Lrrn2 marks rhombomere 4 motor neurons and the second branchial arch. In situ hybridisation with (A-D, F-H) Lrrn2, (E) RXR, (I) Tbx1, (J) CRABP1, (K) Dlx2 at HH14 (A-E) and HH18 (F-K). (A) Wholemount embryo shows Lrrn2 expression in two stripes in ventral r4 (black arrows) and in nearby cells within the presumptive BA2 region (red arrow). (B) Flatmounted hindbrain shows Lrrn2 expression restricted to two columns immediately alongside the ventral midline of r4, with a few scattered cells within the body of r4 (yellow arrows). (C) Transverse section through the hindbrain at the level of r4 shows expression in the outer layer (red arrowheads). Yellow arrows indicate more dorsal cells seen also in (B). (D) Next posterior transverse section of the same series as (C) shows staining in the ventrally located mesoderm (below red dashed line). (E) Neural crest marker RXR labels a dorsally migrating population lying above the same line. (F) At HH18, Lrrn2 is strongly expressed in BA2 but is essentially absent from BA1. Staining is also visible in scattered cells more dorsally in r2 and r4, as well as in the ventral motor column (black arrows) and ventral midbrain. (G, H) Adjacent coronal sections through the branchial arch region at HH18 shows Lrrn2 in the central core area of BA2 (red arrows). (I) Tbx1 expression labels the mesodermal core of the branchial arches; staining in the BA2 core area is indicated by a red arrow. (J) CRABP1 and (K) Dlx2 are neural crest markers and show a peripheral crescent of staining in the branchial arches, unlike Lrrn2. Scale bars: 100 μm (E, I-K). Courtesy of Robyn Quinlan and Anthony Graham.