Differential Molecular Responses of Zebrafish Larvae to Fluoxetine and Norfluoxetine

The occurrence of psychopharmaceuticals in aquatic ecosystems is a growing problem. Fluoxetine (FL) and its metabolite norfluoxetine (NF) are selective serotonin reuptake inhibitors. Although they may be potentially harmful to non-target species, available knowledge on the effects of NF is sparse, relative to FL. This study aimed at contributing to the body of knowledge about the modes-of-action (MoA) of these compounds and their underlying mechanisms eliciting hazardous effects during the early development of the teleost model zebrafish (Danio rerio). One hour post-fertilisation (hpf), embryos were exposed up to 80 hpf to these compounds at levels found in surface waters and higher (FL, 0.0015 and 0.05 µM; NF, 0.00006 and 0.0014 µM). Developmental anomalies were observed at 8, 32 and 80 hpf. Larvae were collected at 80 hpf to assess the expression of 34 genes related to FL and NF MoA and metabolism, using qPCR (quantitative reverse transcription PCR). Results showed that both compounds elicited an increased frequency of embryos exhibiting abnormal pigmentation, relative to controls. Gene expression alterations were more pronounced in FL- than in NF-exposed larvae. Cluster Analysis revealed two groups of genes discriminating between the drugs. for their marked opposing responses. Globally, downregulation of gene expression was typical of FL, whilst upregulation or no alteration was found for NF. These clusters identified were linked to the adrenergic pathway and to the retinoid and peroxisome proliferator-activated nuclear receptors. Overall, our data contradict the prevailing notion that NF is more toxic than FL and unveiled the expression levels of genes drd2b, 5-ht2c and abcc2 as possible markers of exposure to FL.