F. Serras

Is a mosaic embryo also a mosaic of communication compartments

We have studied the pathways of cell communication in embryos of the mollusc Lymnaea stagnalis in which the developmental fate of a cell or a group of cells is known from cell lineage studies. We iontophoretically injected Lucifer Yellow CH and followed the spread of fluorescence between cells interconnected via gap junctions. In early stages all blastomeres appear to be dye-coupled, but later on communication is restricted within compartments. The pattern of cell communication corresponds with the development of compartments with specific cell fates. Dye-spread is limited by communication boundaries which completely or mostly prevent the passage of dye to adjacent compartments with different developmental fates. These boundaries appear progressively during development. Our results suggest that, during the development of Lymnaea, the progressive changes in the pattern of dye spread correspond with the progressive restrictions of the developmental fates of individual cells or groups of cells. We conclude that changes in the pattern of cell communication and in the appearance of communication compartments are not exclusive features of regulative embryos.

Communication compartments in the ectoderm of embryos of Patella vulgata

SummaryPatterns of gap junctional communication in the ectoderm of embryos of Patella vulgata have been studied by intracellular injection of the fluorescent dye Lucifer Yellow, and by analysis of its subsequent spread to adjacent cells (dye-coupling). We found that dye-coupling became progressively restricted to different domains of the ectoderm, forming communication compartments. These communication compartments are characterized by their high coupling abilities within the compartment, and reduction of coupling across their boundaries. During development, the pretrochal (anterior) ectoderm becomes subdivided into two communication compartments, the apical organ and the anlage of the head ectoderm. The posttrochal (posterior) ectoderm becomes subdivided into different communication compartments in two successive phases. Firstly, in the 15-h embryo the dorsal and ventral domains of the ectoderm form separate communication compartments. A dorso-ventral communication boundary restricts the passage of dye between the two domains. Secondly, in the 24-h embryo dye-coupling becomes further compartmentalized in both the dorsal and ventral domains. These compartments correspond to the anlagen of different ectodermal structures. In order to study whether any level of coupling persists between the ectodermal compartments we injected currents through a microelectrode inserted into one cell of one compartment and monitored its spread by means of a second microelectrode inserted into one cell of another compartment (electrical coupling). Despite the absence of dye-coupling, electrical coupling between the ectodermal dye-coupling compartments was detected, which suggests that some level of communication is maintained between compartments. Our results demonstrate that within the ectoderm layer of Patella vulgata the transfer of dyes becomes progressively restricted to communication compartments and, concomitantly with the specification of the different ectodermal anlagen, these compartments become subdivided into smaller communication compartments.

Cell communication compartments in molluscan embryos

Early embryos of Patella vulgata have been injected with Lucifer Yellow. No restriction of dye spread was found. We show that later in the development, the larval trochophore stage present evidence of compartments of cell communication. These dye compartments coincide with different presumptive regions.

Changes in junctional communication associated with cell cycle arrest and differentiation of trochoblasts in embryos of Patella vulgata

In early embryos of molluscs, different clones of successively determined trochoblasts differentiate into prototroch cells and together contribute to the formation of a ciliated ring of cells known as the prototroch. Trochoblasts differentiate after cell cycle arrest, which occurs two cell cycles after the commitment of their stem cell. To study the changes of junctional communication in embryos of Patella vulgata in relation to commitment, cell cycle arrest, and differentiation of the trochoblasts, we have monitored electrical coupling as well as transfer of fluorescent dyes. The appearance of dye coupling in embryos of Patella occurs after the fifth cleavage (at the 32-cell stage), when the cell cycles of all embryonic cells become asynchronous and longer. At the 32- and 64-cell stages all cells are well coupled. However, after the 72-cell stage dye transfer to or from any cell of the four interradial clones of four primary trochoblasts becomes abruptly reduced, whereas electrical coupling between these cells and the rest of the embryo can still be detected. From scanning electron microscopical analysis of the cell pattern we conclude that this change in gap junctional communication coincides with cell cycle arrest and with the development of cilia in all four clones of primary trochoblasts. Similarly, after the 88-cell stage the four radial clones of accessory trochoblasts stop dividing, reduce cell coupling, and become ciliated. By the formation of the prototroch, the embryo becomes subdivided into an anterior (pretrochal) and a posterior (posttrochal) domain which will develop different structures of the adult. At the 88-cell stage, the cells within each of these two domains remain well coupled and form two different communication compartments that are separated from each other by the interposed ring of uncoupled trochoblasts. The relations among control of cell cycle, changes in junctional communication, and differentiation are discussed.

