Fixation of embryonic chick cells (heart, neural retina, and limb bud) in the presence of lanthanum ions shows the presence of an electron-opaque layer, about 50 A thick, external to the cell membrane. This layer, designated LSM (for lanthanum-staining material), is not removable by trypsin, pronase, EDTA, DNase, a-amylase, neuraminidase, or N-acetyl-Lcysteine. However, phospholipase C, in concentrations as low as 0.001 mg/ml, succeeds in stripping the LSM from the cell surface. Heating the enzyme preparation does not inhibit this activity, but removal of divalent cations does; both of these results are consistent with the known properties of phospholipase C. The LSM is present at the cell surface in the control tissues and on cells dissociated from the tissues by proteolytic enzymes and EDTA. These results are interpreted to mean that the LSM is probably an integral part of the cell and not an extraneous coat. How this phenomenon bears on the problem of cellular adhesion is discussed, as is the possible chemical composition of the LSM.
During epiboly stages the cells (called deep blastomeres) which will form the definitive embryo disperse over the surface of the yolk sphere, only later aggregating and developing an embryonic axis. Five different statistical tests were used to study the pattern formed by the deep blastomeres during epiboly and early dispersed stages. The two most reliable tests, based on the distance from each deep blastomere within a selected area to its nearest neighboring cell, indicate that the distribution pattern changes from regular during epiboly stages to random during dispersed stages 1 and 2. Careful observation and time-lapse microphotography revealed some aspects of how the cells set up the regular pattern. The deep blastomeres exhibit a variety of cell extensions, with which they often contact one another. When two deep blastomeres make contact during epiboly stages, they soon break the contact and move apart; they overlap one another only rarely. Deep blastomeres are frequently located at, and are even elongated along, borders of the overlying flat cells (enveloping layer cells). These two mechanisms, one similar to contact inhibition of cell movement, the other to contact guidance, may contribute to the rather regular spacing of the deep blastomeres as well as to their arrangement in rows during epiboly stages.
The deep blastomeres of annual fish embryos emigrate from the blastodisc to wander as single cells, during epiboly and early dispersed stages, between the periblast and the enveloping layer cells. Time-lapse films of the deep blastomere movements were analyzed to determine whether the cells move persistently or randomly and whether or not they show contact inhibition of cell movement. Certain predictions which follow from the random-walk model (uniform distribution of intersegmental angles, exponential distribution of squared segmental lengths, and a straight-line plot -passing through the origin -of mean square displacement against time) were tested and, in general, found to apply to these deep blastomeres during dispersed phase 1 but not during epiboly stages. These results indicate that the cells tend to persist in their direction of movement during epiboly, but not during dispersed phase 1, when their movement is random. Upon completion of epiboly, there is a decrease in cell motility, as measured by the augmented diffusion constant, D*, and an increase in the average collision rate. Numerous cases of cell contact between deep blastomeres were observed (297 during epiboly and 167 during dispersed phase 1); in every case one or both cells changed their direction of movement. The role this contact inhibition might play in the persistent movement of epiboly and the random movement of dispersed phase 1 is discussed.Cells undergo extensive morphogenetic movements in all animal embryos, but in most embryos they cannot be observed as individuals during their migrations because of the close packing of the cells, usually several layers thick, and often because of the opaqueness of the embryo itself andlor of enveloping layers. Therefore, knowledge about these fundamental cell movements and their control is rather limited (for review, see Trinkaus, '76). The transparency of the annual fish embryo and the migration of its deep blastomeres as single cells in the narrow space between the enveloping layer and the periblast (Peters, '63; Wourms, '63, '72a,b) present an almost ideal system for a study of how certain cells move and interact in a living embryo.After flattening and hollowing of the cellular blastodisc, development of annual fishes departs radically from the typical pattern of teleost embryogenesis. In annual fish embryos, the deep blastomeres do not participate
Deep blastomeres of annual f i s h embryos exhi b i t contact inhibition o f c e l l movement during epiboly and early dispersed stages before returning t o self-cohering and overlapping behavior. Deep blastomeres were dissociated from embryos of several d i f f e r e n t ages and cultured in hanging drops and in s i t t i n g drops. In every culture the c e l l s rapidly reaggregated, irrespective o f whether the c e l l s were aggregated o r dispersed in the living embryo. some c e l l s subsequently flattened onto the p l a s t i c surface and assumed a f i b r o b l a s t i c shape. These r e s u l t s indicate t h a t the contact inhibition and the overlapping behavior exhibited by deep blastomeres i n vivo depend upon interactions o f these c e l l s with t h e i r embryonic environment, probably with the c e l l layers on which they a r e moving.
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