Name: Behavioral Characterization of an Angelman Syndrome Mouse Model (Video) | JoVE Journal
Uploaded: 2023-10-20T12:10:00.000+00:00
Duration: 11 min 5 s
Description: Learn behavioral characterization methods for Angelman syndrome using a validated mouse model. This protocol establishes reproducible tests including elevated plus maze, open field, and gait analysis for neuroscience research.

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

This research was supported by the Czech Academy of Sciences RVO 68378050, LM2018126 Czech Centre for Phenogenomics provided by MEYS CR, OP RDE CZ.02.1.01/0.0/0.0/16_013/0001789 (Upgrade of the Czech Centre for Phenogenomics: developing toward translation research by MEYS and ESIF), OP RDE CZ.02.1.01/0.0/0.0/18_046/0015861 (CCP Infrastructure Upgrade II by MEYS and ESIF), and OP RDI CZ.1.05/2.1.00/19.0395 (higher quality and capacity for transgenic models by MEYS and ERDF). In addition, this study received funding from the NGO &#34;Association of Gene Therapy (ASGENT)&#34;, Czechia (https://asgent.org/) and LM2023036 Czech Centre for Phenogenomics provided by Ministry of Education, Youth and Sports of the Czech Republic.

Validating a Murine Model of Angelman Syndrome Using Behavioral Tests

The authors have no conflicts of interest to disclose.

AS models created in different murine strains are commonly validated with tests of animal emotional state, motor functions, and cognitive abilities to facilitate comparison to human symptoms31,32. A motor deficit in AS models is the most consistent finding across laboratories, followed by an unchanged emotionality state of mutants and difficulties building nests31,32,33....

Angelman syndrome (AS) is a rare neurodevelopmental disease. The most common genetic origin of AS is a large deletion of the 15q11-q13 region of the maternally-derived chromosome, which is found in nearly 74% of patients1. Deletion of this region causes the loss of UBE3A, the main causative gene of AS that encodes an E3 ubiquitin ligase. The paternal allele of the UBE3A gene in neurons is silenced in a process known as imprinting. As a consequence, paternal imprinting of the gene allows only maternal expression in the central nervous system (CNS)2. Therefore, UBE3A gene deletion from the maternally-derived chromosome leads to the development of AS symptoms. In humans, AS manifests at around 6 months of age, with developmental retardation that persists throughout all developmental stages and results in severe debilitating symptoms in affected individuals3,4. The core symptoms of the disorder include the deficit of fine and gross motor skills, including jerky ataxic gait, serious speech impairment, and intellectual disability. Approximately 80% of AS patients also suffer from sleep disturbances and epilepsy. To date, the only available treatment are symptomatic drugs, which reduce epileptic seizures and improve sleep quality1. Therefore, the development of robust animal models with reproducible behavioral phenotypes alongside refined phenotyping analysis will be essential to elucidate the pathophysiological mechanisms of the disorder and discover effective medications and treatments.
The complexity of the human disorder affecting the CNS demands model organisms to possess a comparable genome, physiology, and behavior. Mice are popular as a model organism due to their short reproductive cycle, small size, and relative ease of DNA modification. In 1984, Paul Willner proposed three basic disease model validation criteria: the construct, face, and predictive validity, which are used to determine the model&#39;s value5. Simply, construct validity reflects the biological mechanisms responsible for the disorder development, face validity recapitulates its symptoms, and predictive validity describes the model response to therapeutic drugs.
To adhere to the above principles, we have chosen the most common genetic etiology, a large deletion of the maternal 15q11.2-13q locus including the UBE3A gene, to create AS model mice. We used the CRISPR/Cas9 technique to delete a 76,225 bp long region spanning the entire UBE3A gene, encompassing both the coding and non-coding elements of the gene, in mice from a C57BL/6N background6. We then bred the animals to obtain UBE3A+/&#8722; heterozygous mice. For face validation of the model, we used animals from crosses of UBE3A+/&#8722; females and wild-type males to gain UBE3A+/- progeny (strain named C57BL/6NCrl-UBE3A&#60;em1(IMPC)Ccpcz&#62;/Ph and later assigned as UBE3AmGenedel/+) and control littermates. We tested their fine and gross motor skills, emotionality, and affect to recapitulate core AS symptoms. In a previous article, we also evaluated the animals&#39; cognitive functions, as AS patients also suffer from intellectual disability6. However, we found no cognitive impairments in UBE3AmGenedel/+ mice, perhaps due to the young age of the animals at the time of testing7. Later examination of the older animals, around 18 weeks old, revealed a deficit in behavioral flexibility during reversal learning in the place preference paradigm. However, the complexity of the employed equipment for this analysis requires a separate methodological module and it is not included here.
The behavioral tests presented here belong to the common phenotyping tools in genetic research, thanks to their high predictive value and sufficient construct validity8,9,10. We used these tests to validate a mouse model of AS by recapitulating core symptoms of the human disease in a reproducible, age-independent manner. The emotionality of the animal was evaluated in the elevated plus maze and open field tests. Both of these tests are based on the approach-avoidance conflict, where animals explore a new environment in search of food, shelter, or mating opportunities while simultaneously avoiding anxiogenic compartments11. Additionally, the open field test is used to test a mouse&#39;s locomotor activity8. The tail suspension test is widely used in depression research to screen for new antidepressant drugs or depressive-like phenotypes in mouse knockout models12. This test evaluates the despair that animals develop over time in an inescapable situation. Motor learning and detailed gait characteristics were determined on the rotarod and in DigiGait, respectively. Animal endurance on the accelerating rod characterizes its balance and movement coordination skills, while detailed analysis of a mouse&#39;s step patterns is a sensitive evaluation of neuromuscular impairments connected to many neurogenerative movement disorders13,14,15. The nestlet shredding test is part of the standard methodology for detecting impulsive behavior in rodents, and since it utilizes natural rodent building behavior, it indicates the animal&#39;s well-being16,17.
The size of the experimental groups was a result of a compromise to meet the 3R rule demands and efficient usage of colony breeding performance. However, to obtain statistical power, the groups had no fewer than 10 individuals, due to the establishment of a sufficient amount of breeding pairs. Unfortunately, breeding performance did not always result in a sufficient number of animals.

