Fluoxetine is one of the most prescribed psychotropic medications, and is an agent of increasing interest for environmental toxicology. Fish and other aquatic organisms are excellent models to study neuroactive small molecules like fluoxetine. However, prone to variance due to experimental factors, data obtained in these models need to be interpreted with caution, using proper experimental protocols, study designs, validated endpoints as well as well-established models and tests. Choosing the treatment protocol and dose range for fluoxetine and other serotonergic drugs is critical for obtaining valid test results and correct data interpretation. Here we discuss the value of aquatic models to study fluoxetine effects, based on prior high-quality research, and outline the directions of future translational studies in the field. We review fluoxetine-evoked phenotypes in acute vs. chronic protocols, discussing them in the contact of complex role of serotonin in behavioral regulation. We conclude that zebrafish and other aquatic models represent a useful in-vivo tool for fluoxetine pharmacology and (eco)toxicology research.