ai_agent_IP

ai_agent

maintenance

instantTrialDurationMinutes

isBiopharma

enableNewMyJoveSectionDesign

educationFeatures

seoMetaApi

ai_copilot_institution_blacklist

ai_chat_institution_blacklist

freeAccessWindow

ssoRoleCapture

scheduledMaintenanceBanner

joveLabsFeatures

disable_autoplay_for_bots

extended_trial_duration

5 minutes

We thank the Fulbright US Student Program, the Latvian Fulbright Post, and the US Department of State.&#160;This project was supported by a grant (lzp-2024/1-0437) of the Latvian Council of Science.

The forced swim assay was conducted in accordance with the principles of the 3Rs (Replacement, Reduction, and Refinement). Replacement was achieved by using Drosophila melanogaster as an invertebrate model, thereby avoiding vertebrate forced swim paradigms that are increasingly restricted for ethical reasons. Reduction was supported by the assay&#39;s high-throughput nature, which enables robust behavioral inference from relatively small cohorts while minimizing animal use. Refinement was addressed by limiting swim exposure to 2 min, a duration that prevents exhaustion or drowning and allows full recovery of normal locomotor behavior immediately after testing. Together, these considerations ensure that the assay provides meaningful behavioral data while minimizing potential distress and maximizing ethical acceptability, including in educational settings.
1. Induction of a depression-like state in Drosophila (vibration stress protocol)
<ol>
	<li>Collect male D. melanogaster within 24 h of eclosion under light CO&#8322; anesthesia. All the details are mentioned in the Table of Materials.</li>
	<li>Place flies in groups of 10 into polypropylene or acrylic vials (95 mm length &#215; 25 mm inner diameter) sealed with cotton plugs.</li>
	<li>Expose flies to mechanical vibration (300 Hz) using a vibration platform.</li>
	<li>Apply repeated cycles consisting of 45 min vibration followed by 15 min recovery&#160;in vials containing standard food.</li>
	<li>Continue vibration cycles for a total of 6 h per day.</li>
	<li>Repeat the stress protocol for 3 consecutive days.</li>
	<li>Maintain control flies under identical housing conditions without vibration exposure. 
	NOTE: In the present protocol, only male flies were used to reduce variability associated with sex-specific physiological and behavioral differences, including reproductive state and hormonal influences. This standardization facilitates the detection of stress-induced behavioral effects in a controlled setting. However, the protocol is fully applicable to female flies, and sex-specific responses represent an important area for future investigation. Given that sensitivity to stress-related and depression-like phenotypes can vary with sex and age across taxa, including Drosophila, extending this framework to different demographic groups may provide additional biological insight. This paradigm is adapted from established uncontrollable vibration stress models that induce motivational and activity alterations in flies.</li>
</ol>
2. Forced swim test in Drosophila melanogaster
CAUTION: Sodium dodecyl sulfate (SDS) is an irritant. Wear appropriate personal protective equipment (e.g., gloves and eye protection) when preparing and handling SDS solutions. Avoid skin and eye contact. Dispose of SDS-containing waste in accordance with institutional chemical safety guidelines.
<ol>
	<li>Collect adult flies within 24 h of eclosion and maintain them under constant light and controlled environmental conditions.</li>
	<li>Prepare multi-well chamber slides and fill each well with ~2 mL of 0.08% SDS solution at room temperature.</li>
	<li>Transfer a single fly into each well immediately before recording. No anesthesia is required.</li>
	<li>Record behavior using an overhead camera for 2 min to quantify latency to first immobility.</li>
	<li>Quantify latency to first immobility.</li>
	<li>Remove flies after recording and place them on absorbent paper to confirm recovery of normal locomotion. 
