Serotonergic neurons are the first brainstem neurons to emerge during early embryo development. Serotonin has been implicated in a wide variety of behaviors, physiological mechanisms and disease processes. Though most of what is known about serotonin involves its role in neuronal communication, studies have suggested that serotonin may also have non-neuronal functions during embryogenesis as a signaling molecule in the regulation of cleavage, morphogenesis and cell differentiation. Previous studies in our lab showed that serotonin levels peak immediately prior to gastrulation. In the present study I demonstrate that serotonin triggers primary invagination, the initial phase of gastrulation, in Lytechinus pictus sea urchin embryos. p-Chlorophenylalanine methyl ester (mCPA), an inhibitor of the rate-limiting enzyme for serotonin synthesis, tryptophan hydroxylase (TPH), blocks serotonin synthesis in vitro and embryos treated with mCPA fail to initiate gastrulation. Previous studies in the sea urchin identified TPH expression only in postgastrula embryos, which suggested that there was an additional enzyme present in the genome capable of synthesizing the serotonin necessary to initiate the gastrulation process. I identified a gene, Lp PAH/TPH, in L. pictus with homology to the dual function Drosophila Henna gene. In Drosophila, Henna is the source of non-neuronal serotonin. My expression studies of Lp PAH/TPH show that the mRNA is present in blastula and gastrula embryos prior to the expression of TPH, which strongly supports PAH/TPH-mediated synthesis of serotonin necessary for the initiation of gastrulation and any other non-neuronal functions in the sea urchin. Antagonists of 5-HT₂ and 5-HT₇ serotonin receptors block primary invagination. Furthermore, primary invagination blocked by mCPA could be rescued with agonists of 5-HT₂ and 5-HT₇ receptors and with downstream signaling molecules known to be linked to these receptors. In addition, mCPA blocks protein phosphorylation through protein kinase A and C, 5-HT₇ and 5-HT₂ receptor effectors, respectively. Taken together, these studies strongly suggest that serotonin mediates its effects on the gastrulation process in sea urchin embryos through a signaling mechanism that involves the 5-HT ₂ and/or 5-HT₇ serotonin receptors.