Role of intermediate filaments and expression of GFAP and S-100 in astrocytes

Sammanfattning: Astrocytes participate in a variety of important physiologic and pathologic processes in the central nervous system (CNS). One of the most remarkable characteristics of astrocytes is their vigorous response to CNS injuries. Reactive astrocytosis is characterized by increased production of intermediate filament (IF) proteins, such as glial fibrillary acidic protein (GFAP) and vimentin. Such conditions also involve the overexpression of a number of neurotrophic/neuroprotective factors. These alterations contribute significantly to the pathogenesis of CNS injuries, i.e., after ischemia, trauma, seizures and hypoxia. The S-100 protein, which is abundant in astrocytes, has been shown to exert neurotrophic effects. However, only a few studies on the involvement of S-100 has been reported in neurodegenerative disorders. With regard to IF proteins, the lack of knowledge about the function of astrocytic intermediate filaments in normal cells precludes the understanding of the functional implications of certain cytoskeletal changes in reactive astrocytes. The thesis aims to reach a better understanding of the functions of S-100, GFAP and vimentin. An animal model for temporal lobe epilepsy induced by systemic kainic acid (KA) administration and primary cultures of astrocytes prepared from mice lacking GFAP and /or vimentin were used.The immunochemical results show that systemic KA administration induced a reactive gliosis as revealed by increased GFAP content in the hippocampus and the amygdala/pyriform cortex which was preceded by a decrease in NSE (an indication of neuronal density), while the gliosis in the frontal cerebral cortex and the striatum was not preceded by changes in NSE. These events were paralleled by a longlasting decrease in tissue levels of S-100 and a transient increase in cerebrospinal fluid levels of S-100. Immunocytochemically, the reactive astrocytes with overexpressed S-100 immunoreactivity were observed in the layers bordering the regions with a high degree of neuronal degeneration in the hippocampus 3 and 7 days after KA. Regions with a high degree of neuronal degeneration, such as the CA3 pyramidal layer and the hilus of the dentate gyrus showed a loss of both GFAP and S-100 immunoreactivity, suggesting a damage to or loss of astrocytes. S-100 immunoreactivity was also elevated at 3-7 days in a region with axonal sprouting, somewhat distant from the lesioned neurons, i.e. the inner third of molecular layer of the dentate gyrus. Together with the observation that tissue S-100 levels were increased in association with normalised/recovered NSE in the hippocampus at day 27, the present results would give support to the proposed role of S-100 as a neurotrophic factor. The importance of astrocytic IF proteins GFAP and vimentin for astrocyte function was studied by investigating primary cultures of astrocytes from GFAP-/- and/or vimentin -/- mice. A reduction in the GFAP expression led to an increase in glutamine levels of the primary cultured astrocytes. This was not dependent on the presence of vimentin. The total absence of IFs (GFAP-/-vim-/-) induced a substantial reduction of the regulatory volume decrease (RVD)-related taurine release in astrocytes. However, the stimulated taurine efflux was not significantly affected in GFAP-/- or vimentin-/- astrocytes which have decreased amount of IFs or differences in their composition. These results implicate an involvement of IFs in astrocyte volume regulation and in the control of glutamine levels. The effects of IFs may not be only a reflection of generally altered metabolism of astrocytes, since the deficiency of IFs does not alter normal functions of astrocytes with respect to glucose, ascorbate and taurine transport under normal culturing conditions.