Role of Gli3 in the developing mouse forebrain
The mammalian forebrain, which consists of the telencephalon and the diencephalon, is responsible for many higher cognitive functions such as thinking, learning and memory. The cerebral cortex, which is important for language and processing information, is located in the dorsal portion of the telencephalon. The basal ganglia, which are important for movement, are located in the ventral telencephalon. Many genes are involved in patterning and the development of the forebrain. One gene that appears to be crucial for forebrain development is Gli3. Gli3 has been shown to work as both a transcriptional activator and a repressor of the Sonic Hedgehog (Shh) signalling pathway in the developing spinal cord and limb buds. In the telencephalon, Shh has been shown to be important for induction of ventral cell fate, but the exact function of Gli3 in the forebrain and the interactions between Gli3 and Shh are still obscure. Previous studies have shown that Gli3 is required for the formation of the cortical hem area of the telencephalon, which does not form in Gli3Xt/Xt mutant mice lacking functional Gli3. The residual dorsal telencephalon of the Gli3Xt/Xt mutants is partially ‘ventralized’. The main aim of this study was to re-examine the developing forebrain of Gli3Xt/Xt mouse mutants to gain insight into the function of Gli3 during forebrain development. In this thesis, the expression of Gli3 mRNA and protein was examined in the E12.5 and E14.5 wild type telencephalon. The highdorsal-to-lowventral expression pattern of Gli3 corresponds to severedorsal-to-mildventral defects observed in the Gli3Xt/Xt mutants. The ratios between the levels of the cleaved and full length isoforms of Gli3 in dorsal and ventral telencephalon resemble those described in dorsal and ventral spinal cord and in the anterior and posterior limb bud, respectively, suggesting Gli3 in the dorsal telencephalon may act as a repressor of the Shh signalling pathway. The total amount and the ratios of the two isoforms of Gli3 protein were examined in Shh and Foxg1 null mice, which lack ventral telencephalon. The results obtained agree with a role of Gli3 as a repressor of the Shh pathway in the dorsal telencephalon. The forebrains of Gli3Xt/Xt mutants were analysed systematically both anatomically and by molecular markers in this thesis. The border between the telencephalon and the diencephalon was delineated in the Gli3Xt/Xt mutants by using a combination of markers expressed in different areas within the forebrain. This lead to the observation that the previously reported ‘ventralization’ only occurred in the very rostral telencephalic sections of the Gli3Xt/Xt mutant embryos, suggesting a possible shape change of the Gli3Xt/Xt telencephalon. To examine the possible causes of the significant size reduction of Gli3Xt/Xt mutant telencephalon compared to wild type telencephalon from E10.5, cell proliferation and cell death properties studies were undertaken. The changes observed were not sufficient to explain the phenotypic differences between the Gli3Xt/Xt mutant and the wild type embryos indicating that they might be the result of an early patterning defect. The dorsal telencephalon is severely reduced in volume at both E12.5 and E10.5, containing cells from adjacent eminentia thalami, probably due to the loss of the dorso-medial telencephalon. Large clusters of eminentia thalami cells were observed at later developmental stages, when the neocortex becomes highly disorganized, forming rosettes comprising mainly neural progenitors. These results suggest Gli3 is important for the formation of an intact telencephalic-diencephalic boundary and for preventing the abnormal location of diencephalic cells in the dorsal telencephalon. The volume of Gli3Xt/Xt ventral telencephalon was increased compared to that of the wild types at E10.5, but became smaller than that of the wild type littermates at E12.5. This might have been the result of a combination of more cells exiting the cell cycle and increased cell death observed in the Gli3Xt/Xt ventral telencephalon at E10.5, suggesting Gli3 regulates cell differentiation and cell death properties at this age and brain region. The significant expansion of rostro-ventral telencephalon observed in the Gli3Xt/Xt mutant might correlate with the expansion of Fgf8 expression and this hypothesis has been tested in this thesis.