Partitioning maternal and exogenous diet contributions to otolith 87Sr/86Sr in Kokanee Salmon (Oncorhynchus nerka)

Strontium isotopes (87Sr/86Sr) recorded in the otoliths of Pacific Salmon (Oncorhynchus spp.) are commonly used to identify natal origin. For species that migrate at or soon after emergence, the embryonic region of the otolith provides the only record of provenance. However, maternal contribution of Sr from the yolk can confound the isotopic signature of the natal site. We experimentally quantified maternal and exogenous diet contributions to otolith 87Sr/86Sr over embryonic development in Kokanee salmon (O. nerka). Eggs from two populations in isotopically distinct lakes were incubated and reared in a common water source. Timing of developmental events and proportional contribution from yolk to otolith 87Sr/86Sr differed significantly between the two populations. We suggest that the magnitude of difference in 87Sr/86Sr between yolk and water, the relative concentrations of Sr and Ca in these isotopic sources, and population-specific effects on otolith growth and composition contribute to this variation. Understanding how these factors affect otolith 87Sr/86Sr could extend the use of otolith geochemistry for determining provenance to species and populations in which natal site rearing is limited.

Heterochronic shifts and conserved embryonic shape underlie crocodylian craniofacial disparity and convergence

The distinctive anatomy of the crocodylian skull is intimately linked with dietary ecology, resulting in repeated convergence on blunt- and slender-snouted ecomorphs. These evolutionary shifts depend upon modifications of the developmental processes which direct growth and morphogenesis. Here we examine the evolution of cranial ontogenetic trajectories to shed light on the mechanisms underlying convergent snout evolution. We use geometric morphometrics to quantify skeletogenesis in an evolutionary context and reconstruct ancestral patterns of ontogenetic allometry to understand the developmental drivers of craniofacial diversity within Crocodylia. Our analyses uncovered a conserved embryonic region of morphospace (CER) shared by all non-gavialid crocodylians regardless of their eventual adult ecomorph. This observation suggests the presence of conserved developmental processes during early development (before Ferguson stage 20) across most of Crocodylia. Ancestral state reconstruction of ontogenetic trajectories revealed heterochrony, developmental constraint, and developmental systems drift have all played essential roles in the evolution of ecomorphs. Based on these observations, we conclude that two separate, but interconnected, developmental programmes controlling craniofacial morphogenesis and growth enabled the evolutionary plasticity of skull shape in crocodylians.

Female gametophyte development and embryogenesis in Taxus baccata L.

Crassinucellate ovules are initiated in <em>Taxus</em>, directly from the shoot apex. The rudimentary pollen chamber is formed in the nucellus. A linear tetrad of megaspores with a functional chalazal megaspore is formed. A free-nuclear stage is charac-teristic at the beginning of megagametophyte development. Archegonia without ventral canal cell are solitary or in complexes. The embryo has a very long suspensor even after maturation. Two types of polyembryony have been revealed: i) embryogenic redifferentiation of suspensor cells and ii) cleavage of embryonic region in the early embryo. In the northern temperate climate of St. Petersburg one month delay in development of reproductive structures has been noted.

The zebrafish buttonhead-like factor Bts1 is an early regulator of pax2.1 expression during mid-hindbrain development

Little is known about the factors that control the specification of the mid-hindbrain domain (MHD) within the vertebrate embryonic neural plate. Because the head-trunk junction of the Drosophila embryo and the MHD have patterning similarities, we have searched for vertebrate genes related to the Drosophila head gap gene buttonhead (btd), which in the fly specifies the head-trunk junction. We report here the identification of a zebrafish gene which, like btd, encodes a zinc-finger transcriptional activator of the Sp-1 family (hence its name, bts1 for btd/Sp-related-1) and shows a restricted expression in the head. During zebrafish gastrulation, bts1 is transcribed in the posterior epiblast including the presumptive MHD, and precedes in this area the expression of other MHD markers such as her5, pax2.1 and wnt1. Ectopic expression of bts1 combined to knock-down experiments demonstrate that Bts1 is both necessary and sufficient for the induction of pax2.1 within the anterior neural plate, but is not involved in regulating her5, wnt1 or fgf8 expression. Our results confirm that early MHD development involves several genetic cascades that independently lead to the induction of MHD markers, and identify Bts1 as a crucial upstream component of the pathway selectively leading to pax2.1 induction. In addition, they imply that flies and vertebrates, to control the development of a boundary embryonic region, have probably co-opted a similar strategy: the restriction to this territory of the expression of a Btd/Sp-like factor.

Isolation of novel tissue-specific genes from cDNA libraries representing the individual tissue constituents of the gastrulating mouse embryo

A total of 5 conventional, directionally cloned plasmid cDNA libraries have been constructed from the entire embryonic region of the mid-gastrulation mouse embryo and from its four principal tissue constituents (ectoderm, mesoderm, endoderm and primitive streak). These libraries have been validated with respect to the number of independent clones, insert-size and appropriate representation of diagnostic marker genes. Subtractive hybridisation has been used to remove clones common to the Endoderm and Mesoderm cDNA libraries resulting in an Endoderm minus Mesoderm subtracted library. Probe prepared from this subtracted library has been hybridised to a grid containing approximately 18,500 Embryonic Region library clones. Three novel clones have been recovered as well as expected genes already known to be highly expressed in the primitive endoderm lineage at this stage of development. In situ hybridisation to early postimplantation embryos has revealed the expression patterns of these novel genes. One is highly expressed exclusively in visceral endoderm, one is expressed in ectodermal and endodermal tissues, and the third proves to be an early marker of prospective and differentiated surface ectoderm as well as being expressed in endoderm and its derivatives.

