Name: Intermittent Binge-Intake Model in Mice (Video) | JoVE Journal
Uploaded: 2025-01-10T14:40:00.000+00:00
Duration: 5 min 15 s
Description: Learn to establish an intermittent binge-intake model in mice using M&M's as a palatable food source. This validated protocol replicates anxiety-like behaviors and increased caloric intake seen in binge-eating disorder.

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We want to express our gratitude to the Biomedical Sciences PhD Program at the National Autonomous University of Mexico and to the support provided by Consejo Nacional de Humanidades, Ciencias y Tecnolog&#237;as (CONAHCYT), No. CVU: 11025222. This work was partially supported by the UNAM-DGAPA-PAPIIT grant IN207423. We thank Andrea Mondrag&#243;n Garc&#237;a and Jos&#233; Enrique Ram&#237;rez S&#225;nchez for their technical support.

All experiments were conducted according to the Mexican Official Standard of Technical Specifications for the Production and Use of Laboratory Animals (NOM-062-ZOO-1999) and in full compliance with the guidelines set forth by the Internal Committee for the Care and Use of Laboratory Animals (CICUAL) at the National Autonomous University of Mexico (UNAM).
1. Animals
<ol>
	<li>To follow this protocol, individually house 27 C57BLC/6 male mice, aged 1 month and weighing 16-20 g, in cages under controlled environmental conditions: a 12-12 h light-dark cycle (06:00 on, 18:00 off), maintaining the temperature at 21 &#177; 22 &#176;C and humidity levels at 70 &#177; 10%. 
	NOTE: In this study, we exclusively used male mice to eliminate the effects of female hormones during binge-type intake. However, in another study18, we applied the same model to female mice and observed no differences in binge intake.</li>
</ol>
2. Habituation
<ol>
	<li>Provide the 30-day-old mice with ad libitum access to standard chow and water. Place them in individual cages to facilitate measuring water and food consumption.</li>
	<li>Provide an enriched environment, including a paper tube, to all the mice and provide wood shavings as bedding.</li>
	<li>Measure body weight, food, and water consumption every 24 h for 7 days.</li>
</ol>
3. Binge eating model
<ol>
	<li>On day 7 of the habituation period, randomly assign the rodents (N= 27) into one of the following three groups.</li>
	<li>Provide all subjects with access to the HPF (nutritional content per 100: 480 kcal, 19g of fat, 70 g of carbohydrates, 5.1 g of protein, 0.13 g of salt) for 2 h (from 16:00 to 18:00) to minimize neophobia toward the HPF.</li>
	<li>Continue the specified feeding conditions for each group consistently for a period of 26 days.
	<ol>
		<li>Chow Group (CG) n = 9: provide ad libitum access to standard chow and water.</li>
		<li>Continuous Group (CoG) n = 9; provide ad libitum access to standard chow, HPF, and water.</li>
		<li>Intermittent Group (IG) n = 9: provide ad libitum access to standard chow and water but give them HPF only on specific days (1, 3, 5, 8, 10, 12, 15, 17, 19, 22, 24, and 26) for 2 h (from 16:00 to 18:00 h).</li>
	</ol>
	</li>
	<li>From the first session to session 26, measure body weight, standard chow intake, palatable food intake, and water consumption for all subjects at 16:00 h and again at 18:00 h.</li>
</ol>
4. Specifical instructions
<ol>
	<li>Place the standard chow on the cage racks and the HPF in small plastic containers. In each session, randomly place these containers at the corners of the cages (left posterior, right posterior, right anterior, and left anterior).</li>
	<li>Replenish water (280-300 mL) and standard chow (10-15 g) on Mondays, Wednesdays, and Fridays.</li>
	<li>Replace the HPF (4-4.5 g) daily for the CoG. For the IG, place the HPF (4-4.5 g) on Mondays, Wednesdays, and Fridays at 16:00 h. Randomly distribute the HPF by color (brown, blue, red, orange, green, and yellow), with no preference observed for any color.</li>
	<li>On day 7 of habituation, give the mice assigned to the CoG group the HPF at 16:00 h. Measure all the animals for standard chow intake, body weight, and water consumption.</li>
</ol>
5. Data analysis
<ol>
	<li>For 24 h intake measurements, record the values at 16:00 h on the previous day and 16:00 h on the current day. For instance, to evaluate the consumption of day 2, subtract the values for standard chow, the HPF, water, and body weight from day 1 from those of day 2 for the same parameters.</li>
	<li>For 2 h intake measurements, record the values at 16:00 h and 18:00 h on the same day. For example, subtract the values for standard chow, the HPF, water, and body weight at 16:00 h from those at 18:00 h for the same parameters.</li>
	<li>Collect all data daily at two time points (16:00 h and 18:00 h), input them into a spreadsheet, and perform kcal calculations.
	<ol>
		<li>To calculate the kcal, convert the grams consumed of standard chow and HPF to kcal based on the energy value of each food using a proportion (e.g., standard chow: 3.02 kcal/g; HPF: 4.80 kcal/g).</li>
	</ol>
	</li>
	<li>Analyze and plot the data using the software of choice.
	<ol>
		<li>For statistical analysis, use the Kruskal-Wallis and Mann-Whitney U tests.</li>
	</ol>
	</li>
</ol>
6. Open field test
NOTE: A 22-day, 14:00 h, open field test was conducted according to the protocol described by Kraeuter and colleagues19. This protocol consists of placing the animals in the center of a black acrylic sheet box. The floor of the arena was white, and the grid was black. Behavior was recorded and quantified with a video tracking system. The system recorded the distance, speed, and time the animals spent in the center or periphery of the arena.
<ol>
	<li>Preparation of the test room and open field apparatus
	<ol>
		<li>For the arena, use a square chamber of 60 cm (length) x 60 cm (width) x 20 cm (height) made of black nonporous plastic.</li>
		<li>Clean the device before and between uses with 70% vol/vol ethanol to remove odor cues.</li>
		<li>Place the camera above the apparatus for optimal exploration view.</li>
		<li>To minimize stress, maintain temperature and humidity like the normal housing conditions. If the mice are moved from one housing room to another for the experiment, acclimate them to the new room for at least 1 h each day before use.</li>
	</ol>
	</li>
	<li>Performing the open field test
	<ol>
		<li>To remove a single mouse from the cage, gently grasp its tail and place it in the center of the open field maze. Activate the video tracking software using the autostart feature, which detects the investigator and begins recording as soon as the investigator is no longer in focus of the camera.</li>
		<li>Allow the mouse to move freely along the maze for 5 min. During this time, the tracking software will record the movement.</li>
		<li>At the end of the trial period, remove the animal from the arena and return it to its home cage.</li>
		<li>Before cleaning the maze, visually count the feces deposited and record them for later analysis.</li>
		<li>Remove all fecal pellets and clean any urine stains. Spray the apparatus floor and walls with 70% ethanol, then wipe with a clean paper towel. Allow the ethanol solution to dry completely.</li>
		<li>After testing all animals, return to the vivarium, clean the arena, and organize all used materials.</li>
	</ol>
	</li>
	<li>Data analysis 
	NOTE: The video tracking software facilitates the extraction of a range of data, such as the animals&#39; average speed, the distance traveled, and the time spent in the center and periphery of the apparatus.
	<ol>
		<li>Select the distance traveled and the time spent by the animals in the center and the periphery of the apparatus.</li>
		<li>Export the data from the tracking software into a spreadsheet or other preferred format for further statistical analysis.</li>
		<li>Analyze the data with the software of choice.</li>
	</ol>
	</li>
</ol>

