Meeting Abstract
The North Atlantic lobster, Homarus americanus, is a cold-water invertebrate that is important fundamentally and economically. Physiological transepithelial transport processes for any nutrient, ion, or heavy metal cannot be studied with ease due to the lobster’s complex anatomical arrangements. Previous studies on organ dissociation, i.e. hepatopancreas, antennal gland, and/or gills, into cellular suspensions that could be supported in vitro to form functional confluent monolayers provide a technical means to study transepithelial transport. In this preliminary study, 3D culture techniques, and cell density seeding, cellular growth, and morphology of cell colonies were investigated to optimize crustacean cell culture in developing a functional confluent monolayer for transepithelial transport studies. It was found that 3D collagen substrata and regular media changes in vitro can support colony growth of cells that were likely undergoing mitosis, forming functional junctions with neighboring cells, and pseudopodal growth. Cell density appears to play an important role in the development of a confluent monolayer. Cells must not be seeded too densely but must be seeded at a specific density to form functional junctions. Adaptation of colony growth appears to be dependent upon specific seeding density and regular media changes along with supportive 3D substrata. The goal of these investigations is to yield a functional analysis of transepithelial transport in cultured monolayers in Ussing Slider cell culture cup inserts placed into an Ussing chamber that separates the monolayer apical and basal membranes so that the nature and regulation of solute movement across the cell layer can be ascertained.