	NOTE: Immobility is defined as cessation of active escape movements while maintaining surface position. To record additional parameters such as total immobility duration and number of immobility bouts, longer recording durations (e.g., 3 min) are needed. This assay reflects passive coping behavior; reduced latency to immobility is interpreted as decreased motivation to sustain escape responses. Variability in latency to immobility / total immobility across flies. This assay may induce transient stress or fatigue. When used in longitudinal designs, perform it prior to other assays and allow sufficient recovery (e.g., &#8805;24 h) before subsequent testing.</li>
</ol>
3. Long-term locomotor activity monitoring using the Drosophila Activity Monitor (DAM)
<ol>
	<li>Anesthetize fruit flies briefly (5 s) using CO&#8322; for sorting.</li>
	<li>Place individual flies into glass DAM tubes containing food and cotton plugs17.</li>
	<li>Insert tubes into the Drosophila Activity Monitor system.</li>
	<li>Record locomotor activity continuously (48 h) under controlled temperature and light&#8211;dark conditions.</li>
	<li>Store beam-break counts using acquisition software.</li>
	<li>Bin activity data into defined time intervals (e.g., 1&#8211;10 min).</li>
	<li>Exclude flies that die or show no activity after acclimation. 
	NOTE: Continuous locomotor monitoring over a 48-hour period is sufficient to capture circadian activity patterns. DAM: variability in total distance, velocity, and bout structure across individuals.</li>
</ol>
4. Open-field locomotor assay
<ol>
	<li>Briefly (5 s) anesthetize fruit flies using CO&#8322;.</li>
	<li>Transfer individual flies into rectangular 25 x 75 mm arenas (one fly per arena).</li>
	<li>Allow flies to acclimate (20 min) to the arena environment prior to recording under constant, low-intensity diffuse white light.</li>
	<li>Record behavior under constant illumination using an automated video-tracking system.</li>
	<li>Analyze trajectories using tracking software to extract: 
	Total distance moved 
	Average locomotor speed 
	Acceleration and turning metrics 
	NOTE: This assay provides a baseline for interpreting stress-related behavioral changes. The acclimation period minimizes handling-induced stress and allows flies to habituate to the arena, ensuring that recorded locomotor activity reflects baseline behavior. Constant illumination was used to reduce variability associated with circadian fluctuations and to ensure consistent behavioral conditions across trials. Trajectories can be extracted using automated tracking software such as EthoVision XT (Noldus Information Technology), or open-source alternatives including Ctrax, idTracker, and TrackMate (ImageJ/Fiji), depending on availability and user preference. Open-field assay: variability in total distance, velocity, and bout structure across individuals. To reduce order effects, standardize acclimation and lighting conditions across sessions. When multiple assays are conducted on the same individuals, keep arena exposure durations consistent.</li>
</ol>
5. Y-maze turning behavior and behavioral variability assay
<ol>
	<li>Briefly anesthetize (5 s) fruit flies using CO&#8322;.</li>
	<li>Place individual flies into Y-maze chambers8,9. Mazes were cut into 1.6 mm-thick black acrylic using a laser engraver. We placed flies in an array of 95 individual mazes, each consisting of three symmetrical arms (12 mm long and 3.3 mm wide).</li>
	<li>Allow flies to explore freely for extended recording periods (20 min).</li>
	<li>Record turning decisions (left or right) during exploratory locomotion (1 h).</li>
	<li>Compute individual turning bias as the proportion of right turns. Calculate relevant metrics such as velocity, acceleration, motion without movement, time-based variations in activity12,15.</li>
	<li>Quantify between-fly behavioral variability using the median absolute deviation (MAD) of turning bias. 
	NOTE: This assay measures locomotor handedness and behavioral predictability independent of overall activity levels. Turning sequences capture decision structure; turning bias reflects lateralization, while variability across individuals reflects behavioral predictability. bias = proportion of right turns (per fly); variability = dispersion (MADn) of bias across flies. For repeated testing, consider counterbalancing assay order across individuals or cohorts to control for sequence effects.</li>
</ol>
