bluuebunny/crossref_metadata_2025 huggingface dataset:
Ablation of the fetal mouse spinal cord: The effect on soleus muscle cytoarchitecture
Authors: Hughes Donna S. , Schade Robert R. , Ontell Marcia
Published: February 2005

A technique is described whereby it is possible to surgically ablate the lumbosacral spinal cord of a developing mouse fetus without interfering with fetal viability. The lumbosacral spinal cords of 14‐day in utero, 129ReJ mice were ablated with a Cooper Nd‐YAG laser, and the fetuses, enclosed in their membranes and attached to the uterus by their placentae, were allowed to develop in the abdominal cavity of the dam. The cytoarchitecture and the temporal pattern of organogenesis of aneural soleus muscles were studied in spaced, serial, transverse, ultrathin sections of muscles of 16‐ and 18‐day gestation and newborn (20‐day gestation) mice. At the time of surgery, the soleus muscle was a discrete mass consisting of primary myotubes and a pleomorphic population of mononucleated cells. Axon bundles and blood vessels were found at the muscle's periphery, but had not penetrated throughout the muscle mass. The organogenesis of the aneural muscle was remarkably similar to that of the innervated soleus muscle (Ontell et al., Am J Anat 181:267–278, 1988). In the aneural muscle, as in the innervated muscle, significant numbers of secondary myotubes formed all along the lengths of primary myotubes. Moreover, the time course of myotube formation, the dynamics of cluster formation and cluster dispersal, and the ultrastructural appearance of the myotubes mimicked that observed in innervated muscle. The frequency of necrotic myotubes was no greater in the aneural muscle than in the innervated soleus muscle. Myotube maturation was similar in aneural and innervated soleus muscles until 18 days gestation. However, at birth, aneural myotubes appeared to be slightly less mature than innervated myotubes. Thus, the major morphogenic phenomena that characterize the development of the soleus muscle appear to be independent of innervation.
Morphometric analysis of the developing, murine aneural soleus muscle
Authors: Hughes Donna S. , Ontell Marcia
Published: February 2005

The pattern of organogenesis of the aneural soleus muscle of the 129ReJ mouse [rendered aneural by laser ablation of the lumbosacral spinal cord at 14 days in utero (during the period of primary myotube formation, but prior to the formation of secondary myotubes)] was evaluated quantitatively with spaced, serial ultrathin sections and computer‐assisted morphometric analysis. Aneural muscles from 16‐ and 18‐day gestation and newborn mice were analyzed to determine age‐related changes in a number of parameters including: muscles' maximal girths, numbers of myotubes, myotube diameter distributions, and cluster frequency. Data were compared with a similar study of the organogenesis of the normal soleus muscle (Ontell et al: Am J Anat 181: 279–288, 1988). Basic patterns of morphogenesis of the soleus muscle were unchanged by spinal cord ablation, and differences in development between the aneural and innervated muscles were quantitative rather than qualitative. At birth, the aneural muscle contained ∼76% of the myotubes found in the innervated muscle (∼840 myotubes in the innervated muscle and ∼640 in the aneural muscle). Evidence is presented consistent with the hypothesis that primary myotube formation is reduced by ∼32% in the aneural muscles and that while extensive secondary myotube formation occurs (∼78% of the myotubes present at birth in these muscles are secondary myotubes), there is a significant reduction in the number of secondary myotubes in aneural muscles. It is suggested that the reduced numbers of secondary myotubes may be related to the reduction in the number of primary myotubes, which are known to act as scaffolds for secondary myotube formation. The time course of secondary myotube formation and of cluster formation and cluster dispersal and the number of cells per cluster are similar in age‐matched, innervated and aneural muscles. The absence of innervation has little effect on myotube growth until birth, when comparison of the myotube diameter distributions reveals a slight alteration in myotube diameter distributions of aneural as compared with innervated muscles.
The cellular basis of the convergence and extension of the Xenopus neural plate
Authors: Keller Ray , Shih John , Sater Amy
Published: February 2005

There is great interest in the patterning and morphogenesis of the vertebrate nervous system, but the morphogenetic movements involved in early neural development and their underlying cellular mechanisms are poorly understood. This paper describes the cellular basis of the early neural morphogenesis of Xenopus laevis. The results have important implications for neural induction. Mapping the fate map of the midneurula (Eagleson and Harris: J. Neurobiol. 21:427–440, 1990) back to the early gastrula with time‐lapse video recording demonstrates that the prospective hindbrain and spinal cord are initially very wide and very short, and thus at the beginning of gastrulation all their precursor cells lie within a few cell diameters of the inducing mesoderm. In the midgastrula, the prospective hindbrain and spinal cord undergo very strong convergence and extension movements in two phases: In the first phase they primarily undergo thinning in the radial direction and lengthening (extension) in the animal‐vegetal direction, and the second phase is characterized primarily by mediolateral narrowing (convergence) and anterior‐posterior lengthening (extension). These movements also occur in sandwich explants of the gastrula, thus demonstrating the local automomy of the forces producing them. Tracing cell movements with fluorescein dextran‐labeled cells in embryos or explants shows that the initial thinning and extension occurs by radial intercalation of deep cells to form fewer layers of greater area, all of which is expressed as increased length. The subsequent convergence and extension occurs by mediolateral intercalation of deep cells to form a longer, narrower array. These results establish that a similar if not identical sequence of radial and mediolateral cell intercalations underlie convergence and extension of the neural and the mesoderm tissues (Wilson and Keller: Development, 112:289–300, 1991). Moreover, these results establish that radial and mediolateral intercalation are the principal neural cell behaviors induced by the planar signals emanating from the dorsal involuting marginal zone (the Spemann organizer) in the early gastrula (Keller et al: Develop. Dynamics, 193: 218–234, 1992). Radial and mediolateral intercalation are induced among the 5 to 7 rows of cells comprising the prospective hindbrain and spinal cord, thus producing the massive convergence and extension movements that narrow and elongate these regions of the nervous system in the late gastrula. A more general significance of these results is that neural induction is best analyzed and understood in terms of the dynamics of the morphogenetic processes involved. © 1992 Wiley‐Liss, Inc.
Planar induction of convergence and extension of the neural plate by the organizer of Xenopus
Authors: Keller Ray , Shih John , Sater Amy K. , Moreno Cecelia
Published: February 2005

