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

수면 장애 치료의 발전: 초파리 기반 스크리닝 접근법

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.

저자 스포트라이트: DAM 시스템을 사용한 초파리 수면 측정의 과제 극복 

Drosophila melanogaster를 사용하여 나이 관련 수면 장애를 위한 High-throughput 작은 분자 약 검열

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  

MATLAB에서 SCAMP2020 데이터 패키지를 사용하여 Drosophila melanogaster 의 수면 거동에 대한 Reserpine의 효과 분석

먼저, 적외선 모니터링 장치를 사용하여 파리의 수면 행동을 모니터링하고, TXT 포맷으로 수집된 데이터를 DAM 파일 스캔(107) 소프트웨어로 가져와 스캔한다. 모니터를 시작한 후 세 번째 아침에 분할 데이터의 시작 시간을 0800 또는 0801로 설정하여 각각 1분 또는 30분 빈 길이 데이터를 얻습니다. 종료 시간을 설정된 시작 시간 이후 3일째 되는 날 오전 8:00로 설정합니다.bin 길이 옵션을 1분으로 조정하고 출력 파일 유형을 채널 파일로 변경합니다. 파일 이름을 바꾸고 출력을 생성합니다. 이제 MATLAB에서 SCAMP 2020 소프트웨어 패키지를 시작합니다.Vecsey Sleep and Circadian Analysis MATLAB 프로그램을 더블 클릭합니다. Vecsey SCAMP Scripts라는 제목의 하위 폴더를 경로에 추가했습니다. SCAMP를 찾습니다.m 파일을 열고 실행합니다. 다음 팝업 창에서 1분과 30분이라는 폴더를 순차적으로 선택합니다. 그런 다음 모니터를 선택하고 개별 C 플롯을 로드하여 미리 봅니다.나타나는 이미지를 검사하고 죽은 파리의 해당 채널을 선택 취소합니다. 테스트할 해당 약물에 따라 모든 모니터의 각 채널 이름을 바꿉니다. 모든 모니터를 선택하고 분석을 위해 선택한 데이터 분석을 클릭합니다.기본 옵션을 선택한 다음 선택한 빈 기능에 대해 분석을 클릭합니다. 데이터 내보내기 상자를 선택합니다. 선택한 그룹의 모든 날짜에 대한 그래프 30분 데이터 유형을 클릭하고 모든 데이터를 내보내 결과 출력을 생성합니다.CSV 파일에서 S30 파일을 선택하고 각 모니터에 해당하는 평균값과 표준 오류 데이터를 찾습니다. 이 데이터를 GraphPad Prism에 게시하여 절전 상태 다이어그램을 작성하기 전에 Excel에 저장하여 몇 가지 조정을 수행합니다. 그런 다음 S-T-D-U-R이라는 파일을 찾아 3일 동안 각 파리의 주간, 야간 및 총 수면의 평균값을 계산합니다.그래프를 그리기 위해 GraphPad Prism에 데이터를 붙여넣습니다. 3일 연속으로 암컷과 수컷 파리에 대한 Reserpine과 Dimethyl Sulfoxide의 수면 패턴이 표시됩니다. 수면 행동에 대한 정량적 분석은 Reserpine을 먹인 고령 여성의 수면 시간이 크게 증가했음을 보여줍니다.수컷 파리에 대한 실험은 또한 레세르핀 농도와 수면 촉진 사이의 긍정적인 상관관계를 보여줍니다.

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 ...