6. Phototactic choices: activity monitoring in the FlyVac apparatus
<ol>
	<li>Load individual flies into FlyVac chambers6,11 without using CO2 anesthesia.</li>
	<li>Allow acclimation (10 min) before initiating trials.</li>
	<li>Present repeated binary light&#8211;dark choices to each fly.</li>
	<li>Record choices across 40 trials using automated detection.</li>
	<li>Calculate light-choice probability per fly and the average time between light-choices as a measure of activity.</li>
	<li>Quantify behavioral variability across individuals using MAD. 
	NOTE: The FlyVac system enables simultaneous assessment of mean decision bias and inter-individual variability. The behavioral assays described in this protocol are modular and can be conducted in different sequences depending on the experimental design. However, to minimize potential carry-over effects, performing the forced swim test prior to other assays is recommended. To reduce order effects, standardize acclimation and lighting conditions across sessions.</li>
</ol>
7. Data analysis
NOTE: Behavioral analyses were performed using individual flies as the unit of inference unless otherwise stated.
<ol>
	<li>Forced swim test: Analyze the video recordings manually or using behavioral scoring software. 
	NOTE: Immobility was defined as the absence of active escape movements while the fly remained afloat. The following parameters were extracted per fly: latency to first immobility (s). Group comparisons were conducted using non-parametric statistical tests due to the non-normal distribution of behavioral measures.</li>
	<li>Long-term locomotor activity (DAM): Record beam-break counts by the Drosophila Activity Monitor. Export and bin into 10 min intervals. 
	NOTE: For each fly, total activity counts and circadian activity patterns were calculated. These patterns can be calculated in R, Python, or Excel&#8212;whichever is available. Flies exhibiting inactivity due to mortality or technical artifacts were excluded prior to analysis. The assay lasted 48 h.</li>
	<li>Open-field locomotor assay: Process the video-tracking data using automated tracking software. Import the video files into EthoVision XT. Perform tracking using automated detection with contrast-based thresholding. 
	NOTE: Trajectories were smoothed using the default filtering algorithm. Per-fly locomotor parameters (distance, velocity, acceleration) were exported as CSV files for downstream analysis. Per-fly locomotor parameters included: total distance moved (mm), accelerations (mm/s2). Summary statistics were calculated at the individual level prior to group comparison. The duration of this test was 1 h for each group.</li>
	<li>Y-maze turning behavior: Extract the turning decisions from recorded trajectories, classify as left or right, and export as binary sequences (left/right) for each individual. 
	NOTE: For each fly, the turning bias was calculated as the proportion of right turns across all recorded decisions. Between-fly behavioral variability was quantified using the scaled median absolute deviation (MADn&#160;= 1.4826 &#215; MAD) of turning bias. The duration of this test was 1 h for each group.</li>
	<li>FlyVac phototactic choices: Record binary light&#8211;dark choices automatically. For each fly, calculate the light-choice probability (LCP) as the proportion of trials in which the light arm was selected. Express the phototactic bias as the mean of binary trial outcomes (light = +1, dark = &#8722;1). 
	NOTE: Between-fly variability was quantified using MADn&#160;of individual phototaxis indices.</li>
	<li>Statistical analysis: Assess the group differences in behavioral measures using permutation-based tests with 10,000 label randomizations. 
	NOTE: To compare variability, permutation tests were performed on the absolute differences in MADn&#160;between groups. Confidence intervals for MADn&#160;estimates were obtained using bootstrap resampling. A two-tailed unpaired t-test was used to compare the Distance moved by flies (mm) and Maximum acceleration (mm/s2). For data visualization, violin plots (primarily to show density, concentration, and skewness), box plots, and column charts were used. Statistical significance was evaluated at &#945; = 0.05.</li>
</ol>

