Migration of dI5 Reelin-Lmx1b-Zfhx3 and Disabled-1-Lmx1b-Zfhx3 neurons contribute to the superficial dorsal horn and lamina V

In adult superficial dorsal horn, 90% of Reelin (Reln+) and 70% of Disabled-1 (Dab1+) neurons co-express the transcription factor LIM-homeobox 1-beta (Lmx1b+) and therefore are glutamatergic neurons. Here we asked if embryonic Reln+Lmx1b+ and Dab1+Lmx1b+ dorsal horn neurons are derived from Lmx1b-expressing early-born dI5 or late-born dILB dorsal neurons. On Embryonic day (E)11.5, Reln+ and Dab1+ neurons appear to be part of the migration of early-born dI5 Lmx1b-expressing neurons. Between E12.5-E15.5, the lateral Reln+Lmx1b+ and Dab1+Lmx1b+ neurons migrate circumferentially along the rim of what will become the superficial dorsal horn, whereas medial Reln+Lmx1b+ and Dab1+Lmx1b+ neurons move into the dorsal midline and then migrate into lamina V. The small, late-born dILB Reln+Lmx1b+ and Dab1-Lmx1b+ neurons fill the superficial dorsal horn. In Reln mutants, large Dab1+Lmx1b+ neurons were mispositioned in lamina I and at the border between the superficial and deep dorsal horn. To confirm the identity of the circumferential and midline Reln+Lmx1b+ and Dab1+Lmx1b+ neurons, we asked if they expressed the transcription factor Zfhx3, a marker of dI5 projection neurons. We detected examples of Reln+Lmx1b+Zfhx3+ and Dab1+Lmx1b+Zfhx3+ projection neurons that migrated along the outer rim of the superficial dorsal horn and others that migrated from the midline into lamina V. Taken together, our study demonstrates that the larger Reln+Lmx1b+Zfhx3+ and Dab+Lmx1b+Zfhx3+ neurons represent two subsets of dI5 projections neurons, whereas smaller Reln+Lmx1b+ and Dab1+Lmx1b+ neurons concentrated in lamina II are likely dILB interneurons.