The Ectomesenchymal-endodermal Interaction System (EEIS) of Triturus alpestris in Tissue Culture

Differentiation of visceral cartilage was studied in the Ectomesenchymal-endodermal Interaction System (EEIS), a hanging-drop culture system containing a piece of head neural fold tissue from behind the prospective ear region of a Triturus alpestris neurula together with a piece of ventrolateral pharynx endoderm from the same embryo. Cartilage cells differentiated only from those neural crest cells which have been in contact with the pharynx endoderm. Cartilage differentiation occurred neither in cultures containing neural fold or pharynx endoderm alone nor in distance cultures in which both explants were separated from each other at a greater distance than usual. However, in some critical cultures of the EEIS containing pharynx endoderm, excised more ventrally from the regular site, accumulations of ectomesenchymal cells were observed near the endoderm which were not transformed into cartilage.
The differentiation of cartilage was classified into three stages: (1) prechondroblasts (day 1 to 5), (2) chondroblasts (about day 6) and (3) chondrocytes (about day 10).
The data obtained suggested that cellular contact between the two inductively correlated tissues was a prerequisite for the transmission mechanism while mitotic cell divisions were mainly responsible for the spread of the inductive message within the responding ectomesenchymal cells.     

Partial restriction in the developmental potential of late emigrating avian neural crest cells.

Trunk neural crest cells migrate along two major pathways: a ventral pathway through the somites whose cells form neuronal derivatives and dorsolateral pathway underneath the ectoderm whose cells become pigmented. In avian embryos, the latest emigrating neural crest cells move only along the dorsolateral pathway. To test whether late emigrating neural crest cells are more restricted in developmental potential than early migrating cells, cultures were prepared from the neural tubes of embryos at various stages of neural crest cell migration. "Early" and "middle" aged neural crest cells differentiated into many derivatives including pigmented cells, neurofilament-immunoreactive cells, and adrenergic cells. In contrast, "late" neural crest cells differentiated into pigment cells and neurofilament-immunoreactive cells, but not into adrenergic cells even after 10-14 days. To further challenge the developmental potential of early and late emigrating neural crest cells, they were transplanted into embryos during the early phases of neural crest cell migration, known to be permissive for adrenergic neuronal differentiation. The cells were labeled with the vital dye, DiI, and injected onto the ventral pathway at stages 14-17. Two and three days after injection, some early neural crest cells were found to express catecholamines, suggesting they were adrenergic neuroblasts. In contrast, DiI-labeled late neural crest cells never became catecholamine-positive. These results suggest that the late emigrating neural crest cell population has a more restricted developmental potential than the early migrating neural crest cell population.     

Fibronectin in early avian embryos: Synthesis and distribution along the migration pathways of neural crest cells

Summary Immunoperoxidase labelling for fibronectin (FN) in chick embryos showed FN-positive basement membranes surrounding the neural crest cell population prior to crest-cell migration. At cranial levels, crest cells migrated laterally into a large cell-free space. Initially they moved as a tongue of cells contacting the FN-positive basement membrane of the ectoderm, but later the crest cell population expanded into space further from the ectoderm, until eventually the entire cranial cell-free space was occupied by mesenchyme cells. This was accompanied by the appearance of FN among the crest cells. At trunk levels, crest cells entered a relatively small space already containing FN-positive extracellular material. At later stages the migration of trunk crest cells broadly matched the distribution of FN. In vitro, chick and quail embryo ectoderm, endoderm, somites, notochord and neural tube synthesized and organized fibrous FN-matrices, as shown by immunofluorescence. Ectoderm and endoderm deposited this matrix only on the substrate face. The FN content of endoderm and neural tube matrices was transient, the immunofluorescence intensity declining after 1–2 days in culture. Some crest cells of cranial and sacral axial levels synthesized FN. Our data suggests that these were the earliest crest cells to migrate from these levels. This ability may be the first expression of mesenchymal differentiation in these crest cells, and in vivo enable them to occupy a large space. Almost all crest cells from cervico-lumbar axial levels were unable to synthesize FN. In vivo, this inability may magnify the response of these crest cells to FN provided by the neighbouring embryonic tissues.     