<table><tbody><tr><td>Cages, individually ventilated</td><td>Techniplast</td><td></td><td></td></tr><tr><td>DigiGait</td><td>Mouse Specifics, Inc., 2 Central Street Level&lt;br/&gt; Unit 110&lt;br/&gt; Framingham, MA 01701, USA</td><td>Equipment was tendered, no catalogue&amp;nbsp; number was provided, nor could be find on company&#039;s web site</td><td>Detailed analysis of mouse gait, hardware and software provided.&amp;nbsp;</td></tr><tr><td>FDA Nestlet squares</td><td>Datesand Ltd., 7 Horsfield Way, Bredbury, Stockport SK6, UK</td><td>Material was bought from Velaz vendor via direct email request. Velaz do not provide any catalogue no.</td><td>Cotton nestlets for nest building test. Nestlet discription: 2-3 g each, with diameter around 5 x 5 x 0.5cm.</td></tr><tr><td>Mouse chow</td><td>Altramion</td><td></td><td></td></tr><tr><td>Rotarod</td><td>TSE Systems GmbH, Barbara-McClintock-Str.4&lt;br/&gt; 12489 Berlin, Germany</td><td>Equipment was tendered, no catalogue&amp;nbsp; number was provided, nor could be find on company&#039;s web site</td><td>Rotarod for 5 mice, hardware and software provided. Drum dimensions: Diameter: 30 mm, width per lane: 50 mm, falling distance 147 mm.</td></tr><tr><td>Tail Suspension Test</td><td>Bioseb, In Vivo Research Instruments, 13845 Vitrolles&lt;br/&gt; FRANCE</td><td>Reference: BIO-TST5</td><td>Fully automated equipment for immobility time evaluation of 3 mice hanged by tail, hardware and software provided</td></tr><tr><td>Transpore medical tape</td><td>Medical M, Ltd.</td><td>P-AIRO1291</td><td>The tape used to attach an animal to the hook by its tail.</td></tr><tr><td>Viewer - Video Tracking System</td><td>Biobserve GmbH, Wilhelmstr. 23 A&lt;br/&gt; 53111 Bonn, Germany</td><td>Equipment with software were tendered, no catalogue&amp;nbsp; number was provided, nor could be find on company&#039;s web site</td><td>Software with custom made hardware: maze, IR base, IR sensitive cameras. Custom-made OF dimensions: 42 x 42 cm area, 49 cm high wall, central zone area: 39 cm&lt;sup&gt;2&lt;/sup&gt;. A custom-made EPM was elevated 50 cm above the floor, with an open arm 79 cm long,&amp;nbsp; 9 cm wide, and closed arm 77 cm long, 7.6 cm wide.&amp;nbsp;</td></tr></tbody></table>

