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Gender dimorphism in aspartame-induced impairment of spatial cognition and insulin sensitivity.
Previous studies have linked aspartame consumption to impaired retention of learned behavior in rodents. Prenatal exposure to aspartame has also been shown to impair odor-associative learning in guinea pigs; and recently, aspartame-fed hyperlipidemic zebrafish exhibited weight gain, hyperglycemia and acute swimming defects. We therefore investigated the effects of chronic lifetime exposure to aspartame, commencing in utero, on changes in blood glucose parameters, spatial learning and memory in C57BL/6J mice. Morris Water Maze (MWM) testing was used to assess learning and memory, and a random-fed insulin tolerance test was performed to assess glucose homeostasis. Pearson correlation analysis was used to investigate the associations between body characteristics and MWM performance outcome variables. At 17 weeks of age, male aspartame-fed mice exhibited weight gain, elevated fasting glucose levels and decreased insulin sensitivity compared to controls (P<0.05). Females were less affected, but had significantly raised fasting glucose levels. During spatial learning trials in the MWM (acquisition training), the escape latencies of male aspartame-fed mice were consistently higher than controls, indicative of learning impairment. Thigmotactic behavior and time spent floating directionless was increased in aspartame mice, who also spent less time searching in the target quadrant of the maze (P<0.05). Spatial learning of female aspartame-fed mice was not significantly different from controls. Reference memory during a probe test was affected in both genders, with the aspartame-fed mice spending significantly less time searching for the former location of the platform. Interestingly, the extent of visceral fat deposition correlated positively with non-spatial search strategies such as floating and thigmotaxis, and negatively with time spent in the target quadrant and swimming across the location of the escape platform. These data suggest that lifetime exposure to aspartame, commencing in utero, may affect spatial cognition and glucose homeostasis in C57BL/6J mice, particularly in males.
Authors: Cheryl S. Rosenfeld, Sherry A. Ferguson.
Published: 02-26-2014
Spatial learning and memory of laboratory rodents is often assessed via navigational ability in mazes, most popular of which are the water and dry-land (Barnes) mazes. Improved performance over sessions or trials is thought to reflect learning and memory of the escape cage/platform location. Considered less stressful than water mazes, the Barnes maze is a relatively simple design of a circular platform top with several holes equally spaced around the perimeter edge. All but one of the holes are false-bottomed or blind-ending, while one leads to an escape cage. Mildly aversive stimuli (e.g. bright overhead lights) provide motivation to locate the escape cage. Latency to locate the escape cage can be measured during the session; however, additional endpoints typically require video recording. From those video recordings, use of automated tracking software can generate a variety of endpoints that are similar to those produced in water mazes (e.g. distance traveled, velocity/speed, time spent in the correct quadrant, time spent moving/resting, and confirmation of latency). Type of search strategy (i.e. random, serial, or direct) can be categorized as well. Barnes maze construction and testing methodologies can differ for small rodents, such as mice, and large rodents, such as rats. For example, while extra-maze cues are effective for rats, smaller wild rodents may require intra-maze cues with a visual barrier around the maze. Appropriate stimuli must be identified which motivate the rodent to locate the escape cage. Both Barnes and water mazes can be time consuming as 4-7 test trials are typically required to detect improved learning and memory performance (e.g. shorter latencies or path lengths to locate the escape platform or cage) and/or differences between experimental groups. Even so, the Barnes maze is a widely employed behavioral assessment measuring spatial navigational abilities and their potential disruption by genetic, neurobehavioral manipulations, or drug/ toxicant exposure.
24 Related JoVE Articles!
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Use of an Eight-arm Radial Water Maze to Assess Working and Reference Memory Following Neonatal Brain Injury
Authors: Stephanie C. Penley, Cynthia M. Gaudet, Steven W. Threlkeld.
Institutions: Rhode Island College, Rhode Island College.
Working and reference memory are commonly assessed using the land based radial arm maze. However, this paradigm requires pretraining, food deprivation, and may introduce scent cue confounds. The eight-arm radial water maze is designed to evaluate reference and working memory performance simultaneously by requiring subjects to use extra-maze cues to locate escape platforms and remedies the limitations observed in land based radial arm maze designs. Specifically, subjects are required to avoid the arms previously used for escape during each testing day (working memory) as well as avoid the fixed arms, which never contain escape platforms (reference memory). Re-entries into arms that have already been used for escape during a testing session (and thus the escape platform has been removed) and re-entries into reference memory arms are indicative of working memory deficits. Alternatively, first entries into reference memory arms are indicative of reference memory deficits. We used this maze to compare performance of rats with neonatal brain injury and sham controls following induction of hypoxia-ischemia and show significant deficits in both working and reference memory after eleven days of testing. This protocol could be easily modified to examine many other models of learning impairment.
Behavior, Issue 82, working memory, reference memory, hypoxia-ischemia, radial arm maze, water maze
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Elevated Plus Maze for Mice
Authors: Munekazu Komada, Keizo Takao, Tsuyoshi Miyakawa.
