The midbrain–hindbrain boundary region is defined by cellular heterogeneity

We identified two distinct isthmic cell populations – isthmic midbrain (IsMB) and isthmic hindbrain (IsHB) – that share expression of canonical isthmic genes, but that differ in their expression of midbrain vs hindbrain genes. We also uncovered a previously unrecognized heterogeneity within the IsHB, reflecting a canonical fgf8-expressing population anteriorly (IsO/r0a), and a novel fgf8-negative population posteriorly (r0p).

We find that inhibition of Fgf signaling disrupts formation of the isthmic region, leading to loss of cell populations except a residual population categorized by a mixed neural identity. Conclusions: Using transcriptional and epigenetic characterization, we expand on prior anatomical and genetic analyses of the isthmic region to refine our understanding of its cellular organization and demonstrate that it consists of several subdomains.