Fig. 1: Workflow of the psychotropic drug-exposed scRNASEQ study in adult Drosophila brain. | Molecular Psychiatry

Fig. 1: Workflow of the psychotropic drug-exposed scRNASEQ study in adult Drosophila brain.

From: The effects of methylphenidate and atomoxetine on Drosophila brain at single-cell resolution and potential drug repurposing for ADHD treatment

Fig. 1

The workflow of our drug-exposed scRNASEQ study contains two parts. The first part (shown in subfigure A–C) is to conduct a behavioral activity assay in wild-type flies using methylphenidate (MPH) and atomoxetine (ATX) to generate hyperactivity-like behavior, which is video recorded and subsequently used to analyze locomotor activity. The second part (shown in subfigure D–F) is to conduct single-cell RNA sequencing and data analysis, which aims to reveal the effects of the aforementioned drugs at single-cell resolution. A Wild-type flies were fed with different drugs (ATX, MPH, or control) using a modified capillary feeder assay (CAFE). B The locomotor activity of each fly was recorded by camera under different conditions, and each batch was recorded for 2.5 h. Two experiments were performed separately on July 12th (replicate 1) and August 10th 2021 (replicate 2). C Analysis of the video recordings. EasyFlyTracker tracked and calculated the locomotor activity of different treatments. There are two main metrics used to calculate the locomotor activity: ① the average distance traveled per fly in a 10-min time period (2 replications); and ② the average distance traveled by each fly in a 10-min time period at each time point during the entire video. Both were significantly (** 0.001\( < \) P \(\le\)0.01) increased in the MPH-treated or ATX-treated groups as compared with the control group throughout the 2.5-h video. D A total of 20 male fly brains were collected and dissociated to generate single-cell suspensions for each sample. Each replicate contained three samples exposed to MPH, ATX, or control treatment. E After cell dissociation, counts larger than 500 live cells/μL were used to prepare 10X Genomic sequencing libraries. F Analysis of scRNASEQ data for all samples contains three different parts: ① clustering and cell type identification; ② differential gene expression analysis and drug-exposed cell type identification; and potential gene analysis for drug repurposing.

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