x
[quotes_form]
Cytology

Clinical Cytogenetics – Pt.2: A Study of Chromosomal Aberrations

 

Introduction

Since the discovery of chromosomes, the cytogenetic field has evolved rapidly. The techniques have unfolded several hidden mysteries of chromosomes. And later, it gave rise to two branches: molecular and clinical cytogenetics. The studies in clinical cytogenetics suggested that the structural and numerical aberrations of chromosomes (autosomal and sex) are the cause of several abnormalities in human beings. For example, Down’s syndrome, Turner’s syndrome, cancer, etc. The molecular cytogenetic techniques (for example FISH, multicolor karyotyping, CGH, etc.) proved to be an essential tool to study genetic disorders and find their possible treatments.

These techniques have also been widely used in the process of prenatal and postnatal diagnosis by researchers. Moreover, doctors also suggest pregnant women for prenatal cytogenetics analysis and prenatal ploidy analysis to detect any genetic abnormality in the fetus (if they have any family history of genetic diseases)[1]. The analysis is generally done by using the karyotyping and FISH technique.

The advanced cytogenetic techniques such as microarray aid in the high-resolution analysis and have the ability to map the copy number changes in the genome. These techniques have also proved very helpful in the diagnosis, therapy, and monitoring of the effect of the treatment on any cancer patient. We’ll discuss clinical cytogenetics and its relevance in the study of chromosomal aberrations.

 

Clinical Cytogenetics

Few categories[1] of chromosomal aberrations:

  1. Autosomal aneuploidy
  2. Structural chromosome rearrangements
  3. Sex chromosomal disorders
  4. Infertility
  5. Prenatal cytogenetics
  6. Chromosome instability.
  7. Spontaneous Abortion

In Clinical Cytogenetics – Pt.1 we discussed points 1-3 above and learned various chromosomal aberrations that included deletions, inversions, substitution, translocation, and aneuploidy of the chromosomes. We also talked about the mechanisms that lead to these aberrations and the diseases they cause in human beings.

So, for this article, we will continue looking into these categories, particularly categories 4-7, as follows;

 

4. Infertility

Infertility is the inability to conceive a child by a couple, despite having unprotected intercourse for at least one year. Both male and female factors can be responsible for infertility. The causes of infertility in both the sexes include:[1]

 

S.No.Causes of female infertilityCauses of male infertility
1.Fallopian tube blockage or adhesionVaricocele (swelling of veins in the testes)
2.Cervical and uterine abnormalityHormone imbalances
3.Endometriosis (endometrial tissue grows outside of the uterus)Ejaculation issue ( premature ejaculation, genetic disease, blockage of testicles, etc)
4.Early menopauseOligospermia (low sperm count)
5.Ovulation disordersChromosome defects and Cancer

Sometimes, amenorrhea is the cause of female infertility. It is of two types: (a) primary amenorrhea, which is the absence of menstruation in females; and (b) secondary amenorrhea, the occurrence of discontinuous menstruation. Cytogenetics studies have found primary amenorrhea as the major cause of infertility in women.

