Genetic analysis of the roles of Hh, FGF8, and nodal signaling during catecholaminergic system development in the zebrafish brain.

CNS catecholaminergic neurons can be distinguished by their neurotransmitters as dopaminergic or noradrenergic and form in distinct regions at characteristic embryonic stages. This raises the question of whether all catecholaminergic neurons of one transmitter type are specified by the same set of factors. Therefore, we performed genetic analyses to define signaling requirements for the specification of distinct clusters of catecholaminergic neurons in zebrafish. In mutants affecting midbrain- hindbrain boundary (MHB) organizer formation, the earliest ventral diencephalic dopaminergic neurons appear normal. However, after 2 d of development, we observed fewer cells than in wild types, which suggests that the MHB provides proliferation or survival factors rather than specifying ventral diencephalic dopaminergic clusters. In hedgehog (Hh) pathway mutants, the formation of catecholaminergic neurons is affected only in the pretectal cluster. Surprisingly, neither fibroblast growth factor 8 (FGF8) alone nor in combination with Hh signaling is required for specification of early developing dopaminergic neurons. We analyzed the formation of prosomeric territories in the forebrain of Hh and Nodal pathway mutants to determine whether the absence of specific dopaminergic clusters may be caused by early patterning defects ablating corresponding parts of the CNS. In Nodal pathway mutants, ventral diencephalic and pretectal catecholaminergic neurons fail to develop, whereas both anatomical structures form at least in part. This suggests that Nodal signaling is required for catecholaminergic neuron specification. In summary, our results do not support the previously suggested dominant roles for sonic hedgehog and Fgf8 in specification of the first catecholaminergic neurons, but instead indicate a novel role for Nodal signaling in this process.