Ontogeny and Innervation Patterns of Dopaminergic, Noradrenergic, and Serotonergic Neurons in Larval Zebrafish

We report the development of aminergic neurons from 0-10 days postfertilization (dpf) in zebrafish (Danio rerio). This study was prompted by the lack of information regarding patterns of spinal aminergic innervation at early stages, when the fish are accessible to optical, genetic, and electrophysiological approaches toward understanding neural circuit function. Our findings suggest that aminergic populations with descending processes are among the first to appear during development. Descending aminergic fibers, revealed by antibodies to tyrosine hydroxylase (TH) and serotonin (5-hydroxytryptamine; 5-HT), innervate primarily the ventral (TH, 5-HT), but also the dorsal (5-HT) aspects of the spinal cord by 4 dpf, with the extent of innervation not changing markedly up to 10 dpf. By tracking the spatiotemporal expression of TH, 5-HT, and dopamine beta hydroxylase reactivity, we determined that these fibers likely originate from neurons in the posterior tuberculum (dopamine), the raphe region (5-HT) and, possibly, the locus coeruleus (noradrenaline). In addition, spinal neurons positive for 5-HT emerge between 1-2 dpf, with processes that appeared to descend along the ventrolateral cord for only 1-2 muscle segments. Their overall morphology distinguished these cells from previously described "VeMe" (ventromedial) interneurons, which are also located ventromedially, but have long, multisegmental descending processes. We confirmed the distinction between spinal serotonergic and VeMe interneurons using fish genetically labeled with green fluorescent protein. Our results suggest that the major aminergic systems described in adults are in place shortly after hatching, at a time when zebrafish are accessible to a battery of techniques to test neuronal function during behavior.