Examples of chromosomal abnormalities causing infertility in females
  1. 45, X (Turner syndrome): It is estimated that 1 in 2500 girls is born with 45, X; and 2-3 % of women undergo normal puberty and menstruation, but there are chances to suffer from secondary amenorrhea. It has also been found that women with 45, X chromosomes are more likely to give birth to a child without abnormality (though with lesser chances) than the women having 45, X mosaic chromosomes.[1]
  2. X Chromosome Deletions: Deletions in the region of the X-chromosome lead to ovarian failure or amenorrheic conditions. Deletions at the p11 region lead to ovarian failure in some women and in other women, menstruation irregularities have been observed, although with rare chances of fertility even if menstruation occurs. If the deletion occurs in the long arm of the X-chromosome, between Xq13 and Xq26, it is definite to cause ovarian failure. The phenotype of women with Xq13 includes no breast development, ovarian failure, primary amenorrhea, and high levels of FSH and LH. Deletions in the distant region of the X-chromosome show milder effects. These women may have normal menarche but fertility is rare.[1]
  3. Endometriosis: In this condition, the endometrial tissue forms outside the uterus (in the ovary, pelvis, or elsewhere in the body). Endometriosis is the cause of infertility in 6-10% of women of their reproductive age. This condition arises due to the loss of chromosomal regions such as  7p, 1p, and 22q. It can be studied by using molecular cytogenetic techniques such as dual-color FISH and comparative genomic hybridization (CGH).[1]
Examples of chromosomal abnormalities causing infertility in males
  1. The SRY Gene and Genetic Sex: Sex determining region Y (SRY) is present on the short arm of chromosome Y. Its function involves the differentiation of precursor cells into Sertoli cells. These cells secrete the anti-mullerian hormone which functions to repress the development of female genitalia. Moreover, it is also involved in the production of testosterone from Leydig cells, which is involved in the formation of internal male genitalia. Any defect in this region of the chromosome may cause infertility in men due to the absence or poor development of male genitalia.[1]
  2. Oligospermia: This is the condition of having low spermatozoa count in an ejaculation. This can be due to chromosomal abnormality of 47, XXY; 47, XYY; or structural abnormality in the Y chromosome. Oligospermia can also occur due to structural rearrangements of the autosomal chromosome (discussed in Clinical cytogenetics-Pt.1).
  3. Sex Chromosome Abnormalities: The various types of non-mosaic chromosomal abnormalities are: 47, XXY, and 47, XYY. Some other mosaic abnormalities include 47, XXY/46, XY, and 47, XYY/46, XY. The researchers found that the frequency of aneuploidy of the sex chromosome ranges from 0.3 to 15 % which leads to infertility in men.[1] This condition can be studied by semen analysis by using the FISH technique.
  4. Autosomal Abnormalities: Robertsonian translocation, reciprocal translocation, and inversion are some common types of autosomal abnormalities that lead to infertility in men. It has been studied that the incidence of Robertsonian and reciprocal translocation is 0.7% and 0.5%.  These findings are studied by using Karyotyping and FISH techniques.[1]

 

5. Prenatal cytogenetics

The key event in the history of clinical cytogenetics was when a scientist, James, developed a technique to determine the sex of the fetus from the amniotic fluid by the Papanicolaou method using Giemsa stains. Now, three procedures are available to collect amniotic fluid for the analysis and these are; Amniocentesis, Chorionic Villus Sampling, and Percutaneous Umbilical Cord Sampling (PUBS).

The collection of amniotic fluid by transabdominal and transcervical puncturing of the uterus is called amniocentesis. This technique was developed and has been practiced since the 1930s[1] and today it’s a common tool to detect prenatal and postnatal cytogenetic and molecular abnormalities.

In the Chorionic villus sampling technique, developing placental cells are taken from pregnant women through the abdomen or cervix. And, PUBS (also called Cordocentesis) involves the collection of blood through the abdomen from a vein of the umbilical cord of pregnant women. These procedures are very helpful to detect any genetic abnormality in the fetus and to provide genetic counseling. Table [1] below will provide a brief of all three techniques.

TechniquesTime period recommended for testing (weeks)Diagnostic/cytogenetic analysis
Amniocentesis16-18

●      Karyotype

●      Biochemical studies

Chorionic Villus Sampling8-11

●      Karyotype

●      Cytogenetic analysis

●      Enzymatic studies

Percutaneous Umbilical Cord Sampling (PUBS)18-23

●      Karyotype and DNA analysis

●      Detection of disorders and infections.

 

(a) Amniocentesis   (b) Chorionic Villus Sampling   (c) Percutaneous Umbilical Cord Sampling (PUBS)

 

Figure: Image showing three procedures/techniques for prenatal cytogenetic analysis.

Source: https://www.pregnancyhealth.net/chorionic-villus-sampling-cvs-procedure-risks/ , http://pennmedicine.adam.com/content.aspx?productid=14&pid=14&gid=000229

 

 

What are the conditions in which Prenatal Cytogenetic analysis should be performed?