Relationships between mesoderm induction and the embryonic axes in chick and frog embryos

The hypoblast is generally thought to be responsible for inducing the mesoderm in the chick embryo because the primitive streak, and subsequently the embryonic axis, form according to the orientation of the hypoblast However, some cells become specified as embryonic mesoderm very late in development, towards the end of the gastrulation period and long after the hypoblast has left the embryonic region. We argue that induction of embryonic mesoderm and of the embryonic axis are different and separable events, both in amniotes and in amphibians. We also consider the relationships between the dorsoventral and anteroposterior axes in both groups of vertebrates.

Neural induction and regionalisation in the chick embryo

Induction and regionalisation of the chick nervous system were investigated by transplanting Hensen's node into the extra-embryonic region (area opaca margin) of a host embryo. Chick/quail chimaeras were used to determine the contributions of host and donor tissue to the supernumerary axis, and three molecular markers, Engrailed, neurofilaments (antibody 3A10) and XlHbox1/Hox3.3 were used to aid the identification of particular regions of the ectopic axis. We find that the age of the node determines the regions of the nervous system that form: young nodes (stages 2–4) induced both anterior and posterior nervous system, while older nodes (stages 5–6) have reduced inducing ability and generate only posterior nervous system. By varying the age of the host embryo, we show that the competence of the epiblast to respond to neural induction declines after stage 4. We conclude that during normal development, the initial steps of neural induction take place before stage 4 and that anteroposterior regionalisation of the nervous system may be a later process, perhaps associated with the differentiating notochord. We also speculate that the mechanisms responsible for induction of head CNS differ from those that generate the spinal cord: the trunk CNS could arise by homeogenetic induction by anterior CNS or by elongation of neural primordia that are induced very early.

Shell implosion depth and implosion morphologies in three species of Sepia (Cephalopoda) from the Mediterranean Sea

The maximum habitation depths of chambered cephalopods are dictated by the mechanical properties of the shell. All chambered cephalopods have a depth at which ambient pressure is sufficient to implode the gas-filled shell portions. Experiments on Sepia elegans D'Orbigny, S. officinalis Linne and S. orbignyana Férussac from the Mediterranean Sea show these three species to have differing depth limits and modes of shell implosion. Large S. officinalis implode between 150 and 200 m, whereas newly hatched and advanced embryonic specimens implode between 50 and 100 m. The larger S. officinalis are occasionally caught at depths greater than the implosion depth of the juvenile shell parts. They apparently avoid implosion of the early shell portions by refilling these first-formed chambers with cameral liquid later in life. Implosion in S. officinalis generally resulted in the crushing of all or most of the septa in a band extending from the embryonic region to the anterior part of the shell. Implosion was generally accompanied by fatal rupture of underlying tissue into the implosion zones. Implosion of S. orbignyana occurred between 550 and 600 m and was very different in form from that in S. officinalis, occurring mostly within the smooth zone of the last-formed several chambers, and rarely extending back into the siphuncular region (striae zone). Increasing depth caused episodic implosion of sequentially older chambers in the smooth zone. The shallower implosions were accompanied by little soft-tissue damage. Massive internal injury only occurred in the deepest implosions (700 m or greater). Implosion in S. elegans occurred betw en 400 and 600 m. Too few specimens were available to allow generalizations about morphology of implosion in this species.

Localization and synthesis of alphafoetoprotein in post-implantation mouse embryos

The localization and synthesis of alphafoetoprotein (AFP) during mouse embryogenesis were studied by immunoperoxidase and by immunoprecipitation after radioactive labelling, using an antiserum prepared against AFP. AFP is first detectable in embryos on the 7th day of gestation (7th day embryos). In 7th and 8th day embryos AFP is confined to visceral (proximal) endoderm cells around the embryonic region of the egg cylinder. Visceral extra-embryonic and parietal (distal) endoderm cells do not contain AFP. By the 9th day of gestation AFP is also present in the extra-embryonic ectoderm, mesoderm and embryonic ectoderm cells around the three cavities of the embryo. These tissues do not synthesize AFP when cultured in isolation, but can adsorb AFP when it is added to the medium. On the 12th day of gestation AFP synthesis is confined to the endoderm layer of the visceral yolk sac. It is concluded that the ability to synthesize AFP is a property which is restricted to the visceral endoderm during early post-implantation development. The presence of AFP in other tissues of the embryo appears to be due to adsorption.

Endodermal formation in blastocytes of the marsupials Isoodon macrousus and Perameles nasuta

Eleven embryos, ranging from partly to fully bilaminar blastocysts, were obtained from the bandicoots I. macrourus and P. nasuta and were examined by light and electron microscopy. The morphological changes which occurred during the differentiation of the endoderm and ectoderm are described. The shell membrane was thinner than it was in unilaminar blastocysts and had a deposit of material of irregular thickness on its outer surface. The mucoid coat and zona pellucida were absent or discontinuous. Endoderm formation was first observed in blastocysts about 1.0 mm in diameter. Cells migrated inwards from regions of thickened protoderm to form a continuous layer of similar flattened cells - the endoderm - beneath the protoderm, which then became the ectoderm. The blastocysts were fully bilaminar when they were 1.5-1.9 mm in diameter. At this stage the ectoderm was composed of two distinct regions, an embryonic region of cuboidal cells and a non-embryonic region of flattened cells resembling the cells of the endoderm. The formation of the endoderm in bandicoots closely resembles that described in other marsupials, except the American opossum Didelphis virginiana in which endoderm cells are released into the blastocoele before they form a continuous layer.