Thesis

Effects of Sugar Alcohols on Sea Urchin Gastrulation in Low Calcium Sea Water

ABSTRACT THE EFFECTS OF SUGAR ALCOHOLS ON SEA URCHIN GASTRULATION IN LOW CALCIUM SEAWATER By Edward Holmes Master of Science in Biology The sea urchin embryo has been designated by the National Institutes of Health as a model system for studying cellular interactions. Over 25 physiological discoveries were made in sea urchins crucial to understanding their function in many taxonomic groups including humans. The sea urchin embryo was used in this study to examine the effects of sugar alcohols on cell- cell interactions during gastrulation. This study examined six different sugar alcohols in 1.5mM low calcium artificial seawater (LCASW) on 24-26 hour Lytechinus pictus sea urchin embryos and 32-34 hour Lytechinus pictus sea urchin embryos. The embryos were incubated in 1.5mM low calcium artificial sea water with and without (controls) 0.1M, 0.05M, 0.01M, 0.005M, and 0.001M adonitol, L-(-) arabitol, dulcitol, D-mannitol, D-sorbitol, and xylitol in 96 well microplates and were fixed at 48-50 hours, observed, and photographed. Three to six separate experiments were conducted for each sugar alcohol concentration at each of the two times. The embryo morphologies observed were: complete archenteron, incomplete archenteron, not invaginated, exogastrulated and dead. Sugar alcohol concentrations for each experiment were tested in 12 replicate wells yielding hundreds of sea urchin embryos in each experiment. Low calcium artificial sea water was utilized in this study because it was found to loosen septate junctions found in the blastula epithelium, accelerating molecular entry into the blastocoel of the embryos without microinjection. An unpaired t-test was performed for each sugar alcohol concentration and morphology compared to the LCASW control. P values less than 0.05 (p<0.05) suggested that the percentages of morphologies in the sugar alcohol concentration were significantly different than in the LCASW control. The sugar alcohols uniformly had little effect on embryo morphologies. The sugar alcohols causing slight effects on archenteron morphology when added at 24-26 hrs: (most effects) adonitol > L-(-) arabitol > xylitol > dulcitol = D- sorbitol > D-mannitol (least effects). The sugar alcohols causing slight effects on archenteron morphology when added at 32-34 hours (most effects) D-mannitol > dulcitol > adonitol = xylitol > D-sorbitol = L-(-) arabitol (least effects). While effects were minimal, this study indicates that very small differences can be identified quantitatively using this assay system.

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