MDC Mouse

What ethical considerations guide animal experimentation at the Max Delbrück Center?

The Max Delbrück Center (MDC) employs a comprehensive ethical framework that prioritizes animal welfare while recognizing the current necessity of animal models in certain types of medical research. The MDC's Preclinical Research Center creates optimal conditions for animal testing with a focus on minimizing suffering through several practical approaches:

• Implementation of the 3R principles (Replacement, Reduction, Refinement)

• Use of advanced monitoring systems that allow data collection with minimal disruption to normal animal behavior

• Temperature, humidity, and light cycle control that mimics natural conditions (approximately 22°C, 55% humidity, with automated day-night cycles)

• Continuous environmental monitoring that allows for the collection of physiological data without invasive procedures

As Professor Thomas Sommer, interim Scientific Director at MDC, notes, "We will not be able to do away with animal testing entirely for the foreseeable future. This makes it all the more important that we are constantly working to make experiments even more animal friendly and predictive." The center recognizes that while artificial intelligence, advanced sequencing methods, organoids, and cell culture systems are valuable tools, they cannot yet fully simulate an entire organism's complexity.

How do researchers select appropriate mouse strains for behavioral experiments?

Strain selection is a fundamental consideration in behavioral testing that significantly impacts experimental outcomes. A methodological approach to strain selection includes:

• Understanding baseline behavioral characteristics: Each strain and substrain has distinct behavioral profiles that must be characterized before experimental use. For example, C57BL/6 mice are commonly used but have documented behavioral differences between substrains (C57BL/6J vs. C57BL/6N) .

• Considering sex differences: Both male and female mice should be evaluated in preliminary testing to understand sex-specific behavioral patterns before proceeding with experimental cohorts .

• Documenting genetic background: Researchers must maintain detailed records of:

  • Specific substrain used

  • Number of generations bred and backcrossed

  • Hybrid status if applicable

• Accounting for genetic manipulation effects: Reagents used in creating genetically manipulated mice (e.g., neo-cassettes, cre-drivers) may influence behavioral responses of wild-type controls, particularly with minimal backcrossing through generations .

Before using any mouse model, researchers should establish baseline behavioral values for each specific strain in each behavioral test to differentiate between strain-inherent behaviors and experimental effects. In cases without established baselines, a full cohort should be evaluated to determine expected values for each assay .

What specialized monitoring systems are available for behavioral phenotyping of mice?

Modern research facilities employ sophisticated monitoring systems that allow for comprehensive behavioral phenotyping while minimizing stress to the animals. The PhenoMaster environmental chambers at the Max Delbrück Center represent one such system with the following capabilities:

• Automated physiological monitoring: These systems can track weight, food intake, and activity levels without handling the animals. For example, specialized tubes within the cage automatically measure mouse weight, while food dispensers document feeding patterns .

• Movement tracking: Infrared photoelectric sensors record detailed movement patterns that can be retrospectively analyzed, including wheel rotations and general locomotor activity .

• Metabolic assessment: Through air composition analysis within the cages, researchers can measure calorie expenditure and detect specific metabolic markers (such as acetone in breath, which is relevant for diabetes research) .

• Environmental consistency: These systems maintain stable environmental conditions (22°C, 55% humidity, automated light cycles) to eliminate confounding variables while collecting data continuously .

These technologies are particularly valuable because they allow data collection while mice remain in familiar environments, significantly reducing stress-related confounds and improving data reliability. The non-invasive nature of these methods also supports ethical considerations by refining experimental approaches to reduce animal suffering .

How does the Single Mouse Experimental Design differ from conventional approaches, and what are its advantages?

The Single Mouse Experimental Design represents a significant methodological advancement in preclinical testing that addresses limitations of conventional approaches. This design features:

• Structure: Each mouse receives a different patient-derived xenograft, with endpoints being tumor regression and Event-Free Survival (EFS). No control (untreated) tumor is used in this design .

• Efficiency: Using one mouse per treatment group allows inclusion of approximately 20 models for every one used in conventional testing experiments (which typically use 10 mice per treatment and control group). This represents a substantial reduction in animal use while increasing experimental scope .

• Enhanced genetic diversity: The approach enables testing across a much broader range of tumor models, better representing the genetic/epigenetic diversity of cancer types. For example, instead of the 6-8 models per disease type used in the Pediatric Preclinical Testing Program (PPTP), researchers can include 30 or more models, substantially improving representation of genetic heterogeneity .

• Biomarker identification: The increased model diversity enables identification of genetic characteristics that may predict drug response, improving clinical translation. For example, testing in a large panel of melanoma xenografts revealed that BRAF-mutant models respond to BRAF inhibitors while wild-type BRAF models show less sensitivity .

Validation studies have confirmed the feasibility of this approach. For instance, a prospective evaluation using the long-acting PEGylated SN-38 prodrug (PLX038A) successfully identified responsive and non-responsive tumor models, with results correlating with previous data from conventional testing of irinotecan. Statistical analysis of mutation data from responsive versus non-responsive models (using Fisher's exact test) successfully identified genetic markers associated with treatment response .

