This work was funded by internal funds of the University Clinic Freiburg, Department of Psychiatry and Psychotherapy and the Berta-Ottenstein Program for Clinician Scientists (to SV). TS is funded by the grants of Medical Research Foundation (FRM) (AJE201912009450) and the University of Strasbourg Institute of Advance Studies (USIAS) (2020-035), as well as Centre National de la Recherche Scientifique (CNRS), France.

All experiments were performed in agreement with European guidelines (EU 2010/63) and in accordance with the German animal protection law (TierSchG), FELASA (www.felasa.eu/guidelines.php), the national animal welfare body GV-SOLAS (www.gv-solas.de/index.html) guide for the care and use of laboratory animals, and were approved by the animal welfare committee of the University of Freiburg and by the Comite d&#8217;Ethique en Matiere d&#8217;Experimentation Animale de Strasbourg (CREMEAS, CEEA35), as well as local authorities. Both sexes of C57Bl6N wild-type mice aged 10-14 weeks (70-98 post-natal days, PND) were used for wild-type (WT) indicated experiments. As a stress-resilient line, the transgenic mouse line with enhanced expression of adenosine A1 receptors under the forebrain neuronal CaMKII promoter was used9,15. After the experiments, mice were sacrificed by cervical dislocation.
1. Preparation
<ol>
	<li>Obtain an animal research license, including thorough experimental planning.</li>
	<li>Arrival: On arrival, raise the animals in the animal facility to perform the CDM. If the animals are bought from an external supplier, allow them at least 2 weeks to adapt to the new environment.</li>
	<li>Housing: To house the animals, ensure that the cages are not occupied with the maximum number of animals to avoid additional stress. Guarantee that housing conditions are in line with international recommendations of mouse housing (for further information, see22) and constantly maintain them at all times. 
	NOTE: The most important standard housing conditions include individually ventilated cages with 25-120 air changes per hour, 12 h light-dark cycle, temperature as stable as possible (at least constant between 20-24 &#176;C), humidity as stable as possible (at least between 45%-65%), gnawing material and shelter present, no individual housing.</li>
	<li>Time point: Perform all experiments at the same time of the day. 
	NOTE: No direct assessment has been made to verify the influence of the daytime on CDM, but most behavioral tests evaluating depressive-like states show variations depending on the time of the day23,24,25, and it is highly probable that daytime also influences CDM.</li>
	<li>Nesting material: Reduce the nesting material to a minimum. Ensure there are no running wheels, etc., present in the cage. 
	NOTE: Enriched environment prevents the induction of a depressed state.</li>
	<li>Group composition: Allow the animals to remain in the same group throughout the whole experiment. Group the female mice together even from different litters; group the male mice together with littermate male animals. Due to upcoming aggressiveness, especially of males, biting and barbering may become a problem, therefore give special emphasis to the group composition. Avoid single housing as deprivation is a major additional stressor.</li>
	<li>Animals: Use different mouse strains, even though specific differences have been observed9,10. A frequently used mouse strain is C57Bl6N. Label mice in order to perform paired statistical analysis (see step 3.2.4).</li>
	<li>Animal sex: Equally use both male and female mice.</li>
	<li>Animal age: Ensure that the animals are at least 10 weeks (70 PND) old. Do not use younger animals due to the exhaustion caused by swimming.</li>
	<li>Equipment: Use a transparent glass cylinder/beaker with a capacity of at least 2 L, a diameter of 24-26 cm, and a minimum height of 30 cm. Further requirements include a thermometer to check the water temperature, paper towels, red light heating lamp/heating mat or comparable source of heating, timer, stopwatch, quiet surroundings. Videotape the swim sessions for offline analysis and documentation. Ensure that the date and the time are continuously visible on the tape/file, together with an identification code number for the individual animal. Store the files for later analysis and further reference. Film from the side of the glass cylinder, not from above, to facilitate analysis.</li>
</ol>
2. Induction phase
<ol>
	<li>Before starting
	<ol>
		<li>Visually observe the animals for abnormalities, including signs of biting or barbering. Exclude the whole cage from the experimental series if an animal shows any minimal injuries. Ensure that a veterinarian is available at any time as injuries will worsen during the experiment and will prevent continuation as mice become more aggressive under the influence of stress.</li>
		<li>Obtain the bodyweight for each animal before starting the experiment. Ensure that the weight loss often observed does not exceed 20% of the initial body weight. Exclude animals with a weight loss of over 20% and immediately euthanize them due to the assumed high suffering.</li>
		<li>Fill up a beaker or cylinder with water at room temperature (22-23 &#176;C) to a height of at least 20 cm from the bottom, leaving a minimum of 10 cm between the water surface and the upper border of the vessel.</li>
	</ol>
	</li>
	<li>Performance
	<ol>
		<li>Gently transfer the animals into the water. During the swim phase, keep the animal under continuous observation to prevent drowning. Observe from a position where the animal cannot see the experimenter (for instance, video observation from a room next door).</li>
		<li>Set a chronometer at the beginning of the experiment. Take the animals out of the water after 10 min by simply grabbing their tails. Gently dry them with a paper towel and put them either under a heating light or on a heating mat.</li>
		<li>Evaluate only one animal at a time. Ensure that animals cannot see each other (for example, separate the housing cage from the experimental set-up by a room-divider).</li>
		<li>Perform the forced swim session for 10 min each day for 5 consecutive days.</li>
	</ol>
	</li>
	<li>Finishing
	<ol>
		<li>Transfer the animals back to their home cages after five swim sessions and allow them to rest for at least 2 days. Start specific treatment interventions subsequently.</li>
	</ol>
	</li>
</ol>
3. Evaluation of an anti-depressive treatment
<ol>
	<li>Time course
	<ol>
		<li>Assess the acute and subchronic treatments with the CDM. Depending on the scientific question, adapt the resting period between the induction phase and the read-out.</li>