Progressive Restrictions in Gap Junctional Communication during Development

One of the most fascinating challenges in developmental biology is to elucidate the mechanisms that control the organization of the body plan during embryonic development. In this regard intercellular transduction of signals is of particular importance for a coordinated pattern of cell differentiation. Because of their ability to allow the exchange of ions and small molecules up to a molecular weight of about 1200 D (Simpson et al., 1977), gap junctions have been proposed as a putative pathway for the intercellular transfer of developmentally important signals (Furshpan and Potter, 1968). In most of the embryos so far studied gap junctions appear in the early stages of development, when important developmental decisions take place (see Caveney, 1985; Guthrie, 1987 for reviews). More striking results have shown that differences in junctional communication may be found between embryonic tissues with different developmental programmes. The changes of functional gap junctional communication between cells with divergent developmental fates suggest an involvement of gap junctional-mediated signal transduction in the specialization and organization of the different domains of the embryo (Van den Biggelaar, 1988).

F-Actin is a marker of dorsal induction in earlyPatella embryos

SummaryThe dorsal-ventral axis inPatella vulgata embryos is established at the 32-cell stage by an inductive interaction between the animal micromeres and one vegetal macromere. This vegetal macromere, once induced, is called the 3D macromere, and marks the future dorsal side of the embryo. We examined the pattern of filamentous (F) actin in such embryos using fluorescent phalloidin and found that this dorsal 3D macromere contains more F-actin than the remainder of the cells. In addition, only one of its two daughter cells, i.e. the 4D macromere, retains this higher density. In embryos in which the establishment of the dorsal-ventral axis has been experimentally inhibited via treatment with monensin, such differences in F-actin were not found. These results suggest that the appearance of an increased density of F-actin in the dorsal 3D and 4D macromeres of normal embryos requires the inductive interactions that establish the dorsal-ventral axis. We therefore conclude that F-actin is an early marker for dorsal induction in thePatella embryo.

Communication compartments in the post-trochal ectoderm of the mollusc Lymnaea stagnalis

Cell-to-cell communication via gap junctions provides a pathway for the transfer of small molecules and ions which may be significant for control of metabolic cooperation, cell proliferation, and differentiation. We have assessed the patterns of gap junctional communication in embryos of the mollusc Lymnaea stagnalis during the subdivision of the post-trochal ectoderm into developmental domains. We have microinjected the tracer Lucifer Yellow CH and subsequently analyzed its transfer to other cells. The post-trochal ectoderm of mollucs develops the shell field, the foot, and the stomodeum anlagen. We have found that the cells within the separate anlagen are well dye-coupled but poorly coupled to cells of adjacent anlagen. These results indicate that in Lymnaea embryos the specification of the different developmental domains is associated with the development of corresponding dye-coupling compartments.

Gap Junctional Communication and Cell Cycle Duration in the Early Molluscan Embryo

The most remarkable aspect of the early molluscan development is the extremely regular succession and pattern of cleavages. Without any variance the embryos of the same species exactly reproduce the same cell pattern. Initially, all blastomeres are in phase, and divide synchronously. After a species specific number of simultaneous division cycles, the cells in the different lineages become out of phase and an in time and space well defined program of cell divisions is executed. In this chapter the possible influence of intercellular communication on the duration of the cell cycle in early molluscan development will be discussed. Although extracellular factors may also influence progress through the cell cycle, these will be left out of consideration.