This paper demonstrates that convergence and extension within the neural plate of Xenopus laevis are regulated by planar inductive interactions with the adjacent Spemann organizer. The companion article (Keller et al.: Developmental Dynamics 193:199–217, 1992) showed that the prospective hindbrain and spinal cord occupy a very short and very wide area just above the Spemann organizer in the early gastrula and that these regions converge and extend greatly during gastrulation and neurulation, using a sequence of radial and mediolateral cell intercalations. In this article, we show that “planar” contact of these regions with the organizer at their vegetal edge until stage 11 is sufficient to induce convergence and extension, after which their convergence and extension become autonomous. Grafts of the organizer in planar contact with uninduced ectodermal tissues induce these ectodermal tissues to converge and extend by a planar inductive signal from the organizer. Labeling of the inducing or responding tissues confirms that only planar interactions occur. Neural convergence and extension are actually hindered in explants deliberately constructed so that vertical interactions occur. These results show unambiguously that the Spemann organizer induces the extraordinary and precocious convergence and extension movements of the Xenopus neural plate by planar interactions acting over short distances.
Mesoderm movement and fate during avian gastrulation and neurulation
Authors: Schoenwolf Gary C. , Garcia‐Martinez Virginio , Dias Mark S.
Published: February 2005

Quail/chick transplantation chimeras were constructed during stages of gastrulation and neurulation to follow the subsequent movement and fate of cells of the primitive streak. All grafts were placed solely within the confines of the primitive streak to prevent confusion between cells that had not yet ingressed and those that had already ingressed, and transplanted cells were distinguished from host cells on the basis of a naturally occurring cell marker. Pathways of movement of ingressing cells corresponded to their level of residence within the primitive streak. Cells residing within Hensen's node (the cranial end of the primitive streak) initially migrated mainly cranially, remaining on or near the midline, and then extended caudally along the midline as regression of Hensen's node occurred. Cells residing within the nodus posterior (the caudal end of the primitive streak) migrated caudally. Cells residing at levels of the primitive streak between Hensen's node and the nodus posterior typically migrated bilaterally, confirming that such cells had not already ingressed prior to their transplantation (in which case, they would have migrated unilaterally). Subsets of these cells residing at progressively more caudal levels of the primitive streak migrated incrementally more laterally. Hensen's node contributed cells to the gut endoderm, head mesenchyme, notochord, and median hinge‐point (MHP) cells of the neural tube (future floor plate). At younger stages (i.e., stages 3a, 3b) Hensen's node contributed cells to principally the foregut endoderm and head mesenchyme, where‐as at older stages (i.e., stages 3c, 3d, 4), it contributed cells to principally the notochord and MHP region. The remaining segments of the cranial half of the primitive streak contributed cells to the various mesodermal subdivisions of the embryo, and the lengths of the segments forming these subdivisions were estimated. The most cranial level of the streak (directly behind Hensen's node) contributed cells to the most medial mesodermal subdivisions (head mesenchyme, somites) and consecutively more caudal levels of the streak contributed cells to sequentially more lateral mesodermal subdivisions (intermediate mesoderm, lateral plate mesoderm). The caudal half of the primitive streak contributed cells to the extraembryonic mesoderm, with the nodus posterior contributing cells to the most caudal extraembryonic mesoderm, including the blood islands. Our results confirm and extend the previous avian prospective fate maps, increasing our understanding of the movement and fate of cells of the gastrula and neurula stages. © 1992 Wiley‐Liss, Inc.
Expression of sarcomeric myosin in the presumptive myocardium of chicken embryos occurs within six hours of myocyte commitment
Authors: Han Y. , Dennis J. E. , Cohen‐Gould L. , Bader D. M. , Fischman D. A.
Published: February 2005

