Chapter 2 46 Figure 3. Cortical interneuron changes in 4H. (A-C) QPCR analysis shows PDPR, ITGA11 and ARX expression in 4H and control neurons. RNA was isolated from cortical neurons at day 18 of the differentiation protocol, which represents an immature state. In 4H neurons, ARX expression is significantly downregulated (C), while there is no change in PDPR or ITGA11 expression (A-B). Immunohistochemical analysis for GAD65/67 on primary cortical tissue of a control individual and two 4H patients showed that the pan interneuron marker is increased in 4H brain tissue (D-E). In iPSC-derived cortical neurons the proportion of glutamatergic and GABAergic cells (based on GAD65/67 and MAP2 staining) was not changed between control and 4H cells (F). A significant increase in the number of CTIP2 and GAD65/67 double positive cells was observed at 14 weeks post plating (G). *** = P < 0.001, * = P < 0.05, AU = arbitrary units. Open circles represent average per individual patient/control, filled circles represent the mean per genotype. Error bars represent standard error of the mean (SEM). Bars in F represent average proportion of GABAergic (dark colours) and glutamatergic (light colours) neurons, with circles representing values of individual lines. Scale bar = 100 µm.
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