		<li>To evaluate the acute and rapid-acting potency of ketamine, choose a short resting period (a few days) after the induction phase of CDM. Apply the treatment (i.e., intraperitoneal injection), and then perform the evaluation (additional swim session or different evaluation method) shortly afterward.</li>
		<li>To evaluate the effects of a subchronic treatment, increase the treatment period up to 4 weeks (there is no data available for longer treatment periods). For example, give the oral treatment with imipramine to the animals during 4 weeks after the induction phase and evaluate thereafter.</li>
		<li>Start to evaluate the depressed state right after the end of the treatment period, e.g., the following day. Always choose an identical time period for control and experimental conditions.</li>
	</ol>
	</li>
	<li>Immobility time
	<ol>
		<li>Proof-of-concept
		<ol>
			<li>To use immobility time as a read-out method, evaluate each day of the induction phase and the test day to provide a proof-of-concept (see Figure 1). For further experimental series, reduce the assessments to day 1, day 5, and the test day (see Figure 1C).</li>
			<li>Videotape each experiment. Allow two trained observers who are blinded to the experimental conditions to perform the analysis independently. Video analysis enables the experimenter to observe the behavior from a different room, therefore minimizing the interference with the test (for example, see the video file in the supplementary material).</li>
		</ol>
		</li>
		<li>Conditions: Observe and identify the three different behavioral conditions during the swim test: struggling, swimming, and immobility. Most researchers focus on immobility; a further differentiation between struggling and swimming is rarely useful and dramatically increases the complexity and duration of the analysis.
		<ol>
			<li>Struggling: The animal actively tries to escape from the threatening situation. This involves pawing the side of the cylinder with the head oriented toward the wall and movements of all limbs. The water surface is typically slightly turbulent.</li>
			<li>Swimming: The animal moves at least both hind paws and travels a distance throughout the water. It actively searches for a way out but does not try to overcome the glass wall of the vessel. Swimming does not involve lifting the paws above the water surface, and the body is usually oriented parallel to the walls of the cylinder. In this condition, animals frequently turn around or move in circles.</li>
			<li>Immobility: The animal keeps still, in a freezing-like position, and does not move at all or only moves the tail, or the forepaws to keep its head above the water surface. No distance is actively traveled except for passive floating, and no directed movement of the front paws is observed.</li>
		</ol>
		</li>
		<li>Tracking
		<ol>
			<li>Perform the assessment using offline video recordings. Use blinded ratings by two independent and experienced examiners and calculate averages between the two ratings.</li>
			<li>Repeat the ratings if the results of the two raters differ above a previously determined range. Continuously observe the mice as the different conditions frequently change between struggling, swimming, and immobility.</li>
			<li>Use a stopwatch to measure the total time spent in a focused stage (usually immobility) over the 10 min the mouse stays in the water. Consider a short latency of about a second before changing the ongoing time measurement (e.g., if an animal remains for 20 s in immobility and only moves once for less than a second and returns to immobility for another 10 s, the total immobility time is 30 s).</li>
		</ol>
		</li>
		<li>Statistics: Due to the relatively high inter-individual standard deviation (probably caused by a transfer of hierarchy-depending behavior from the cage to the swim test), mark or label the animals to perform paired (instead of unpaired) parametric tests afterward. Evaluate the normality distribution and, depending on the specific question, perform analysis of variance (ANOVA) with post-hoc t-tests or paired t-tests to compare the different groups. Perform the analysis using absolute values of immobility time (s) or as normalized values.
		<ol>
			<li>Absolute values: Give mean values of the immobility time from day 1 to day 5 and for the test day &#177; SEM (see Figure 1A). Compare the averaged values for day 1 and day 5, preferably using a paired t-test to validate the induction of a depressed state. If there is a significant difference between day 1 and 5, compare the mean values of day 5 to the averaged results of the test day. Ensure that a typical group size in one experiment is between 6 and 10 animals and expect significant differences between baseline and post-induction immobility times in wild-type animals. Comparing different groups with an unpaired t-test is difficult if absolute values are used because of baseline differences; therefore, use normalized values.</li>
			<li>Relative/Normalized values: Compare the different treatment effects by normalization to the individual result on day 5, and then express the values as a percentage of day 5 (see Figure 1B).</li>
		</ol>
		</li>
		<li>Control experiments 
		NOTE: The swimming performance might be correlated with locomotion. Substances that cause a hyper-locomotion could induce false-positive results (namely, a decrease of immobility time); as well as sedative agents could artificially increase immobility time.
		<ol>
			<li>Evaluate the changes in locomotion for unknown substances before performing the swim analysis. Use Open Field Test (OFT) in a separate group of animals for at least 10 min.</li>
			<li>Choose the same observation time (10 min) in the OFT as in the CDM to detect unspecific hyper-locomotive effects of the tested compound that might influence CDM read-out via measurement of immobility-time with high validity.</li>
			<li>In case of significant hyper-locomotive effects, do not evaluate the swim session to assess the anti-depressive potency but use different read-out methods (for instance, sucrose preference, tail suspension test, etc.).</li>
		</ol>
		</li>
	</ol>
	</li>
</ol>
4. Evaluation of the development of a depressive-like state
<ol>
	<li>To evaluate the development of a depressive disorder, assess each day of the induction phase to measure the immobility-time. 
	NOTE: In this case, a minor increase of immobility-time between each day describes resilience, whereas a stronger and earlier increase compared to untreated or wild-type animals represents an enhanced vulnerability to stress-induced despair. By treating mice before the swimming event, the preventive intervention or transgenic mouse lines could be assessed concerning the development of behavioral despair.</li>
</ol>