FGF8 signaling is chemotactic for cardiac neural crest cells

Cardiac neural crest cells migrate into the pharyngeal arches where they support development of the pharyngeal arch arteries. The pharyngeal endoderm and ectoderm both express high levels of FGF8. We hypothesized that FGF8 is chemotactic for cardiac crest cells. To begin testing this hypothesis, cardiac crest was explanted for migration assays under various conditions. Cardiac neural crest cells migrated more in response to FGF8. Single cell tracing indicated that this was not due to proliferation and subsequent transwell assays showed that the cells migrate toward an FGF8 source. The migratory response was mediated by FGF receptors (FGFR) 1 and 3 and MAPK/ERK intracellular signaling. To test whether FGF8 is chemokinetic and/or chemotactic in vivo, dominant negative FGFR1 was electroporated into the premigratory cardiac neural crest. Cells expressing the dominant negative receptor migrated slower than normal cardiac neural crest cells and were prone to remain in the vicinity of the neural tube and die. Treating with the FGFR1 inhibitor, SU5402 or an FGFR3 function-blocking antibody also slowed neural crest migration. FGF8 over-signaling enhanced neural crest migration. Neural crest cells migrated to an FGF8-soaked bead placed dorsal to the pharynx. Finally, an FGF8 producing plasmid was electroporated into an ectopic site in the ventral pharyngeal endoderm. The FGF8 producing cells attracted a thick layer of mesenchymal cells. DiI labeling of the neural crest as well as quail-to-chick neural crest chimeras showed that neural crest cells migrated to and around the ectopic site of FGF8 expression. These results showing that FGF8 is chemotactic and chemokinetic for cardiac neural crest adds another dimension to understanding the relationship of FGF8 and cardiac neural crest in cardiovascular defects.     

Interactions between Hox-negative cephalic neural crest cells and the foregut endoderm in patterning the facial skeleton in the vertebrate head

The vertebrate face contains bones that differentiate from mesenchymal cells of neural crest origin, which colonize the median nasofrontal bud and the first branchial arches. The patterning of individual facial bones and their relative positions occurs through mechanisms that remained elusive. During the early stages of head morphogenesis, an endodermal cul-de-sac, destined to become Sessel's pouch, underlies the nasofrontal bud. Reiterative outpocketings of the foregut then form the branchial pouches. We have tested the capacity of endoderm of the avian neurula to specify the facial skeleton by performing ablations or grafts of defined endodermal regions. Neural crest cells that do not express Hox genes respond to patterning cues produced regionally in the anterior endoderm to yield distinct skeletal components of the upper face and jaws. However, Hox-expressing neural crest cells do not respond to these cues. Bone orientation is likewise dependent on the position of the endoderm relative to the embryonic axes. Our findings thus indicate that the endoderm instructs neural crest cells as to the size, shape and position of all the facial skeletal elements, whether they are cartilage or membrane bones.     

Morphology and behaviour of neural crest cells of chick embryo in vitro

Summary Neural primordia of chick embryos were cultured for three days and the behaviour of migrating neural crest cells studied. Somite cells were used as a comparison. Crest cells were actively multipolar with narrow projections which extended and retracted rapidly, contrasting to the gradual extension of somite-cell lamellae. On losing cell contact, somite cells were also more directionally persistent. The rate of displacement of isolated crest cells was particularly low when calculated over a long time base. Both crest and somite cells were monolayered; contact paralysis occurred in somite cell collisions but was not ascertained for crest cells. However, crest cells in a population were far more directionally persistent than isolated cells. Contact duration between crest cells increased with time and they formed an open network. Eventually, retraction clumping occurred, initially and chiefly at the periphery of the crest outgrowth. Crest cells did not invade cultured embryonic mesenchymal or epithelial populations but endoderm underlapped them. No effects were observed on crest cells prior to direct contact. Substrate previously occupied by endoderm or ectoderm caused crest cells to flatten while substrate previously occupied by the neural tube caused them to round up and clump prematurely.     

Formation and distribution of neural crest mesenchyme to the first pharyngeal arch region of the mouse embryo