Investigating Binge Eating Disorder with a Palatable Food Access Mouse Model

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<li>Gainey, S. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Short-term+high-fat+diet+%28HFD%29+induced+anxiety-like+behaviors+and+cognitive+impairment+are+improved+with+treatment+by+glyburide%5BTitle%5D)+AND+%22Front+Behav+Neurosci%22%5BJournal%5D)">Short-term high-fat diet (HFD) induced anxiety-like behaviors and cognitive impairment are improved with treatment by glyburide.</a> Front Behav Neurosci. 10, 156(2016).</li>
<li>Bahi, A., Dreyer, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Effects+of+chronic+psychosocial+stress+on+%27binge-like%27+sucrose+intake+in+mice%5BTitle%5D)+AND+%22Prog+Neuropsychopharmacol+Biol+Psychiatry%22%5BJournal%5D)">Effects of chronic psychosocial stress on 'binge-like' sucrose intake in mice.</a> Prog Neuropsychopharmacol Biol Psychiatry. 120, 110625(2023).</li>
<li>Blanco-Gandía, M. C., Miñarro, J., Rodríguez-Arias, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Behavioral+profile+of+intermittent+vs+continuous+access+to+a+high+fat+diet+during+adolescence%5BTitle%5D)+AND+%22Behav+Brain+Res%22%5BJournal%5D)">Behavioral profile of intermittent vs continuous access to a high fat diet during adolescence.</a> Behav Brain Res. 368, s111891(2019).</li>
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<li>Awad, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Altered+reward+processing+following+sucrose+bingeing+in+male+and+female+mice%5BTitle%5D)+AND+%22Nutr+Neurosci%22%5BJournal%5D)">Altered reward processing following sucrose bingeing in male and female mice.</a> Nutr Neurosci. 27 (11), 1269-1282 (2024).</li>
<li>Chen, L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Ventral+tegmental+area+GABAergic+neurons+induce+anxiety-like+behaviors+and+promote+palatable+food+intake%5BTitle%5D)+AND+%22Neuropharmacology%22%5BJournal%5D)">Ventral tegmental area GABAergic neurons induce anxiety-like behaviors and promote palatable food intake.</a> Neuropharmacology. 173, 108114(2020).</li>
<li>Passeri, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Linking+drug+and+food+addiction%3A+an+overview+of+the+shared+neural+circuits+and+behavioral+phenotype%5BTitle%5D)+AND+%22Front+Behav+Neurosci%22%5BJournal%5D)">Linking drug and food addiction: an overview of the shared neural circuits and behavioral phenotype.</a> Front Behav Neurosci. 17, 1240748(2023).</li>
</ol>