<ol>
	<li>James, S. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Global,+regional,+and+national+incidence,+prevalence,+and+years+lived+with+disability+for+354+diseases+and+injuries+for+195+countries+and+territories,+1990-2017:+a+systematic+analysis+for+the+Global+Burden+of+Disease+Study+2017.">Global, regional, and national incidence, prevalence, and years lived with disability for 354 diseases and injuries for 195 countries and territories, 1990-2017: a systematic analysis for the Global Burden of Disease Study 2017.</a> The Lancet. 392 (10159), 1789-1858 (2018).</li><li>Aleman, A., Denys, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mental+health:+A+road+map+for+suicide+research+and+prevention.">Mental health: A road map for suicide research and prevention.</a> Nature. 509 (7501), 421-423 (2014).</li><li>Greenberg, P. E., Fournier, A. -. A., Sisitsky, T., Pike, C. T., Kessler, R. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+economic+burden+of+adults+with+major+depressive+disorder+in+the+United+States+(2005+and+2010).">The economic burden of adults with major depressive disorder in the United States (2005 and 2010).</a> The Journal of Clinical Psychiatry. 76 (2), 155-162 (2015).</li><li>Nestler, E. J., Hyman, S. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Animal+Models+of+Neuropsychiatric+Disorders.">Animal Models of Neuropsychiatric Disorders.</a> Nature Neuroscience. 13 (10), 1161-1169 (2010).</li><li>Porsolt, R. D., Le Pichon, M., Jalfre, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Depression:+a+new+animal+model+sensitive+to+antidepressant+treatments.">Depression: a new animal model sensitive to antidepressant treatments.</a> Nature. 266 (5604), 730-732 (1977).</li><li>Can, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+mouse+forced+swim+test.">The mouse forced swim test.</a> Journal of Visualized Experiments: JoVE. (59), e3638 (2012).</li><li>Chatterjee, M., Jaiswal, M., Palit, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparative+evaluation+of+forced+swim+test+and+tail+suspension+test+as+models+of+negative+symptom+of+schizophrenia+in+rodents.">Comparative evaluation of forced swim test and tail suspension test as models of negative symptom of schizophrenia in rodents.</a> ISRN Psychiatry. 2012, 595141 (2012).</li><li>Reardon, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Depression+researchers+rethink+popular+mouse+swim+tests.">Depression researchers rethink popular mouse swim tests.</a> Nature. 571 (7766), 456-457 (2019).</li><li>Serchov, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Increased+signaling+via+adenosine+A1+receptors,+sleep+deprivation,+imipramine,+and+ketamine+inhibit+depressive-like+behavior+via+induction+of+Homer1a.">Increased signaling via adenosine A1 receptors, sleep deprivation, imipramine, and ketamine inhibit depressive-like behavior via induction of Homer1a.</a> Neuron. 87 (3), 549-562 (2015).</li><li>Holz, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enhanced+mGlu5+signaling+in+excitatory+neurons+promotes+rapid+antidepressant+effects+via+AMPA+receptor+activation.">Enhanced mGlu5 signaling in excitatory neurons promotes rapid antidepressant effects via AMPA receptor activation.</a> Neuron. 104 (2), 338-352 (2019).</li><li>Sun, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Increase+in+cortical+pyramidal+cell+excitability+accompanies+depression-like+behavior+in+mice:+A+transcranial+magnetic+stimulation+study.">Increase in cortical pyramidal cell excitability accompanies depression-like behavior in mice: A transcranial magnetic stimulation study.</a> Journal of Neuroscience. 31 (45), 16464-16472 (2011).</li><li>Hellwig, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Altered+microglia+morphology+and+higher+resilience+to+stress-induced+depression-like+behavior+in+CX3CR1-deficient+mice.">Altered microglia morphology and higher resilience to stress-induced depression-like behavior in CX3CR1-deficient mice.</a> Brain, Behavior, and Immunity. 55, 126-137 (2016).</li><li>Serchov, T., Heumann, R., van Calker, D., Biber, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Signaling+pathways+regulating+Homer1a+expression:+implications+for+antidepressant+therapy.">Signaling pathways regulating Homer1a expression: implications for antidepressant therapy.</a> Biological Chemistry. 397 (3), 207-214 (2016).</li><li>van Calker, D., Serchov, T., Normann, C., Biber, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recent+insights+into+antidepressant+therapy:+Distinct+pathways+and+potential+common+mechanisms+in+the+treatment+of+depressive+syndromes.">Recent insights into antidepressant therapy: Distinct pathways and potential common mechanisms in the treatment of depressive syndromes.</a> Neuroscience and Biobehavioral Reviews. 88, 63-72 (2018).