Thesis (M.S.)--University of Michigan, 1968.

Thesis (M.S.)--University of Michigan, 1968.

Thesis (Ph. D.)--University of Wisconsin--Madison, 1964. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Bibliography: leaves 120-133.

Targeted radiotherapy (TRT) is a technique which allows for individual cancer cells to be targeted by radiation. However, there is variation in uptake at the whole body, organ, cellular and subcellular levels. This distribution affects the biological efficacy of the TRT agents. To address this problem, novel techniques have been developed and demonstrated. These aim to provide quantitative information about the spatial distribution of Auger electron (AE) emitting radiopharmaceuticals at the subcellular level. Two methods have been developed. The first, photoresist autoradiography (PAR), uses photoresists as an autoradiography substrate, and the second uses microautoradiography (MAR) and a transmission electron microscope (TEM). The techniques have been demonstrated using the AE emitter indium-111. Firstly, PAR is demonstrated using poly (methyl methacrylate) (PMMA). Photoresists were exposed to indium-111 which had been internalised into cells, and the photoresists were analysed using atomic force microscopy (AFM). The technique has a theoretical resolution in the nanometre range and was able to demonstrate cellular patterns on the micron scale. To gain quantitative information, the photoresist response (depth of pattern) was calibrated as a function of electron fluence and a model of the patterns was created. Combining the calibration data with the point source model allowed the position and intensity of the internalised source terms to be estimated using the PAR method. Secondly, a technique for electron microscope-microautoradiography (EM-MAR) was developed. The processing conditions of the MAR technique were determined and staining techniques developed, to produce high quality TEM micrographs. A time course experiment showed the distribution and variation in the uptake of the radiopharmaceutical at the cellular level. Both techniques are able to provide information about the subcellular distribution of the radioactivity at a higher resolution than current techniques. Both enable the collection of information which can be used in microdosimetric calculations.

Methods of measuring primary productivity are discussed in terms of what information they provide concerning energy flow in the phytoplankton community. A track autoradiographic technique is presented which measures species-specific phytoplankton productivity and its use is demonstrated under laboratory conditions. Controls and standardized processing conditions are established which permit the use of the track technique in quantitative evaluations of primary productivity.

The binding of low concentrations of actinomycin D (Act D) to fixed human metaphase chromosomes was studied using both autoradiographic and immunofluorescent techniques. At the concentration range of 0.001 - 0.1 μg/ml Act D is known to selectively inhibit rRNA synthesis. Although it was previously suggested that at these low concentrations Act D would selectively bind to the ribosomal cistrons, evidence also exists that the drug binds to non-ribosomal DNA, and inhibits rRNA transcription in an indirect fashion. Because of the conflicting data on Act D binding and a lack of focus on biologically relevant concentrations of drug, it was decided to systematically investigate the distribution of the drug binding in low concentrations to chromosomes from 72-hr human lymphocyte cultures. Autoradiographic detection of [³H]Act D bound to chromosomes showed no selective binding of the drug at concentrations that maximally inhibit rRNA synthesis. A new technique was employed using Formvar and potassium chromium sulfate as a pretreatment to autoradiography. This technique permitted simultaneous detection of silver grains and chromosome identification by G-banding. With autoradiographic exposure times of 1 and 7 days, there was a positive correlation of autoradiographic grains with chromosome length. To increase sensitivity in detection of Act D bound to chromosomes, a specific anti-Act D antibody was generated in rabbits. Antibody avidity was evaluated on the basis of a rapid charcoal assay. This charcoal assay was then used in development of a radioimmunoassay for Act D which is sensitive in quantitating the drug down to 0.005 μg/ml. The anti-Act D antibody was characterized to be IgG, and was shown to be specific for the pentapeptide lactone portion of the Act D molecule. Indirect immunofluorescence of Protein A-purified IgG containing anti-Act D was used to detect drug bound to fixed human chromosomes. The antibody was shown to be specific for drug bound to chromatin. When 0.1 μg/ml Act D was bound to chromosomes, the drug was observed bound throughout the genome, with no selective binding at the ribosomal cistrons. This confirms the autoradiographic data and supports the model of extranucleolar regulation of rRNA synthesis. Preliminary results suggest that Act D binds to GC-rich DNA, since an R-banding pattern was observed in 5% of the immunofluorescent metaphases examined.