The authors have no conflicts of interest to declare.

We used the binge-eating model proposed by Corwin in 2006 as a basis for developing this protocol. However, the present study modified the variable of providing palatable food for 2 h for the CoG8. Instead, we offered palatable food ad libitum to CoG, as it better simulates a reality where we have 24 h access to any food. We also used M&#38;M&#39;s as an HPF because they combine fats and sugars, unlike other binge-eating models that use either fat or sugar separately8</s...

According to data from the World Health Organization1, one in eight people worldwide is obese. Although eating is a physiological necessity, overeating without a physiological need is known to be a factor associated with overweight and obesity2. It is well known that humans prefer palatable foods high in calories, such as fats and sugars, over healthier foods3.
From the perspective of complexity science, eating disorders, particularly binge eating disorder (BED), are multifaceted phenomena that emerge from the interaction of various biological, psychological, and social factors exhibiting emerging behaviors such as cyclic binging patterns, complex compensatory behaviors, and dysfunctional interpersonal relationships4. BED is characterized by the excessive intake of high-calorie foods within a short period, approximately 2 h, according to the Diagnostic and Statistical Manual of Mental Disorders, 5th ed.5 (DSM-V). Additionally, BED is associated with behavioral comorbidities such as anxiety and cravings6,7. The complexity of these disorders requires better experimental models that can capture this interplay of factors and provide a more comprehensive understanding of binge eating behavior and its implications for obesity.
The most used models for binge-like behavior in basic research have primarily been rats. These models typically use limited and intermittent access to high-fat or high-sugar foods to induce binge-like behaviors8,9,10. An alternative approach involves providing palatable food for a short period (2 h) every 8 days, known as a cyclical model, using a highly palatable food (HPF) paste containing a mix of fats and sugars11. Another method presents the HPF as cookies unpredictably12. In mouse models, binge-like behaviors are often studied using the same cyclical model13. Another study with mice uses limited access to sucrose to induce binge-like behaviors14.
Despite extensive research on binge-like eating behaviors in rats, models for binge eating in mice remain limited. This study aims to propose a binge-like eating model using M&#38;M&#39;s (hereafter referred to as &#34;HPF&#34;) as an HPF according to the nutritional criteria of HPF proposed by Fazzino and colleagues15. Additionally, we aim to evaluate the rewarding properties of the HPF based on whether they are presented continuously or intermittently. In our research, we aimed to observe similarities between specific diagnostic DSM-V criteria for binge eating in humans within a murine model: consuming food within a short period (2 h), ingesting a large amount of food than usual, eating substantial amount despite being in a state of satiety, and experiencing binge episodes at the least twice a week.
The use of laboratory mouse models is an invaluable tool in biomedical research, especially for studying human diseases, as mice share 99% of their genes with humans16. Utilizing mice also offers significant advantages in understanding transcriptomic information and modifying therapeutic targets related to cellular functions and neural circuits associated with various diseases17.
Therefore, this study proposes a continuous and intermittent access model to M&#38;M&#39;s (an HPF made of fat and sugar) to observe binge-like eating in C57BL/6 mice without standard chow or water deprivation. This model is novel and necessary for biomedical research, as it represents a better approach to studying these complex problems. Standardizing animal models is essential to obtain reproducible and comparable results, which is fundamental to advancing translational research and developing effective therapeutic interventions for BED and obesity. The approach described here will expand the scope of binge-like eating behavior research in future studies, facilitating the testing of various drugs as potential therapeutic targets for BED. The implementation of more realistic and standardized models in mice will allow for a better understanding of the complex dynamics involved in eating disorders and their metabolic consequences, thus offering new alternatives for the treatment and prevention of BED.

