Factors regulating the migration and positioning of interneurons during cortical development
AuthorNg, Hui Xuan
AffiliationFlorey Department of Neuroscience and Mental Health
MetadataShow full item record
Document TypePhD thesis
Access StatusOnly available to University of Melbourne staff and students, login required
© 2015 Dr. Hui Xuan Ng
Interneurons of differing birth-dates migrate in varying routes from the ventral telencephalon to finally reside in specific positions within the six-layered cortex during development. Work during my honours degree revealed the varying migratory schedules used by early-, mid- and late-born interneurons to reach a final position. This project further explores the preliminary findings with a global aim to identify the intrinsic and extrinsic factors that regulate migration and positioning of interneuron birth-date cohorts. The differing final positioning schedules lead to the hypothesis of varying intracortical migratory behaviour among birth-date cohorts. This was tested in Chapter 3 using MGE explants and organotypic cortical slice co-culture system where the contribution of the cortical substrate was concomitantly investigated. Early- and late-born interneurons prefer radial migration from the pial and ventricular surface, respectively and this highlights the combination of intrinsic (birth-date) and extrinsic regulation (cortical substrate) of migratory behaviour. To further explore the intrinsic regulatory mechanisms for migration, Chapter 4 aims to develop techniques to analyse gene expression of birth-date cohorts along the migratory routes. A protocol that combines EdU labelling, FACS and RNA extraction was developed, however, gene expression can only be analysed via candidate gene screening. A microarray between early- and late-born MGE interneurons was conducted to identify the candidate genes. Early-born interneurons show upregulation of patterning genes while late-born interneurons the cell surface receptors, indicative of an ability to respond rapidly to extrinsic guidance cues and this may mediate the shortest time-frame for intracortical migration. The final placement of mid-born interneurons coincides temporally and spatially with the formation of the barrel field. To understand mid-born interneuron recruitment and subtype specification, two cortical mutants with varying barrel field defects were characterized in Chapter 5. The study revealed altered recruitment of mid-born interneurons in adenylyl cyclase 1 (AC1) knockout (KO) that leads to the loss of parvalbumin-positive (PV+) interneurons in layer IV; whereas phospholipase C-beta1 (PLC-β1) KO develops a loss of PV+ interneurons at adult stage that coincides with an age-dependent disruption of the barrel field cytoarchitecture. The findings suggest the role of AC1 and PLC-β1 and highlight the susceptibility of PV+ interneurons to perturbation during migration. Previous characterization of the human cortex at mid-gestation showed a large number of calretinin-positive (CR+) cells with a migratory morphology, however, migration remains unexplored due to the lack of tool to label human interneurons for live imaging. Chapter 6 uses in vitro dissociated primary neuronal culture and in situ hybridization to identify new interneuron marker. The study revealed distal-less homeobox 5 (Dlx5) expression at mid-gestation and that Dlx5/6-Cre can label at least two subpopulations of Dlx5/6+ cells with varying morphology and coexpression of CR. Together, this study has advanced understanding of the factors that regulate interneuron migration and positioning. Birth-date is a criterion to be taken into account in future investigation of migration. The birth-date specific gene expression patterns may correlate with the pathophysiology of clinical disorders related to GABAergic dysfunctions. Loss of AC1 or PLC-β1 functions contributes to the abnormalities in mid-born cohort recruitment and interneuron subtype specification. The ability to label human interneuron for live imaging will further advance study of human interneuron migration.
Keywordsinterneuron; migration; positioning; birth-date; genes; cortical substrate; human
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