There are some situations in which significant risks of chromosomal abnormalities have been studied by researchers. Some indications of the risks[1] are discussed below:

  • Maternal Age: It has been found in various studies that women undergoing pregnancy at the age of 35 or more (or pregnant with twins at the age of 31 or more), are at higher risk of fetal chromosomal abnormality. For example, Down’s syndrome, Klinefelter syndrome, Turner syndrome, and structural rearrangements.
  • Nuchal folds and cystic hygromas: These are fluid-filled sac-like abnormal structures observed in the fetus during the second or third trimester. There are 22-70% chances of chromosomal abnormality in the fetus if any of these abnormal structures are observed in the ultrasound image.
  • Cardiac anomaly: It has been estimated that the frequency of chromosomal abnormality in a child with a cardiac anomaly is 5–10% (indicated by postnatal data). The prenatal data showed that there are 32–48% chances of chromosomal abnormality in the fetus with the cardiac anomaly.
  • Renal Pyelectasis: In this condition, the renal pelvis is mildly dilated. Researchers found the association between renal pyelectasis (observed in the ultrasound image of the fetus) with the trisomy of chromosomes and other mosaic conditions such as 47, XYY/46, and XY.

 

6. Chromosome instability

The instability of the chromosome is categorized at two levels: nucleotide level and chromosome level. The instability at the nucleotide level includes deletions or substitutions of a few nucleotides that cause frameshift mutations. However, instability at the chromosome level includes structural rearrangements of chromosomes (deletions, duplications, inversions, and translocation) which occurs due to breaks in the chromosomes. Another cause of chromosome instability is the numerical changes (aneuploidy and polyploidy) in the chromosome. This occurs due to the dysregulation of genes involved in cell division.

Mutations in cellular processes such as impairment of DNA replication, DNA recombination, and DNA repair lead to some other instabilities that are: the formation of fragile sites, sister chromatid exchange, and chromosome breakage.[1]

We are already familiar with the structural and numerical instability of the chromosomes. So, here we will discuss fragile sites and associated diseases.

 

What are fragile sites?

Fragile sites are considered as the points/regions on the chromosome which form gaps or breaks when the cell is exposed to stress conditions. These sites are rare but they are inherited as codominant traits. Some of the commonly known fragile sites are 3p14.2 (FRA3B), 6q26 (FRA6E), and Xp22 (FRAXB).[1]

It has been found that fragile sites are induced by chemicals such as aphidicolin, 5-azacytidine, and bromodeoxyuridine (BrdU).

 

Figure: The image shows fragile sites on human chromosomes (shown by arrow)-induced by folate.

Source: The Principles of Clinical Cytogenetics (2013).[1]

 

Examples of syndromes due to chromosome breakage
  1. Fanconi anemia (FA): This is a rare disease that occurs due to a homozygous mutation of the FANCD1 gene. The phenotype of this disease includes congenital anomaly, bone marrow failure, malformation of organs such as the heart and kidney, and overlapping of skeleton and limbs. The frequency of occurrence of this disease is 1-5/million. The types of chromosomal abnormalities observed in the cells of the patients are chromatid breaks, acentric and dicentric fragments, endoreduplication chromosomes, and telomere shortening.[1]
  2. Ataxia Telangiectasia: It is an autosomal recessive disorder. Breakage in the ATM gene is responsible for causing this disease. The phenotype includes cerebellar degeneration, immunodeficiency (diminished level of IgG2 and IgA), radiosensitivity, and cancer. The frequency of occurrence of this disease is 1 in 89,000.[1]
  3. Xeroderma Pigmentosum:  This disease is inherited in an autosomal recessive manner and is caused due to impairment in DNA repair ( defective excision of pyrimidine dimers) or replication of damaged DNA. The phenotype includes sensitive skin to sunlight (even can develop skin cancer), severe sunburn with blistering, persistent erythema, neurologic degeneration, and sensorineural deafness.[1]
  4. Robert Syndrome (RS): This is the rarest disease that occurs due to mutation in the ESCO2 gene, which causes loss of acetyltransferase activity. The phenotype includes growth retardation, abnormal limbs, craniofacial defects, hypertelorism, and cleft lip and palate. In some patients, heterochromatin repulsion and premature centromere separation are observed in chromosomes 1, 9, and 16. Prenatal diagnosis and cytogenetic analysis are done by ultrasound examination and by using the C-banding technique.[1]

 

7. Spontaneous Abortion

The frequency of occurrence of abortion is 15-20% in the early gestation period or second and third trimester of pregnancy. Multiple studies suggested that cytogenetic abnormalities are significant factors contributing to spontaneous abortion. It has been also found that the chances of abortion or chromosomal abnormality increase with the maternal age(at the age of 35 or more). Some of the errors[1] leading to abortion are given below:

  1. Trisomies: The most frequent type of trisomies that have been observed are trisomy 21, 22, and 16 chromosomes. The frequency of occurrence of trisomy 21 is 1 in 700 live births. The studies have found an association between maternal age and down syndrome. So, it is generally recommended for pregnant women of age 35 or above for prenatal diagnosis.
  2. Sex Chromosome Aneuploidy: It is the most common chromosomal abnormality responsible for abortion. In the majority of cases 45, X (Turner syndrome) arrangement is found to be responsible for miscarriages. The frequency of occurrence of turner syndrome is 1 in 1,000 female live births. In some cases, trisomy of sex chromosomes, such as 47, XXX, 47, XXY, and 47, XYY, found to be responsible for infertility.
  3. Structural rearrangements: The most common type of structural rearrangements leading to abortions are Robertsonian translocation, reciprocal translocation, and inversions. These rearrangements lead to the production of unbalanced chromosomes due to the abnormal segregation of chromosomes during meiosis. It has been estimated that the frequency of abortions due to structural rearrangements is 1-2%.
  4. Errors in fertilization: Due to errors in fertilization, pregnant women may have a triploid or abnormal diploid condition. Both mother (digyny) and father (diandry) can contribute to an extra set of chromosomes in the fetus. The 69, XYY arrangement indicated diandry; and 69, XXX or 69, XXY arrangement is demonstrated either by diandry or digyny. The frequency of occurrence of triploidy ranges from 1-3%.

 

Conclusion

As seen, clinical cytogenetics is the study of the relation of chromosomal aberrations with genetic diseases. The study provides a deep insight into structural and numerical chromosomal abnormalities leading to variant genetic disorders. The most important aspect of clinical cytogenetics is that it helps in the analysis of fetal chromosomes for any genetic disorders and genetic counseling of parents. So, clinical cytogenetics provides a whole picture of chromosomal disorders and their causes, which proves to be a useful tool for disease diagnosis and treatment.

 

References

Gersen Steven L. and Keagle Martha B. (2013). The Principles of Clinical Cytogenetics (3rd ed.), Springer, New York.

Learn More about our Services and how can we help you with your research!

Introduction

In behavioral neuroscience, the Open Field Test (OFT) remains one of the most widely used assays to evaluate rodent models of affect, cognition, and motivation. It provides a non-invasive framework for examining how animals respond to novelty, stress, and pharmacological or environmental manipulations. Among the test’s core metrics, the percentage of time spent in the center zone offers a uniquely normalized and sensitive measure of an animal’s emotional reactivity and willingness to engage with a potentially risky environment.

This metric is calculated as the proportion of time spent in the central area of the arena—typically the inner 25%—relative to the entire session duration. By normalizing this value, researchers gain a behaviorally informative variable that is resilient to fluctuations in session length or overall movement levels. This makes it especially valuable in comparative analyses, longitudinal monitoring, and cross-model validation.

Unlike raw center duration, which can be affected by trial design inconsistencies, the percentage-based measure enables clearer comparisons across animals, treatments, and conditions. It plays a key role in identifying trait anxiety, avoidance behavior, risk-taking tendencies, and environmental adaptation, making it indispensable in both basic and translational research contexts.

Whereas simple center duration provides absolute time, the percentage-based metric introduces greater interpretability and reproducibility, especially when comparing different animal models, treatment conditions, or experimental setups. It is particularly effective for quantifying avoidance behaviors, risk assessment strategies, and trait anxiety profiles in both acute and longitudinal designs.

What Does Percentage of Time in the Centre Measure?

This metric reflects the relative amount of time an animal chooses to spend in the open, exposed portion of the arena—typically defined as the inner 25% of a square or circular enclosure. Because rodents innately prefer the periphery (thigmotaxis), time in the center is inversely associated with anxiety-like behavior. As such, this percentage is considered a sensitive, normalized index of:

  • Exploratory drive vs. risk aversion: High center time reflects an animal’s willingness to engage with uncertain or exposed environments, often indicative of lower anxiety and a stronger intrinsic drive to explore. These animals are more likely to exhibit flexible, information-gathering behaviors. On the other hand, animals that spend little time in the center display a strong bias toward the safety of the perimeter, indicative of a defensive behavioral state or trait-level risk aversion. This dichotomy helps distinguish adaptive exploration from fear-driven avoidance.

