Research Summary:
During embryogenesis, neurons learn of their cell-fates and are assigned to specific brain nuclei (nucleogenesis), which organize brain connectivity and function. Little is known about how such complex patterning is achieved during vertebrate development. We study these processes in the mouse and chick midbrain, where clinically important nuclei such as the oculomotor complex (involved in eye movements) and midbrain dopaminergic neurons (involved in Parkinson's disease and addictive behaviors) reside. I have previously shown that the signaling molecule, Sonic Hedgehog, is sufficient to specify the entire ventral midbrain (organogenesis). We are now examining the downstream transcriptional mechanisms that allow this single molecule to specify multiple cell and nuclear fates. Experimental approaches include in vivo gene misexpression by electroporation (in chick and mouse), protein misexpression using coated-beads, and gene silencing by novel strategies such as RNA interference and morpholino oligonucleotides and microarray analyses. Manipulations in the chick are complemented with gene expression analyses of mutant chicks and mice with defects in midbrain patterning.

- In vivo RNA intereference in a chick midbrain. The left side of the brain is the control. Rnai constructs targeted against gli2 (blue) reduce its expression and expand a FOXA2 (brown) as a result. In vitro gene misexpression in mice - Misexpression of GFP in a mouse embryo (top) followed by an emergence of embryonic midbrain pattern after 2 days in vitro. Agarwala Laboratory