Institutions: Graduate School of Medicine, Kyoto University, Fujita Health University.
Although the mouse genome is now completely sequenced, the functions of most of the genes expressed in the brain are not known. The influence of a given gene on a specific behavior can be determined by behavioral analysis of mutant mice. If a target gene is expressed in the brain, behavioral phenotype of the mutant mice could elucidate the genetic mechanism of normal behaviors. The elevated plus maze test is one of the most widely used tests for measuring anxiety-like behavior. The test is based on the natural aversion of mice for open and elevated areas, as well as on their natural spontaneous exploratory behavior in novel environments. The apparatus consists of open arms and closed arms, crossed in the middle perpendicularly to each other, and a center area. Mice are given access to all of the arms and are allowed to move freely between them. The number of entries into the open arms and the time spent in the open arms are used as indices of open space-induced anxiety in mice. Unfortunately, the procedural differences that exist between laboratories make it difficult to duplicate and compare results among laboratories. Here, we present a detailed movie demonstrating our protocol for the elevated plus maze test. In our laboratory, we have assessed more than 90 strains of mutant mice using the protocol shown in the movie. These data will be disclosed as a part of a public database that we are now constructing. Visualization of the protocol will promote better understanding of the details of the entire experimental procedure, allowing for standardization of the protocols used in different laboratories and comparisons of the behavioral phenotypes of various strains of mutant mice assessed using this test.
Neuroscience, Issue 22, Knockout mice, genetically engineered mice, behavioral test, phenotyping
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Development of a Virtual Reality Assessment of Everyday Living Skills
Authors: Stacy A. Ruse, Vicki G. Davis, Alexandra S. Atkins, K. Ranga R. Krishnan, Kolleen H. Fox, Philip D. Harvey, Richard S.E. Keefe.
Institutions: NeuroCog Trials, Inc., Duke-NUS Graduate Medical Center, Duke University Medical Center, Fox Evaluation and Consulting, PLLC, University of Miami Miller School of Medicine.
Cognitive impairments affect the majority of patients with schizophrenia and these impairments predict poor long term psychosocial outcomes.  Treatment studies aimed at cognitive impairment in patients with schizophrenia not only require demonstration of improvements on cognitive tests, but also evidence that any cognitive changes lead to clinically meaningful improvements.  Measures of “functional capacity” index the extent to which individuals have the potential to perform skills required for real world functioning.  Current data do not support the recommendation of any single instrument for measurement of functional capacity.  The Virtual Reality Functional Capacity Assessment Tool (VRFCAT) is a novel, interactive gaming based measure of functional capacity that uses a realistic simulated environment to recreate routine activities of daily living. Studies are currently underway to evaluate and establish the VRFCAT’s sensitivity, reliability, validity, and practicality. This new measure of functional capacity is practical, relevant, easy to use, and has several features that improve validity and sensitivity of measurement of function in clinical trials of patients with CNS disorders.
Behavior, Issue 86, Virtual Reality, Cognitive Assessment, Functional Capacity, Computer Based Assessment, Schizophrenia, Neuropsychology, Aging, Dementia
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Perceptual and Category Processing of the Uncanny Valley Hypothesis' Dimension of Human Likeness: Some Methodological Issues
Authors: Marcus Cheetham, Lutz Jancke.
Institutions: University of Zurich.
Mori's Uncanny Valley Hypothesis1,2 proposes that the perception of humanlike characters such as robots and, by extension, avatars (computer-generated characters) can evoke negative or positive affect (valence) depending on the object's degree of visual and behavioral realism along a dimension of human likeness (DHL) (Figure 1). But studies of affective valence of subjective responses to variously realistic non-human characters have produced inconsistent findings 3, 4, 5, 6. One of a number of reasons for this is that human likeness is not perceived as the hypothesis assumes. While the DHL can be defined following Mori's description as a smooth linear change in the degree of physical humanlike similarity, subjective perception of objects along the DHL can be understood in terms of the psychological effects of categorical perception (CP) 7. Further behavioral and neuroimaging investigations of category processing and CP along the DHL and of the potential influence of the dimension's underlying category structure on affective experience are needed. This protocol therefore focuses on the DHL and allows examination of CP. Based on the protocol presented in the video as an example, issues surrounding the methodology in the protocol and the use in "uncanny" research of stimuli drawn from morph continua to represent the DHL are discussed in the article that accompanies the video. The use of neuroimaging and morph stimuli to represent the DHL in order to disentangle brain regions neurally responsive to physical human-like similarity from those responsive to category change and category processing is briefly illustrated.
