Jacqueline N. Crawley

Atm-Deficient Mice: A Paradigm of Ataxia Telangiectasia

A murine model of ataxia telangiectasia was created by disrupting the Atm locus via gene targeting. Mice homozygous for the disrupted Atm allele displayed growth retardation, neurologic dysfunction, male and female infertility secondary to the absence of mature gametes, defects in T lymphocyte maturation, and extreme sensitivity to gamma-irradiation. The majority of animals developed malignant thymic lymphomas between 2 and 4 months of age. Several chromosomal anomalies were detected in one of these tumors. Fibroblasts from these mice grew slowly and exhibited abnormal radiation-induced G1 checkpoint function. Atm-disrupted mice recapitulate the ataxia telangiectasia phenotype in humans, providing a mammalian model in which to study the pathophysiology of this pleiotropic disorder.

Behavioral phenotypes of inbred mouse strains: implications and recommendations for molecular studies

Abstract Choosing the best genetic strains of mice for developing a new knockout or transgenic mouse requires extensive knowledge of the endogenous traits of inbred strains. Background genes from the parental strains may interact with the mutated gene, in a manner which could severely compromise the interpretation of the mutant phenotype. The present overview summarizes the literature on a wide variety of behavioral traits for the 129, C57BL/6, DBA/2, and many other inbred strains of mice. Strain distributions are described for open field activity, learning and memory tasks, aggression, sexual and parental behaviors, acoustic startle and prepulse inhibition, and the behavioral actions of ethanol, nicotine, cocaine, opiates, antipsychotics, and anxiolytics. Using the referenced information, molecular geneticists can choose optimal parental strains of mice, and perhaps develop new embryonic stem cell progenitors, for new knockouts and transgenics to investigate gene function, and to serve as animal models in the development of novel therapeutics for human genetic diseases.

Preliminary report of a simple animal behavior model for the anxiolytic effects of benzodiazepines

A simple system is described to analyze the possibility that increased exploratory behavior is an index for the anxiolytic effects of benzodiazepines in laboratory rodents. Mice were allowed free run in a two-chambered arena, where two-thirds of the area was illuminated and one-third was darkened. The two chambers were separated by a black partition equipped with photocells across the opening, and the entire cage rested on an Animex activity monitor. Transitions across the partition between the light and dark chambers, and total Animex locomotor activity, were increased by clonazepam and chlordiazepoxide, in dose-dependent ranges consistent with previously reported behavior models. The increased exploratory activity with benzodiazepines does not appear to be a non-specific increase in general motor activity, as locomotion in clonazepam and chlordiazepoxide treated mice placed in a bare, undifferentiated cage was not significantly different from vehicle treated mice.

Behavioural phenotyping assays for mouse models of autism

Autism is a heterogeneous neurodevelopmental disorder of unknown aetiology that affects 1 in 100–150 individuals. Diagnosis is based on three categories of behavioural criteria: abnormal social interactions, communication deficits and repetitive behaviours. Strong evidence for a genetic basis has prompted the development of mouse models with targeted mutations in candidate genes for autism. As the diagnostic criteria for autism are behavioural, phenotyping these mouse models requires behavioural assays with high relevance to each category of the diagnostic symptoms. Behavioural neuroscientists are generating a comprehensive set of assays for social interaction, communication and repetitive behaviours to test hypotheses about the causes of austism. Robust phenotypes in mouse models hold great promise as translational tools for discovering effective treatments for components of autism spectrum disorders.

