The overall aims of this research are severalfold: (1) to elucidate and time the events in the cell division, fissioning, and regeneration of Stenostomum-virginianum; (2) to study the effects of mitotic agents, co-carcinogens, and pollutants on these processes, and (3) to develop Stenostomum as an animals model for drug and pollution, research. Stenostomum offers ideal material for the study of drug and pollution effects on cell growth, fissioning, and regeneration because the animal fissions and regenerates within a day if induced by decapitation or transectioning. The animal will be obtained from Carolina Biological, subcultured into a qenetically uniform clone, and maintained in 200-ml pond water cultures containing several wheat seed supplemented often with dilute yeast. To sequence the stages of cell division, fissioning, and regeneration, during the first year, animals will be decapitated or transected and allowed to fission or regenerate for 0, 1, 2, 4, 8, 16, 24, and 32 h post-treatment. The animals will then be fixed in 1-3% phosphate-buffered glutaraldehyde; postfixed in buffered 1% osmium tetroxide; en bloc stained in 1% uranyl acetate; dehydrated in an acetone series; embedded in Araldite; and thick and thin sections cut and stained for LM and EM. About 20 animals will be processed for each interval. To follow the effects of mitotic agents (colchicine, vinblastine, captoethanol) during the second year; cocarcinogens (cadmium sulfate/TBA) during the third year; and pollutants (2, 4-D, Alar, etc) in the 4th year, dose response curves will be determined and geometrical series of concentrations will be prepared from routine dilutions of the minimal lethal dosages. Exposure times will vary in the same geometric series as above. Differences in the rates of division, fissioning, and regeneration will be followed as well as histopathological differences in cytology and tumorigenesis.