Behavior, Issue 76, Neuroscience, Neurobiology, Molecular Biology, Psychology, Neuropsychology, uncanny valley, functional magnetic resonance imaging, fMRI, categorical perception, virtual reality, avatar, human likeness, Mori, uncanny valley hypothesis, perception, magnetic resonance imaging, MRI, imaging, clinical techniques
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The Multiple Sclerosis Performance Test (MSPT): An iPad-Based Disability Assessment Tool
Authors: Richard A. Rudick, Deborah Miller, Francois Bethoux, Stephen M. Rao, Jar-Chi Lee, Darlene Stough, Christine Reece, David Schindler, Bernadett Mamone, Jay Alberts.
Institutions: Cleveland Clinic Foundation, Cleveland Clinic Foundation, Cleveland Clinic Foundation, Cleveland Clinic Foundation.
Precise measurement of neurological and neuropsychological impairment and disability in multiple sclerosis is challenging. We report a new test, the Multiple Sclerosis Performance Test (MSPT), which represents a new approach to quantifying MS related disability. The MSPT takes advantage of advances in computer technology, information technology, biomechanics, and clinical measurement science. The resulting MSPT represents a computer-based platform for precise, valid measurement of MS severity. Based on, but extending the Multiple Sclerosis Functional Composite (MSFC), the MSPT provides precise, quantitative data on walking speed, balance, manual dexterity, visual function, and cognitive processing speed. The MSPT was tested by 51 MS patients and 49 healthy controls (HC). MSPT scores were highly reproducible, correlated strongly with technician-administered test scores, discriminated MS from HC and severe from mild MS, and correlated with patient reported outcomes. Measures of reliability, sensitivity, and clinical meaning for MSPT scores were favorable compared with technician-based testing. The MSPT is a potentially transformative approach for collecting MS disability outcome data for patient care and research. Because the testing is computer-based, test performance can be analyzed in traditional or novel ways and data can be directly entered into research or clinical databases. The MSPT could be widely disseminated to clinicians in practice settings who are not connected to clinical trial performance sites or who are practicing in rural settings, drastically improving access to clinical trials for clinicians and patients. The MSPT could be adapted to out of clinic settings, like the patient’s home, thereby providing more meaningful real world data. The MSPT represents a new paradigm for neuroperformance testing. This method could have the same transformative effect on clinical care and research in MS as standardized computer-adapted testing has had in the education field, with clear potential to accelerate progress in clinical care and research.
Medicine, Issue 88, Multiple Sclerosis, Multiple Sclerosis Functional Composite, computer-based testing, 25-foot walk test, 9-hole peg test, Symbol Digit Modalities Test, Low Contrast Visual Acuity, Clinical Outcome Measure
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Training Synesthetic Letter-color Associations by Reading in Color
Authors: Olympia Colizoli, Jaap M. J. Murre, Romke Rouw.
Institutions: University of Amsterdam.
Synesthesia is a rare condition in which a stimulus from one modality automatically and consistently triggers unusual sensations in the same and/or other modalities. A relatively common and well-studied type is grapheme-color synesthesia, defined as the consistent experience of color when viewing, hearing and thinking about letters, words and numbers. We describe our method for investigating to what extent synesthetic associations between letters and colors can be learned by reading in color in nonsynesthetes. Reading in color is a special method for training associations in the sense that the associations are learned implicitly while the reader reads text as he or she normally would and it does not require explicit computer-directed training methods. In this protocol, participants are given specially prepared books to read in which four high-frequency letters are paired with four high-frequency colors. Participants receive unique sets of letter-color pairs based on their pre-existing preferences for colored letters. A modified Stroop task is administered before and after reading in order to test for learned letter-color associations and changes in brain activation. In addition to objective testing, a reading experience questionnaire is administered that is designed to probe for differences in subjective experience. A subset of questions may predict how well an individual learned the associations from reading in color. Importantly, we are not claiming that this method will cause each individual to develop grapheme-color synesthesia, only that it is possible for certain individuals to form letter-color associations by reading in color and these associations are similar in some aspects to those seen in developmental grapheme-color synesthetes. The method is quite flexible and can be used to investigate different aspects and outcomes of training synesthetic associations, including learning-induced changes in brain function and structure.
Behavior, Issue 84, synesthesia, training, learning, reading, vision, memory, cognition
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Assessment of Age-related Changes in Cognitive Functions Using EmoCogMeter, a Novel Tablet-computer Based Approach
Authors: Philipp Fuge, Simone Grimm, Anne Weigand, Yan Fan, Matti Gärtner, Melanie Feeser, Malek Bajbouj.
Institutions: Freie Universität Berlin, Charité Berlin, Freie Universität Berlin, Psychiatric University Hospital Zurich.
The main goal of this study was to assess the usability of a tablet-computer-based application (EmoCogMeter) in investigating the effects of age on cognitive functions across the lifespan in a sample of 378 healthy subjects (age range 18-89 years). Consistent with previous findings we found an age-related cognitive decline across a wide range of neuropsychological domains (memory, attention, executive functions), thereby proving the usability of our tablet-based application. Regardless of prior computer experience, subjects of all age groups were able to perform the tasks without instruction or feedback from an experimenter. Increased motivation and compliance proved to be beneficial for task performance, thereby potentially increasing the validity of the results. Our promising findings underline the great clinical and practical potential of a tablet-based application for detection and monitoring of cognitive dysfunction.