Sociability and preference for social novelty in five inbred strains: An approach to assess autistic-like behavior in mice

Deficits in social interaction are important early markers for autism and related neurodevelopmental disorders with strong genetic components. Standardized behavioral assays that measure the preference of mice for initiating social interactions with novel conspecifics would be of great value for mutant mouse models of autism. We developed a new procedure to assess sociability and the preference for social novelty in mice. To quantitate sociability, each mouse was scored on measures of exploration in a central habituated area, a side chamber containing an unfamiliar conspecific (stranger 1) in a wire cage, or an empty side chamber. In a secondary test, preference for social novelty was quantitated by presenting the test mouse with a choice between the first, now‐familiar, conspecific (stranger 1) in one side chamber, and a second unfamiliar mouse (stranger 2) in the other side chamber. Parameters scored included time spent in each chamber and number of entries into the chambers. Five inbred strains of mice were tested, C57BL/6J, DBA/2J, FVB/NJ, A/J and B6129PF2/J hybrids. Four strains showed significant levels of sociability (spend‐ ing more time in the chamber containing stranger 1 than in the empty chamber) and a preference for social novelty (spending more time in the chamber containing stranger 2 than in the chamber containing the now‐familiar stranger 1). These social preferences were observed in both male and female mice, and in juveniles and adults. The exception was A/J, a strain that demonstrated a preference for the central chamber. Results are discussed in terms of potential applications of the new methods, and the proper controls for the interpretation of social behavior data, including assays for health, relevant sensory abilities and motor functions. This new standardized procedure to quantitate sociability and preference for social novelty in mice provides a method to assess tendencies for social avoidance in mouse models of autism.

Behavioral phenotyping of transgenic and knockout mice: experimental design and evaluation of general health, sensory functions, motor abilities, and specific behavioral tests

Rigorous experimental design can minimize the high risk of false positives and false negatives in the behavioral phenotyping of a new transgenic or knockout mouse. Use of well established, quantitative, reproducible behavioral tasks, appropriate Ns, correct statistical methods, consideration of background genes contributed by the parental strains, and attention to litter and gender issues, will maximize meaningful comparisons of -/-, +/-, and +/+ genotypes. Strategies developed and used by our laboratory are described in this review. Preliminary observations evaluate general health and neurological reflexes. Sensory abilities and motor functions are extensively quantitated. Specific tests include observations of home cage behaviors, body weight, body temperature, appearance of the fur and whiskers, righting reflex, acoustic startle, eye blink, pupil constriction, vibrissae reflex, pinna reflex, Digiscan open field locomotion, rotarod motor coordination, hanging wire, footprint pathway, visual cliff, auditory threshold, pain threshold, and olfactory acuity. Hypothesis testing then focuses on at least three well-validated tasks within each relevant behavioral domain. Specific tests for mice are described herein for the domains of learning and memory, feeding, nociception, and behaviors relevant to discrete symptoms of human anxiety, depression, schizophrenia, and drug addiction. An example of our approach is illustrated in the behavioral phenotyping of C/EBPdelta knockout mice, which appear to be normal on general health, neurological reflexes, sensory and motor tasks, and the Morris water task, but show remarkably enhanced performance on contextual fear conditioning.

Biological actions of cholecystokinin

Cholecystokinin (CCK) has emerged as an important mammalian neuropeptide, localized in peripheral organs and in the central nervous system. This review presents an overview of the molecular aspects of CCK peptides and CCK receptors, the anatomical distribution of CCK, the neurophysiological actions of CCK, release of CCK and effects of CCK on release of other neurotransmitters, and the actions of CCK on digestion, feeding, cardiovascular function, respiratory function, neurotoxicity and seizures, cancer cell proliferation, analgesia, sleep, sexual and reproductive behaviors, memory, anxiety, and dopamine-mediated exploratory and rewarded behaviors. Human clinical studies of CCK in feeding disorders and panic disorders are described. New findings are presented on potent, nonpeptide CCK antagonists, selective for the two CCK receptor subtypes, which demonstrate that endogenous CCK has biologically important effects on physiology and behavior.