The distribution of sarcomeric myosin heavy chain (MyHC) has been examined immunoctochemically in the presumptive myocardial cells of chicken embryos (stages 6–10) prior to the onset of the heart beat. Embryos were stained with monoclonal antibody MF20, a reagent which recognizes all chicken sarcomeric MyHCs (Bader et al., 1982), and then examined both in whole mount by immunofluorescence and in semithin, plastic‐embedded sections following immunoperoxidase labeling. We observed that myosin could be detected as early as stage 7 (0–2 pairs of somites) in 29% of the 31 embryos examined, and by stage 8 (4 pairs of somites) more than 80% of the embryos were MF20+, Every embryo with 5 pairs of somites (stage 8+) labeled strongly with MF20. Labeling was first detected at stage 7 to 7+ as a diffuse fluorescent signal within pleomorphic cells of the splanchnic mesoderm located in two crescent‐shaped regions bordering each side of the anterior intestinal portal (AIP). With progressive development, the two crescent‐shaped regions merged at the apex of the AIP, and as the two heart tubes began fusion at stage 9, the MyHC+ regions extended cranially and medially. By somite stages 9–10, the myosin‐positive cells completely encircled the heart tube. From stages 7 to 9 the myosin signal had no sarcomeric distribution; i.e., there were no MyHC striations nor periodic repeats evident in the presumptive myocytes until late stage 9 and stage 10. Semithin sections revealed that myosin was first distributed in apical regions of the myocytes, adjacent to the pericardial coelom. The implications of these findings for myocyte determination, differentiation and morphogenesis are discussed.© 1992 Wiley‐Liss, Inc.
The influence of bone and marrow on cartilage hypertrophy and degradation during 30‐day serum‐free culture of the embryonic chick tibia
Authors: Cole Ada A. , Luchene Lawrence J. , Linsenmayer Thomas F. , Schmid Thomas M.
Published: February 2005

In this study, an organ culture system is defined which demonstrates complete loss of cartilage matrix from embryonic chick tibiae. Efficient loss of the cartilage matrix occurs within 30 days of serum‐free culture only when the intact tibiae containing bone, marrow, and cartilage tissue are cultured. During organ culture nonhypertrophic chondrocytes become hypertrophic and stain positively for type X collagen and alkaline phosphatase. The cartilage loses Safranin O staining, and finally all cartilage matrix disappears leaving the bony collar and marrow cells. If the tibial cartilage is separated from the bony collar and cultured alone in serum‐free medium, the nonhypertrophic chondrocytes also hypertrophy; the matrix loses Safranin O staining; however, some components of the matrix including type X collagen still remain after 30 days. In the presence of serum, the chondrocytes will hypertrophy but cartilage degradation is not evident. The results of this study support the conclusions that (1) hypertrophy is inherently programmed in the chondrocyte and (2) while Safranin O staining of cartilage cultured alone is diminished in serum‐free organ culture, the degradation of cartilage is complete only when bone and marrow are also present. © 1992 Wiley‐Liss, Inc.
Developmental expression of two antigens associated with mouse intestinal crypts
Authors: Beaulieu J.‐F. , Millane G. , Calvert R.
Published: February 2005

Two monoclonal antibodies were prepared against the duodenal mucosa of fourday‐old mice (MIM‐1/39 and MIM 1/130). The expression of the antigens was associated with the crypts of the small and large intestine in the fetus and adult. MIM‐1/39 was present in epithelial cells of the intervillous areas in the small intestine at 17 and 18 days of gestation; afterwards its expression was detected only in crypt cells from birth to adulthood. Transition from the mouth of the crypts to intestinal villi was abrupt. Expression of MIM‐1/39 was first detected at time of birth in the colon: In the adult, only crypt cells expressed the antigen and goblet cells were negative. Antigen MIM‐1/130 was detected from 16 to 18 days of gestation in the small intestine, in the mesenchymal matrix lying under the intervillous epithelium. After birth, it was present in the pericryptal mesenchymal matrix. This antigen was also expressed at birth in the colon and remained in the pericryptal matrix in the adult. In vivo, multiple injections of an organic extract of rat amniotic fluid to mothers, starting at 14 days of gestation, induced a profound modulation in the pattern of expression of both antigens at 17 days of gestation: The pattern of expression was comparable to that observed at least 5 days after birth in untreated animals. The expression of both antigens before crypt appearance may reflect some molecular differentiation in preparation for the formation of crypts, while their association with differentiated crypts may indicate that they have a role in the maintenance of crypt functional and/or morphological integrity. Finally, the fact that their expression can be modulated experimentally may prove to be a breakthrough for the study of crypt formation. © 1992 Wiley‐Liss, Inc.
Expression of α‐cardiac and α‐skeletal actin mRNAs in relation to innervation in regenerating and non‐regenerating rat skeletal muscles
Authors: Toyofuku Toshihiko , Hoffman John R. , Zak Radovan , Carlson Bruce M.
Published: February 2005

The expression of α‐cardiac and α‐skeletal actin mRNA in regenerating muscle was examined. Changes in mRNA levels were analyzed in autografted extensor digitorum longus (EDL) muscles in rats using α‐isoform specific synthetic oligonucleotides and β‐actin cDNA as probes. After autografting, the expression of α‐cardiac actin mRNA was induced; concomitantly that of α‐skeletal actin mRNA was reduced. The pattern of α‐actin mRNA expression appeared to be similar to that seen in embryonic skeletal muscle. In order to evaluate the effects of innervation on α‐actin mRNA expression in regenerating muscle, nerveless, standard, and nerve‐intact autografted muscles were examined. More complete innervation facilitated the recovery of α‐skeletal actin mRNA to control levels, but had little effect on the amount of α‐cardiac actin mRNA. We found that regenerating muscle shows the embryonic pattern of α‐actin mRNAs in the early stage and concluded that the recovery of α‐skeletal actin mRNA expression to the adult pattern is influenced by innervation, while α‐cardiac actin mRNA expression is nerve independent. © 1992 Wiley‐Liss, Inc.
Colocalization of thrombospondin and syndecan during murine development
Authors: Corless Christopher L. , Mendoza Arturo , Collins Tucker , Lawler Jack
Published: February 2005

