Fig. 2.

Loss of Isl1 leads to TCA pathfinding defects in the prethalamus and thalamus. (A) Immunohistochemistry for GFP on coronal sections of control (Gbx2^GFP;Dlx5/6-Cre; Isl1^F/+) (A1-A6) and mutant (Gbx2^GFP;Dlx5/6-Cre; Isl1^F/F) (A7-A12) embryos at E16.5. The red arrows in A10 and A11 indicate abnormal TCA projections in the prethalamus of Isl1 mutant embryos. White asterisks in A10, A11, and A12 indicate abnormally large bundles of TCAs crossing the prethalamus and entering the ventral telencephalon in Isl1 mutants compared to controls. GFP⁺ TCAs misrouted caudally to the habenula (white arrows in A9). Red asterisks in A10-A12 mark abnormal fasciculation of TCAs within the thalamus. GFP⁺ TCAs misrouted ventrally in the ventral telencephalon (A7-A11, red arrowheads) and hypothalamus (A12, yellow arrowhead). In the cortex, Isl1 mutant TCAs were shorter than those of control embryos (A7-A11, white arrowheads). (A13) Examples of the measurements taken for the quantification. Yellow dashed lines in A4, A5, A10, A11 indicate the position of the lines used to quantify the number of GFP⁺ fluorescent signals crossing the prethalamus. (n = 5 embryos; Student’s t-test; *p < 0.001). (B) Double immunofluorescence for GFP and NF-M on sagittal sections of control and Isl1 mutant embryos at E14.5. Control TCAs extended in parallel and oriented ventrolaterally, forming a thin fasciculus in the thalamus and prethalamus, including the TRN. In contrast, Isl1 mutant TCAs formed an abnormally thick fasciculus and extended randomly in the thalamus (white arrowheads) and prethalamus (white arrows). (C) Double immunofluorescence for GFP and NF-M on sagittal sections of control and Isl1 mutant embryos at E18.5. In the thalamus, GFP⁺ TCAs formed abnormally thick axon bundles orienting randomly and misrouting caudally into the pretectum (white arrowheads in C4’,C4”). Cx, cerebral cortex; Eth, epithalamus; Hyp, hypothalamus; Prt, pretectum; Pt, prethalamus; Th, thalamus. Scale bars, 200 μm