<ol>
	<li>Chan HY, Bonini NM. Drosophila models of human neurodegenerative disease. Cell Death &#38; Differentiation. 2000 Nov;7(11):1075-80.</li>
	<li>McGurk L, Berson A, Bonini NM. Drosophila. as an in vivo model for human neurodegenerative disease. Genetics. 2015 Oct 1;201(2):377-402.</li>
	<li>Honegger KS, Smith MA, Churgin MA, Turner GC, de Bivort BL. Idiosyncratic neural coding and neuromodulation of olfactory individuality in Drosophila. Proceedings of the National Academy of Sciences. 2020 Sep 22;117(38):23292-7.</li>
	<li>Neckameyer WS, R. Nieto-Romero A. Response to stress in Drosophila. is mediated by gender, age and stress paradigm. Stress. 2015 Mar 4;18(2):254-66.</li>
	<li>Hibicke M, Nichols CD. Validation of the forced swim test in Drosophila., and its use to demonstrate psilocybin has long-lasting antidepressant-like effects in flies. Scientific Reports. 2022 Jun 15;12(1):10019.</li>
	<li>Kain JS, Stokes C, de Bivort BL. Phototactic personality in fruit flies and its suppression by serotonin and white. Proceedings of the National Academy of Sciences. 2012 Nov 27;109(48):19834-9.</li>
	<li>Ayroles JF et al. Behavioral idiosyncrasy reveals genetic control of phenotypic variability. Proceedings of the National Academy of Sciences. 2015 May 26;112(21):6706-11.</li>
	<li>Buchanan SM, Kain JS, De Bivort BL. Neuronal control of locomotor handedness in Drosophila. Proceedings of the National Academy of Sciences. 2015 May 26;112(21):6700-5.</li>
	<li>Krama T et al. Development under predation risk increases serotonin-signaling, variability of turning behavior and survival in adult fruit flies Drosophila melanogaster. Frontiers in Behavioral Neuroscience. 2023 May 25;17:1189301.</li>
	<li>Kain JS et al. Variability in thermal and phototactic preferences in Drosophila. may reflect an adaptive bet-hedging strategy. Evolution. 2015 Dec 1;69(12):3171-85.</li>
	<li>Krams IA et al. Serotoninergic modulation of phototactic variability underpins a bet-hedging strategy in Drosophila melanogaster. Frontiers in Behavioral Neuroscience. 2021 Apr 16;15:659331.</li>
	<li>Popovs S et al. Explaining the survival of the sickest: altered walking patterns are linked with improved adult survival in Drosophila melanogaster. grown with predators during larval development. Behaviour. 2024 Jan 2;161(2):133-48.</li>
	<li>Neckameyer WS, Bhatt P. Protocols to study behavior in Drosophila. Drosophila.: Methods and Protocols. 2016 Oct 12:303-20.</li>
	<li>Ali YO, Escala W, Ruan K, Zhai RG. Assaying locomotor, learning, and memory deficits in Drosophila. models of neurodegeneration. Journal of Visualized Experiments: JoVE. 2011 Mar 11(49):2504.</li>
	<li>Krama T et al. A diabetes-like biochemical and behavioural phenotype of Drosophila. induced by predator stress. Proceedings of the Royal Society B: Biological Sciences. 2023 Jul 1;290(2002).</li>
	<li>Ries AS, Hermanns T, Poeck B, Strauss R. Serotonin modulates a depression-like state in Drosophila. responsive to lithium treatment. Nature Communications. 2017 Jun 6;8(1):15738.</li>
	<li>Rozato E, Kyriacou CP. Analysis of locomotor activity rhythms in Drosophila. Nature Protocols. 2006. Jul 6;1(2):559-568</li>
	<li>Yankelevitch-Yahav R, Franko M, Huly A, Doron R. The forced swim test as a model of depressive-like behavior. Journal of Visualized Experiments: JoVE. 2015 Mar 2(97):52587.</li>
</ol>