Thrombospondin is an adhesive glycoprotein that is thought to play a role in tissue genesis and repair. We have used a monoclonal anti‐thrombospondin antibody, designated 5G11, to localize thrombospondin in paraformaldehyde fixed, paraffin‐embedded sections of developing mouse embryos. Thrombospondin expression is observed in uterine smooth muscle, endometrial glands, the decidua, and trophoblastic giant cells during the initial phase of post‐implantation development in the embryo. Cardiac myocytes and neuroepithelial cells show positive staining for thrombospondin at day 8.5 of gestation, and this expression continues throughout the development of the myocardium and central nervous system. Strong staining for thrombospondin is seen in developing bone and in the liver. Thrombospondin is also observed in developing smooth muscle and skeletal muscle, as well as in a variety of epithelia, including the epidermis, small intestinal epithelium, lens epithelium, renal tubular epithelium, and the epithelium of the developing tooth. Comparison of thrombospondin staining with that of two known cell surface receptors for thrombospondin, syndecan and the vitronectin receptor, reveals remarkable colocalization of thrombospondin and syndecan in all tissues, but almost no coexpression with the vitronectin receptor. Coexpression of thrombospondin and syndecan may play an important role in cell‐cell or cell‐matrix interactions during development. © 1992 Wiley‐Liss, Inc.
Expression patterns of engrailed‐like proteins in the chick embryo
Authors: Gardner Charles A. , Barald Kate F.
Published: February 2005

The protein products of both of the identified chick engrailed‐like (En) genes, chick En‐1 and chick En‐2, are localized in cells of the developing brain, mandibular arch, spinal cord, dermatome, and ventral limb bud ectoderm, as demonstrated by labeling with the polyclonal antiserum αEnhb‐1 developed by Davis et al. (Development 111:281–298, 1991). A subpopulation of cephalic neural crest cells is also En‐protein‐positive. The monoclonal antibody 4D9 recognizes the chick En‐2 gene product exclusively (Patel et al.: Cell 58:955–968, 1989; Davis et al., 1991) and colocalizes with chick En‐2 mRNA in the developing head region of the chick embryo as shown by in situ hybridization (Gardner et al.: J. Neurosci. Res. 21:426–437, 1988). In the present study we examine the pattern of αEnhb‐1 and 4D9 localization throughout the chick embryo from the first appearance of antibody (Ab)‐positive cells at stage 8 (Hamburger and Hamilton: J. Morphol. 88:49–92, 1951) through stage 28 (1–5.5 days). We compare the localization patterns of the two Abs to each other, as well as to the localization of the monoclonal Ab, HNK‐1, which recognizes many neural crest cells, using double‐and triple‐label fluorescence immunohistochemistry. Most En protein‐positive cells in the path of neural crest cell migration are not HNK‐1 positive. In detailed examination of αEnhb‐1 and 4D9 localization, we find previously undetected patterns of En protein localization in the prechordal plate, hindbrain, myotome, ventral body‐wall mesoderm, and extraembryonic membranes. Based upon these observations we propose: (1) that En expression in the mesoderm may be induced through interaction with En expressing cells in the neuroectoderm; (2) that En expression in the head mesenchyme is associated with somitomere 4; and (3) that En expression may be involved in epithelial‐mesenchymal cell transformations. © 1992 Wiley‐Liss, Inc.
Changes in dorsoventral but not rostrocaudal regionalization of the chick neural tube in the absence of cranial notochord, as revealed by expression of Engrailed‐2
Authors: Darnell Diana K. , Schoenwolf Gary C. , Ordahl Charles P.
Published: February 2005

Notochord has been implicated in previous studies in both the dorsoventral and rostrocaudal patterning of the developing neural tube. This possibility has been further explored by analyzing the expression of Engrailed‐2 in chick embryos developing with cranial notochord defects. Control embryos containing intact notochords expressed Engrailed‐2 protein within the neural tube and in a subset of the neural crest and overlying surface ectoderm at the future mesencephalon and cranial metencephalon levels. Within the neural tube, expression was confined to cell nuclei in the roof plate and lateral walls; floor plate nuclei directly overlying the notochord typically failed to show expression. After surgical removal of Hensen's node, the source of notochord precursor cells, embryos were cultured through neurulation and assayed for expression of Engrailed‐2 protein. All embryos that partially or completely lacked cranial notochord expressed Engrailed‐2 in a pattern similar to that of control embryos containing intact notochords, except that when notochord and floor plate were absent, Engrailed‐2 was also expressed in the most ventral part of the neural tube. These results indicate that (1) Engrailed‐2 expression is suppressed in the most ventral neural tube owing to induction of the floor plate by the notochord, and (2) that the presence of an underlying notochord is not required for correct rostrocaudal expression, suggesting that multiple pathways act in the patterning of the rudiment of the central nervous system. © 1992 Wiley‐Liss, Inc.
Expression of cell adhesion molecules during initiation and cessation of neural crest cell migration
Authors: Akitaya Tatsuo , Bronner‐Fraser Marianne
Published: February 2005