Murine neural crest mesenchyme begins its escape from columnar epithelium near the tips of the midbrain-rostral hindbrain neural folds at 4+ to 5 somites of age. At that time the tip of each fold is located dorsolateral to the pharynx. Once crest formation is complete at this earliest site, it leaves behind both crest mesenchyme and overlying squamous epithelium. Crest formation then progresses medially, into the lateral margin of the neural plate. At the same time, this lateral margin elevates as the tip of the neural fold. By the time crest formation ceases at approximately 10 somites, the result of these simultaneous activities is to passively distribute the earliest mesenchyme, formed from the lateralmost epithelium, dorsolateral to the pharynx and the later, more medially derived mesenchyme lateral to the neural tube. Once formed, the crest mesenchyme dorsolateral to the pharynx is displaced ventromedially in a narrow, transient subectodermal space functionally similar to that observed in the chick embryo. Displacement might result from cell motility or the formation of matrix-filled spaces between cells of the mesenchyme. Displaced cells are closely associated with the overlying columnar epithelium. This association precedes their subsequent induction and may reflect preliminary patterning. The crest mesenchyme passively distributed lateral to the neural tube is subsequently displaced medially. Here the formation of enlarged (matrix-filled?) spaces is clearly involved in the initial displacement. Displaced cells proliferate to form the anlage of the trigeminal ganglion. The other major contributor to this ganglion is the trigeminal placode. The placodal epithelium is located dorsolateral to the pharynx of the 12-somite embryo. If the epithelia of the head maintain their relative positions, this placode is derived from the squamous epithelium formed together with the earliest crest mesenchyme. If not, an alternative source is the columnar epithelium located ventromedial to the tip of the 4+- to 5-somite neural fold.     

A vital dye analysis of the timing and pathways of avian trunk neural crest cell migration

To permit a more detailed analysis of neural crest cell migratory pathways in the chick embryo, neural crest cells were labelled with a nondeleterious membrane intercalating vital dye, DiI. All neural tube cells with endfeet in contact with the lumen, including premigratory neural crest cells, were labelled by pressure injecting a solution of DiI into the lumen of the neural tube. When assayed one to three days later, migrating neural crest cells, motor axons, and ventral root cells were the only cells types external to the neural tube labelled with DiI. During the neural crest cell migratory phase, distinctly labelled cells were found along: (1) a dorsolateral pathway, under the epidermis, as well adjacent to and intercalating through the dermamyotome; and (2) a ventral pathway, through the rostral portion of each sclerotome and around the dorsal aorta as described previously. In contrast to those cells migrating through the sclerotome, labelled cells on the dorsolateral pathway were not segmentally arranged along the rostrocaudal axis. DiI-labelled cells were observed in all truncal neural crest derivatives, including subepidermal presumptive pigment cells, dorsal root ganglia, and sympathetic ganglia. By varying the stage at which the injection was performed, neural crest cell emigration at the level of the wing bud was shown to occur from stage 13 through stage 22. In addition, neural crest cells were found to populate their derivatives in a ventral-to-dorsal order, with the latest emigrating cells migrating exclusively along the dorsolateral pathway.     

Plasminogen activator activity is associated with neural crest cell motility in tissue culture

We have examined the possibility that proteases such as plasminogen activator (PA) contribute to the extraordinary motile capability of neural crest cells. We show that trunk neural crest cells that migrate from isolated neural tubes in vitro produce PA and that the level of cell-associated PA increases dramatically after 8 days in culture. This increase is not the result of differentiation or time in culture, because neural crest cell clusters that form on top of the neural tube and differentiate into pigment cells but are immotile produce very low levels of PA. If these clusters are removed from the neural tube and replated on a plastic substratum where they migrate, the level of PA associated with the cells increases dramatically, suggesting that PA production is associated with motility. Inhibitors of PA/plasmin activity significantly reduce neural crest cell motility in vitro, further supporting the idea that proteases are important in neural crest cell migration.     

Neural crest cells provide species-specific patterning information in the developing branchial skeleton

The skeletal elements of the branchial region are made by neural crest cells, following tissue interactions with the pharyngeal endoderm. Previous transplantation experiments have claimed that the cranial neural crest is morphogenetically prespecified in respect of its branchial skeletal derivatives, that is, that information for the number, size, shape, and position of its individual elements is already determined in these cells when they are still in the neural folds. This positional information would somehow be preserved during delamination from the neural tube and migration into the branchial arches, before being read out as a spatial pattern of chondrogenesis and osteogenesis. However, it now appears that signals from the endoderm are able to specify not only the histogenic differentiation state of neural crest cells but also the identity and orientation of the branchial skeletal elements. It is therefore important to ask whether fine details of branchial skeletal pattern such as those that exist between different species are also governed by extrinsic factors, such as the endoderm, or by the neural crest itself. We have grafted neural crest between duck and quail embryos and show that the shape and size of the resulting skeletal elements is donor derived. The ability to form species-specific patterns of craniofacial skeletal tissue thus appears to be an inherent property of the neural crest, expressed as species-specific responses to endodermal signals.     

