Using High-Performance Liquid Chromatography to Study Alterations in Biogenic Amines in Drosophila Models of Brain Disorders

Many neuropsychiatric and neurodegenerative disorders are characterized by disruptions in brain neurotransmitters, including alterations in the levels of the biogenic amines (BAs) dopamine (DA) and serotonin (5-hydroxytryptamine; 5-HT), as well as in their release or reuptake. These neurochemical changes correlate with distinct key behavioral phenotypes, such as locomotor deficits, sleep fragmentation, and altered stress responses, among others. Because the principal BA systems interact, disease models frequently show coordinated shifts across multiple amines rather than isolated changes in a single neurotransmitter. Therefore, accurate, simultaneous quantification of several BAs is essential for interpreting behavioral and genetic findings. Drosophila melanogaster offers a genetically tractable system for modeling such disorders and neurotransmitter changes, but BAs occur in fly brain tissue at very low, nanomolar-to-femtomolar concentrations, and the small size of the Drosophila central nervous system makes their detection technically challenging. This study describes a protocol to detect and quantify DA, 5-HT, and octopamine in fly brains using high-performance liquid chromatography coupled with electrochemical detection (HPLC-ECD). HPLC-ECD was applied to detect changes in brain DA and 5-HT in a fly model of Parkinson's disease based on dopaminergic overexpression of human alpha-synuclein. Applying this protocol to the alpha-synuclein Parkinson's disease model reveals significantly elevated dopamine, but unchanged serotonin, relative to controls, consistent with early compensatory dopaminergic mechanisms. This approach provides a sensitive, low-cost, and readily accessible method for multiplexed BA quantification from limited Drosophila tissue. It also enables systematic screening of genetic modifiers and candidate therapeutics targeting monoaminergic pathways relevant to human brain disorders.