Because of their distribution and known ability to promote neuronal adhesion, it has been proposed that N‐CAM and N‐cadherin are involved in the formation of the nervous system. Here, we examine the expression of these molecules during the initiation and cessation of trunk neural crest cell migration during the formation of the peripheral nervous system. Whereas other neural tube cells express N‐cadherin, the dorsal neural tube containing neural crest precursors has little or no N‐cadherin immunoreactivity. In contrast, N‐CAM is expressed in the dorsal neural tube and on early migrating neural crest cells, from which it gradually disappears during migration. Both N‐CAM and N‐cadherin are absent from neural crest cells at advanced stages of migration. As neural crest cells cease migration and condense to form dorsal root and sympathetic ganglia, N‐cadherin but not N‐CAM is observed on the forming ganglia, identified by neurofilament expression and the aggregation of HNK‐1 reactive cells. The results demonstrate that the absence of N‐cadherin correlates with the onset of neural crest migration and its reappearance correlates with cessation of migration and precedes ganglio‐genesis. © 1992 Wiley‐Liss, Inc.
Coincident loss of mitochondria and nuclei during lens fiber cell differentiation
Authors: Bassnett Steven , Beebe David C.
Published: February 2005

During normal differentiation, lens fiber cells lose their nuclei, mitochondria, and other membrane‐bound organelles. In the present study, a slice preparation of the embryonic chicken lens was used with laser scanning confocal microscopy to study the spatial and temporal patterns of organelle breakdown during embryonic development. At all stages examined, mitochondria in lens epithelial cells were present in perinuclear clusters. In contrast, early in development, lens fiber cells contained extremely elongated mitochondria (>100 μm) that were distributed throughout the cytoplasm and oriented along the long axis of the cells. By the 8th day of embryonic development (E8), the mitochondria in the central fiber cells began to fragment. At the same time, the nuclei in these cells became smaller and more spherical. By E10, mitochondrial staining in the central fibers became punctate. Electron microscopy of this region revealed swollen mitochondria with disrupted cristae. By E12, cells in the central region of the lens lacked mitochondria and nuclei. The loss of nuclei and mitochondria from a given cell was coincident and abrupt (2–4 hr), occurring in a previously unsuspected domain situated about 300 μm from the anterior surface of the lens. A cytoskeletal component, actin, persisted in the central cells indicating that organelle degradation represents a selective process and not simply the global degradation of supramolecular structures. Throughout embryonic development, the organelle‐free region grew at approximately the same rate as the lens and, by the time of hatching, had expanded to match the diameter of the pupil. The present results suggest that the embryonic lens will be a useful model system with which to study mechanisms of organelle degradation. © 1992 Wiley‐Liss, Inc.
Expression of NCAM isoforms during skeletal myogenesis in the mouse embryo
Authors: Lyons Gary E. , Moore Robert , Yahara Osamu , Buckingham Margaret E. , Walsh Frank S.
Published: February 2005

We have examined the developmental patterns of neural cell adhesion molecule (NCAM) gene expression in embryonic mouse skeletal muscle cells by in situ hybridization. Moreover, by utilising exon‐specific cRNA probes, we have examined tissue specific splicing of the NCAM gene. We show that there is a distinct sequence of NCAM isoform expression during skeletal muscle development. Since NCAMs are also expressed in other cell types, particularly neurons, NCAM mRNAs have been colocalised with acetylcholine receptor α (AChRα) gene transcripts to identify muscle‐specific expression. NCAM is first detected in somites as they first form, prior to their differentiation into muscle and nonmuscle compartments. Myotomes, the first skeletal muscle masses to form in the embryo, express mRNAs for the transmembrane 180 and 140 kDa isoforms of NCAM. Both of these transcripts are also detected in the neural tube, and their spatial pattern of expression changes with development. Transcripts containing the muscle‐specific domain (MSD) of the NCAM gene are not detected prior to 11 days postcoitum (p.c.), at a time when rostral somites already contain well‐developed myotomes. As the level of MSD mRNAs increases at 12 days p.c., the 140 and 180 kDa transcript levels decrease in skeletal muscle masses. The level of all NCAM isoform transcripts declines between 13 and 15 days p.c. in muscle. However, the 180 and 140 kDa NCAM isoforms are expressed at a high level in neural tissue and in other locations in the developing embryo such as in smooth muscle, around vibrissae follicles, and in the perichondrial zone of digits. © 1992 Wiley‐Liss, Inc.
Cardiac expression of polysialylated NCAM in the chicken embryo: Correlation with the ventricular conduction system
Authors: Watanabe Michiko , Timm Maureen , Fallah‐Najmabadi Hessam
Published: February 2005