The authors declare no conflicts of interest.

The present protocol demonstrates a multimodal approach for inducing and quantifying depression-like behavioral states in D. melanogaster. Using a vibration-based stress paradigm combined with complementary behavioral assays, this study shows that depression-like induction can be detected across measures of coping behavior, locomotor activity, exploratory dynamics, decision-making, and behavioral variability16. The stress regime employed in this study serves as a practical tool to elicit ...

Behavioral alterations such as changes in locomotion, decision-making, and exploratory activity are widely used as organism-level indicators of neural and physiological state. Across taxa, neural dysfunction often manifests as altered movement structure, reduced exploratory motivation, impaired sensory-guided decision behavior, and changes in behavioral predictability. Because these organism-level outputs integrate neural, metabolic, and physiological state, they provide scalable readouts for detecting disease-relevant phenotypes. However, despite the extensive use of Drosophila melanogaster in molecular genetics and neuroscience, standardized multimodal behavioral frameworks that simultaneously capture locomotor performance, decision structure, and behavioral variability remain limited.
Model organisms are essential for studying mechanisms underlying stress-related and neuropsychiatric phenotypes because controlled experimental manipulation is often impossible in humans. Stress-related behavioral syndromes occur across many taxa, including insects, and involve conserved neuromodulatory systems, such as serotonin and dopamine, as well as metabolic signaling pathways. The fruit fly has therefore emerged as a powerful system for linking molecular dysfunction to organism-level behavioral outcomes due to its short generation time, genetic tractability, and conservation of disease-relevant pathways1,2.
At the same time, many behavioral assays in Drosophila are typically applied in isolation, making it difficult to distinguish between general sickness effects, motor impairment, motivational changes, and decision-level behavioral alterations. Moreover, mean behavioral measures alone may obscure biologically meaningful phenotypes, as neuromodulatory and developmental perturbations often alter behavioral variability rather than average behavioral output. A multimodal behavioral approach combining locomotor, exploratory, sensory, and decision-making assays is therefore necessary to generate robust organism-level phenotypic signatures3.
Here, a multimodal behavioral pipeline integrating forced swim exposure4,5, Y-maze turning behavior and handedness6,7,8,9, phototaxis and activity assessment using the FlyVac system10,11, open-field exploration12, and long-term locomotor monitoring using the Drosophila Activity Monitor5,13 is presented (Figure 1). Each assay in this framework captures a distinct behavioral dimension relevant to stress-related phenotypes. The forced swim test measures passive coping behavior, where an earlier transition to immobility reflects reduced motivation to sustain escape responses. The open-field and DAM assays quantify general locomotor activity and circadian structure, providing baseline measures of motor output and arousal. The Y-maze assay captures decision-making structure through sequences of left&#8211;right choices, allowing estimation of both turning bias (lateralization) and behavioral variability as a measure of predictability. Finally, the FlyVac assay evaluates sensory-guided decision-making and approach&#8211;avoidance behavior through repeated phototactic choices. Together, these assays provide complementary readouts that distinguish changes in activity, motivation, and decision-level behavior.
This protocol captures complementary aspects of motor output, motivation, decision structure, and behavioral variability, providing a scalable framework for detecting stress-, metabolic-, and neurodegeneration-related behavioral phenotypes in D. melanogaster14. The primary aim of this study is to establish a multimodal behavioral phenotyping framework that integrates complementary assays to capture multiple dimensions of stress-induced behavioral change. The stress paradigm is used here as a standardized perturbation to demonstrate the framework&#39;s sensitivity and applicability.