</li><li>Serchov, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enhanced+adenosine+A1+receptor+and+Homer1a+expression+in+hippocampus+modulates+the+resilience+to+stress-induced+depression-like+behavior.">Enhanced adenosine A1 receptor and Homer1a expression in hippocampus modulates the resilience to stress-induced depression-like behavior.</a> Neuropharmacology. 162, 107834 (2020).</li><li>Quitkin, F. M., Rabkin, J. G., Ross, D., Stewart, J. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Identification+of+true+drug+response+to+antidepressants.+Use+of+pattern+analysis.">Identification of true drug response to antidepressants. Use of pattern analysis.</a> Archives of General Psychiatry. 41 (8), 782-786 (1984).</li><li>Normann, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Antidepressants+rescue+stress-induced+disruption+of+synaptic+plasticity+via+serotonin+transporter-independent+inhibition+of+L-type+calcium+channels.">Antidepressants rescue stress-induced disruption of synaptic plasticity via serotonin transporter-independent inhibition of L-type calcium channels.</a> Biological Psychiatry. 84 (1), 55-64 (2018).</li><li>Zanos, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=NMDAR+inhibition-independent+antidepressant+actions+of+ketamine+metabolites.">NMDAR inhibition-independent antidepressant actions of ketamine metabolites.</a> Nature. 533 (7604), 481-486 (2016).</li><li>Alboni, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fluoxetine+effects+on+molecular,+cellular,+and+behavioral+endophenotypes+of+depression+are+driven+by+the+living+environment.">Fluoxetine effects on molecular, cellular, and behavioral endophenotypes of depression are driven by the living environment.</a> Molecular Psychiatry. 22 (4), 552-561 (2017).</li><li>Niehusmann, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Coincidence+detection+and+stress+modulation+of+spike+time-dependent+long-term+depression+in+the+hippocampus.">Coincidence detection and stress modulation of spike time-dependent long-term depression in the hippocampus.</a> The Journal of Neuroscience: The Official Journal of the Society for Neuroscience. 30 (18), 6225-6235 (2010).</li><li>Schreiber, S. S., Tocco, G., Shors, T. J., Thompson, R. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Activation+of+immediate+early+genes+after+acute+stress.">Activation of immediate early genes after acute stress.</a> Neuroreport. 2 (1), 17-20 (1991).</li><li>. National centre for the replacement refinement and reduction of animals in research Available from: <a target="_blank" href="https://www.nc3rs.org.uk">https://www.nc3rs.org.uk</a> (2021)</li><li>Loss, C. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Influence+of+environmental+enrichment+vs.+time-of-day+on+behavioral+repertoire+of+male+albino+Swiss+mice.">Influence of environmental enrichment vs. time-of-day on behavioral repertoire of male albino Swiss mice.</a> Neurobiology of Learning and Memory. 125, 63-72 (2015).</li><li>Walker, W. H., Walton, J. C., DeVries, A. C., Nelson, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Circadian+rhythm+disruption+and+mental+health.">Circadian rhythm disruption and mental health.</a> Translational Psychiatry. 10 (1), 1-13 (2020).</li><li>Merrow, M., Spoelstra, K., Roenneberg, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+circadian+cycle:+daily+rhythms+from+behaviour+to+genes.">The circadian cycle: daily rhythms from behaviour to genes.</a> EMBO Reports. 6 (10), 930-935 (2005).</li><li>Holderbach, R., Clark, K., Moreau, J. -. L., Bischofberger, J., Normann, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enhanced+long-term+synaptic+depression+in+an+animal+model+of+depression.">Enhanced long-term synaptic depression in an animal model of depression.</a> Biological Psychiatry. 62 (1), 92-100 (2007).</li><li>Nissen, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Learning+as+a+model+for+neural+plasticity+in+major+depression.">Learning as a model for neural plasticity in major depression.</a> Biological Psychiatry. 68 (6), 544-552 (2010).</li><li>Kuhn, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=State-dependent+partial+occlusion+of+cortical+LTP-like+plasticity+in+major+depression.">State-dependent partial occlusion of cortical LTP-like plasticity in major depression.</a> Neuropsychopharmacology. 41 (6), 1521-1529 (2016).</li><li>Schwabe, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Stress+and+the+engagement+of+multiple+memory+systems:+integration+of+animal+and+human+studies.">Stress and the engagement of multiple memory systems: integration of animal and human studies.</a> Hippocampus. 23 (11), 1035-1043 (2013).</li><li>Ballan, R., Gabay, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Does+acute+stress+impact+declarative+and+procedural+learning.">Does acute stress impact declarative and procedural learning.</a> Frontiers in Psychology. 11, 342 (2020).</li></ol>