What methodological steps ensure reliable and reproducible behavioral phenotyping in mice?

Establishing reliable behavioral phenotyping requires systematic methodology to ensure data reproducibility across technicians and laboratories. Key methodological considerations include:

• Standardized training and validation:

  • Behavioral testing should be conducted exclusively by trained technicians with demonstrated proficiency

  • Inter-rater reliability assessments should be conducted regularly

  • Standardized protocols should be documented in detail

• Environmental control:

  • Testing should occur at consistent times of day to minimize circadian influences

  • Temperature, humidity, and lighting conditions must be standardized

  • Background noise and other environmental variables should be minimized or controlled

  • Testing rooms should be separate from housing facilities

• Test battery design:

  • Order of testing should progress from least to most stressful to minimize carryover effects

  • Adequate recovery periods between tests (typically 2-3 days minimum)

  • Consistent handling procedures across all animals and tests

  • Validation of each assay with positive and negative controls

• Data collection standardization:

  • Use of automated systems where possible to reduce observer bias

  • Blinding of experimenters to treatment conditions

  • Consistent scoring criteria applied across all animal subjects

  • Multiple behavioral endpoints to provide convergent evidence

How do researchers effectively measure temperature perception in mice for neurosensory studies?

Measuring temperature perception in mice presents unique challenges since mice cannot verbally report sensations. Researchers at the MDC have developed methodological approaches to overcome this limitation:

• Behavioral conditioning technique:

  • Mice are trained to provide an immediate behavioral response (licking) when they detect a cooling sensation on their forepaw

  • This creates a direct, measurable report of perception rather than relying on indirect evidence

  • The approach allows precise measurement of minimum detectable temperature changes (approximately two degrees below skin temperature)

• Advantages over traditional methods:

  • Previous approaches relied on preference tests (e.g., observing where mice choose to spend time on surfaces of different temperatures), which only provide indirect evidence of perception

  • The conditioned response method provides immediate temporal data about perception

  • This approach enables more direct comparison with human perceptual abilities

• Neural correlates:

  • This behavioral paradigm can be combined with neurophysiological recording to correlate perception with cortical activity

  • Research has revealed that temperature sensing and touch perception are processed in the same region of the brain

  • The method enables investigation of specific neural circuits involved in thermal perception

This methodological approach has demonstrated that mice can perceive cooling of just two degrees below skin temperature, which is remarkably similar to human perceptual thresholds. This provides valuable validation for using mice as models for studying somatosensory processing with direct translational relevance to human perception .

How do researchers identify and utilize molecular markers for specific brain regions in mouse models?

Identifying molecular markers for specific brain regions requires systematic methodological approaches, particularly for complex structures like the mediodorsal thalamus (MD). The process typically involves:

• Developmental timing assessment:

  • Researchers must first establish when distinct subdivisions become anatomically identifiable

  • For example, in the mediodorsal thalamus, subdivision differentiation becomes apparent through Nissl staining at around postnatal day 15

• In situ hybridization (ISH) analysis:

  • This technique allows visualization of gene expression patterns in tissue sections

  • Systematic screening of candidate genes across brain regions identifies subdivision-specific expression

  • For the MD thalamus, researchers examine expression patterns along both medial-lateral and anterior-posterior axes

• Subdivision-specific marker identification:

  • Genes uniquely expressed in specific subdivisions are cataloged

  • Combinations of genes that together define a region are documented

  • For the MD, regions are classified as medial MD (MDm), central MD (MDc), and lateral MD (MDl)

• Region-specific mouse line development:

  • Once markers are identified, region-specific Cre driver mouse lines can be developed

  • For example, researchers developed MDm-specific and MDc-specific Cre mouse lines to enable targeted manipulation

• Projection mapping:

  • These region-specific mouse lines allow detailed mapping of neural projections

  • For the MD thalamus, this revealed diverse region- and layer-specific prefrontal cortex projection patterns

This methodology has revealed important species differences in gene expression patterns. When comparing classified MD marker genes between mice and common marmosets (a non-human primate model), researchers identified diverging gene expression patterns, highlighting the species-specific organization of cell types and their projections . This finding underscores the importance of careful cross-species validation when translating findings from mouse models to human applications.

What approach should researchers take when analyzing contradictory experimental results in mouse studies?

