In this study we analyze 5-hydroxytryptamine [5-HT]; serotonin) signaling in zebrafish, an increasingly popular vertebrate disease model. We compare and contrast expression of the 5-HT transporter genes slc6a4a and slc6a4b, which identify 5-HT-producing neurons and three novel 5-HT receptors, htr1aa, htr1ab, and htr1bd. slc6a4a and slc6a4b are expressed in the raphe nuclei, retina, medulla oblongata, paraventricular organ, pretectal diencephalic complex, and caudal zone of the periventricular hypothalamus, in line with the expression profiles of homologues from other vertebrates. Our analysis of serotonin transporter (SERT)-encoding genes also identifies parallel genetic pathways used to build the 5-HT system in zebrafish. In cells in which 5-HT is synthesized by tph1, slc6a4b is used for re-uptake, whereas tph2-positive cells utilize slc6a4a. The receptors htr1aa, htr1ab, and htr1bd also show widespread expression in both the larval and adult brain. Receptor expression is seen in the superior raphe nucleus, retina, ventral telencephalon, optic tectum, thalamus, posterior tuberculum, cerebellum, hypothalamus, and reticular formation, thus implicating 5-HT signaling in several neural circuits. We also examine larval brains double-labeled with 5-HTergic and dopaminergic pathway-specific antibodies, to uncover the identity of some 5-HTergic target neurons. Furthermore, comparison of the expression of transporter and receptor genes also allows us to map sites of autoreceptor activity within the brain. We detect autoreceptor activity in the pretectal diencephalic cluster (htr1aa-, htr1ab-, htr1bd-, and slc6a4a-positive), superior raphe nucleus (htr1aa-, htr1ab-, and slc6a4a-positive), paraventricular organ (htr1aa-, htr1ab-, htr1bd-, and slc6a4b-positive), and the caudal zone of the periventricular hypothalamus (htr1ab- and slc6a4b-positive).