All the authors declare no conflicts of interest.

The CDM model represents a relevant and established model for testing the anti-depressive potency of new interventions and opens an extended time window for molecular or electrophysiological experiments to elucidate the pathophysiology of depression. Especially when combined with other tests to assess a depression-like state, CDM has a high face and concept validity. It combines subchronic stress and acquired helplessness for induction and produces a long-lasting depressive-like state. It is insensitive to the single app...

Affective disorders, such as major depressive disorder, are among the most frequent and challenging mental illnesses and are associated with high individual suffering1, an increase of suicide risk2, and cause a considerable socioeconomic burden3 for society. Despite its impact, treatment options are limited, and there is an urgent need for the development of novel antidepressive interventions, especially due to the innovation crisis in psychopharmacology over the last decades. In order to understand the pathophysiology of depression and test potential new agents, rational and valid animal models are urgently needed4. For almost half a century, the classical forced swim test (FST), originally described by Porsolt5, was used as induction and read-out for screening of potential novel antidepressants. It consists of a forced swim period for 5-15 min on day 1, subsequent one-time drug application, and evaluation of the portion mice spend immobile in water in another swim period on the following day. The immobility time was considered to represent a missing natural escape behavior and was thought to correlate with the degree of a depression-like state in the mice5.
The classical FST has been heavily criticized, not only in the scientific community6,7,8 but also in public media8. Most controversies around the FST are due to the short induction and treatment periods of only 1 day in the classical paradigm. It was argued that FST represents rather an acute trauma model than a state comparable to human depression. Moreover, the Porsolt test might be suitable as a screening tool for potential antidepressive agents, but it ignores the delayed onset of action of many antidepressants.
The chronic despair model (CDM)9,10,11,12,13,14,15, which is derived from the original FST, represents a more appropriate animal model for depression. In CDM, repeated swim stress over 5 consecutive days avoids acute traumatic effects. By failing to escape from a repeated and ongoing stressful situation, mice are thought to develop a state of helplessness, surrender, and ultimately despair. This paradigm is more comparable to current psychological theories for the development of depression in humans than a single acute trauma, which is commonly experienced at the onset of a posttraumatic stress disorder. The resulting depression-like state in CDM is stable for up to 4 weeks9 and therefore opens the possibility for longer treatment periods, which are better comparable to clinical conditions, where antidepressants usually need 2-4 weeks to show a benefit16.
The evaluation of the depressive-like state should then be multidimensional. The measurement of immobility time, such as in the classical FST, is useful, but should not be used as the only outcome parameter. Various methods, which are described below, should be able to map different dimensions of a depressive state in line with symptoms usually found in depressed humans. Suitable read-out assessments could include escape behavior (immobility time9,10,17), tail suspension test (TST)9, anhedonia (classical sucrose preference test (SPT)18), motivation-oriented behavior (nose-poke sucrose preference test (NPSPT)10), expectation/exploration-behavior (response to ambiguous signal19; Y-maze test9), electrophysiology (measurements of long-term plasticity (long-term potentiation, LTP; long-term depression, LTD)20), molecular assessments (activation patterns of immediate early genes (IEGs); further stress patterns21).
Theoretically, a repeated swim test can be used to induce a depressed state without any assessment of immobility time. However, it is strongly recommended to provide at least a proof-of-concept experimental series with immobility times. Additionally, CDM represents a suitable model to assess the development of a depressive-like state by measuring immobility time during the induction phase. Specific mouse strains or mice treated before swimming can be evaluated with respect to resilience or vulnerability to stress and the induction of behavioral despair.

<table><tbody><tr><td>Beaker, 2000 mL</td><td>Kimble Kimax</td><td>14000-2000</td><td>any vessel &gt;2000ml and diameter of 24-26 cm possible</td></tr><tr><td>Digital Thermometer</td><td>Hanna Instruments</td><td>846-4708</td><td>any digital thermometer possible</td></tr><tr><td>Digitalwaage 200 g Dipse</td><td>DIPSE</td><td>tp200</td><td>any digital scale possible</td></tr><tr><td>Lenovo ThinkCentre V50a-24IMB AiO 11FJ00DVGE - 60,5 cm</td><td>Lenovo</td><td>A 908278</td><td>any standard Personalcomputer possible</td></tr><tr><td>Logitech PTZ Pro</td><td>Logitech</td><td>1000005246</td><td>any high resolution camera possible</td></tr><tr><td>Stopwatch ROTILABO</td><td>Carl Roth</td><td>L423.1</td><td>any stopwatch possible</td></tr><tr><td>Timer ROTILABO</td><td>Carl Roth</td><td>A802.1</td><td>any timer possible</td></tr></tbody></table>

animal models of depression - chronic despair model (cdm)