  • Emotional reactivity: Fluctuations in center time percentage serve as a sensitive behavioral proxy for changes in emotional state. In stress-prone or trauma-exposed animals, decreased center engagement may reflect hypervigilance or fear generalization, while a sudden increase might indicate emotional blunting or impaired threat appraisal. The metric is also responsive to acute stressors, environmental perturbations, or pharmacological interventions that impact affective regulation.

  • Behavioral confidence and adaptation: Repeated exposure to the same environment typically leads to reduced novelty-induced anxiety and increased behavioral flexibility. A rising trend in center time percentage across trials suggests successful habituation, reduced threat perception, and greater confidence in navigating open spaces. Conversely, a stable or declining trend may indicate behavioral rigidity or chronic stress effects.

  • Pharmacological or genetic modulation: The percentage of time in the center is widely used to evaluate the effects of pharmacological treatments and genetic modifications that influence anxiety-related circuits. Anxiolytic agents—including benzodiazepines, SSRIs, and cannabinoid agonists—reliably increase center occupancy, providing a robust behavioral endpoint in preclinical drug trials. Similarly, genetic models targeting serotonin receptors, GABAergic tone, or HPA axis function often show distinct patterns of center preference, offering translational insights into psychiatric vulnerability and resilience.

Critically, because this metric is normalized by session duration, it accommodates variability in activity levels or testing conditions. This makes it especially suitable for comparing across individuals, treatment groups, or timepoints in longitudinal studies.

A high percentage of center time indicates reduced anxiety, increased novelty-seeking, or pharmacological modulation (e.g., anxiolysis). Conversely, a low percentage suggests emotional inhibition, behavioral avoidance, or contextual hypervigilance. reduced anxiety, increased novelty-seeking, or pharmacological modulation (e.g., anxiolysis). Conversely, a low percentage suggests emotional inhibition, behavioral avoidance, or contextual hypervigilance.

Behavioral Significance and Neuroscientific Context

1. Emotional State and Trait Anxiety

The percentage of center time is one of the most direct, unconditioned readouts of anxiety-like behavior in rodents. It is frequently reduced in models of PTSD, chronic stress, or early-life adversity, where animals exhibit persistent avoidance of the center due to heightened emotional reactivity. This metric can also distinguish between acute anxiety responses and enduring trait anxiety, especially in longitudinal or developmental studies. Its normalized nature makes it ideal for comparing across cohorts with variable locomotor profiles, helping researchers detect true affective changes rather than activity-based confounds.

2. Exploration Strategies and Cognitive Engagement

Rodents that spend more time in the center zone typically exhibit broader and more flexible exploration strategies. This behavior reflects not only reduced anxiety but also cognitive engagement and environmental curiosity. High center percentage is associated with robust spatial learning, attentional scanning, and memory encoding functions, supported by coordinated activation in the prefrontal cortex, hippocampus, and basal forebrain. In contrast, reduced center engagement may signal spatial rigidity, attentional narrowing, or cognitive withdrawal, particularly in models of neurodegeneration or aging.

3. Pharmacological Responsiveness

The open field test remains one of the most widely accepted platforms for testing anxiolytic and psychotropic drugs. The percentage of center time reliably increases following administration of anxiolytic agents such as benzodiazepines, SSRIs, and GABA-A receptor agonists. This metric serves as a sensitive and reproducible endpoint in preclinical dose-finding studies, mechanistic pharmacology, and compound screening pipelines. It also aids in differentiating true anxiolytic effects from sedation or motor suppression by integrating with other behavioral parameters like distance traveled and entry count (Prut & Belzung, 2003).

4. Sex Differences and Hormonal Modulation

Sex-based differences in emotional regulation often manifest in open field behavior, with female rodents generally exhibiting higher variability in center zone metrics due to hormonal cycling. For example, estrogen has been shown to facilitate exploratory behavior and increase center occupancy, while progesterone and stress-induced corticosterone often reduce it. Studies involving gonadectomy, hormone replacement, or sex-specific genetic knockouts use this metric to quantify the impact of endocrine factors on anxiety and exploratory behavior. As such, it remains a vital tool for dissecting sex-dependent neurobehavioral dynamics.
The percentage of center time is one of the most direct, unconditioned readouts of anxiety-like behavior in rodents. It is frequently reduced in models of PTSD, chronic stress, or early-life adversity. Because it is normalized, this metric is especially helpful for distinguishing between genuine avoidance and low general activity.

