Thesis (Ph. D.)--West Virginia University, 2003. / Title from document title page. Document formatted into pages; contains viii, 153 p. : ill. (some col.). Vita. Includes abstract. Includes bibliographical references (p. 107-135).

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The mutant mouse stargazer shows both ataxia and absence epilepsy from P14 onwards. A PGR amplification strategy was utilised to identify adult (i.e. > 3 months) +/stg from +/+ mice, which share the same phenotype, for use for breeding purposes. The same technique was employed to identify +/+, +/stg and stg/stg neonates (i.e. < 7 days old) for cell culture purposes, since the stargazer phenotype is not apparent at this age. GABA(_A) receptor α(_6) subunit expression levels were significantly decreased in adult stargazer (stg/stg) cerebella when compared to control (+/+ and +/stg) cerebella. Interestingly, autoradiography using [(^3)H] Ro15-4513 revealed an apparent upregulation in α(_4)γ-containing receptors in the adult stargazer dentate gyrus. No significant differences in the expression of NMDAR subunits were detected between adult control and stargazer brain membranes. A significant decrease was observed in AMPAR subunit expression within the adult stargazer cerebellum, particularly with the GIuR2 subunit, which was reduced by 73 %. This decrease was replicated in cerebellar granule cells cultured from stargazer neonates, which also expressed at the cell surface only 18 % of the total GluR2 found in control granule cells. Inmunohistochemistry analyses using mouse anti-stargazin antibodies revealed stargazin to be found throughout the adult control brain, with highest levels of expression being within the hippocampus and cerebellum. Stargazin protein, however, was not expressed in adult stargazer forebrain nor in adult stargazer cerebellar membranes. Finally, immunoaffinity columns using the anti-stargazin antibodies were prepared and demonstrated that stargazin could be purified from adult control mouse brain extracts. Moreover, AMPAR subunits co-immunoprecipitated, indicating an association in vivo.

Thesis (doctoral)--University of Tartu, 2006. / Thesis is based on 3 papers.

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Dystroglycanopathies are muscular dystrophies caused by mutations in genes involved the in O-linked glycosylation of alpha-dystroglycan. Severe forms exhibit brain and ocular developmental abnormalities in addition to muscular dystrophy. While cerebellar dysplasia is a common feature of dystroglycanopathy, its pathogenesis has not been thoroughly investigated. Here we evaluate the role of dystroglycan during cerebellar development. Brain-selective deletion of dystroglycan does not affect overall cerebellar growth, but causes malformations associated with glia limitans disruptions and granule cell heterotopia that recapitulate phenotypes found in dystroglycanopathy patients. Cerebellar pathology in these mice is not evident until birth even though dystroglycan is lost during the second week of embryogenesis. The severity and spatial distribution of glia limitans disruption, Bergmann glia disorganization, and granule cell heterotopia rapidly increase during postnatal development. Astrogliosis becomes prominent at these same sites by the time cerebellar development is complete. Interestingly, there is spatial heterogeneity in the glia limitans and granule neuron migration defects that spares the tips of lobules IV-V and VI. The full spectrum of developmental pathology is caused by loss of dystroglycan from Bergmann glia, as neither granule cell- nor Purkinje cell-specific deletion of dystroglycan results in similar pathology. These data illustrate the importance of dystroglycan function in radial/Bergmann glia, but not neurons, during cerebellar histogenesis. The spatial heterogeneity of pathology shows that the dependence on dystroglycan is not uniform. Cognitive deficits are constant features of severe dystroglycanopathies, yet the precise molecular mechanism leading to neuronal dysfunction in these diseases is not known. Here, we show that dystroglycan interaction with dystrophin is required for the normal clustering of a subset of inhibitory synapses in Purkinje neurons. Using mouse models of dystroglycan mutants, we demonstrate that the number of gamma-aminobutyric acid receptor-containing synapses is significantly reduced in the absence of dystroglycan or portions of dystroglycan; a similar result is attained in dystrophin-deficient mice. Finally, we verify that the number of these receptors is retained when dystroglycan and dystrophin are preserved exclusively in Purkinje neurons. Our findings substantiate the notion that brain dystroglycan is important for neuronal function and suggest a molecular mechanism that may underline cognitive impairments in dystroglycanopathies.

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