Exploratory behavior models of anxiety in mice

Parameters of exploratory behaviors responsive to anti-anxiety drugs are reviewed with respect to their sensitivity and specificity for anxiolytics in mice. Mouse models appear to rest on a disinhibition of natural exploratory tendencies by anxiolytic treatments. Analysis of agonists of the brain benzodiazepine binding site, such as chlordiazepoxide and diazepam, significantly increase exploration of a hole-board, of a two-chambered light in equilibrium dark apparatus, increase social interaction under high levels of illumination, increase consumption of a novel food in an unfamiliar environment, and increase punished crossings in a footshock conflict paradigm. These tests detect anxiolytic responses at doses of benzodiazepines well within the clinically effective range. Pharmacological specificity was established for the hole-board and light in equilibrium transition tests, showing that non-anxiolytic categories of psychoactive drugs did not produce false positives. Open field behaviors and isolation-induced aggression were reduced by anxiolytics, at doses which may be within the sedative-hypnotic range. Analysis of antagonists of the brain benzodiazepine binding site did not show active antagonist properties in the light in equilibrium transitions model, although the antagonist Ro-15-1788 appeared to have partial agonist properties in the open field test, suggesting that rat models may be more sensitive to anxiogenic compounds than are mouse models. The wide separation between anxiolytic and sedative doses in mouse models recommend these exploration paradigms as good predictive screens for the testing of novel anxiolytic compounds.

Mouse behavioral tasks relevant to autism: Phenotypes of 10 inbred strains

Three defining clinical symptoms of autism are aberrant reciprocal social interactions, deficits in social communication, and repetitive behaviors, including motor stereotypies and insistence on sameness. We developed a set of behavioral tasks designed to model components of these core symptoms in mice. Male mice from 10 inbred strains were characterized in assays for sociability, preference for social novelty, and reversal of the spatial location of the reinforcer in T-maze and Morris water maze tasks. Six strains, C57BL/6J, C57L/J, DBA/2J, FVB/NJ, C3H/HeJ, and AKR/J, showed significant levels of sociability, while A/J, BALB/cByJ, BTBR T(+)tf/J, and 129S1/SvImJ mice did not. C57BL/6J, C57L/J, DBA/2J, FVB/NJ, BALB/cByJ, and BTBR T(+)tf/J showed significant preference for social novelty, while C3H/HeJ, AKR/J, A/J, and 129S1/SvImJ did not. Normal scores on relevant control measures confirmed general health and physical abilities in all strains, ruling out artifactual explanations for social deficits. Elevated plus maze scores confirmed high anxiety-like behaviors in A/J, BALB/cByJ, and 129S1/SvImJ, which could underlie components of their low social approach. Strains that showed high levels of performance on acquisition of a T-maze task were also able to reach criterion for reversal learning. On the Morris water maze task, DBA/2J, AKR/J, BTBR T(+)tf/J, and 129S1/SvImJ failed to show significant quadrant preference during the reversal probe trial. These results highlight a dissociation between social task performance and reversal learning. BTBR T(+)tf/J is a particularly interesting strain, displaying both low social approach and resistance to change in routine on the water maze, consistent with an autism-like phenotype. Our multitask strategy for modeling symptoms of autism will be useful for investigating targeted and random gene mutations, QTLs, and microarray analyses.

A Proposed Test Battery and Constellations of Specific Behavioral Paradigms to Investigate the Behavioral Phenotypes of Transgenic and Knockout Mice

Behavioral phenotyping of transgenic and knockout mice requires rigorous, formal analyses. Well-characterized paradigms can be chosen from the established behavioral neuroscience literature. This review describes (1) a series of neurological and neuropsychological tests which are effectively used as a first screen for behavioral abnormalities in mutant mice, and (2) a series of specific behavioral paradigms, clustered by category. Included are multiple paradigms for each category, including learning and memory, feeding, analgesia, aggression, anxiety, depression, schizophrenia, and drug abuse models. Examples are given from the experiences of the authors, in applying these experimental designs to transgenic and knockout mice. Extensive references for each behavioral paradigm are provided, to allow new investigators to access the relevant literature on behavioral methodology.