Control of neural crest cell dispersion in the trunk of the avian embryo

Many hypotheses have been advanced to explain the orientation and directional migration of neural crest cells. These include positive and negative chemotaxis, haptotaxis, galvanotaxis, and contact inhibition. To test directly the factors that may control the directional dispersion of the neural crest, I have employed a variety of grafting techniques in living embryos. In addition, time-lapse video microscopy has been used to study neural crest cells in tissue culture. Trunk neural crest cells normally disperse from their origin at the dorsal neural tube along two extracellular pathways. One pathway extends laterally between the ectoderm and somites. When either pigmented neural crest cells or neural crest cells isolated from 24-hr cultures are grafted into the space lateral to the somites, they migrate: (1) medially toward the neural tube in the space between the ectoderm and somites and (2) ventrally along intersomitic blood vessels. Once the grafted cells contact the posterior cardinal vein and dorsal aorta they migrate along both blood vessels for several somite lengths in the anterior-posterior axis. Neural crest cells grafted lateral to the somites do not immediately move laterally into the somatic mesoderm of the body wall or the limb. Dispersion of neural crest cells into the mesoderm occurs only after blood vessels and nerves have first invaded, which the grafted cells then follow. The other neural crest pathway extends ventrally alongside the neural tube in the intersomitic space. When neural crest cells were grafted to a ventral position, between the notochord and dorsal aorta, in this intersomitic pathway at the axial level of the last somite, the grafted cells migrate rapidly within 2 hr in two directions: (1) dorsally, in the intersomitic space, until the grafted cells contact the ventrally moving stream of the host neural crest and (2) laterally, along the dorsal aorta and endoderm. All of the above experiments indicate that neither a preestablished chemotactic nor adhesive (haptotactic) gradient exists in the embryo since the grafted neural crest cells will move in the reverse direction along these pathways toward the dorsal neural tube. For the same reason, these experiments also show that dispersal of the neural crest is not directed passively by other environmental controls, since the cells can clearly move counter to their usual pathway and against such putative passive mechanisms.(ABSTRACT TRUNCATED AT 400 WORDS)     

The effects of embryonic retinal neurons on neural crest cell differentiation into Schwann cells

Amongst the many cell types that differentiate from migratory neural crest cells are the Schwann cells of the peripheral nervous system. While it has been demonstrated that Schwann cells will not fully differentiate unless in contact with neurons, the factors that cause neural crest cells to enter the differentiative pathway that leads to Schwann cells are unknown. In a previous paper (Development 105: 251, 1989), we have demonstrated that a proportion of morphologically undifferentiated neural crest cells express the Schwann cell markers 217c and NGF receptor, and later, as they acquire the bipolar morphology typical of Schwann cells in culture, express S-100 and laminin. In the present study, we have grown axons from embryonic retina on neural crest cultures to see whether this has an effect on the differentiation of neural crest cells into Schwann cells. After 4 to 6 days of co-culture, many more cells had acquired bipolar morphology and S-100 staining than in controls with no retinal explant, and most of these cells were within 200 microns of an axon, though not necessarily in contact with axons. However, the number of cells expressing the earliest Schwann cell markers 217c and NGF receptor was not affected by the presence of axons. We conclude that axons produce a factor, which is probably diffusible, and which makes immature Schwann cells differentiate. The factor does not, however, influence the entry of neural crest cells into the earliest stages of the Schwann cell differentiative pathway.     

Neural fold and neural crest movement in the Mexican salamander Ambystoma mexicanum

In studies of amphibian neurulation, the terms "neural ridge," "neural fold," and "neural crest" are sometimes used as synonyms. This has occasionally led to the misconception that grafting of the neural crest is equivalent to grafting of the neural fold. The neural fold, however, is composed of three parts: the neural crest, prospective neural tube tissue, and epidermis. In order to investigate how these neural fold components move during neurulation, time-lapse photography, electron microscopy, and grafting were performed. Ambystoma mexicanum embryos were photographed during neurulation at regular intervals. The photographs were analyzed to find the position of those cells at beginning of neurulation that end up on the line of fusion as the neural folds close. Posteriorly, these cells are already on the emerging neural fold. In the anterior neural folds, however, these cells are located in the lateral epidermis. Electron microscopy of the neural folds confirms the presence of epidermis. To follow the movement of the cells differentiating into melanophores (neural crest), neural fold parts were grafted into albino hosts. The crest cells differentiating into melanophores following ectopic grafting are located in the flank of the neural fold that is in contact with the neural plate. In grafts from the outside (distal) flank, no melanophores developed. Semithin sections show that the third part of the neural fold consists of apically constricted cells known to differentiate into neural tissue. Because the neural folds consist of epidermis, neural tissue, and neural crest, neural fold and neural crest cannot be used as synonyms.     

