摘要：Tryptophan hydroxylase (TRH) is the rate limiting enzyme in the serotonin synthesis. CRISPR-Cas9 technology was used to generate seven indel TRH mutants in Daphnia magna. Mono-allelic indel TRH-/+ clones showed normal levels of serotonin, measured by both immunohistochemistry and mass spectrometry (LC-MS/MS), whereas bi-allelic indel TRH-/- clones showed no detectable levels of serotonin. Life history and behavioral responses of TRH-/- clones showed the anti-phenotype of those exposed to selective serotonin reuptake inhibitors (SSRI). Mutants lacking serotonin grew less and hence reproduced latter, produced smaller clutches of smaller offspring and responded to a greater extent to light than wild type individuals. Mono-allelic indel TRH-/+ individuals showed the intermediate phenotype. The SSRI fluoxetine enhanced offspring production in all clones and decreased the response to light only in those clones having serotonin, thus indication that behavioral effects of this drug in D. magna are associated to serotonin. Results obtained with the TRH mutants are in line with reported ones in TRH knockouts of Caenorhabditis elegans, Drosophila and mice, indicating that there is one gene encoding TRH, which is the serotonin limiting enzyme in both the central and the periphery nervous system in Daphnia and that deprivation of serotonin increases anxiety-like behavior.