Major depressive disorder is one of the most prevalent forms of mental illnesses and causes tremendous individual suffering and socioeconomic burden. Despite its importance, current pharmacological treatment is limited, and novel treatment options are urgently needed. One key factor in the search for potential new drugs is evaluating their anti-depressive potency in appropriate animal models. The classical Porsolt forced swim test was used for this purpose for decades to induce and assess a depressive-like state. It consists of two short periods of forced swimming: the first to induce a depressed state and the second on the following day to evaluate the antidepressant effect of the agent given in between the two swim sessions. This model might be suitable as a screening tool for potential antidepressive agents but ignores the delayed onset of action of many antidepressants. The CDM was recently established and represented a modification of the classical test with notable differences. Mice are forced to swim for 5 consecutive days, following the idea that in humans, depression is induced by chronic rather than by acute stress. In a resting period of several days (1-3 weeks), animals develop sustained behavioral despair. The standard read-out method is the measurement of immobility time in an additional delayed swim session, but several alternative methods are proposed to get a broader view of the mood status of the animal. Multiple analysis tools can be used targeting behavioral, molecular, and electrophysiological changes. The depressed phenotype is stable for at least 4 weeks, providing a time window for rapid but also subchronic antidepressant treatment strategies. Furthermore, alterations in the development of a depressive-like state can be addressed using this approach. CDM, therefore, represents a useful tool to better understand depression and to develop novel treatment interventions.

In the first swim session of the induction phase of CDM, mice usually show a mean immobility time between 190 s and 230 s, which constantly rises with every additional swim session (Figure 1A). This increase is more pronounced in the first 3 days and reaches a plateau-like phase during the last 2-3 days. The immobility-time measured on day 5 remains stable over up to 4 weeks, indicating stable behavioral despair. The antidepressant potency of an intervention ...

Watch this Scientific Journal Video about Animal Models of Depression - Chronic Despair Model CDM at JoVE.com

Animal Models of Depression - Chronic Despair Model CDM

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)

Major depressive disorder is one of the most prevalent forms of mental illnesses and causes tremendous individual suffering and socioeconomic burden. ...

animal-models-of-depression-chronic-despair-model-cdm

Research

JoVE Journal

Behavior

7.1K Views.  University of Freiburg. 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.

Video: Animal Models of Depression - Chronic Despair Model CDM

Using Chronic Social Stress to Model Postpartum Depression in Lactating Rodents

Exposure to chronic stress is a reliable predictor of depressive disorders, and social stress is a common ethologically relevant stressor in both animals and humans. However, many animal models of depression were developed in males and are not applicable or effective in studies of postpartum females. Recent studies have reported significant effects of chronic social stress during lactation, an ethologically relevant and effective stressor, on maternal behavior, growth, and behavioral neuroendocrinology. This manuscript will describe this chronic social stress paradigm using repeated exposure of a lactating dam to a novel male intruder, and the assessment of the behavioral, physiological, and neuroendocrine effects of this model. Chronic social stress (CSS) is a valuable model for studying the effects of stress on the behavior and physiology of the dam as well as her offspring and future generations. The exposure of pups to CSS can also be used as an early life stress that has long term effects on behavior, physiology, and neuroendocrinology.

This article describes the use of chronic resident intruder social stress as an ethologically relevant paradigm to model postpartum depression and anxiety in lactating rodents.

Exposure to chronic stress is a reliable predictor of depressive disorders, and social stress is a common ethologically relevant stressor in both animals ...

The Forced Swim Test as a Model of Depressive-like Behavior

The goal of the present protocol is to describe the forced swim test (FST), which is one of the most commonly used assays for the study of depressive-like behavior in rodents. The FST is based on the assumption that when placing an animal in a container filled with water, it will first make efforts to escape but eventually will exhibit immobility that may be considered to reflect a measure of behavioral despair. This test has been extensively used because it involves the exposure of the animals to stress, which was shown to have a role in the tendency for major depression. Additionally, the FST has been shown to share some of the factors that are influenced or altered by depression in humans, including changes in food consumption, sleep abnormalities and drug-withdrawal-induced anhedonia. The main advantages of this procedure are that it is relatively easy to perform and that its results are easily and quickly analyzed. Moreover, its sensitivity to a broad range of antidepressant drugs that makes it a suitable screening test is one of the most important features leading to its high predictive validity. Despite its appeal, this model has a number of disadvantages. First, the issue of chronic augmentation is problematic in this test because in real life patients need to be treated for at least several weeks before they experience any relief from their symptoms. Last, due to the aversiveness of the FST, it is important to take into account possible influences it might have on brain structure/function if brain analyses are to be carried out following this procedure.

This protocol describes the forced swim test, which is used for the study of depressive-like behavior in rodents. This procedure involves placing an animal in a container filled with water that eventually will lead to the exhibition of immobility behavior, which is considered to reflect behavioral despair.

The goal of the present protocol is to describe the forced swim test (FST), which is one of the most commonly used assays for the study of depressive-like ...

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

Stressful life events often increase the incidence of depression in humans. To study the mechanisms of depression, the development of animal models of depression is essential. Because there are several types of depression, various animal models are needed for a deeper understanding of the disorder. Previously, a mouse model of subchronic and mild social defeat stress (sCSDS) using a modified chronic social defeat stress (CSDS) paradigm was established. In the paradigm, to reduce physical injuries from aggressors, the duration of physical contact between the aggressor and a subordinate was reduced compared to in the original CSDS paradigm. sCSDS mice showed increased body weight gain, food intake, and water intake during the stress period, and their social behaviors were suppressed after the stress period. In terms of the face validity of the stress-induced overeating and overdrinking following the increased body weight gain, the sCSDS mice may show some features related to atypical depression in humans. Thus, a mouse model of sCSDS may be useful for studying the pathogenic mechanisms underlying depression. This protocol will help establish the sCSDS mouse model, especially for studying the mechanisms underlying stress-induced weight gain and polydipsia- and hyperphagia-like symptoms.

