Rana arvalis and Rana temporaria that have a very similar ecology and potentially compete) were enclosed by erecting a fence around the plots. Frogs were captured, measured, marked and partly relocated to create two high density and two low density plots. In early autumn the frogs were again captured and their individual summer growth determined. Growth effects were evaluated in relation to two density measures: density by design (high/low manipulation), and actual (numerical) density. R. arvalis in plots with low density by design grew faster than those in high density plots. No such effect was found for R. temporaria. For none of the species was growth related to actual summer density, determined by the Lincoln index and including the density manipulation. The result suggests that R. arvalis initially settled according to an ideal free distribution and that density had a regulatory effect (mediated through growth). The fact that there were no density effects on R. temporaria (and a significant difference in its response to that of R. arvalis) suggests it is a superior competitor to R. arvalis during the terrestrial phase. There were no density effects on frog condition index, suggesting that the growth rate modifications may actually be an adaptive trait of R. arvalis. The study demonstrates that density regulation may be dependent on resources in frogs' summer habitat.     

Unequal first cleavage in the Tubifex egg: involvement of a monastral mitotic apparatus

The first cleavage in the freshwater oligochaete Tubifex hattai is unequal and meridional, and produces a smaller cell AB and a larger cell CD. This study traces the process of furrow formation, reorganization of cortical F-actin and the assembly of a mitotic apparatus during this unequal division. Cleavage furrow formation consists of two stages: (i) when eggs are viewed from the animal pole, meridionally running furrows emerge at two points of the egg's equator that are 90° apart from each other and approach the egg axis as they deepen; and (ii) at the midpoint between the equator and the egg center, the bottoms of these furrows link to each other on the animal and vegetal surfaces of the egg and form a continuous ring of constriction in a plane parallel to the egg axis. Egg cortices, isolated during the first step and stained with rhodamine-phalloidin, show that the bottoms of recently formed furrows are underlaid by a belt of tightly packed actin bundles (i.e. a contractile arc). The transition to the second stage of furrow formation coincides with the conversion of these actin belts into a continuous ring of F-actin. Whole-mount immunocytochemistry of microtubules reveals that the first cleavage in Tubifex involves an asymmetric mitotic spindle, which initially possesses an aster at one pole but not the other. This ‘monastral’ spindle is located at the egg's center and orients itself perpendicular to the egg axis. During anaphase, astral rays elongate to reach the cell surface, so that the array of astral microtubules in the plane of the egg's equator covers a sector of 270–300°. In contrast, it is not until the transition to telophase that microtubules emanating from the anastral spindle pole approach the cell margin. If eggs are compressed along the egg axis or forced to elongate, they form monastral spindles and divide unequally. In living compressed eggs, mitotic spindles, which are recognizable as bright streaks at the egg's center, appear not to shift their position along the spindle axis during division, suggesting that without eccentric migration of spindles Tubifex eggs are able to divide unequally. These results suggest that mechanisms that translocate the mitotic spindle eccentrically do not operate in Tubifex eggs during the first cell cycle. The mechanisms that generate asymmetry in spindle organization are discussed in the light of the present results.     

Constitutive and stress‐inducible expression of the endoplasmic reticulum heat shock protein 70 gene family member,immunoglobulin‐binding protein (BiP), during Xenopus laevis early development

We have characterized the constitutive and stress‐inducible pattern of immunoglobulin‐binding protein (BiP) gene expression during Xenopus early development. Whole mount in situ hybridization analysis revealed that BiP mRNA was detected in unfertilized eggs, cleavage and blastula stage embryos. In gastrulae, BiP mRNA was present across the surface of the embryo, while in neurulae BiP mRNA was enriched in the neural plate, neural fold, and around the blastopore. In early and late tailbud embryos, BiP mRNA was found primarily in the dorsal region. Tunicamycin and A23187, the calcium ionophore, enhanced BiP mRNA accumulation first at the neurula stage, while heat shock induced BiP mRNA accumulation first at the gastrula stage. Compared to control, A23187‐ and heat shock‐treated neurulae displayed relatively high levels of BiP mRNA in selected tissues, including the neural plate, neural folds, around the blastopore, and ectoderm. At the early tailbud stage, A23187 and heat shock enhanced BiP mRNA accumulation primarily in the head, somites, tail, and along the spinal cord. A similar situation was found with A23187‐ and heat shock‐treated late tailbud embryos, except that heat‐shocked embryos also displayed enhanced BiP mRNA accumulation in the epidermis. These studies demonstrate a preferential accumulation of BiP mRNA in selected tissues during development and in response to stress. Dev. Genet. 25:31–39, 1999. © 1999 Wiley‐Liss, Inc.     