Directed differentiation of pluripotent cells to neural lineages: homogeneous formation and differentiation of a neurectoderm population

During embryogenesis the central and peripheral nervous systems arise from a neural precursor population, neurectoderm, formed during gastrulation. We demonstrate the differentiation of mouse embryonic stem cells to neurectoderm in culture, in a manner which recapitulates embryogenesis, with the sequential and homogeneous formation of primitive ectoderm, neural plate and neural tube. Formation of neurectoderm occurs in the absence of extraembryonic endoderm or mesoderm and results in a stratified epithelium of cells with morphology, gene expression and differentiation potential consistent with positionally unspecified neural tube. Differentiation of this population to homogeneous populations of neural crest or glia was also achieved. Neurectoderm formation in culture allows elucidation of signals involved in neural specification and generation of implantable cell populations for therapeutic use.     

Anterior/Posterior Influences on Neural Crest-Derived Pigment Cell Differentiation

The neural crest of vertebrate embryos has been used to elucidate steps involved in early embryonic cellular processes such as differentiation and migration. Neural crest cells form a ridge along the dorsal midline and subsequently they migrate throughout the embryo and differentiate into a wide variety of cell types. Intrinsic factors and environmental cues distributed along the neural tube, along the migratory pathways, and/or at the location of arrest influence the fate of neural crest cells. Although premigratory cells of the cranial and trunk neural crest exhibit differences in their differentiation potentials, premigratory trunk neural crest cells are generally assumed to have equivalent developmental potentials. Axolotl neural crest cells from different regions of origin, different stages of development, and challenged with different culture media have been analyzed for differentiation preferences pertaining to the pigment cell lineages. We report region-dependent differentiation of chromatophores from trunk neural crest at two developmental stages. Also, dosage with guanosine produces region-specific influences on the production of xanthophores from wild-type embryos. Our results support the hypothesis that spatial and temporal differences among premigratory trunk neural crest cells found along the anteroposterior axis influence developmental potentials and diminish the equivalency of axolotl neural crest cells.     

sucker encodes a zebrafish Endothelin-1 required for ventral pharyngeal arch development

Mutation of sucker (suc) disrupts development of the lower jaw and other ventral cartilages in pharyngeal segments of the zebrafish head. Our sequencing, cosegregation and rescue results indicate that suc encodes an Endothelin-1 (Et-1). Like mouse and chick Et-1, suc/et-1 is expressed in a central core of arch paraxial mesoderm and in arch epithelia, both surface ectoderm and pharyngeal endoderm, but not in skeletogenic neural crest. Long before chondrogenesis, suc/et-1 mutant embryos have severe defects in ventral arch neural crest expression of dHAND, dlx2, msxE, gsc, dlx3 and EphA3 in the anterior arches. Dorsal expression patterns are unaffected. Later in development, suc/et-1 mutant embryos display defects in mesodermal and endodermal tissues of the pharynx. Ventral premyogenic condensations fail to express myoD, which correlates with a ventral muscle defect. Further, expression of shh in endoderm of the first pharyngeal pouch fails to extend as far laterally as in wild types. We use mosaic analyses to show that suc/et-1 functions nonautonomously in neural crest cells, and is thus required in the environment of postmigratory neural crest cells to specify ventral arch fates. Our mosaic analyses further show that suc/et-1 nonautonomously functions in mesendoderm for ventral arch muscle formation. Collectively our results support a model for dorsoventral patterning of the gnathostome pharyngeal arches in which Et-1 in the environment of the postmigratory cranial neural crest specifies the lower jaw and other ventral arch fates.     

The endoderm plays an important role in patterning the segmented pharyngeal region in zebrafish (Danio rerio)