Behavior, Issue 84, Neuropsychological Testing, cognitive decline, age, tablet-computer, memory, attention, executive functions
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Identification of Disease-related Spatial Covariance Patterns using Neuroimaging Data
Authors: Phoebe Spetsieris, Yilong Ma, Shichun Peng, Ji Hyun Ko, Vijay Dhawan, Chris C. Tang, David Eidelberg.
Institutions: The Feinstein Institute for Medical Research.
The scaled subprofile model (SSM)1-4 is a multivariate PCA-based algorithm that identifies major sources of variation in patient and control group brain image data while rejecting lesser components (Figure 1). Applied directly to voxel-by-voxel covariance data of steady-state multimodality images, an entire group image set can be reduced to a few significant linearly independent covariance patterns and corresponding subject scores. Each pattern, termed a group invariant subprofile (GIS), is an orthogonal principal component that represents a spatially distributed network of functionally interrelated brain regions. Large global mean scalar effects that can obscure smaller network-specific contributions are removed by the inherent logarithmic conversion and mean centering of the data2,5,6. Subjects express each of these patterns to a variable degree represented by a simple scalar score that can correlate with independent clinical or psychometric descriptors7,8. Using logistic regression analysis of subject scores (i.e. pattern expression values), linear coefficients can be derived to combine multiple principal components into single disease-related spatial covariance patterns, i.e. composite networks with improved discrimination of patients from healthy control subjects5,6. Cross-validation within the derivation set can be performed using bootstrap resampling techniques9. Forward validation is easily confirmed by direct score evaluation of the derived patterns in prospective datasets10. Once validated, disease-related patterns can be used to score individual patients with respect to a fixed reference sample, often the set of healthy subjects that was used (with the disease group) in the original pattern derivation11. These standardized values can in turn be used to assist in differential diagnosis12,13 and to assess disease progression and treatment effects at the network level7,14-16. We present an example of the application of this methodology to FDG PET data of Parkinson's Disease patients and normal controls using our in-house software to derive a characteristic covariance pattern biomarker of disease.
Medicine, Issue 76, Neurobiology, Neuroscience, Anatomy, Physiology, Molecular Biology, Basal Ganglia Diseases, Parkinsonian Disorders, Parkinson Disease, Movement Disorders, Neurodegenerative Diseases, PCA, SSM, PET, imaging biomarkers, functional brain imaging, multivariate spatial covariance analysis, global normalization, differential diagnosis, PD, brain, imaging, clinical techniques
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Quantitative Autonomic Testing
Authors: Peter Novak.
Institutions: University of Massachusetts Medical School.
Disorders associated with dysfunction of autonomic nervous system are quite common yet frequently unrecognized. Quantitative autonomic testing can be invaluable tool for evaluation of these disorders, both in clinic and research. There are number of autonomic tests, however, only few were validated clinically or are quantitative. Here, fully quantitative and clinically validated protocol for testing of autonomic functions is presented. As a bare minimum the clinical autonomic laboratory should have a tilt table, ECG monitor, continuous noninvasive blood pressure monitor, respiratory monitor and a mean for evaluation of sudomotor domain. The software for recording and evaluation of autonomic tests is critical for correct evaluation of data. The presented protocol evaluates 3 major autonomic domains: cardiovagal, adrenergic and sudomotor. The tests include deep breathing, Valsalva maneuver, head-up tilt, and quantitative sudomotor axon test (QSART). The severity and distribution of dysautonomia is quantitated using Composite Autonomic Severity Scores (CASS). Detailed protocol is provided highlighting essential aspects of testing with emphasis on proper data acquisition, obtaining the relevant parameters and unbiased evaluation of autonomic signals. The normative data and CASS algorithm for interpretation of results are provided as well.
Medicine, Issue 53, Deep breathing, Valsalva maneuver, tilt test, sudomotor testing, Composite Autonomic Severity Score, CASS
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Low-stress Route Learning Using the Lashley III Maze in Mice
Authors: Amanda Bressler, David Blizard, Anne Andrews.
Institutions: Pennsylvania State University, Pennsylvania State University, Pennsylvania State University, Pennsylvania State University, University of California, Los Angeles, University of California, Los Angeles.
