Gerald W. Eagleson

Origin and segregation of cranial placodes in Xenopus laevis.

Cranial placodes are local thickenings of the vertebrate head ectoderm that contribute to the paired sense organs (olfactory epithelium, lens, inner ear, lateral line), cranial ganglia and the adenohypophysis. Here we use tissue grafting and dye injections to generated fate maps of the dorsal cranial part of the non-neural ectoderm for Xenopus embryos between neural plate and early tailbud stages. We show that all placodes arise from a crescent-shaped area located around the anterior neural plate, the pre-placodal ectoderm. In agreement with proposed roles of Six1 and Pax genes in the specification of a panplacodal primordium and different placodal areas, respectively, we show that Six1 is expressed uniformly throughout most of the pre-placodal ectoderm, while Pax6, Pax3, Pax8 and Pax2 each are confined to specific subregions encompassing the precursors of different subsets of placodes. However, the precursors of the vagal epibranchial and posterior lateral line placodes, which arise from the posteriormost pre-placodal ectoderm, upregulate Six1 and Pax8/Pax2 only at tailbud stages. Whereas our fate map suggests that regions of origin for different placodes overlap extensively with each other and with other ectodermal fates at neural plate stages, analysis of co-labeled placodes reveals that the actual degree of overlap is much smaller. Time lapse imaging of the pre-placodal ectoderm at single cell resolution demonstrates that no directed, large-scale cell rearrangements occur, when the pre-placodal region segregates into distinct placodes at subsequent stages. Our results indicate that individuation of placodes from the pre-placodal ectoderm does not involve large-scale cell sorting in Xenopus.

Spatial distribution of abundant proteins in oocytes and fertilized eggs of the Mexican axolotl (Ambystoma mexicanum).

Two-dimensional protein patterns were compared from sections along the longitudinal axis of oocytes and fertilized eggs of the Mexican axolotl (Ambystoma mexicanum). Only a few differences were observed between four different sections through both oocyte and fertilized eggs. A set of proteins (14 out of 120 proteins) were found that reside only in the germinal vesicles (GV) of the fully grown oocyte. Two of these were observed exclusively in the vegetal half, and one in the animal half after GV breakdown, while other proteins were randomly distributed within the fertilized egg. One cytoplasmic protein was present only in the vegetal half of the mature oocyte and became present also in the animal half of the fertilized egg. Additional proteins were observed in all transverse sections of both mature oocyte and fertilized eggs. It is proposed that these proteins are modified rather than newly synthesized proteins.

Forebrain Differentiation and Axonogenesis in Amphibians: I. Differentiation of the Suprachiasmatic Nucleus in Relation to Background Adaptation Behavior

Functional forebrain development is the result of a complex series of early developmental processes which include cell division, cellular rearrangements, tissue-tissue interactions, cellular determinative and differentiation events, and axonogenesis. In these studies, Xenopus laevis embryos were examined for early forebrain neuronal determination, differentiation and axonogenesis with special emphasis on the hypothalamic area known to be involved in regulating pars intermedia function. Whole brain acetylcholine esterase (AChE) histochemistry was used to follow the early pattern of forebrain neuronal differentiation, and whole brain acetylated-tubulin immunocytochemistry was done to follow early forebrain axonogenesis. AChE histochemistry indicated that the source of the tract of the postoptic commissure (stpoc) was the first forebrain area to begin differentiation (stage 22). Whole brain immunocytochemistry for acetylated-tubulin indicated that the tpoc is also the first forebrain tract to develop (at stage 25/26). The main forebrain tracts have developed and become interconnected by stage 35/36. The forebrain undergoes a pronounced extension, with much cellular mixing and rearrangement during stages 37/38 to 43/44. This results in bending and contortions in the already developed tracts. Whole brain immunocytochemistry for tyrosine hydroxylase and extirpation of the stage 14 presumptive suprachiasmatic (SC) area indicated that the dopaminergic cells of the SC are determined by stage 14 and initially undergo differentiation between stages 37/38 and 40. Tadpoles with stage 14 presumptive SC extirpated lacked TH-positive tracts to the pars intermedia, lacked most midline TH-positive forebrain cells, and also failed to background adapt to white background. Thus, the SC tracts to the pars intermedia that inhibit melanotrope secretion probably form during the extension stages of 37/38 and contact the pars intermedia by stage 40 when animals are first capable of background adaptation.

Stage-specific effects of retinoic acid on gene expression during forebrain development

Treatment of early gastrula- and neurula-staged Xenopus embryos with all-trans retinoic acid (RA) results in truncation of the anterior structures of the forebrain and head. The extent of truncation is dependent upon both the stage of immersion and the RA concentration used. As a method to investigate genes important during early forebrain regionalization, late gastrula and neurula embryos were immersed for 2h within low (1x10(-9)M to 5x10(-8)M) concentrations of RA. Embryos were allowed to develop to tadpole stages and forebrain marker genes were assessed for any alteration in patterns of expression. Comparisons of controls to experimental groups indicated that the greatest sensitivity to low levels of RA occurred just before the initial expression of the forebrain-specific markers investigated. We concluded that forebrain regionalization and gene expression occurred in the following order: Xotx2-->Xsix3-->Xrx (&Xfez1)-->Xbf1-->Xemx1. Xsix3 seems to be very important for the initial parcellation of telencephalon, retinal and diencephalon areas.

Paraboloid of axolotl retinal photoreceptor and bovine pituitary thyrotropin fraction share an antigen

Lenses of newts (genera Notophthalmus, Triturus, Cynops) regenerate from irises in the presence of retinae of larval frogs (Rana) or adult salamanders (Hynobius, Ambystoma), species which are themselves incapable of lens regeneration from the iris. In newts, bovine pituitary thyrotropin preparation NIH-TSH-B8 can also stimulate lens regeneration from the iris. An antiserum against NIH-TSH-B7 (purified as is NIH-TSH-B8), absorbed with bovine lutropin preparation NIH-LH-B9, cross-reacts with bovine retinal glycoprotein extracts in immunodiffusion tests, and with retinal photoreceptor cells of the axolotl (Ambystoma mexicanum), as evidenced by immunofluorescence. In normal adult eyes and in eyes 21 days after lens removal, the paraboloid portion of the photoreceptor inner segments, and in some cases the perinuclear cytoplasm of the photoreceptor cells, contained the antigen. The cross-reacting antigen appears to be different from thyrotropin, and also different from the basic and acidic retinal fibroblast growth factors. However, immunodiffusion reveals a precipitation arc with retina-derived growth factor fraction III (EDGF III). If bovine pituitary thyrotropin preparations produce lens regeneration, and if these preparations cross-react with an antigen in the retinal photoreceptors, the retinal antigen may be involved in the stimulation of lens regeneration as well.