Hoxb1 is required for the glutamatergic identity of dB2 neurons in r4.

<p>(<b>A–F</b>) Transverse sections through r4 of E12.5 control and Hoxb1^−/− embryos immunolabeled for Phox2b and the glutamatergic marker Vglut2. The expression of Phox2a in the dB2 domain is lost or dramatically reduced in the Hoxb1^−/− embryo compared to the control (A, B). Vglut2 staining in the cellular domain occupied by dB2 and dB3 neurons is unchanged in the Hoxb1^−/− embryo compared to the control embryo (C, D). Panels E and F are composite of images A–D that were separated for clarity. (<b>G, H</b>) Transverse sections through r4 of E12.5 control and Hoxb1^−/− embryos immunolabeled for VGLUT2 (green) and Gad1 (red). The complementary expression of Vglut2 and Gad1 is unperturbed in the absence of Hoxb1. (<b>I–M</b>) Transverse sections through r4 of E12.5 control and Hoxb1^−/− embryos immunolabeled for Tlx3 and the glutamatergic marker Vglut2. Tlx3 is ectopically expressed in the dB2 domain in the Hoxb1^−/− embryo compared to the control (I, J). Again, the staining for Vglut2 in the cellular domain occupied by dB2 and dB3 neurons is unchanged in and the Hoxb1^−/− embryo compared to the control embryo (C, D). Panels M and N are composite of images I–L that were separated for clarity. (<b>O, P</b>) Transverse sections through r4 of E12.5 control and Hoxb1^−/− embryos immunolabeled for Tlx3 (green) and Gad1 (red). Tlx3 is ectopically expressed in the dB2 domain while the expression of Gad1 remains complementary to this domain in the Hoxb1^−/− embryo. This suggests that the ectopic expression of Tlx3 in the dB2 domain does not perturb the neurotransmitter phenotype from this cellular domain. Scale bars (A–P = 50 µm).