Many behavior tests designed to assess learning and memory in rodents, particularly mice, rely on visual cues, food and/or water deprivation, or other aversive stimuli to motivate task acquisition. As animals age, sensory modalities deteriorate. For example, many strains of mice develop hearing deficits or cataracts. Changes in the sensory systems required to guide mice during task acquisition present potential confounds in interpreting learning changes in aging animals. Moreover, the use of aversive stimuli to motivate animals to learn tasks is potentially confounding when comparing mice with differential sensitivities to stress. To minimize these types of confounding effects, we have implemented a modified version of the Lashley III maze. This maze relies on route learning, whereby mice learn to navigate a maze via repeated exposure under low stress conditions, e.g. dark phase, no food/water deprivation, until they navigate a path from the start location to a pseudo-home cage with 0 or 1 error(s) on two consecutive trials. We classify this as a low-stress behavior test because it does not rely on aversive stimuli to encourage exploration of the maze and learning of the task. The apparatus consists of a modular start box, a 4-arm maze body, and a goal box. At the end of the goal box is a pseudo-home cage that contains bedding similar to that found in the animal’s home cage and is specific to each animal for the duration of maze testing. It has been demonstrated previously that this pseudo-home cage provides sufficient reward to motivate mice to learn to navigate the maze1. Here, we present the visualization of the Lashley III maze procedure in the context of evaluating age-related differences in learning and memory in mice along with a comparison of learning behavior in two different background strains of mice. We hope that other investigators interested in evaluating the effects of aging or stress vulnerability in mice will consider this maze an attractive alternative to behavioral tests that involve more stressful learning tasks and/or visual cues.
Neuroscience, Issue 39, mouse, behavior testing, learning, memory, neuroscience, phenotyping, aging
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Behavioral and Locomotor Measurements Using an Open Field Activity Monitoring System for Skeletal Muscle Diseases
Authors: Kathleen S. Tatem, James L. Quinn, Aditi Phadke, Qing Yu, Heather Gordish-Dressman, Kanneboyina Nagaraju.
Institutions: Children's National Medical Center, George Washington University School of Medicine and Health Sciences.
The open field activity monitoring system comprehensively assesses locomotor and behavioral activity levels of mice. It is a useful tool for assessing locomotive impairment in animal models of neuromuscular disease and efficacy of therapeutic drugs that may improve locomotion and/or muscle function. The open field activity measurement provides a different measure than muscle strength, which is commonly assessed by grip strength measurements. It can also show how drugs may affect other body systems as well when used with additional outcome measures. In addition, measures such as total distance traveled mirror the 6 min walk test, a clinical trial outcome measure. However, open field activity monitoring is also associated with significant challenges: Open field activity measurements vary according to animal strain, age, sex, and circadian rhythm. In addition, room temperature, humidity, lighting, noise, and even odor can affect assessment outcomes. Overall, this manuscript provides a well-tested and standardized open field activity SOP for preclinical trials in animal models of neuromuscular diseases. We provide a discussion of important considerations, typical results, data analysis, and detail the strengths and weaknesses of open field testing. In addition, we provide recommendations for optimal study design when using open field activity in a preclinical trial.
Behavior, Issue 91, open field activity, functional testing, behavioral testing, skeletal muscle, congenital muscular dystrophy, muscular dystrophy
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Morris Water Maze Test for Learning and Memory Deficits in Alzheimer's Disease Model Mice
Authors: Kelley Bromley-Brits, Yu Deng, Weihong Song.
Institutions: University of British Columbia.
The Morris Water Maze (MWM) was first established by neuroscientist Richard G. Morris in 1981 in order to test hippocampal-dependent learning, including acquisition of spatial memoryand long-term spatial memory 1. The MWM is a relatively simple procedure typically consisting of six day trials, the main advantage being the differentiation between the spatial (hidden-platform) and non-spatial (visible platform) conditions 2-4. In addition, the MWM testing environment reduces odor trail interference 5. This has led the task to be used extensively in the study of the neurobiology and neuropharmacology of spatial learning and memory. The MWM plays an important role in the validation of rodent models for neurocognitive disorders such as Alzheimer’s Disease 6, 7. In this protocol we discussed the typical procedure of MWM for testing learning and memory and data analysis commonly used in Alzheimer’s disease transgenic model mice.
Neuroscience, Issue 53, Morris Water Maze, spatial memory testing, hippocampal dependent learning, Alzheimer's Disease
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Shallow Water (Paddling) Variants of Water Maze Tests in Mice
Authors: Robert M.J. Deacon.
Institutions: University of Oxford.