<table><tbody><tr><td>Anesthesia equipment: e.g., Benchtop Flowbuddy, Complete System w/ Ultimate Flypad</td><td>Genesis Scientific, El Cajon, CA, USA</td><td>59-122BCU</td><td></td></tr><tr><td>BioSan MSV-3500 multispeed vortex</td><td>Biosan SIA, Riga, Latvia</td><td>BS-010210-TAH</td><td>With all platforms</td></tr><tr><td>Cotton closures for narrow vials&amp;nbsp;</td><td>Flystuff by Genesee Scientific, El Cajon, CA 92020 USA</td><td>Catalog number: 51-101</td><td></td></tr><tr><td>Drosophila Activity Monitor (DAM2)</td><td>TriKinetics Inc, Waltham, MA, USA</td><td>DAM2; RRID: not available&amp;nbsp;</td><td>32 tubes</td></tr><tr><td>Drosophila melanogaster (Oregon-R-modENCODE)</td><td>Drosophila melanogaster (Oregon-R-modENCODE)</td><td>BDSC:25211; RRID:BDSC_25211</td><td></td></tr><tr><td>GraphPad Prism</td><td>GraphPad Software, Boston, MA, USA;</td><td>Catalog name: GraphPad Prism, RRID:SCR_002798</td><td>Version 11.0.1</td></tr><tr><td>Logitech C920 HD Pro Webcam, HD 1080p lensCatalog name: HD Pro Webcam C920</td><td>Logitech Europe S.A., Lausanne, Switzerland</td><td>RRID: not available</td><td>The overhead recording camera used for FST</td></tr><tr><td>Narrow Fly Vial, Polypropylene</td><td>Flystuff by Genesee Scientific, El Cajon, CA 92020 USA</td><td>Catalog number: 32-120BF</td><td></td></tr><tr><td>Noldus EthoVision XT&amp;nbsp;</td><td>Noldus Information Technology, Wageningen, The Netherlands</td><td>RRID: SCR_000441</td><td>v. 15.0</td></tr><tr><td>Sodium dodecyl sulfate (SDS, &amp;ge;99%)&amp;nbsp;</td><td>Merck KGaA (Sigma-Aldrich), Darmstadt, Germany</td><td>Catalog name: Sodium dodecyl sulfate (ACS reagent, &amp;ge;99% purity); RRID: not available</td><td></td></tr><tr><td>ZEISS Stemi 508 Stereomicroscope&amp;nbsp;</td><td>Carl Zeiss AG, Oberkochen, Baden-W&amp;uuml;rttemberg, Germany</td><td>15634448</td><td></td></tr></tbody></table>

multimodal behavioral phenotyping of stress-induced depression-like states in drosophila melanogaster

Multimodal Behavioral Phenotyping of Stress-Induced Depression-Like States in Drosophila melanogaster

Metabolic, neurodegenerative, and stress-related disorders are frequently accompanied by altered locomotion, impaired decision-making, and reduced behavioral flexibility. However, accessible multimodal behavioral frameworks for quantifying these phenotypes in Drosophila melanogaster remain limited. Here, a protocol integrating forced swim exposure, Y-maze turning behavior and handedness, phototaxis, and activity assessment in the FlyVac system, open-field exploration, and long-term locomotor monitoring using the Drosophila Activity Monitor is presented. These complementary assays capture multiple dimensions of behavior, including motor output, motivation, decision structure, and behavioral variability, as functional readouts of neural and metabolic states. The pipeline is scalable, reproducible, and adaptable to pharmacological, genetic, and environmental manipulations, providing a versatile framework for detecting stress-, metabolic-, and neurodegeneration-related behavioral phenotypes in Drosophila.

Application of the vibration stress protocol produced measurable behavioral alterations across multiple assays (Figure 1) designed to capture activity, coping behavior, and decision-making in D. melanogaster.
Depression-like induction
Flies exposed to repeated mechanical vibration (Figure 1A) displayed reduced exploratory activity and altered behavioral responses compared to unstre...

This protocol provides a framework for assessing stress-induced behavioral changes in Drosophila melanogaster. Combining complementary assays enables the quantification of activity, exploration, and decision-making for individual flies. The approach is flexible and can be adapted to a wide range of studies investigating stress biology, metabolism, and neurobehavioral function.

Metabolic, neurodegenerative, and stress-related disorders are frequently accompanied by altered locomotion, impaired decision-making, and reduced ...