The work presented in this thesis is divided into two parts. The first part is the sorption of Ni and Eu to granitic materials, and cation exchange capacity measurements for powdered and intact samples. The second part is method development on autoradiography. In the first part, static batch sorption experiments were carried out to study the relative sorption properties of different granitic rocks and minerals. Experimental data were described using non-electrostatic correction models such as the Langmuir, Freundlich and Linear models. Sorption data obtained for sorption in a constant pH environment and variable metal concentration were used to test the Component Additive Model (CAM). Sorption test studies carried out using energy dispersive X-ray microanalysis were used to map the sorption of Eu on an intact sample. The results showed the CAM was applicable for Ni sorption to BG but that it was not applicable for Eu sorption to any of the granitic rocks studied. The sorption data fitted the CAM in the following order; BG (1) > GA (0.7) > RG (0.5) > GG (0.2), GrG (0.2) for Ni sorption and RG (0.7) > BG (0.4) > GA (0.2), GG (0.2), GrG (0.2) for Eu sorption to the different granitic rocks. Values in brackets represent the ratio of Rd-predicted/Rd-calculated. Results from the application of the CAM showed it was not possible to predict the Rd of the bulk sample from the component minerals. Desorption studies at constant pH were analysed by calculating the hysteresis H. The results showed that the higher the Rd the higher the hysteresis. Surface complexation using JChess Geochemical Code was used to obtain surface complexation parameters for the metal-solid complex for sorption in variable pH and constant metal concentration. Experimental data were described by the mass action law to obtained proton stoichiometry at which the sorption edge is defined. Results showed the presence of NaCl decreased the sorption of Ni, and increased the sorption of Eu. Sorption kinetics experiments in different carbonate complexing environments were carried out to study the effect of carbonate on Eu sorption capacity and rate of sorption. Data were fitted to first and second order kinetic models to investigate the sorption rates. Results showed the sorption to be fast initially before reaching a steady state after more than 200 hours of equilibration. Kinetic data confirmed the low sorption capacity observed for quartz. Data obtained for sorption in a mixed radionuclide system were modelled using the Linear model and the surface complexation model. The surface complexation constants are correlated to the Rd values obtained from the linear sorption isotherms. Modelling the results using Rdmix and Rdsing showed that sorption was suppressed in a mixed system, with no effect observed for sorption to feldspar in single and mixed systems. Cation exchange capacity (CEC) measurements were undertaken to deduce a correlation between the CEC of powdered rock samples and intact sample using rock beakers developed from the British Geological Survey by applying the Bascomb method in which the pH was buffered to pH 8.1. Normalising the results using the surface area showed that the CEC of the rock beakers was 6 orders of magnitude greater than that of the powdered sample. In the second part, a method for differentiating two or more radionuclides using storage phosphor imaging plates coupled with the Storm Scanner system was tested. Initial results showed that it is possible to differentiate one radionuclide from another in a mixed system using different levels of shielding.

Doctor of Philosophy (PhD) / GABAA receptors are implicated in the pathology of psychiatric disorders such as schizophrenia and depression. They are rapidly affected by stress in a sex-dependent fashion, suggesting that GABAA receptors may be relevant to understanding the association between stress and psychiatric disorders. Thus, this thesis examined how GABAA receptors are affected in both male and female mice exposed to stress in adulthood (Chapter 2), early-life (Chapter 3-5) and a combination of both early-life and adulthood stress (Chapter 6). 2. The effects of acute adulthood stress (3 minute warm swim stress) on GABAA receptor binding in the brains of male and female mice were examined using quantitative receptor autoradiography. The total number of GABAA receptor [3H]GABA binding sites was increased following swim stress in specific forebrain cortical regions of female mice swum individually or in a group, but decreased in male mice when swum in a group only. These findings confirm and extend previous studies, identifying the cortical regions involved in rapid stress-induced changes in GABAA receptors. 3. Post-natal handling models in rodents comparing control (brief handling sessions; EH) with no intervention stress conditions (NH), indicate that the NH condition results in an anxious adulthood phenotype and this was confirmed in the present thesis using the elevated plus-maze behavioural test. Using this model the effects of early-life stress on adulthood GABAA receptors were then examined. 4. Regional densities of GABAA receptor α1 and α2 subunit proteins were observed in the adult brain of male and female mice using immunoperoxidase histochemistry. NH males showed a loss of the α2 subunit from the thalamus and the lower layers (IV-VI) of the primary somatosensory cortex, whilst NH females showed a reduction of α2 but an increase in α1 protein in the lower layers of the primary somatosensory cortex only. These regionally specific alterations in the α1:α2 subunit ratio suggest that early-life stress disrupts the developmental α subunit switch, which occurs in a regionally-dependent fashion over the first two weeks of rodent life. 5. Double-labelling immunofluorescence and confocal microscopy were used to examine the effects of sex and early-life stress on GABAA receptor synaptic clustering. Regardless of sex, mice exposed to early-life stress (NH) showed reduced colocalisation of the GABAA receptor α2 subunit with the synaptic marker protein gephyrin relative to the control condition (EH). This suggests that early-life stress impairs adulthood inhibitory synaptic strength and is consistent with the increased anxiety of the stressed relative to control mice. 6. Finally, the effects of early-life stress on adulthood swim stress-induced changes in GABAA receptor binding were examined using quantitative receptor autoradiography in forebrain cortical regions. Findings showed that the effect of adulthood stress on the total number of GABAA receptor binding sites for [3H]GABA in forebrain cortical regions was altered by early-life stress in both male and female mice, suggesting that the rapid adulthood stress response of GABAA receptors is affected by early-life experience. 7. Together these results show that GABAA receptors are sensitive to subtle changes in the environment in both early-life and adulthood and that these neurochemical responses to stress in adulthood are sex-dependent. The short and long-term stress-sensitivity of the GABAergic system implicates GABAA receptors in the non-genetic aetiology of psychiatric illnesses in which sex and stress are important factors.

No description available.

Thesis (Ph. D.)--Institute of Paper Chemistry, 1975. / Includes bibliographical references (p. 60-67).