Name: Analyzing the Effect of Reserpine on the Sleep Behavior of Drosophila melanogaster Using SCAMP2020 Data Package in MATLAB
Uploaded: 2023-10-20T14:15:00
Description: Watch this Scientific Journal Video about Analyzing the Effect of Reserpine on the Sleep Behavior of Drosophila melanogaster Using SCAMP2020 Data Package in MATLAB at JoVE.com

analyzing the effect of reserpine on the sleep behavior of drosophila melanogaster using scamp2020 data package in matlab

Fremme af behandlinger af søvnforstyrrelser: En Drosophila-baseret screeningstilgang

Sleep, one of the essential behaviors necessary for human survival, is characterized by two main states: rapid eye movement (REM) sleep and non-rapid eye movement (NREM) sleep1. NREM sleep comprises three stages: N1 (the transition between wakefulness and sleep), N2 (light sleep), and N3 (deep sleep, slow wave sleep), representing the progression from wakefulness to deep sleep1. Sleep plays a crucial role in both physical and mental health2. However, aging reduces total sleep duration, sleep efficiency, slow-wave sleep percentage, and REM sleep percentage in adults3. Older individuals tend to spend more time in light sleep compared to slow-wave sleep, making them more sensitive to nocturnal awakenings. As the number of awakenings increases, average sleep time decreases, resulting in a fragmented sleep pattern in the elderly, which may be associated with excessive excitation of Hcrt neurons in mice4. Additionally, age-related declines in circadian mechanisms contribute to an earlier shift in sleep duration5,6. In combination with physical illness, psychological stress, environmental factors, and medication use, these factors make older adults more susceptible to sleep disorders, such as insomnia, REM sleep behavior disorder, narcolepsy, periodic leg movements, restless legs syndrome, and sleep-disordered breathing7,8.
Epidemiological studies have shown that sleep disorders are closely linked to chronic diseases in the elderly9, including depression10, cardiovascular disease11, and dementia12. Addressing sleep disorders plays a crucial role in improving and treating chronic diseases and enhancing the quality of life for older adults. Currently, patients primarily rely on drugs such as benzodiazepines, non-benzodiazepines, and melatonin receptor agonists to enhance sleep quality13. However, benzodiazepines can lead to downregulation of receptors and dependence after long-term use, causing severe withdrawal symptoms upon discontinuation14,15. Non-benzodiazepine drugs also carry risks, including dementia16, fractures17, and cancer18. The commonly used melatonin receptor agonist, ramelteon, reduces sleep latency but does not increase sleep duration and has hepatic function-related concerns due to extensive first-pass elimination19. Agomelatine, a melatonin receptor agonist and serotonin receptor antagonist, improves depression-related insomnia but also poses a risk of liver damage20. Consequently, there is an urgent need for safer drugs to treat or alleviate sleep disorders. However, current drug screening strategies, based on molecular and cellular experiments combined with automated systems and computer analysis, are expensive and time-consuming21. Structure-based drug design strategies, relying on receptor structure and properties, require a clear understanding of receptor three-dimensional structure and lack predictive capabilities for drug effects22.
In 2000, based on the sleep criteria proposed by Campbell and Tobler in 198423, researchers established simple animal models to study sleep24, including Drosophila melanogaster, which exhibited sleep-like states25,26. Despite anatomical differences between Drosophila and humans, many neurochemical components and signaling pathways regulating sleep in Drosophila are conserved in mammalian sleep, facilitating the study of human neurological diseases27,28. Drosophila is also extensively used in circadian rhythm studies, despite differences in core oscillators between flies and mammals29,30,31. Therefore, Drosophila serves as a valuable model organism for studying sleep behavior and conducting sleep-related drug screening.
This study proposes a cost-effective and simple phenotype-based approach for screening small-molecule drugs to treat sleep disorders using aged flies. Sleep regulation in Drosophila is highly conserved25, and the decline in sleep observed with age may be reversible through drug administration. Thus, this sleep phenotype-based screening method can intuitively reflect drug efficacy. We feed the flies with a mixture of the drug under investigation and food, monitor and record sleep behavior using the Drosophila Activity Monitor (DAM)32, and analyze the acquired data using the open-source SCAMP2020 data package in MATLAB (Figure1). Statistical analysis is performed using statistics and graphing software (see Table of Materials). As an example, we demonstrate the effectiveness of this protocol by presenting experimental data on Reserpine, a small-molecule inhibitor of the vesicular monoamine transporter reported to increase sleep33. This protocol provides a valuable approach to identify drugs for treating age-related sleep problems.