Our research aims to establish multimodal behavioral framework for quantifying stress-induced depression-like states in Drosophila Melanogaster. This protocol applies to screening antidepressant compounds, investigating gene environment interactions, and exploring neurobiological mechanisms behind behavioral variability in drosophila. To begin, anesthetize the male flies under light carbon dioxide anesthesia within 24 hours of occlusion.Place the flies in groups of 10 into polypropylene or acrylic vials and seal them with cotton plugs. Place the fly vials on a vibration platform to expose them to mechanical vibration at 300 RPM. Apply 45 minutes of vibration followed by 15 minutes of recovery.Repeat this cycle for six hours per day for three consecutive days. Maintain the control flies under identical housing conditions without exposure to vibration. Collect adult flies within 24 hours of occlusion and maintain them under controlled environmental conditions with 12 hours of light, followed by 12 hours of darkness.Prepare multi-well chamber slides and the overhead camera to record behavior for two minutes and calculate the latency to first immobility. Fill each well with approximately two milliliters of 0.08%sodium dodecyl sulfate solution at room temperature. Without anesthesia, transfer a single fly into each well immediately before recording.Briefly, anesthetize fruit flies for five seconds using carbon dioxide for sorting. Place individual flies into glass drosophila activity monitor or DAM tubes containing food. Seal with cotton plugs.Insert the tubes into the drosophila activity monitor system and record locomotor activity continuously for 48 hours under controlled temperature and light dark conditions. Briefly anesthetize fruit flies for five seconds using carbon dioxide. Transfer individual flies into rectangular 25 by 75 millimeter arenas with one fly per arena.Allow the flies to acclimate to the constant low-intensity diffuse white-light arena environment for 20 minutes before recording. Record behavior under constant illumination using an automated video tracking system. Briefly, anesthetize fruit flies for five seconds using carbon dioxide.Place individual flies into Y-Maze chambers. Cut into 1.6 millimeter thick-black acrylic with a laser engraver. Allow the flies to explore freely for an extended recording period of 20 minutes.Record left or right turning decisions during exploratory locomotion for one hour. For phototactic assay, load individual flies into flyvac chambers without using carbon dioxide anesthesia and allow them to acclimate for 10 minutes before initiating trials. Present repeated binary light dark choices to each fly.Record choices across 40 trials using automated detection. For data analysis, process the behavioral data from stressed and control groups across all assays and use permutation-based tests to evaluate group differences. In the forced swim assay, stressed flies adopted immobility more rapidly and showed prolonged passive episodes compared to control flies.In terms of long-term locomotor activity, continuous DAM recordings revealed that control flies were significantly more active than stressed flies. In the open-field locomotor assay, control flies displayed sustained locomotion, frequent directional changes, and traveled significantly greater distances in one hour compared to stressed flies that showed reduced displacement and slower movement. In contrast, the stressed flies showed significantly higher maximum acceleration than control flies.In the Y-Maze assay, control flies covered significantly longer distances in one hour than stressed flies. Contrastingly, the stressed flies reached significantly higher maximum acceleration than control flies. In terms of turning bias, the group difference between control and stressed flies was non-significant.In the flyvac phototactic test, the groups did not differ in the number of completed trials. The stressed flies showed a lower-light choice probability than control flies, indicating a shift toward the dark arm. The variability in phototactic choice among individual flies did not differ significantly between control and stressed groups.It measures motor output, passive coping motivation, sensory-guided decision structure, circadian structure, and inter-individual behavioral variability. Controlling environmental factors like temperature, humidity and handling is critical to minimize carryover effects and reducing and pounding locomotive suppression. Future studies can investigate sex-specific and age-related stress responses as well as integrate pharmacological manipulations to clarify underlying neurochemical mechanisms.

multimodal-behavioral-phenotyping-stress-induced-depression-like

Research

JoVE Journal

Medicine

This video demonstrates the method of inducing chronic stress in a mouse through repeated immobilization, leading to corticosterone release and reduced glutamate receptor activity in neurons, which results in depression-like behavior.

Inducing Depression-like Behavior in a Mouse via Immobilization Stress

Here, we present a protocol to quantify the avoidance of stressed individuals. This paradigm is powerful yet user-friendly and can be used to assess the influence of genes and environment on one kind of social interaction in Drosophila melanogaster.