When Richard Morris devised his water maze in 19817, most behavioral work was done in rats. However, the greater understanding of mouse genetics led to the mouse becoming increasingly important. But researchers found that some strains of mutant mice were prone to problems like passively floating or diving when they were tested in the Morris water maze11. This was unsurprising considering their natural habitat; rats swim naturally (classically, the "sewer rat"), whereas mice evolved in the dry areas of central Asia. To overcome these problems, it was considered whether shallow water would be a sufficient stimulus to provide escape motivation for mice. This would also avoid the problems of drying the small creatures with a towel and then putting them in a heated recovery chamber to avoid hypothermia, which is a much more serious problem than with rats; the large ratio of surface area to volume of a mouse makes it particularly vulnerable to rapid heat loss. Another consideration was whether a more natural escape strategy could be used, to facilitate learning. Since animals that fall into water and swim away from the safety of the shore are unlikely to pass on their genes, animals have evolved a natural tendency to swim to the edge of a body of water. The Morris water maze, however, requires them to swim to a hidden platform towards the center of the maze - exactly opposite to their evolved behavior. Therefore the paddling maze should incorporate escape to the edge of the apparatus. This feature, coupled with the use of relatively non-aversive shallow water, embodies the "Refinement" aspect of the "3 Rs" of Russell and Burch8. Various types of maze design were tried; the common feature was that the water was always shallow (2 cm deep) and escape was via a tube piercing the transparent wall of the apparatus. Other tubes ("false exits") were also placed around the walls but these were blocked off. From the inside of the maze all false exits and the single true exit looked the same. Currently a dodecagonal (12-sided) maze is in use in Oxford, with 12 true/false exits set in the corners. In a recent development a transparent paddling Y-maze has been tested successfully.
Behavior, Issue 76, Neuroscience, Neurobiology, Medicine, Psychology, Mice, hippocampus, paddling pool, Alzheimer's, welfare, 3Rs, Morris water maze, paddling Y-maze, Barnes maze, animal model
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Automated, Quantitative Cognitive/Behavioral Screening of Mice: For Genetics, Pharmacology, Animal Cognition and Undergraduate Instruction
Authors: C. R. Gallistel, Fuat Balci, David Freestone, Aaron Kheifets, Adam King.
Institutions: Rutgers University, Koç University, New York University, Fairfield University.
We describe a high-throughput, high-volume, fully automated, live-in 24/7 behavioral testing system for assessing the effects of genetic and pharmacological manipulations on basic mechanisms of cognition and learning in mice. A standard polypropylene mouse housing tub is connected through an acrylic tube to a standard commercial mouse test box. The test box has 3 hoppers, 2 of which are connected to pellet feeders. All are internally illuminable with an LED and monitored for head entries by infrared (IR) beams. Mice live in the environment, which eliminates handling during screening. They obtain their food during two or more daily feeding periods by performing in operant (instrumental) and Pavlovian (classical) protocols, for which we have written protocol-control software and quasi-real-time data analysis and graphing software. The data analysis and graphing routines are written in a MATLAB-based language created to simplify greatly the analysis of large time-stamped behavioral and physiological event records and to preserve a full data trail from raw data through all intermediate analyses to the published graphs and statistics within a single data structure. The data-analysis code harvests the data several times a day and subjects it to statistical and graphical analyses, which are automatically stored in the "cloud" and on in-lab computers. Thus, the progress of individual mice is visualized and quantified daily. The data-analysis code talks to the protocol-control code, permitting the automated advance from protocol to protocol of individual subjects. The behavioral protocols implemented are matching, autoshaping, timed hopper-switching, risk assessment in timed hopper-switching, impulsivity measurement, and the circadian anticipation of food availability. Open-source protocol-control and data-analysis code makes the addition of new protocols simple. Eight test environments fit in a 48 in x 24 in x 78 in cabinet; two such cabinets (16 environments) may be controlled by one computer.
Behavior, Issue 84, genetics, cognitive mechanisms, behavioral screening, learning, memory, timing
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Methods for the Modulation and Analysis of NF-κB-dependent Adult Neurogenesis
Authors: Darius Widera, Janine Müller, Yvonne Imielski, Peter Heimann, Christian Kaltschmidt, Barbara Kaltschmidt.
Institutions: University of Bielefeld, University of Bielefeld.
The hippocampus plays a pivotal role in the formation and consolidation of episodic memories, and in spatial orientation. Historically, the adult hippocampus has been viewed as a very static anatomical region of the mammalian brain. However, recent findings have demonstrated that the dentate gyrus of the hippocampus is an area of tremendous plasticity in adults, involving not only modifications of existing neuronal circuits, but also adult neurogenesis. This plasticity is regulated by complex transcriptional networks, in which the transcription factor NF-κB plays a prominent role. To study and manipulate adult neurogenesis, a transgenic mouse model for forebrain-specific neuronal inhibition of NF-κB activity can be used. In this study, methods are described for the analysis of NF-κB-dependent neurogenesis, including its structural aspects, neuronal apoptosis and progenitor proliferation, and cognitive significance, which was specifically assessed via a dentate gyrus (DG)-dependent behavioral test, the spatial pattern separation-Barnes maze (SPS-BM). The SPS-BM protocol could be simply adapted for use with other transgenic animal models designed to assess the influence of particular genes on adult hippocampal neurogenesis. Furthermore, SPS-BM could be used in other experimental settings aimed at investigating and manipulating DG-dependent learning, for example, using pharmacological agents.