<table><tbody><tr><td>ANY-maze Video Tracking Software</td><td>Stoelting Co.</td><td>4.73 version</td><td></td></tr><tr><td>C270 HD Webcam</td><td>Logitech</td><td>C270 HD</td><td>It is not necessary to use a specific video recorder; any webcam or camcorder with satisfactory image quality and resolution will suffice.</td></tr><tr><td>Gloves Ambiderm (black)</td><td>Ambiderm</td><td>7.50222E+12</td><td></td></tr><tr><td>GraphPad Prism 9</td><td>GraphPad</td><td></td><td></td></tr><tr><td>Laboratory Rodent Diet 5001</td><td>LabDiet</td><td>1320</td><td>https://di.facmed.unam.mx/comisiones/Composici%C3%B3n%20del%20alimento%20Laboratory%20Rodent%20Diet%205001.pdf</td></tr><tr><td>M&amp;amp;Ms 43.8g (Brown packaging)</td><td>M&amp;amp;M&amp;acute;s&amp;nbsp;</td><td></td><td>Nutritional content per 100 : 480 kcal, 19g of fat, 70 g of carbohydrates, 5.1 g of protein, 0.13 g of saltIngredients:&amp;nbsp;sugar, cocoa mass, skimmed&amp;nbsp;milk&amp;nbsp;powder, cocoa butter,&amp;nbsp;lactose, starch,&amp;nbsp;milk&amp;nbsp;fat, palm fat, glucose syrup, shea fat, stabiliser (gum arabic), dextrin, glazing agents (beeswax, carnauba wax), colours (E100, carmine, E132, E133, E150a, E150c, E150d, E153, E160a, E160e, E162, E163, E170), beetroot concentrate, emulsifiers (soya&amp;nbsp;lecithin, E445), salt, flavourings, palm kernel oil, antioxidant (E306). (May contain:&amp;nbsp;peanut, hazelnut, almond). Milk chocolate contains milk solids 14% minimum.&amp;nbsp;Milk chocolate contains vegetable fats in addition to cocoa butter.</td></tr><tr><td>Open Field maze&amp;nbsp;</td><td></td><td></td><td></td></tr><tr><td>Polypropylene Mice Cage</td><td>Orchid Scientific</td><td>SMP 01</td><td></td></tr><tr><td>Rodent drinkers</td><td>Sunnypet</td><td>SP-3656</td><td></td></tr><tr><td>Uline Balanza-220 g x 0.01 g</td><td>unike.mx</td><td>H-9884</td><td></td></tr><tr><td>UniMask4</td><td>Uniseal</td><td></td><td></td></tr><tr><td>Plastic bottles</td><td>unike.mx</td><td>S-14487</td><td></td></tr><tr><td>Sanitary Bed, Sawdust for Animals</td><td>BIOINVERT</td><td>No number</td><td>10 kg</td></tr></tbody></table>

intermittent binge-intake model in mice

Intermittent Binge-Intake Model in Mice

Obesity affects one in eight individuals globally. Overeating, especially high-calorie foods, plays a significant role in obesity. Binge eating disorder (BED) is a complex condition caused by a combination of biological, psychological, and social factors. It involves excessive consumption of high-calorie foods in a short period and is often linked to anxiety and cravings.
Here, we present a protocol employing continuous and intermittent access to a new highly palatable food (HPF) -- M&amp;M&#39;s -- to develop a mouse model for investigating binge eating disorder (BED). The HPF, chosen for its high fat and sugar content, represents an optimal food source to study due to its strong palatability, which makes it particularly suited for examining compulsive eating behaviors. Using C57BL/6 mice, we provided continuous or intermittent access to M&amp;M&#39;s while allowing unrestricted access to standard chow and water. By the 8th day, the mice in the intermittent access group exhibited pronounced binge-eating behaviors, which persisted through the 26th day. These mice also consumed significantly more calories -- predominantly from the confectionary -- than those in the continuous access and control groups. The contrast between continuous and intermittent access underscores the critical role of feeding schedules in promoting the overconsumption of palatable foods.
Furthermore, behavioral assessments revealed intermittent access to the HPF-induced anxiety-like behaviors, highlighting the psychological impact of access patterns on both eating behavior and emotional states. By incorporating these two distinct feeding paradigms, this study offers valuable insights into how the availability of highly palatable foods can exacerbate binge-eating tendencies. This model provides a more realistic approach to studying binge-eating behaviors and their metabolic consequences. It highlights the importance of standardized models in biomedical research. Our findings offer insights into the physiological and neural mechanisms underlying BED, which could pave the way for developing effective therapeutic interventions for BED and obesity.

