The evolutionary origin of the vertebrate brain is still a major subject of debate. Its distinctive dorsal position and development from a tubular neuroepithelium are unique to the chordate phylum. Conversely, apical organs (AO) are larval sensory/neurosecretory centers found in many invertebrate taxa, including in animals without a brain. Previous studies have shown that AOs are specified by a conserved set of genes under the influence of Wnt signalling. Although most of these genes are expressed in chordate nervous systems (including vertebrates), no AOs have ever been described in this group of animals. Here we have exploited single-cell genomic approaches to characterize cells showing AO profiles in sea urchin (ambulacrarian), amphioxus (invertebrate chordate) and zebrafish (vertebrate chordate). This, in combination with co-expression analysis in amphioxus embryos, has allowed us to identify an active and dynamic anterior Gene Regulatory Network (aGRN) in the three deuterostome species. We have further discovered that as well as controlling AO specification in sea urchin, this aGRN is involved in the formation of the hypothalamic region in amphioxus and zebrafish. Using a functional approach, we find that the aGRN is controlled by Wnt signalling in amphioxus, and that suppression of the aGRN by Wnt overactivation leads to a loss of forebrain cell types. The loss of the forebrain does not equate to a reduction of neuronal tissue, but to a loss of identity, suggesting a new role for Wnt in amphioxus in specifically positioning the forebrain. We thus propose that the aGRN is conserved throughout bilaterians and that in the chordate lineage was incorporated into the process of neurulation to position the brain, thereby linking the evolution of the AO to that of the chordate forebrain. Overall design: Differential gene expression analysis between Azakenpaullone treated and untreated Brachiostoma lanceolatum embryos at 12 and 21 hours post fertilistaion (hpf) - 3 biological replicates per group