Neuroscience, Issue 84, NF-κB, hippocampus, Adult neurogenesis, spatial pattern separation-Barnes maze, dentate gyrus, p65 knock-out mice
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A Proboscis Extension Response Protocol for Investigating Behavioral Plasticity in Insects: Application to Basic, Biomedical, and Agricultural Research
Authors: Brian H. Smith, Christina M. Burden.
Institutions: Arizona State University.
Insects modify their responses to stimuli through experience of associating those stimuli with events important for survival (e.g., food, mates, threats). There are several behavioral mechanisms through which an insect learns salient associations and relates them to these events. It is important to understand this behavioral plasticity for programs aimed toward assisting insects that are beneficial for agriculture. This understanding can also be used for discovering solutions to biomedical and agricultural problems created by insects that act as disease vectors and pests. The Proboscis Extension Response (PER) conditioning protocol was developed for honey bees (Apis mellifera) over 50 years ago to study how they perceive and learn about floral odors, which signal the nectar and pollen resources a colony needs for survival. The PER procedure provides a robust and easy-to-employ framework for studying several different ecologically relevant mechanisms of behavioral plasticity. It is easily adaptable for use with several other insect species and other behavioral reflexes. These protocols can be readily employed in conjunction with various means for monitoring neural activity in the CNS via electrophysiology or bioimaging, or for manipulating targeted neuromodulatory pathways. It is a robust assay for rapidly detecting sub-lethal effects on behavior caused by environmental stressors, toxins or pesticides. We show how the PER protocol is straightforward to implement using two procedures. One is suitable as a laboratory exercise for students or for quick assays of the effect of an experimental treatment. The other provides more thorough control of variables, which is important for studies of behavioral conditioning. We show how several measures for the behavioral response ranging from binary yes/no to more continuous variable like latency and duration of proboscis extension can be used to test hypotheses. And, we discuss some pitfalls that researchers commonly encounter when they use the procedure for the first time.
Neuroscience, Issue 91, PER, conditioning, honey bee, olfaction, olfactory processing, learning, memory, toxin assay
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Contextual and Cued Fear Conditioning Test Using a Video Analyzing System in Mice
Authors: Hirotaka Shoji, Keizo Takao, Satoko Hattori, Tsuyoshi Miyakawa.
Institutions: Fujita Health University, Core Research for Evolutionary Science and Technology (CREST), National Institutes of Natural Sciences.
The contextual and cued fear conditioning test is one of the behavioral tests that assesses the ability of mice to learn and remember an association between environmental cues and aversive experiences. In this test, mice are placed into a conditioning chamber and are given parings of a conditioned stimulus (an auditory cue) and an aversive unconditioned stimulus (an electric footshock). After a delay time, the mice are exposed to the same conditioning chamber and a differently shaped chamber with presentation of the auditory cue. Freezing behavior during the test is measured as an index of fear memory. To analyze the behavior automatically, we have developed a video analyzing system using the ImageFZ application software program, which is available as a free download at Here, to show the details of our protocol, we demonstrate our procedure for the contextual and cued fear conditioning test in C57BL/6J mice using the ImageFZ system. In addition, we validated our protocol and the video analyzing system performance by comparing freezing time measured by the ImageFZ system or a photobeam-based computer measurement system with that scored by a human observer. As shown in our representative results, the data obtained by ImageFZ were similar to those analyzed by a human observer, indicating that the behavioral analysis using the ImageFZ system is highly reliable. The present movie article provides detailed information regarding the test procedures and will promote understanding of the experimental situation.
Behavior, Issue 85, Fear, Learning, Memory, ImageFZ program, Mouse, contextual fear, cued fear
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Using an Automated 3D-tracking System to Record Individual and Shoals of Adult Zebrafish
Authors: Hans Maaswinkel, Liqun Zhu, Wei Weng.
Institutions: xyZfish.
Like many aquatic animals, zebrafish (Danio rerio) moves in a 3D space. It is thus preferable to use a 3D recording system to study its behavior. The presented automatic video tracking system accomplishes this by using a mirror system and a calibration procedure that corrects for the considerable error introduced by the transition of light from water to air. With this system it is possible to record both single and groups of adult zebrafish. Before use, the system has to be calibrated. The system consists of three modules: Recording, Path Reconstruction, and Data Processing. The step-by-step protocols for calibration and using the three modules are presented. Depending on the experimental setup, the system can be used for testing neophobia, white aversion, social cohesion, motor impairments, novel object exploration etc. It is especially promising as a first-step tool to study the effects of drugs or mutations on basic behavioral patterns. The system provides information about vertical and horizontal distribution of the zebrafish, about the xyz-components of kinematic parameters (such as locomotion, velocity, acceleration, and turning angle) and it provides the data necessary to calculate parameters for social cohesions when testing shoals.
Behavior, Issue 82, neuroscience, Zebrafish, Danio rerio, anxiety, Shoaling, Pharmacology, 3D-tracking, MK801
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Construction of Vapor Chambers Used to Expose Mice to Alcohol During the Equivalent of all Three Trimesters of Human Development
Authors: Russell A. Morton, Marvin R. Diaz, Lauren A. Topper, C. Fernando Valenzuela.