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.

Stressful life events often increase the incidence of depression in humans. To study the mechanisms of depression, the development of animal models of ...

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

Chronic, unresolved stress is a major risk factor for the development of clinical depression. While many preclinical models of stress-induced depression have been reported, the unpredictable chronic mild stress (UCMS) protocol is an established translationally-relevant model for inducing behavioral symptoms commonly associated with clinical depression, such as anhedonia, altered grooming behavior, and learned helplessness in rodents. The UCMS protocol also induces physiological (e.g., hypercortisolemia, hypertension) and neurological (e.g., anhedonia, learned helplessness) changes that are clinically associated with depression. Importantly, UCMS-induced depressive symptoms can be ameliorated through chronic, but not acute, treatment with common SSRIs. As such, the UCMS protocol offers many advantages over acute stress protocols or protocols that utilize more extreme stressors. Our protocol involves randomized, daily exposures to 7 distinct stressors: damp bedding, removal of bedding, cage tilt, alteration of light/dark cycles, social stresses, shallow water bath, and predator sounds/smells. By subjecting rodents 3-4 hr daily to these mild stressors for 8 weeks, we demonstrate both significant behavioral changes and poor health outcomes to the cardiovascular system. This approach allows for in-depth interrogation of the neurological, behavioral, and physiological alterations associated with chronic stress-induced depression, as well as for testing of new potential therapeutic agents or intervention strategies.

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.

Chronic, unresolved stress is a major risk factor for the development of clinical depression. While many preclinical models of stress-induced depression ...

The Mouse Forced Swim Test

The forced swim test is a rodent behavioral test used for evaluation of antidepressant drugs, antidepressant efficacy of new compounds, and experimental manipulations that are aimed at rendering or preventing depressive-like states. Mice are placed in an inescapable transparent tank that is filled with water and their escape related mobility behavior is measured. The forced swim test is straightforward to conduct reliably and it requires minimal specialized equipment. Successful implementation of the forced swim test requires adherence to certain procedural details and minimization of unwarranted stress to the mice. In the protocol description and the accompanying video, we explain how to conduct the mouse version of this test with emphasis on potential pitfalls that may be detrimental to interpretation of results and how to avoid them. Additionally, we explain how the behaviors manifested in the test are assessed.

The forced swim test is validated as an experimental approach to assess potential antidepressant efficacy in rodents. Experimental animals are placed in a tank of water and escape-related mobility behavior is quantified. The common procedures for the mouse version of this test are described.

The forced swim test is a rodent behavioral test used for evaluation of antidepressant drugs, antidepressant efficacy of new compounds, and experimental ...

Neuroscience

A New Method for Inducing a Depression-Like Behavior in Rats

Contagious depression is a phenomenon that is yet to be fully recognized and this stems from insufficient material on the subject. At the moment, there is no existing format for studying the mechanism of action, prevention, containment, and treatment of contagious depression. The purpose of this study, therefore, was to establish the first animal model of contagious depression.
Healthy rats can contract depressive behaviors if exposed to depressed rats. Depression is induced in rats by subjecting them to several manipulations of chronic unpredictable stress (CUS) over 5 weeks, as described in the protocol. A successful sucrose preference test confirmed the development of depression in the rats. The CUS-exposed rats were then caged with na&#239;ve rats from the contagion group (1 na&#239;ve rat/2 depressed rats in a cage) for an additional 5 weeks. 30 social groups were created from the combination of CUS-exposed rats and na&#239;ve rats.
This proposed depression-contagion protocol in animals consists mainly of cohabiting CUS-exposed and healthy rats for 5 weeks. To ensure that this method works, a series of tests are carried out - first, the sucrose preference test upon inducing depression to rats, then, the sucrose preference test, alongside the open field and forced-swim tests at the end of the cohabitation period. Throughout the experiment, rats are given tags and are always returned to their cages after each test.
A few limitations to this method are the weak differences recorded between the experimental and control groups in the sucrose preference test and the irreversible traumatic outcome of the forced swim test. These may be worth considering for suitability before any future application of the protocol. Nonetheless, following the experiment, na&#239;ve rats developed contagion depression after 5 weeks of sharing the same cage with the CUS-exposed rats.

This protocol describes a new model by which healthy rats could contract depression over a given time periodthrough contagion by exposure to chronic unpredictable stressed (CUS) rats.

Contagious depression is a phenomenon that is yet to be fully recognized and this stems from insufficient material on the subject. At the moment, there is ...

The Unpredictable Chronic Mild Stress Protocol for Inducing Anhedonia in Mice

Depression is a highly prevalent and debilitating condition, only partially addressed by current pharmacotherapies. The lack of response to treatment by many patients prompts the need to develop new therapeutic alternatives and to better understand the etiology of the disorder. Pre-clinical models with translational merits are rudimentary for this task. Here we present a protocol for the unpredictable chronic mild stress (UCMS) method in mice. In this protocol, adolescent mice are chronically exposed to interchanging unpredictable mild stressors. Resembling the pathogenesis of depression in humans, stress exposure during the sensitive period of mice adolescence instigates a depressive-like phenotype evident in adulthood. UCMS can be used for screenings of antidepressants on the variety of depressive-like behaviors and neuromolecular indices. Among the more prominent tests to assess depressive-like behavior in rodents is the sucrose preference test (SPT), which reflects anhedonia (core symptom of depression). The SPT will also be presented in this protocol. The ability of UCMS to induce anhedonia, instigate long-term behavioral deficits and enable reversal of these deficits via chronic (but not acute) treatment with antidepressants strengthens the protocol&#39;s validity compared to other animal protocols for inducing depressive-like behaviors.

