The first cleavage of the mouse zygote predicts the blastocyst axis

One of the unanswered questions in mammalian development is how the embryonic-abembryonic axis of the blastocyst is first established. It is possible that the first cleavage division contributes to this process, because in most mouse embryos the progeny of one two-cell blastomere primarily populate the embryonic part of the blastocyst and the progeny of its sister populate the abembryonic part. However, it is not known whether the embryonic-abembryonic axis is set up by the first cleavage itself, by polarity in the oocyte that then sets the first cleavage plane with respect to the animal pole, or indeed whether it can be divorced entirely from the first cleavage and established in relation to the animal pole. Here we test the importance of the orientation of the first cleavage by imposing an elongated shape on the zygote so that the division no longer passes close to the animal pole, marked by the second polar body. Non-invasive lineage tracing shows that even when the first cleavage occurs along the short axis imposed by this experimental treatment, the progeny of the resulting two-cell blastomeres tend to populate the respective embryonic and abembryonic parts of the blastocyst. Thus, the first cleavage contributes to breaking the symmetry of the embryo, generating blastomeres with different developmental characteristics.     

A simple method for counting nuclei in the preimplantation mouse embryo

An easy and rapid method of counting the number of cells in the preimplantation mouse embryo is described. The procedure increases the speed with which large numbers of embryos can be processed using a simple squash technique. Cell numbers are determined by exposing the embryos to the fluorescent DNA-binding dye, Hoechst 33258, removing the zona pellucida and simply squashing the embryo and counting the number of fluorescent nuclei. An increase in fluorescent intensity and maintenance of nuclear conformation of the squashed preparations are greatly improved by the use of the non-ionic detergent Triton X-100. Viability of dye-treated fertilized one-cell and blastocyst stage embryos is maintained at least up to day 13 of pregnancy following transfer of the embryos to the uteri of pseudopregnant recipients. Additional uses for this staining technique are discussed.     

Tbx6-dependent Sox2 regulation determines neural or mesodermal fate in axial stem cells

The classical view of neural plate development held that it arises from the ectoderm, after its separation from the mesodermal and endodermal lineages. However, recent cell-lineage-tracing experiments indicate that the caudal neural plate and paraxial mesoderm are generated from common bipotential axial stem cells originating from the caudal lateral epiblast. Tbx6 null mutant mouse embryos which produce ectopic neural tubes at the expense of paraxial mesoderm must provide a clue to the regulatory mechanism underlying this neural versus mesodermal fate choice. Here we demonstrate that Tbx6-dependent regulation of Sox2 determines the fate of axial stem cells. In wild-type embryos, enhancer N1 of the neural primordial gene Sox2 is activated in the caudal lateral epiblast, and the cells staying in the superficial layer sustain N1 activity and activate Sox2 expression in the neural plate. In contrast, the cells destined to become mesoderm activate Tbx6 and turn off enhancer N1 before migrating into the paraxial mesoderm compartment. In Tbx6 mutant embryos, however, enhancer N1 activity persists in the paraxial mesoderm compartment, eliciting ectopic Sox2 activation and transforming the paraxial mesoderm into neural tubes. An enhancer-N1-specific deletion mutation introduced into Tbx6 mutant embryos prevented this Sox2 activation in the mesodermal compartment and subsequent development of ectopic neural tubes, indicating that Tbx6 regulates Sox2 via enhancer N1. Tbx6-dependent repression of Wnt3a in the paraxial mesodermal compartment is implicated in this regulatory process. Paraxial mesoderm-specific misexpression of a Sox2 transgene in wild-type embryos resulted in ectopic neural tube development. Thus, Tbx6 represses Sox2 by inactivating enhancer N1 to inhibit neural development, and this is an essential step for the specification of paraxial mesoderm from the axial stem cells.     

Demographic characteristics,seasonal range and habitat topography of Balkan chamois population in its southernmost limit of its distribution (Giona mountain,Greece)

The annual range of Balkan chamois (Rupicapra rupicapra balcanica) in Giona mountain was found to be 5502 ha, with a low population density (2 individuals/100 ha). Seasonal range patterns varied significantly, with a minimum extent in summer and a maximum in winter (30% and 79% of the annual range, respectively). Summer stress and the rutting period might be associated with the observed aggregated distributions during the summer and autumn (core areas of 28% and 22% of seasonal ranges, respectively, defined after the Fixed Kernel Density Estimator). Chamois were found to use significantly lower altitude habitats in winter (1212 m) than in summer (2223 m), and significantly steeper slopes in winter (35°); aspect was not found to have a significant effect on habitat use. Population structure consisted of kids (21%), yearlings (8%), females (35%) and males (36%). Conservation management for the species should consider poaching, livestock competition and global warming.     

A simple method for counting nuclei in the preimplantation mouse embryo

Summary An easy and rapid method of counting the number of cells in the preimplantation mouse embryo is described. The procedure increases the speed with which large numbers of embryos can be processed using a simple squash technique. Cell numbers are determined by exposing the embryos to the fluorescent DNA-binding dye, Hoechst 33258, removing the zona pellucida and simply squashing the embryo and counting the number of fluorescent nuclei. An increase in fluorescent intensity and maintenance of nuclear conformation of the squashed preparations are greatly improved by the use of the non-ionic detergent Triton X-100. Viability of dye-treated fertilized one-cell and blastocyst stage embryos is maintained at least up to day 13 of pregnancy following transfer of the embryos to the uteri of pseudopregnant recipients. Additional uses for this staining technique are discussed.Acknowledgment. Financial support was provided by NIH Grant HDD-06210 (KME) and by Basil O'Connor Starter Research Grant No. 5-379 from the March of Dimes Birth Defects Foundation (VEP). We thank Steven Halpern for help with the photography and Jon Flax for expert technical assistance.     

‘Handedness’ in mice: Some behavioural correlates

Résumé Nous avons observé que les souris manifestent une préférence pour l'usage de l'une ou de l'autre de leurs pattes antérieures. Cette préférence ne dépend ni de la capacité d'apprendre ni de la tendence à prendre l'une ou l'autre pour tourner. On peut modifier cette préférence par des lésions corticales.