The neural cell adhesion molecule (NCAM) and its polysialic acid moeity (PSA) affect cellular interactions during the development of the nervous system and skeletal muscle. NCAM has also been identified in the embryonic heart of various species including humans. However, knowledge regarding the role of NCAM and its function‐modulating PSA in cardiogenesis is limited. The distribution of NCAM and its PSA in the ventricular myocardium of chicken embryos was determined by indirect immunofluorescence staining. The NCAM polypeptide was found throughout the cardiac myocardium. In contrast PSA was located in discrete regions in stage 20 to 44 embryos (during and after septation). Myocardium at the subendocardial regions of the atrioventricular canal and ventricular trabeculae were PSA positive by stage 20. At later stages, transverse sections of the postseptation heart just below the level of the atrioventricular interface revealed a PSA‐positive bundle of myocardium in the septum. This bundle was continuous with two branches at a more apicallevel which in turn were continuous with the PSA‐positive subendocardial myocardium lining the left and right ventricles. This pattern of PSA in the myocardium was similar to that of the ventricular conduction system configuration defined in the adult heart. Electron micrographs of the subendocardium of the ventricular septum revealed PSA positivity on myofibril‐containing cells with the ultrastructural location of Purkinje fibers. At later stages (35–44) a subset of cells within PSA‐positive regions was stained by an antibody against an isoform of the myosin heavy chain found in adult Purkinje fibers. These cells and surrounding tissue lacked PSA in the adult heart. Thus polysialylated NCAM may be modulating cell–cell interactions during the development of the ventricular conduction system. © 1992 Wiley‐Liss, Inc.
Notochord of chick embryos secretes short‐form type IX collagen prior to the onset of vertebral chondrogenesis
Authors: Hayashi Masando , Hayashi Kimiko , Iyama Ken‐Ichi , Trelstad Robert L. , Linsenmayer Thomas F. , Mayne Richard
Published: February 2005

The notochord of embryonic chicks produces type IX collagen, as well as type II collagen, prior to the onset of vertebral chondrogenesis. To address the question of whether the notochord secretes the “long‐form” type IX collagen found in cartilage or the “short‐form” type IX found in the cornea and vitreous humor, we examined immunoreactivity of the notochordal type IX collagen using two different monoclonal antibodies. The antibody 2C2 recognizes an epitope close to the carboxyl‐terminus of the HMW fragment, which is present in both the long‐ and short‐form type IX collagens, whereas another antibody 4D6 recognizes an epitope in the NC4 domain of the long‐form type IX collagen, which is absent in the short‐form type IX collagen. Therefore, the long‐form is recognized by its reaction with both 2C2 and 4D6, while the short‐form by its reaction with only 2C2 and no reaction with 4D6. Immunostaining of vertebral sections with 2C2 shows an identical distribution of staining with that for type II collagen, although the staining with 2C2 is less intense. The 2C2‐reactive type IX collagen is found within the notochord at stage 14 and in the notochordal sheath at stage 20. Deposition of this collagen in the perinotochordal matrix increases with time and reaches a level comparable with that for type II at stage 31. In contrast, the 4D6‐reactive type IX collagen is not found within the notochord nor in the notochordal sheath. The collagen becomes detectable, however, in the perinotochordal matrix at stages 27 to 28 and is markedly increased at stage 29, reaching the levels for the 2C2‐reactive type IX at stage 31. A highly sensitive immuno‐dot blot assay has confirmed that the notochord from chick embryos at stages 19 to 20 produces type IX collagen that reacts with 2C2 but does not react with 4D6. The present study indicates (1) that the short‐form type IX collagen is secreted from the notochord and also from the sclerotome cells, and (2) that the switching in the production from the short‐form type IX collagen to the long‐form type IX occurs during stage 27 and stage 31 when sclerotome cells differentiate into chondrocytes in the developing vertebral body. © 1992 Wiley‐Liss, Inc.
The changing architecture of the neonatal rat ovary during histogenesis
Authors: Rajah Roopa , Glaser Edmund M. , Hirshfield Anne N.
Published: February 2005

The purpose of this study was to describe the changing histological organization of the rat ovary during postpartum days one through three (p1‐p3). A PC‐based image‐combining microscope system was used to reconstruct the ovary in three dimensions. On p1, cylindrical pocket‐like structures radiated from the core of the ovary that were open toward the surface epithelium. The walls of the pockets contained connective tissue cells and capillaries (stroma). By p2, these pockets had completely closed; each pocket enclosed a small nest of oocytes and a few presumptive granulosa cells. By p3, the pocket‐like organization had disappeared. On p1, only one or two primordial follicle‐like structures were observed in the core and toward the periphery of the ovary; most oocytes were not enclosed in follicles. By p3, very few naked oocytes remained; primordial follicles predominated in all the regions of the ovary and some of the follicles had multiple layers of granulosa cells. There were changes in location, area, and volume of the rete tubules during these postnatal days. The extraovarian rete was visible on all 3 days but changed its orientation relative to the ovary. The connecting rete was found beneath the epithelial layer of the ovary on all 3 days and showed dramatic increase in area on p2. The wide lumen of the intraovarian rete was in direct contact with some of the oocytes near by on all 3 days, but these “communication points” were most abundant on p2. Based on our observations of different cell–cell associations during this time period, we hypothesize (1) that the mesenchymal–presumptive granulosa cell association is essential for the completion of folliculogenesis, and (2) the rete ovarii may have an inductive role in follicle assembly. These observations suggest that the first 3 days postpartum are critically important for studying the heterogeneous cell interactions that lead to the assembly of primordial follicles. The regional differences in tissue organization during this formative period may have significant implications on later aspects of follicular development. © 1992 Wiley‐Liss, Inc.
Insertional mutation of a gene involved in growth regulation of the early mouse embryo
Authors: Iannaccone P. M. , Zhou X. , Khokha M. , Boucher D. , Kuehn M. R.
Published: February 2005