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.

Depression is a highly prevalent and debilitating condition, only partially addressed by current pharmacotherapies. The lack of response to treatment by ...

Evaluating the Anti-depression Effect of Xiaoyaosan on Chronically-stressed Mice

In addition to the standardized use of antidepressant medications and psychotherapy, the usage of traditional Chinese medicine has lead to an overall improvement of patients with major depressive disorder (MDD). Therefore, the purpose of this study was to establish the mouse depressive model, observe the behavior changes associated with chronic unpredictable mild stress (CUMS), and then evaluate the anti-depression effect of Xiaoyaosan. Mice were randomly divided into four groups: a control group, a model group, a treatment group with Xiaoyaosan, and a treatment group with fluoxetine. All mice were individually kept in cages, and depression was induced in the mice by exposing them to several designed manipulations of CUMS for 21 days, as described in the protocol. Mice in the control group and model group received 0.5 mL of distilled water, while mice in the treatment groups received either Xiaoyaosan (0.25 g/kg/day) or fluoxetine (2.6 mg/kg/day). The drugs used in the study were given intragastrically daily during the entire three weeks. To estimate the depressive-like behaviors, a series of parameters including the coat state, body weight, open field test score, and sucrose preference test score were recorded. Data analysis showed that behaviors of model mice were significantly changed compared to behaviors of mice in the control group, which were improved by the treatment of Xiaoyaosan and fluoxetine. The current findings demonstrated the anti-depression effects of Xiaoyaosan on the behaviors of CUMS-induced mice and revealed that compounds from the Xiaoyaosan prescription may be worthwhile for treating depression, considering their beneficial effects on depressive-like behaviors.

Here, we present a protocol to establish the mouse depressive model, observe the behavior changes associated with chronic unpredictable mild stress (CUMS), and evaluate the anti-depression effect of Xiaoyaosan.

In addition to the standardized use of antidepressant medications and psychotherapy, the usage of traditional Chinese medicine has lead to an overall ...

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

Depression is not yet fully understood, but various causative factors have been reported. Recently, the prevalence of depression has increased. However, therapeutic treatments for depression or research on depression is scarce. Thus, in the present paper, we propose a mouse model of depression induced by movement restriction. Chronic mild stress (CMS) is a well-known technique to induce depressive-like behavior. However, it necessitates a complex procedure consisting of a combination of various mild stresses. In contrast, chronic immobilization stress (CIS) is a readily accessible chronic stress model, modified from a restraint model that induces depressive behavior by restricting movement using a restrainer for a certain period. To evaluate the depressive-like behaviors, the sucrose preference test (SPT), the tail suspension test (TST), and the ELISA assay to measure stress marker corticosterone levels are combined in the present experiment. The described protocols illustrate the induction of CIS and evaluation of the changes in behavior and physiological factors for the validation of depression.

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.

Depression is not yet fully understood, but various causative factors have been reported. Recently, the prevalence of depression has increased. However, ...

Developing a Rat Model for Bipolar Disorder

Bipolar disorder is a mental health condition characterized by extreme mood swings, including periods of emotional highs (mania) and lows (depression). While the exact underlying neurobiology is not yet fully understood, imbalances in neurotransmitter systems, particularly dopamine, appear to play a central role. For this reason, manipulations of dopaminergic pathways have been used to model mania or depression in rodents. However, models that accurately represent the typical switch between these two episodes are rare, limiting face validity. In a unique model, modern techniques are used to temporarily increase dopamine D1 receptor expression, which has been implicated in the pathology of bipolar disorder. A tetracycline-inducible lentiviral construct that expresses the dopamine D1 receptor under the control of the calmodulin kinase II alpha promoter is stereotactically injected into the medial prefrontal cortex of adult rats. Dopamine D1 receptor overexpression is achieved by adding the tetracycline analog doxycycline to the animals&#39; drinking water, leading to an increase in reward-related, impulsive, and risk-taking behaviors and a decrease in anxiety. These behaviors resemble a mania-like phenotype. By removing doxycycline from the drinking water, a depressive-like phenotype, characterized by increased helplessness and anhedonia, can be induced within the same animal. This article provides a step-by-step protocol for performing the surgery, as well as procedures for inducing the bipolar disorder-like phenotype. Additionally, considerations for assessing behavioral changes associated with mania-like and depressive-like behavior are described. This promising model, which demonstrates good construct and face validity, offers a valuable tool for further investigating the pathophysiological mechanisms of bipolar disorder.

This article presents a protocol for the induction of a unique rat model of bipolar disorder that captures both mania-like and depressive-like behavior.

Bipolar disorder is a mental health condition characterized by extreme mood swings, including periods of emotional highs (mania) and lows (depression). ...