The development of the vertebrate head is a highly complex process involving tissues derived from all three germ layers. The endoderm forms pharyngeal pouches, the paraxial mesoderm gives rise to endothelia and muscles, and the neural crest cells, which originate from the embryonic midbrain and hindbrain, migrate ventrally to form cartilage, connective tissue, sensory neurons, and pigment cells. All three tissues form segmental structures: the hindbrain compartmentalizes into rhombomeres, the mesoderm into somitomeres, and the endoderm into serial gill slits. It is not known whether the different segmented tissues in the head develop by the same molecular mechanism or whether different pathways are employed. It is also possible that one tissue imposes segmentation on the others. Most recent studies have emphasized the importance of neural crest cells in patterning the head. Neural crest cells colonize the segmentally arranged arches according to their original position in the brain and convey positional information from the hindbrain into the periphery. During the screen for mutations that affect embryonic development of zebrafish, one mutant, called van gogh (vgo), in which segmentation of the pharyngeal region is absent, was isolated. In vgo, even though hindbrain segmentation is unaffected, the pharyngeal endoderm does not form reiterated pouches and surrounding mesoderm is not patterned correctly. Accordingly, migrating neural crest cells initially form distinct streams but fuse when they reach the arches. This failure to populate distinct pharyngeal arches is likely due to the lack of pharyngeal pouches. The results of our analysis suggest that the segmentation of the endoderm occurs without signaling from neural crest cells but that tissue interactions between the mesendoderm and the neural crest cells are required for the segmental appearance of the neural crest-derived cartilages in the pharyngeal arches. The lack of distinct patches of neural crest cells in the pharyngeal region is also seen in mutants of one-eyed pinhead and casanova, which are characterized by a lack of endoderm, as well as defects in mesodermal structures, providing evidence for the important role of the endoderm and mesoderm in governing head segmentation.     

The neural crest and neural crest cells: discovery and significance for theories of embryonic organization

The neural crest has long fascinated developmental biologists, and, increasingly over the past decades, evolutionary and evolutionary developmental biologists. The neural crest is the name given to the fold of ectoderm at the junction between neural and epidermal ectoderm in neurula-stage vertebrate embryos. In this sense, the neural crest is a morphological term akin to head fold or limb bud. This region of the dorsal neural tube consists of neural crest cells, a special population(s) of cell, that give rise to an astonishing number of cell types and to an equally astonishing number of tissues and organs. Neural crest cell contributions may be direct — providing cells — or indirect — providing a necessary, often inductive, environment in which other cells develop. The enormous range of cell types produced provides an important source of evidence of the neural crest as a germ layer, bringing the number of germ layers to four — ectoderm, endoderm, mesoderm, and neural crest. In this paper I provide a brief overview of the major phases of investigation into the neural crest and the major players involved, discuss how the origin of the neural crest relates to the origin of the nervous system in vertebrate embryos, discuss the impact on the germ-layer theory of the discovery of the neural crest and of secondary neurulation, and present evidence of the neural crest as the fourth germ layer. A companion paper (Hall, Evol. Biol. 2008) deals with the evolutionary origins of the neural crest and neural crest cells.     

Effect of lectins and sugars on primary sperm attachment in the horseshoe crab, Limulus polyphemus L

The in vitro differentiation of quail neural crest cells into serotoninergic neurons is reported. Serotoninergic neurons were identified by two independent methods, formaldehyde-induced histofluorescence and indirect staining with antiserotonin antibodies. Serotonin-positive cells first appeared on the third day in culture, simultaneously, or slightly prior to the first pigmented cells and adrenergic neurons. Comparable numbers of serotoninergic cells were found in crest cell cultures derived from vagal, thoracic/upper lumbar, and lumbosacral levels of the neuraxis. The neural crest origin of the serotonin neurons was further corroborated by the demonstration that cultures of somites, notochords, and neural tubes (three tissues adjacent to the neural crest and thus the most likely contaminants of crest cell cultures) did not contain serotonin-producing cells, and that mast cells were absent in crest cell cultures. The identification of serotoninergic neurons in quail neural crest cell cultures makes an important addition to the number of neural crest derivatives that are capable of differentiating in culture. Furthermore, it suggests that the in vitro culture system will prove a valid approach to the elucidation of the cellular and molecular mechanisms that govern neural crest cell differentiation.     

Environmental factors affecting neural crest differentiation: Melanocyte differentiation by crest cells exposed to cell-free (deoxycholate-extracted) dermal mesenchyme matrix

Summary Deoxycholate-extracted, cell-free matrices were prepared from primary expiants or dispersed cell cultures of embryonic avian dermis, ectoderm, gut mesenchyme, endoderm, pharynx, or umbilical artery. Neural crest cells in association with matrices from dermal expiants or monolayers formed melanocytes after six days. Crest cells in association with matrices from all other tissues or grown on plastic did not form melanocytes. It is concluded that a deoxycholate-resistant structural component of the dermal extracellular matrix induces melanocyte differentiation.Abbreviations used ECM extracellular matrix - GAG glycosaminoglycan - HANKS Hank's balanced salt solution - FCS fetal calf serum - DME Dulbecco's modified Eagle's medium - DOC deoxycholate - PMSF phenyl methyl sulfonyl fluoride