Figure 2

Effect of ketamine on protein levels of the GABAergic lineage markers Nkx2.1, DLX1 and LHX6. Quantification by western blotting of Nkx2.1 (a), DLX1 (b) and LHX6 (c) levels in cultured E15 GEs after 6 and 24 h of ketamine treatment (100 μM). Double labeling of cleaved caspase-3- (d) and Nkx2.1-positive cells (e) in the developing cortex of E15 brain slices after a 24-h treatment with ketamine (100 μM). Note that Nkx2.1 labeling partially overlaps with the cleaved caspase-3 signal (f), which is preferentially localized in the previously characterized migratory routes of GABAergic precursors (dashed arrow). Visualization of cleaved caspase-3 and Nkx2.1 double-labeled cells in the developing cortex from control (g) and ketamine-treated (h) E15 slices. Note the strong co-localization of the two signals after ketamine exposure. The total number of nuclei was visualized by Hoechst labeling. (i) High magnification confocal acquisition visualizing Nkx2.1 cells immunopositive (arrows) or immunonegative (arrow heads) for the cleaved caspase-3. (j) Quantification of the effect of a 24 h (100 μM) ketamine exposure on Nkx2.1/cleaved caspase-3 immunoreactive cells. For western blot studies, each value represents the mean (±S.E.M.) of at least six independent experiments. For cell quantification, each value represents the mean (±S.E.M.) of 18 ROI from 6 different slices. *P<0.05; **P<0.01 for ketamine versus control groups, Mann–Whitney U-test