A transgenic mouse strain derived from embryonic stem (ES) cells infected with multiple copies of a retroviral vector carries a recessive insertional mutation resulting in prenatal lethality. A detailed histological analysis of developing embryos has shown that the mutation results in hyperplasia of both embryonic and extraembryonic ectoderm and failure of mesoderm formation in the egg cylinder stage embryo. The number of cells in each lineage of normal and mutant embryos was estimated using stereological analysis of serial sections taken from implantation sites. We observed a 2‐fold increase in the number of embryonic ectoderm cells in mutant embryos at 7.5 days postcoitum (dpc). In addition, we found that mutant embryonic ectoderm cells are only 0.6 times as large as normal cells. The number of extraembryonic ectoderm cells in mutant embryos at 7.5 dpc is also increased, by almost 4‐fold. Mutant extraembryonic ectoderm cells are also smaller than normal, being only two‐thirds the size of wild‐type cells. The mutant phenotype suggests that the gene identified by this insertional mutation plays an important role in the growth control of early embryonic lineages. © 1992 Wiley‐Liss, Inc.
Development of olfactory nerve glia defined by a monoclonal antibody specific for schwann cells
Authors: Norgren Robert B. , Ratner Nancy , Brackenbury Robert
Published: February 2005

Although there is considerable interest in the possible role of olfactory glia in the pathfinding abilities of olfactory nerve axons, the complete development of these glia in vivo has not been described. Using a specific Schwann cell marker, the 1E8 antibody, we have found that olfactory nerve glia can be identified throughout development. These glia appear to originate in the olfactory placode and migrate initially into the periphery of the olfactory nerve, and later into the center of the nerve. Olfactory nerve glia enter the presumptive olfactory bulb with the olfactory receptor neuron axons and distribute themselves along the edge of the olfactory nerve layer. The fact that olfactory nerve glia are specifically immunostained by the 1E8 monoclonal antibody, which recognizes the Schwann cell‐specific protein P0, suggests that these cells more closely resemble Schwann cells than astrocytes or enteric glia. These results support and extend previous findings suggesting that olfactory nerve glia have distinctive developmental and anatomical features which may be important to the regenerative capacity of the olfactory system. © 1992 Wiley‐Liss, Inc.
Puzzle of crystallin diversity in eye lenses
Authors: Piatigorsky Joram
Published: February 2005

Crystallins have evolved by various mechanisms that are associated with high expression of their genes in the eye lens. The diversity and pattern of crystallins among different species indicate that independent events have occurred at the molecular level during the evolution of the lens in different invertebrates (jellyfish, squid, and octopus) and vertebrates. Although it is possible that different crystallins are needed to fulfill the specific needs of individual species, the unexpectedly large array of proteins that function as crystallins and their abundance in the lens raise the possibility that selective pressures optimizing the function of certain transcription factors in the lens contribute to the recruitment of crystallins. © 1993 wiley‐Liss, Inc.
Molecular control of glutamine synthetase expression in the developing retina tissue
Authors: Vardimon Lily , Ben‐dror Iris , Havazelet Nadav , Fox Lyle E.
Published: February 2005

Glutamine synthetase is a differentiation marker of the neural retina, whose expression is restricted to Müller glia cells, is inducible by glucocorticoids and is dependent on tissue development. The retina tissue acquires the competence to express GS in response to glucocorticoids with development, although the level of hormone binding activity in the cells does not alter with age. Using CAT constructs that are controlled by “simple GRE” promoters we demonstrated that glucocorticoid receptor transcription activity in retina cells increases with development. The increase in receptor activity correlates directly with the increase in inducibility of the glutamine synthetase gene and inversely with the rate of retina cell proliferation. At early developmental ages, when retina cells are still proliferating, the glucocorticoid receptor is transcriptionally inactive and glutamine synthetase expression cannot be induced. Receptor activity increases progressively with development and by day 12, when cell proliferation ceases, competence for glutamine synthetase induction is high. This competence for glutamine synthetase induction can be repressed by overexpressing the oncogene v‐src, which stimulates retina cell proliferation. We discuss possible mechanisms for developmental‐dependent modulation of glucocorticoid receptor transcriptional activity. © 1993 wiley‐Liss, Inc.
Dynamics of actin and assembly of connectin (titin) during myofibrillogenesis in embryonic chick cardiac muscle cells in vitro
Authors: Komiyama Masatoshi , Kouchi Katsunori , Maruyama Koscak , Shimada Yutaka
Published: February 2005

Immunogold electron microscopy of cardiac myocytes microinjected with biotin‐labeled actin showed that gold labeling was first found around the A band level of myofibrils at their proximal parts. This observation suggests that polymerization of actin and/or the addition of newly formed actin filaments occurs preferentially in association with myosin filaments to increase the myofibrillar girth. At the distal portions of developing myofibrils, their terminal ends were initially labeled, suggesting that continued reorganization and/or de novo formation of myofibrils occurs at these locations. Soon, gold particles were seen along the termini of growing myofibrils. This appears to indicate that actin subunits are added at the membrane‐associated ends of preexisting actin filaments to increase the length of myofibrils. Adhesion plaque proteins, e.g., vinculin, do not appear to play any role in assembling actin monomers at these sites on the inner surface of the sarcolemma.