Straightforward Assay for Quantification of Social Avoidance in Drosophila melanogaster

Here, methods for developing a mouse model of subchronic and mild social defeat stress are described and used to investigate the pathogenic features of depression including hyperphagia- and polydipsia-like symptoms following increased body weight.

A Mouse Model of Subchronic and Mild Social Defeat Stress for Understanding Stress-induced Behavioral and Physiological Deficits

The unpredictable chronic mild stress (UCMS) protocol is a validated method for studying behavioral and physiological changes associated with chronic stress and depressive symptoms. Eight weeks of imposition of the UCMS protocol induces behavioral changes and poor health outcomes in rodents of either gender.

An Unpredictable Chronic Mild Stress Protocol for Instigating Depressive Symptoms, Behavioral Changes and Negative Health Outcomes in Rodents

Here we present the unpredictable chronic mild stress protocol in mice. This protocol induces a long-term depressive-like phenotype and enables to assess the efficacy of putative antidepressants in reversing the behavioral and neuromolecular depressive-like deficits.

The Unpredictable Chronic Mild Stress Protocol for Inducing Anhedonia in Mice

This article provides a simplified and standardized protocol for induction of depressive-like behavior in chronically immobilized mice by using a restrainer. In addition, behavior and physiological techniques to verify induction of depression are explained.

A Chronic Immobilization Stress Protocol for Inducing Depression-Like Behavior in Mice

The Y-maze barrier task is a behavior test that examines motivation to expend effort for reward. Here, we discuss testing multiple well-validated chronic stressors including chronic corticosterone and social defeat stress with this behavior, as well as the novel chronic non-discriminatory social defeat stress (CNSDS), which is effective in females.

Chronic Stress Shifts Effort-Related Choice Behavior in a Y-Maze Barrier Task in Mice

The chronic despair mouse model (CDM) of depression consists of repetitive forced swim sessions and another delayed swim phase as a read-out. It represents a suitable model for induction of a chronic depressive-like state stable for at least 4 weeks, amendable to evaluate subchronic and acute treatment interventions.

Animal Models of Depression - Chronic Despair Model (CDM)

Autism Spectrum Disorder (ASD) is associated with impaired social and communicative behavior and the emergence of repetitive behavior. For studying the interrelation between ASD genes and behavioral deficits in the Drosophila model, five behavioral paradigms are described in this paper for assaying social spacing, aggression, courtship, grooming, and habituation behavior.

Paradigms for Behavioral Assessment in Drosophila Model of Autism Spectrum Disorder

By integrating four daily social stressors, the present study describes a multiple integrated social stress (MISS) model for psychiatric disorders in female C57BL/6J mice. After ten days of repeated MISS exposure, the mice developed depressive- and anxiety-like phenotypes, thus providing a natural, etiology-based animal model for studying psychiatric disorders in female subjects.

A Multiple Integrated Social Stress Model for Psychiatric Disorders in Female C57BL/6J Mice

The protocol presents a high-throughput, cost-effective method for assessing neurotoxicity in the fruit fly through the quantification of locomotor dysfunction as an alternative to traditional mammalian models. It aims to evaluate the neurotoxic effects of pharmaceutical compounds, environmental agents, or genetic modifications using a sensitive, reproducible, and ethically favorable model.

Behavioral Analysis of Locomotor Dysfunction in Drosophila melanogaster as a Readout for Neurotoxicity

Latvian Biomedical Research and Study Center.  This protocol provides a framework for assessing stress-induced behavioral changes in Drosophila melanogaster. Combining complementary assays enables the quantification of activity, exploration, and decision-making for individual flies. The approach is flexible and can be adapted to a wide range of studies investigating stress biology, metabolism, and neurobehavioral function.

Multimodal Behavioral Phenotyping of Stress-Induced Depression-Like States in Drosophila melanogaster

Explore More Videos