Methodological Considerations

  • Zone Definition: Accurately defining the center zone is critical for reliable and reproducible data. In most open field arenas, the center zone constitutes approximately 25% of the total area, centrally located and evenly distanced from the walls. Software-based segmentation tools enhance precision and ensure consistency across trials and experiments. Deviations in zone parameters—whether due to arena geometry or tracking inconsistencies—can result in skewed data, especially when calculating percentages.

     

  • Trial Duration: Trials typically last between 5 to 10 minutes. The percentage of time in the center must be normalized to total trial duration to maintain comparability across animals and experimental groups. Longer trials may lead to fatigue, boredom, or habituation effects that artificially reduce exploratory behavior, while overly short trials may not capture full behavioral repertoires or response to novel stimuli.

     

  • Handling and Habituation: Variability in pre-test handling can introduce confounds, particularly through stress-induced hypoactivity or hyperactivity. Standardized handling routines—including gentle, consistent human interaction in the days leading up to testing—reduce variability. Habituation to the testing room and apparatus prior to data collection helps animals engage in more representative exploratory behavior, minimizing novelty-induced freezing or erratic movement.

     

  • Tracking Accuracy: High-resolution tracking systems should be validated for accurate, real-time detection of full-body center entries and sustained occupancy. The system should distinguish between full zone occupancy and transient overlaps or partial body entries that do not reflect true exploratory behavior. Poor tracking fidelity or lag can produce significant measurement error in percentage calculations.

     

  • Environmental Control: Uniformity in environmental conditions is essential. Lighting should be evenly diffused to avoid shadow bias, and noise should be minimized to prevent stress-induced variability. The arena must be cleaned between trials using odor-neutral solutions to eliminate scent trails or pheromone cues that may affect zone preference. Any variation in these conditions can introduce systematic bias in center zone behavior. Use consistent definitions of the center zone (commonly 25% of total area) to allow valid comparisons. Software-based segmentation enhances spatial precision.

Interpretation with Complementary Metrics

Temporal Dynamics of Center Occupancy

Evaluating how center time evolves across the duration of a session—divided into early, middle, and late thirds—provides insight into behavioral transitions and adaptive responses. Animals may begin by avoiding the center, only to gradually increase center time as they habituate to the environment. Conversely, persistently low center time across the session can signal prolonged anxiety, fear generalization, or a trait-like avoidance phenotype.

Cross-Paradigm Correlation

To validate the significance of center time percentage, it should be examined alongside results from other anxiety-related tests such as the Elevated Plus Maze, Light-Dark Box, or Novelty Suppressed Feeding. Concordance across paradigms supports the reliability of center time as a trait marker, while discordance may indicate task-specific reactivity or behavioral dissociation.

Behavioral Microstructure Analysis

When paired with high-resolution scoring of behavioral events such as rearing, grooming, defecation, or immobility, center time offers a richer view of the animal’s internal state. For example, an animal that spends substantial time in the center while grooming may be coping with mild stress, while another that remains immobile in the periphery may be experiencing more severe anxiety. Microstructure analysis aids in decoding the complexity behind spatial behavior.

Inter-individual Variability and Subgroup Classification

Animals naturally vary in their exploratory style. By analyzing percentage of center time across subjects, researchers can identify behavioral subgroups—such as consistently bold individuals who frequently explore the center versus cautious animals that remain along the periphery. These classifications can be used to examine predictors of drug response, resilience to stress, or vulnerability to neuropsychiatric disorders.

Machine Learning-Based Behavioral Clustering

In studies with large cohorts or multiple behavioral variables, machine learning techniques such as hierarchical clustering or principal component analysis can incorporate center time percentage to discover novel phenotypic groupings. These data-driven approaches help uncover latent dimensions of behavior that may not be visible through univariate analyses alone.

