Thesis (M.S.)--Virginia Polytechnic Institute and State University, 1994. / Vita. Abstract. Includes bibliographical references (leaves 61-73). Also available via the Internet.

Thesis (M.S.)--Brigham Young University. Dept. of Physiology and Developmental Biology, 2007. / Includes bibliographical references (p. 19-25).

Thesis (Ph. D.)--University of Missouri-Columbia, 2006. / The entire dissertation/thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file (which also appears in the research.pdf); a non-technical general description, or public abstract, appears in the public.pdf file. Title from title screen of research.pdf file (viewed on August 10, 2007) Vita. Includes bibliographical references.

The role of cyclic AMP and a differentiation-indueing factor (DIF) in the differentiation of stalk cells was investigated in the cellular slime mold Dictyostelium discoideum. In this organism, starvation triggers the aggregation of amoebae into multicellular masses within which a simple, well-regulated pattern of partially differentiated cells is formed and which ultimately form fruiting bodies comprised of spore and stalk cells. In a monolayer system at low cell densities, stalk cell formation is dependent on the presence of both cyclic AMP and DIF. Both factors act within a short time of each other, induction by cyclic AMP preceding induction by DIF, beginning between 8 to 10 hours of incubation in monolayers, and progressively committing an increasing proportion of the cells in monolayer to form stalk cells. The relative effectiveness of analogues of cyclic AMP to induce stalk cell formation in monolayers indicates that the well-characterized cell surface cyclic AMP receptor most probably mediates the action of cyclic AMP. Although this receptor appears early during aggregation, it does not become activated until later during development in vivo, probably because the cyclic AMP concentrations within developing cell masses must build up to levels higher than those in aggregation streams. The finding that caffeine inhibits stalk cell formation in low density monolayers and that the permeable analogue 8-Bromo-cyclic AMP can partially reverse this inhibition suggests that activation of this receptor leads to an increase in internal cyclic AMP levels as one of the steps in stalk cell differentiation. The finding that the expression in low density monolayers of AP IV, a cell-type non-specific isozyme of acid phosphatase, was cyclic AMP-dependent is consistent with the view that cyclic AMP induces non-specific postaggregative gene expression during development in vivo. The findings that the expression of pre-stalk arid stalk cell specific antigens and of the pre-stalk cell specific isozyme AP II was DIF-dependent provide good evidence for the idea that both pre-stalk and stalk cell formation are induced by DIF. The fact that isolated pre-stalk cells require DIF for stalk cell formation in low density monolayers further supports this idea. Whereas cells independent of DIF for stalk cell formation in monolayers appear immediately after cyclic AMP-independent cells during differentiation in low density monolayers, DIF-independent cells appear considerably later during development in vivo. This evidence and the fact that developing cell masses contain elevated levels of DIF lead to the postulate that the factor(s) which triggers the formation of fruiting bodies also controls the pre-stalk to stalk cell conversion. / Science, Faculty of / Botany, Department of / Zoology, Department of / Graduate

Morphological, electrophysiological and biochemical changes have been shown to occur in the retina, lateral geniculate nucleus, and visual cortex of light deprived animals. We attempted to determine whether the dark-rearing of rats from birth to 15, 30 and 60 days of age alters the ability of noradrenaline (NA) 30 μM, potassium chloride (KCI) 50 μM, adenosine 30 μM and combinations of NA and KCI with adenosine to stimulate the _in vitro formation of cyclic AMP (cAMP) in visual cortical slices and, as an internal control, in frontal cortical slices. At 15 and 30 days of age there was an 11% and 2170 reduction, respectively, compared to normally reared controls, in the stimulation of cAMP formation in a 5 minute incubation with NA in both frontal and visual cortical slices. After 60 days of dark-rearing, however, this was reversed in that the NA stimulation of cAMP formation was 23% and 357» higher than controls in frontal and visual cortical slices. In frontal cortical slices of rats dark-reared for 15 and 30 days there was a significant reduction in the stimulation of cAMP formation in a 20 minute incubation with NA. No differences were observed between 30 day old experimental and control animals in studies of the accumulation of cAMP in frontal and visual cortical slices incubated for various times with KCI. The stimulation of cAMP formation induced by KCI and adenosine in a 5 minute incubation was 5770 and 397o higher, respectively, in frontal cortical slices of 60 day old experimental animals than controls while the response in visual cortical slices was unaffected. The differences found between 60 day old experimental and control animals were abolished in both visual and frontal cortical slices when adenosine was used in combination with NA or KCI. Studies of the accumulation of cAMP in slices incubated for various times with NA revealed that the effect observed in the visual cortex after 30 days of light deprivation was due to a decrease in the maximum level of cAMP reached within a 20 minute incubation period, whereas in the frontal cortex the maximum level attained within a 20 minute incubation period was unaffected. These results are discussed in terms of our present knowledge concerning supersensitivity and plasticity in the central nervous system and the role of cAMP in nerve. / Medicine, Faculty of / Biochemistry and Molecular Biology, Department of / Graduate

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The application of microanalytical techniques was used to elucidate the enzyme profiles of adenylate cyclase and cAMP-dependent protein kinase within a series of malignant and benign human mammary tumors. The methods allow for determination of enzyme activity in histologically-confirmed neoplastic areas within individual tumors which permits comparisons to be made with particle and cytosolic fractions prepared from mammary tumors. The potential for cAMP formation by adenylate cyclase and mediation of cAMP effects by cAMP-dependent protein kinase were studied to provide information on the levels of enzyme activity and the extent and type(s) of regulation of these enzymes in a cell-free system. Adenylate cyclase (AC) activity levels were higher in benign than malignant tumor tissue on a dry wt basis. A 2 to 3-fold activation by Gpp(NH)p and a 4 to 5-fold stimulation by fluoride was demonstrated. Calmodulin activation of tumor-associated AC was not observed. Membrane-associated AC could be stimulated by PGE₁, prolactin and TSH. Thermal stability studies suggested that AC from malignant tissue was more labile than that from benign tissue. Cyclic AMP-dependent protein kinase associated with human mammary tumors was primarily type II kinase based upon DEAE-Sephacel chromatography and demonstrated a preference for histone VS as a substrate. Cyclic AMP-dependency was supported by enzyme activation by cAMP, by cAMP binding, and inhibition by protein kinase inhibitors and regulatory subunit. The activity ratio was slightly higher in benign than malignant tissue. Histone kinase activity was higher in benign than malignant tumor microsections. Cyclic AMP-binding activity was higher in malignant than benign tumor cytosolic fractions, although cAMP affinity was similar. Catalytic subunit was prepared from tumor histone kinase by cAMP affinity chromatography. Reassociation of tumor catalytic subunit and heterologous regulatory subunit was shown. A cAMP-independent casein kinase was also partially characterized in human mammary tumors. Endogenous phosphorylation in tumor cytosolic fractions was examined and polypeptides identified which showed phosphorylation differences in the presence of cAMP. / Ph. D.