Using HPF, we successfully established a binge-eating model in C57BL/6 mice. By day 8, binge-eating behavior (consuming excessive food for 2 h) was firmly established and maintained through day 26 (Figure 1A). During the intermittent sessions, the IG group exhibited significantly higher total kcal consumption (from both standard chow and the HPF than the CG and CoG) groups (Figure 1A, ### p &lt; 0.0001, *** p &lt; 0.0001, ** p &lt; 0.001, Kruskal-Wallis test). N...

A binge-eating model was established in mice using intermittent access to M&amp;M's (a highly palatable food made of fat and sugar). Mice with intermittent access to this high-fat, high-sugar food showed stress-like behaviors, increased caloric intake, and a preference for that food over standard chow compared to mice with continuous or no access to the food.

Obesity affects one in eight individuals globally. Overeating, especially high-calorie foods, plays a significant role in obesity. Binge eating disorder ...

Research

JoVE Journal

Behavior

Here we present a protocol to induce activity-based anorexia (ABA) in female adolescent mice. ABA is a condition of hyperactivity evoked by imposing food restriction on rodents with access to a running wheel. This phenomenon is being used as a model to study the underlying neurobiology of anorexia nervosa.

Using the Activity-based Anorexia Rodent Model to Study the Neurobiological Basis of Anorexia Nervosa

Alcohol Use Disorder (AUD) is a major national health problem and the development of more effective treatments is required to offset the needs of this patient population. To this end, the following protocol utilizes two simple rodent drinking models to assess the preclinical efficacy of lead anti-alcohol compounds.

Murine Drinking Models in the Development of Pharmacotherapies for Alcoholism: Drinking in the Dark and Two-bottle Choice

Mice individually housed with a running wheel while given limited access to food develop reductions in food consumption and increase activity on the running wheel. This experimental phenomenon is called activity-based anorexia. This paradigm provides an experimental tool for studying the neurobiology and behaviors underlying aspects of anorexia nervosa.

Assessing Activity-based Anorexia in Mice

The current article describes a detailed protocol for isocaloric 2:1 intermittent fasting to protect and treat against obesity and impaired glucose metabolism in wild-type and ob/ob mice.

Assessment of the Metabolic Effects of Isocaloric 2:1 Intermittent Fasting in Mice

Restricting the timing of food intake has emerged as a promising intervention to attenuate diet-induced metabolic diseases. This manuscript details the construction and use of an efficient system built in-house for measuring and manipulating rhythmic food intake in mice.

Manipulation of Rhythmic Food Intake in Mice Using a Custom-Made Feeding System

Presented here is a protocol for the 2BC/CIE model of alcohol dependence in mice to study alcohol use disorder.

Chronic Intermittent Ethanol Vapor Exposure Paired with Two-Bottle Choice to Model Alcohol Use Disorder

This protocol describes a standard intermittent-access two-bottle choice home cage drinking paradigm to model alcohol consumption in rats. In addition, it provides step-by-step instructions to augment the standard protocol with a DIY lick detection system that enables microstructural analysis of drinking behavior.

Modeling Alcohol Consumption in Rodents Using Two-Bottle Choice Home Cage Drinking and Microstructural Analysis

In the following experiment we describe a protocol for trace fear conditioning in mice. This type of associative memory includes a trace period that separates the neutral stimulus and the unconditioned stimulus.

Trace Fear Conditioning in Mice

This protocol describes the modified hole board, which is a behavioral test set-up that comprises the characteristics of an open field and a traditional hole board. This set-up enables the differential analysis of unconditioned behavior of small laboratory mammals as well as the analysis of cognitive abilities.

The Modified Hole Board - Measuring Behavior, Cognition and Social Interaction in Mice and Rats

This manuscript describes a protocol to examine the olfactory system of rodents. The olfactory habituation/dishabituation test will allow investigators to determine whether a mouse habituates to a repeatedly presented odor and whether the mouse demonstrates dishabituation when presented a novel odor.

 Testing for Odor Discrimination and Habituation in Mice

Learn to establish an intermittent binge-intake model in mice using M&M's as a palatable food source. This validated protocol replicates anxiety-like behaviors and increased caloric intake seen in binge-eating disorder.

Intermittent Binge-Intake Model in Mice

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