When researchers encounter contradictory experimental results in mouse studies, a systematic analytical approach is essential:

• Strain and genetic background analysis:

  • Verify complete documentation of strain, substrain, and genetic background

  • Assess whether contradictions may arise from subtle genetic differences between supposedly similar mouse lines

  • Consider genetic drift in laboratory mouse colonies as a potential factor

• Methodological standardization assessment:

  • Compare detailed experimental protocols between contradictory studies

  • Identify variations in testing conditions, handling procedures, or data collection methods

  • Evaluate potential environmental confounds (time of day, lighting, noise, experimenter effects)

• Cross-laboratory validation:

  • Implement standardized protocols across multiple laboratories

  • Use identical equipment, environmental conditions, and handling procedures

  • Employ blinded analysis to minimize experimenter bias

• Multi-modal data integration:

  • Correlate behavioral findings with biochemical, electrophysiological, or imaging data

  • Seek convergent evidence from multiple methodological approaches

  • Apply statistical methods specifically designed to assess reproducibility

• Species-specific considerations:

  • For translational research, evaluate whether contradictions reflect species-specific differences

  • Compare findings with data from other model organisms and human studies when available

  • Consider evolutionary divergence in neural circuitry, as seen in comparative studies of mouse and marmoset brain organization

Contradictory results often highlight important biological variables rather than experimental failures. In the case of mediodorsal thalamus research, for example, contradictory findings from lesion studies led to more sophisticated analyses revealing that "a subpopulation of MD projects to PFC and plays distinctive roles on cognitive function" . This understanding advanced the field beyond simple lesion approaches to more targeted optogenetic and chemogenetic manipulations of specific neural circuits.

How can researchers optimize pharmacological testing protocols in mouse models for improved translational outcomes?

Optimizing pharmacological testing in mouse models requires careful methodological considerations to enhance translational relevance:

• Comprehensive behavioral phenotyping:

  • Establish baseline behavioral profiles for your specific mouse strain before drug testing

  • Include multiple behavioral tests that assess different aspects of the target condition

  • Ensure technicians conducting tests demonstrate proficiency through standardized training

• Pharmacokinetic characterization:

  • Determine drug metabolism, distribution, and blood-brain barrier penetration in mice

  • Calculate appropriate dosing based on mouse-specific pharmacokinetics rather than simple weight-based scaling from human doses

  • Consider using long-acting formulations (like PLX038A, the PEGylated SN-38 prodrug) that provide sustained drug exposure

• Genetic diversity incorporation:

  • Implement testing across multiple mouse strains or patient-derived xenograft models

  • Consider single mouse experimental designs to increase model diversity within resource constraints

  • Correlate drug response with genetic/molecular characteristics to identify potential biomarkers

• Endpoint optimization:

  • Select endpoints with clear translational relevance to human clinical outcomes

  • For cancer studies, consider both tumor regression and Event-Free Survival (EFS)

  • For neurobehavioral research, employ endpoints that closely model human symptoms or functional outcomes

• Biomarker identification:

  • Correlate treatment response with molecular/genetic characteristics

  • For example, PLX038A sensitivity correlated with wild-type TP53 or specific mutations in DNA damage response pathways

  • These correlations can identify patient populations most likely to benefit in subsequent clinical trials

The single mouse experimental design has demonstrated particular value for pharmacological testing. In a validation study with PLX038A, researchers found that tumor sensitivity correlated with sensitivity to irinotecan, validating the single mouse approach for identifying agents with the same mechanism of action. This approach enables testing across a much broader range of genetic backgrounds while using fewer animals, potentially improving predictive validity for heterogeneous human diseases .

How do researchers balance statistical power requirements with ethical considerations in determining mouse cohort sizes?

Balancing statistical power with ethical animal use presents a fundamental challenge in experimental design. Researchers can address this through several methodological approaches:

• Power analysis refinement:

  • Conduct detailed power analyses based on expected effect sizes and variability

  • Consider alternative statistical approaches that may require fewer animals

  • Utilize historical control data when appropriate to reduce control group sizes

• Experimental design optimization:

  • Implement factorial designs to assess multiple variables simultaneously

  • Consider repeated measures designs when appropriate to reduce animal numbers

  • Evaluate whether alternative model systems can address certain research questions

• Single mouse experimental design implementation:

  • For certain research questions (particularly in oncology), consider single mouse designs

  • This approach allows testing in 20 different tumor models using the same number of mice as a conventional design with just one model

  • Endpoints focus on tumor regression and Event-Free Survival rather than relative tumor growth comparisons

• Advanced monitoring technology utilization:

  • Employ systems like the PhenoMaster environmental chambers that collect multiple data points from each animal

  • These systems enable continuous monitoring of weight, activity, feeding, and metabolic parameters

  • The multiparametric approach maximizes data obtained from each experimental subject

• Cross-laboratory standardization:

  • Implement consistent protocols across research groups to reduce study replication

  • Share raw data to enable meta-analyses with increased statistical power

  • Establish tissue and data repositories to maximize utility of each experimental animal

Research at the Max Delbrück Center exemplifies this balanced approach: "Here at the Max Delbrück Center, we want to find new therapies for sick people that doctors have so far been unable to treat effectively," while simultaneously working "to make experiments even more animal friendly and predictive." This dual focus on scientific advancement and ethical animal use drives methodological innovation in experimental design.

What methodological approaches help researchers address the challenge of behavioral variability in mouse models?

Behavioral variability presents a significant challenge in mouse research. Effective methodological approaches to address this issue include:

Researchers should recognize that some degree of behavioral variability reflects natural biological diversity rather than methodological weakness. The goal should be to understand and account for this variability rather than eliminating it entirely, as excessive standardization can actually reduce the translational relevance of findings .