The objectives of this project are 1) to use an embryonic epithelium, the enveloping layer of the medaka, Oryzias latipas, a teleost fish, to test two hypotheses of a model for epithelial morphogenesis; 2) to determine the roles of calcium ion and microfilaments in the contraction of the enveloping layer; and 3) to determine the physiological and cellular bases of rhythmic contractions of the enveloping layer. The origin and propagation of the contractile waves will be viewed using Nomarski optics and a time lapse video tape recorder. Myosin and actin will be localized by using fluorescently-labelled antibodies and phalloidin. The role of calcium ion will be assessed by using specific calcium channel blocking agents and by using the calcium indicator substance, quin-2. The role of gap junctions in the propagation of rhythmic contractions will be assessed by viewing the diffusion of fluorescein from cell to cell; by using o-nitrobenzyl acetate (and other permeant esters) to acidify the cytoplasm and thus decrease gap junction permeability; and by assessing the effect of o-NBA on the diffusion of fluorescein from cell to cell. The research promise to further our understanding of how embryonic epithelia change their form. It specifically addresses the question of how the activities of individual cells are orchestrated into coordinated movements of entire populations of cells-movements that occur throughout embryonic development.