behavioral characterization of an angelman syndrome mouse model

Behavioral Characterization of an Angelman Syndrome Mouse Model

This manuscript describes a battery of behavioral tests available to characterize Angelman syndrome (AS)-like phenotypes in an established murine model of AS. We use the rotarod learning paradigm, detailed gait analysis, and nest building test to detect and characterize animal motor impairments. We test animal emotionality in the open field and elevated plus maze tests, as well as the affect in the tail suspension test. When AS mice are tested in the open field test, the results should be interpreted with care, since motor dysfunctions influence mouse behavior in the maze and alter activity scores.
The reproducibility and effectiveness of the presented behavioral tests has already been validated in several independent Uba3a mouse lines with different knockout variants, establishing this set of tests as an excellent validation tool in AS research. Models with the relevant construct and face validity will warrant further investigations to elucidate the pathophysiology of the disease and grant the development of causal treatments.

Elevated plus maze and open field tests 
The EPM and OF tests use the natural tendency of rodents to explore new environments18,19. The exploration is governed by an approach-avoidance conflict, where rodents choose between the exploration of a new environment and avoidance of possible danger. Animals explore unknown places in search for shelter, social contact, or foraging. However, new places may involve risk factors such as predators or com...

This manuscript presents a set of highly reproducible behavioral tests to validate an Angelman syndrome mouse model.

This manuscript describes a battery of behavioral tests available to characterize Angelman syndrome (AS)-like phenotypes in an established murine model of ...

behavioral-characterization-of-an-angelman-syndrome-mouse-model

Research

JoVE Journal

Behavior

Inchworming is a highly repetitive synchronous digging motion displayed by BTBR T+ Itpr3tf/J (BTBR) mice when placed in a testing cage with sufficient sawdust bedding.  The procedure is a modification of the juvenile social interaction protocol and is used here to assess repetitive motor stereotypies relevant to Autism Spectrum Disorder.

Inchworming: A Novel Motor Stereotypy in the BTBR T+ Itpr3tf/J Mouse Model of Autism

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

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

Footprint analysis is a low-cost alternative to digitized gait analysis programs for researchers quantifying movement abnormalities in mice. Because of its speed, simplicity, and longitudinal potential, it is ideal for behavioral phenotyping of mouse models.

Low-Cost Gait Analysis for Behavioral Phenotyping of Mouse Models of Neuromuscular Disease

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

Rodent models are valuable tools for studying core behaviors related to autism spectrum disorder (ASD). In this article, we expound on two behavioral tests for modeling the core features of ASD in mice: self-grooming, which assesses repetitive behavior, and the three-chamber social interaction test, which documents social impairments.

Strategies for Assessing Autistic-Like Behaviors in Mice

We demonstrate the utility of nest building behavior in laboratory mice as an indicator of welfare. Nest scoring is a sensitive technique that is altered by temperature, illness, and aggression. The time to integrate into nest test (TINT) is a simple cage-side assessment that can detect postoperative pain.

Nest Building as an Indicator of Health and Welfare in Laboratory Mice

Prolonged and comprehensive monitoring of mice in a home-cage environment provides a deeper understanding of aberrant behavior in murine models of brain diseases. This paper describes the Integrated Behavioral Station (INBEST) as the key component of contemporary behavioral analysis.

Behavioral Phenotyping of Murine Disease Models with the Integrated Behavioral Station (INBEST)

Mouse models have become increasingly popular in studies of behavioral neuroscience.  As models advance, it is important to develop sensitive behavioral measures specific to the mouse.  This protocol describes the Pasta Matrix Reaching Task, which is a skilled motor task for use in mouse models of stroke.

Compensatory Limb Use and Behavioral Assessment of Motor Skill Learning Following Sensorimotor Cortex Injury in a Mouse Model of Ischemic Stroke

Motor control and balance performance are known to deteriorate with age. This paper presents a number of standard noninvasive behavioral tests with the addition of a simple rotary stimulus to challenge the vestibular system and show changes in balance performance in a murine model of aging.

Behavioral Assessment of the Aging Mouse Vestibular System

Learn behavioral characterization methods for Angelman syndrome using a validated mouse model. This protocol establishes reproducible tests including elevated plus maze, open field, and gait analysis for neuroscience research.

Behavioral Characterization of an Angelman Syndrome Mouse Model

Explore More Videos