Immunofluorescence and immunoelectron microscopy of cardiomyocytes double‐stained with antibodies against two distant domains of connecttin (titin) filaments and other sarcomeric proteins showed that these domains of connectin filaments and myosin were synthesized almost simultaneously on large polyribosomes and/or associated immediately after the synthesis of these molecules. Connectin and myosin bands were formed after α‐actinin striations (Z bands) were seen on preformed I‐Z‐I‐like structures. The observation that the development and distribution of connectin were tightly linked with those of myosin suggests the possible role of connectin for integrating myosin filaments with the early formed I‐Z‐I complexes of myofibrils. © 1993 wiley‐Liss, Inc.
Distribution of FGF‐2 suggests it has a role in chick limb bud growth
Authors: Savage Mary Pfann , Fallon John F. , Hart Charles E. , Riley Bruce B. , Sasse Joachim , Olwin Bradley B.
Published: February 2005

We developed and characterized antibodies specific for FGF‐2 and used them to locate FGF‐2 during chick embryo development. A series of micrographs demonstrated the progression of FGF‐2 staining during development of the different tissues and organs. FGF‐2 was present in the ectoderm covering the entire embryo, muscle cells, nervous system, neural crest cells, and mesonephros. FGF‐2 was also present in the limb from initiation of budding through differentiation. The limb ectoderm and subjacent mesoderm showed the strongest immunostaining, with lower levels in the center of the bud. However, the distribution of FGF‐2 positive cells in the mesoderm was not homogeneous. This heterogeneity was not due to cell cycle specific distribution of FGF‐2 protein, as flow cytometric analysis showed that FGF‐2‐positive cells were distributed throughout the cell cycle. However, the amount of anti‐FGF‐2 fluorescence varied most during G1, consistent with the possibility that FGF‐2 is low after M phase and increases during G1. A bioassay was used to demonstrate FGF‐2 levels in the wing ectoderm were approximately 2.7‐fold greater than in the mesoderm. We propose that the location of FGF‐2 in the embryo is consistent with a role in epithelial‐mesenchymal interactions; in the limb bud it may prevent differentiation and permit limb outgrowth and subsequent expression of patterning events. © 1993 Wiley‐Liss, Inc.
Spatiotemporal expression of pregnancy‐specific glycoprotein gene rnCGM1 in rat placenta
Authors: Rebstock Sabine , Lucas Kurt , Weiss Martina , Thompson John , Zimmermann Wolfgang
Published: February 2005

As a basis towards a better understanding of the role of the pregnancy‐specific glycoprotein (PSG) family in the maintenance of pregnancy, detailed investigations are described on the expression of a recently identified rat PSG gene (rnCGM1) at the mRNA and protein levels. Using specific oligonucleotide primers, rnCGM1 transcripts were identified after reverse transcription, polymerase chain reaction, and hybridization with a radiolabelled, internal oligonucleotide. Transcripts were only found in significant amounts in placenta. In situ hybridization visualized rnCGM1 transcripts at day 14 post coitum (p.c.), in secondary trophoblast giant cells and in the spongiotrophoblast. Only those secondary giant cells lining the maternal decidua were positive. In contrast, primary giant cells did not contain rnCGM1 mRNA. At day 18 p.c., rnCGM1. transcripts were almost exclusively detectable in the spongiotrophoblast. No rnCGM1 transcripts were found in rat embryos of these two developmental stages. Rabbit antisera were generated against the amino‐terminal immunoglobulin variable‐like domain and against a synthetic peptide containing the last 13 carboxy‐terminal amino acids of rnCGM1. Bothe antisera recognized a 124 kDa protein in day 18 rat placental extracts as identified by Western blot analysis. The anti‐peptide antiserum recognized a 116 kDa protein in the serum of a 14 day p.c. pregnant rat that is absent from the sera of non‐pregnant females. Taken together, these results confirm exclusive expression of rnCGM1 in the rat trophoblast, but unlike human PSG, negligible or no expression is found in other organs, such as fetal liver or salivary glands, indicating a more specialized function of rnCGM1. Its spatiotemporal expression pattern is conducive with a potential role of PSG in protecting the fetus against the maternal immune system and/or in regulating the invasive growth of trophoblast cells. © 1993 Wiley‐Liss, Inc.
Proliferation and migration of primordial germ cells in We/We mouse embryos
Authors: Buehr Mia , McLaren Anne , Bartley Aine , Darling Susan
Published: February 2005

We have examined the numbers and distribution of primordial germ cells in We/We, We/+, and +/+ mouse embryos using Southern blotting to determine embryo genotypes. At early somite stages (5–7 somites: approximately 81/2 days post coitum [dpc]) there are 50 to 100 germ cells in embryos of all genotypes. The number of germ cells in We/+ and +/+ embryos then begins to increase: at later somite stages (17–19 somites: approximately 91/2 dpc) they number about 200, and by 101/2 dpc there are approximately 725We/+ and 850 +/+ germ cells. During this time, however, the number of germ cells in We/We embryos remains less than 100. At 81/2 dec, the distribution of germ cells in the hindgut endoderm is the same in all genotypes. By 91/2 dpc, 30% of We/We germ cells are found in ectopic sites (allantois and vitelline artery); germ cell distribution along the length of the hindgut appears normal, but germ cells remain confined to the floor of the gut in We/We embryos, rather than being distributed around its circumference as in the other two genotypes. By 101/2 days, the migration of We/We germ cells through the dorsal mesentery lags behind that of the other genotypes, and a larger proportion remains in the gut wall. © 1993 Wiley‐Liss, Inc.