Forfatter Spotlight: Overvindelse af udfordringer i Drosophila-søvnmåling ved hjælp af DAM-system 

High-throughput Small Molecule Drug Screening for aldersrelaterede søvnforstyrrelser ved hjælp af Drosophila melanogaster

Drug screening techniques include high-throughput screening, which uses automation for drug discovery, structure-based designs, and find receptor binding molecules based on the structure, and a fragment-based design, which uses nuclear magnetic resonance and mass spectrometry to optimize molecular fragments and identify the compounds. The DAM system measures fly sleep by detecting infrared beam bricks. However, this method can overestimate sleep if the flies remain static at one end of the tube, or if the fly is not highly mobile.Furthermore, the order of any tested drug may impact the fly fruit intake, potentially skewing experimental results. Therefore, our experiment will incorporate various stretches to mitigate these factors and enhance accuracy. Compared with video analysis based methods, our method offers high-throughput capabilities, reduce the cost, and simplify operation.So process of experimental data is also relatively straightforward, where the resulting graph visually represents the drug's effects.

Watch this Scientific Journal Video about Analyzing the Effect of Reserpine on the Sleep Behavior of Drosophila melanogaster Using SCAMP2020 Data Package in MATLAB at JoVE.com

Analyzing the Effect of Reserpine on the Sleep Behavior of Drosophila melanogaster Using SCAMP2020 Data Package in MATLAB  

Analyse af effekten af reserpin på søvnadfærd af Drosophila melanogaster ved hjælp af SCAMP2020 datapakke i MATLAB

Til at begynde med skal du overvåge fluernes søvnadfærd ved hjælp af en infrarød overvågningsenhed og importere de indsamlede data i TXT-format til DAM-filscanning 107-softwaren til scanning. Indstil starttidspunktet for segmenteringsdataene til enten 0800 eller 0801 den tredje morgen efter start af skærmene for at hente henholdsvis en eller 30 minutters placeringslængdedata. Indstil opsigelsestidspunktet til kl. 8:00 på den tredje dag efter det indstillede starttidspunkt.Juster beholderlængden til et minut, og skift outputfiltypen til kanalfiler. Omdøb filen, og generer outputtet. Start nu SCAMP 2020-softwarepakken i MATLAB.Dobbeltklik på Vecsey Sleep and Circadian Analysis MATLAB-program. Tilføjede den undermappe med titlen Vecsey SCAMP Scripts i stien. Find SCAMP.m fil i den mappe og kør den. I det efterfølgende popup-vindue skal du sekventielt vælge mappen med navnet et minut og 30 minutter. Vælg derefter en skærm, og klik på indlæs individuelle C-plots for at forhåndsvise.Undersøg det billede, der vises, og fravælg den tilsvarende kanal for eventuelle døde fluer. Omdøb hver kanal i hver skærm baseret på det tilsvarende lægemiddel, der skal testes. Vælg alle skærme, og klik på Analysér valgte data til analysen.Vælg standardindstillingen, og klik derefter på analyser for den valgte bin-funktion. Markér afkrydsningsfeltet Eksportér data. Klik på graf 30 minutters datatyper for alle dage for udvalgte grupper, og eksporter alle data for at generere output af resultaterne.Vælg S30-filen fra CSV-filen, og find den tilsvarende middelværdi og standardfejldata for hver skærm. Gem disse data i Excel for at foretage nogle justeringer, før du sender dem til GraphPad Prism for at formulere et søvnstatusdiagram. Find derefter filen med navnet S-T-D-U-R og beregn gennemsnitsværdierne for dagtimerne, natten og den samlede søvn for hver flyvning over tre dage.Indsæt data i GraphPad Prism for at tegne grafer. Søvnmønstrene for Reserpin og Dimethylsulfoxid på kvindelige og mandlige fluer over tre på hinanden følgende dage vises. Kvantitativ analyse af søvnadfærden viser en signifikant stigning i søvntiden hos ældre kvinder fodret med Reserpine.Forsøget med hanfluer viser også en positiv sammenhæng mellem Reserpinkoncentration og fremme af søvn.

Research

JoVE Journal

Behavior

Analyzing the Effect of Reserpine on the Sleep Behavior of Drosophila melanogaster Using SCAMP2020 Data Package in MATLAB

High-Throughput Small Molecule Drug Screening For Age-Related Sleep Disorders Using Drosophila melanogaster

Sleep, an essential component of health and overall well-being, often presents challenges for older individuals who frequently experience sleep disorders ...