On the mechanism of electrical coupling between cells of earlyXenopus embryos

Summary The mechanism of electrical coupling between cells of earlyXenopus embryos has been studied by examination of the nonjunctional membrane resistances and capacitances as a function of cleavage stage, the junctional and nonjunctional membrane resistances as functions of time during the first cleavage, and the electrical properties of the primitive blastocoel. The changes in membrane resitances and capacitances during the first two cleavages may be completely explained by the addition of new membrane, identical in specific resistance and capacitance to the original membrane, at a constant rate to furrows which are electrically connected to the perivitelline space. Microelectrode recording from the primitive blastocoel indicates that there is no electrical difference detectable between it and the perivitelline space. These results are discussed in the context of current theories of the mechanism of intercellular electrotonic coupling.     

Experimental change of cytoplasmic composition can convert determination of blastomeres inPlatynereis dumerilii (Annelida,Polychaeta)

Summary Early development ofPlatynereis dumerilii is characterized by an extremely constant cleavage pattern in which the volumes and cytoplasmic contents of the blastomeres show remarkably little variability (Dorresteijn 1990). In order to test the necessity of a precise partitioning of the cytoplasm, we have stratified the ooplasm by mild centrifugation (10 min at 300 g) after completion of meiosis but before first cleavage. The cytoplasm of the zygote stratifies randomly with respect to the pre-existing animal-vegetal axis, but first cleavage follows the animal-vegetal axis dividing the plasm before it has rearranged to its normal distribution. As usual, first cleavage is unequal in the majority of centrifuged eggs. Different sorts of cytoplasm are always distributed abnormally in comparison to normal two-cell stages. Under two circumstances this leads to the formation of double trunk structures in the young worm. Such double monsters either originate from zygotes whose clear cytoplasm has been distributed equally to the two daughter blastomeres at first cleavage, or from unequal two-cell embryos whose larger blastomere cleaved equally at second cleavage forming blastomeres with equal lots of clear cytoplasm. Cell-lineage could be followed in an individual embryo of the latter category and showed the existence of two adjacent D-quadrants, giving rise to a double monster with a forked trunk. Embryos of the former category give rise to two opponent D-quadrants and double monsters with a four-sided trunk. As in the normal embryo, the amount of clear cytoplasm in a blastomere is positively correlated with the speed of its cell cycle, and endows the cell with D-quadrant developmental capacities as can be judged by the cleavage pattern.     

Morphometric differentiation of the moor frog (Rana arvalis Nilss.) in Central Europe

Morphometric variation of 11 characters was studied in 14 samples of Rana arvalis from allopatric population groups in Central Europe, representing two subspecies formerly recognised: R. a. arvalis and R. a. wolterstorffi. All samples from Poland (nine) were collected from the area, which is believed to be populated by the nominal form. In addition, the isolated population of R. arvalis from the Eastern Carpathians of Romania was classified to that form. All the Hungarian samples fall into the range of R. a. wolterstorffi. No significant differences in body size between sexes and among the populations from the three geographic regions (Poland, Hungary and Romanian Eastern Carpathians) were found. Multivariate analysis of the differences in body shape among groups showed that the populations from Hungary differed from the nominal form mainly in the relative hindlimb length. However, substantial overlap in overall shape differences resulted in a relatively low percentage of correct classifications to the respective geographic groups in the Discriminant Analysis. Large intrapopulation variation in the values of the two ratios (SVL/TL and TL/IMT) formerly used for differentiate the two European subspecies of R. arvalis makes them unreliable characters for distinguishing the two forms. The pattern of morphometric variation does not match the overall genetic divergence of the R. arvalis populations, which suggests that the body shape differences in this species result from the phenotypic plasticity correlated with local climatic factors.