Total Distance Traveled

Total locomotion helps contextualize center time. Low percentage values in animals with minimal movement may reflect sedation or fatigue, while similar values in high-mobility subjects suggest deliberate avoidance. This metric helps distinguish emotional versus motor causes of low center engagement.

Number of Center Entries

This measure indicates how often the animal initiates exploration of the center zone. When combined with percentage of time, it differentiates between frequent but brief visits (indicative of anxiety or impulsivity) versus fewer but sustained center engagements (suggesting comfort and behavioral confidence).

Latency to First Center Entry

The delay before the first center entry reflects initial threat appraisal. Longer latencies may be associated with heightened fear or low motivation, while shorter latencies are typically linked to exploratory drive or low anxiety.

Thigmotaxis Time

Time spent hugging the walls offers a spatial counterbalance to center metrics. High thigmotaxis and low center time jointly support an interpretation of strong avoidance behavior. This inverse relationship helps triangulate affective and motivational states.

Applications in Translational Research

  • Drug Discovery: The percentage of center time is a key behavioral endpoint in the development and screening of anxiolytic, antidepressant, and antipsychotic medications. Its sensitivity to pharmacological modulation makes it particularly valuable in dose-response assessments and in distinguishing therapeutic effects from sedative or locomotor confounds. Repeated trials can also help assess drug tolerance and chronic efficacy over time.
  • Genetic and Neurodevelopmental Modeling: In transgenic and knockout models, altered center percentage provides a behavioral signature of neurodevelopmental abnormalities. This is particularly relevant in the study of autism spectrum disorders, ADHD, fragile X syndrome, and schizophrenia, where subjects often exhibit heightened anxiety, reduced flexibility, or altered environmental engagement.
  • Hormonal and Sex-Based Research: The metric is highly responsive to hormonal fluctuations, including estrous cycle phases, gonadectomy, and hormone replacement therapies. It supports investigations into sex differences in stress reactivity and the behavioral consequences of endocrine disorders or interventions.
  • Environmental Enrichment and Deprivation: Housing conditions significantly influence anxiety-like behavior and exploratory motivation. Animals raised in enriched environments typically show increased center time, indicative of reduced stress and greater behavioral plasticity. Conversely, socially isolated or stimulus-deprived animals often show strong center avoidance.
  • Behavioral Biomarker Development: As a robust and reproducible readout, center time percentage can serve as a behavioral biomarker in longitudinal and interventional studies. It is increasingly used to identify early signs of affective dysregulation or to track the efficacy of neuromodulatory treatments such as optogenetics, chemogenetics, or deep brain stimulation.
  • Personalized Preclinical Models: This measure supports behavioral stratification, allowing researchers to identify high-anxiety or low-anxiety phenotypes before treatment. This enables within-group comparisons and enhances statistical power by accounting for pre-existing behavioral variation. Used to screen anxiolytic agents and distinguish between compounds with sedative vs. anxiolytic profiles.

Enhancing Research Outcomes with Percentage-Based Analysis

By expressing center zone activity as a proportion of total trial time, researchers gain a metric that is resistant to session variability and more readily comparable across time, treatment, and model conditions. This normalized measure enhances reproducibility and statistical power, particularly in multi-cohort or cross-laboratory designs.

For experimental designs aimed at assessing anxiety, exploratory strategy, or affective state, the percentage of time spent in the center offers one of the most robust and interpretable measures available in the Open Field Test.

Explore high-resolution tracking solutions and open field platforms at

References

  • Prut, L., & Belzung, C. (2003). The open field as a paradigm to measure the effects of drugs on anxiety-like behaviors: a review. European Journal of Pharmacology, 463(1–3), 3–33.
  • Seibenhener, M. L., & Wooten, M. C. (2015). Use of the open field maze to measure locomotor and anxiety-like behavior in mice. Journal of Visualized Experiments, (96), e52434.
  • Crawley, J. N. (2007). What’s Wrong With My Mouse? Behavioral Phenotyping of Transgenic and Knockout Mice. Wiley-Liss.
  • Carola, V., D’Olimpio, F., Brunamonti, E., Mangia, F., & Renzi, P. (2002). Evaluation of the elevated plus-maze and open-field tests for the assessment of anxiety-related behavior in inbred mice. Behavioral Brain Research, 134(1–2), 49–57.

Written by researchers, for researchers — powered by Conduct Science.