Institutions: University of New Mexico Health Sciences Center.
Exposure to alcohol during development can result in a constellation of morphological and behavioral abnormalities that are collectively known as Fetal Alcohol Spectrum Disorders (FASDs). At the most severe end of the spectrum is Fetal Alcohol Syndrome (FAS), characterized by growth retardation, craniofacial dysmorphology, and neurobehavioral deficits. Studies with animal models, including rodents, have elucidated many molecular and cellular mechanisms involved in the pathophysiology of FASDs. Ethanol administration to pregnant rodents has been used to model human exposure during the first and second trimesters of pregnancy. Third trimester ethanol consumption in humans has been modeled using neonatal rodents. However, few rodent studies have characterized the effect of ethanol exposure during the equivalent to all three trimesters of human pregnancy, a pattern of exposure that is common in pregnant women. Here, we show how to build vapor chambers from readily obtainable materials that can each accommodate up to six standard mouse cages. We describe a vapor chamber paradigm that can be used to model exposure to ethanol, with minimal handling, during all three trimesters. Our studies demonstrate that pregnant dams developed significant metabolic tolerance to ethanol. However, neonatal mice did not develop metabolic tolerance and the number of fetuses, fetus weight, placenta weight, number of pups/litter, number of dead pups/litter, and pup weight were not significantly affected by ethanol exposure. An important advantage of this paradigm is its applicability to studies with genetically-modified mice. Additionally, this paradigm minimizes handling of animals, a major confound in fetal alcohol research.
Medicine, Issue 89, fetal, ethanol, exposure, paradigm, vapor, development, alcoholism, teratogenic, animal, mouse, model
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A Video Demonstration of Preserved Piloting by Scent Tracking but Impaired Dead Reckoning After Fimbria-Fornix Lesions in the Rat
Authors: Ian Q. Whishaw, Boguslaw P. Gorny.
Institutions: Canadian Centre for Behavioural Neuroscience, University of Lethbridge.
Piloting and dead reckoning navigation strategies use very different cue constellations and computational processes (Darwin, 1873; Barlow, 1964; O’Keefe and Nadel, 1978; Mittelstaedt and Mittelstaedt, 1980; Landeau et al., 1984; Etienne, 1987; Gallistel, 1990; Maurer and Séguinot, 1995). Piloting requires the use of the relationships between relatively stable external (visual, olfactory, auditory) cues, whereas dead reckoning requires the integration of cues generated by self-movement. Animals obtain self-movement information from vestibular receptors, and possibly muscle and joint receptors, and efference copy of commands that generate movement. An animal may also use the flows of visual, auditory, and olfactory stimuli caused by its movements. Using a piloting strategy an animal can use geometrical calculations to determine directions and distances to places in its environment, whereas using an dead reckoning strategy it can integrate cues generated by its previous movements to return to a just left location. Dead reckoning is colloquially called "sense of direction" and "sense of distance." Although there is considerable evidence that the hippocampus is involved in piloting (O’Keefe and Nadel, 1978; O’Keefe and Speakman, 1987), there is also evidence from behavioral (Whishaw et al., 1997; Whishaw and Maaswinkel, 1998; Maaswinkel and Whishaw, 1999), modeling (Samsonovich and McNaughton, 1997), and electrophysiological (O’Mare et al., 1994; Sharp et al., 1995; Taube and Burton, 1995; Blair and Sharp, 1996; McNaughton et al., 1996; Wiener, 1996; Golob and Taube, 1997) studies that the hippocampal formation is involved in dead reckoning. The relative contribution of the hippocampus to the two forms of navigation is still uncertain, however. Ordinarily, it is difficult to be certain that an animal is using a piloting versus a dead reckoning strategy because animals are very flexible in their use of strategies and cues (Etienne et al., 1996; Dudchenko et al., 1997; Martin et al., 1997; Maaswinkel and Whishaw, 1999). The objective of the present video demonstrations was to solve the problem of cue specification in order to examine the relative contribution of the hippocampus in the use of these strategies. The rats were trained in a new task in which they followed linear or polygon scented trails to obtain a large food pellet hidden on an open field. Because rats have a proclivity to carry the food back to the refuge, accuracy and the cues used to return to the home base were dependent variables (Whishaw and Tomie, 1997). To force an animal to use a a dead reckoning strategy to reach its refuge with the food, the rats were tested when blindfolded or under infrared light, a spectral wavelength in which they cannot see, and in some experiments the scent trail was additionally removed once an animal reached the food. To examine the relative contribution of the hippocampus, fimbria–fornix (FF) lesions, which disrupt information flow in the hippocampal formation (Bland, 1986), impair memory (Gaffan and Gaffan, 1991), and produce spatial deficits (Whishaw and Jarrard, 1995), were used.
Neuroscience, Issue 26, Dead reckoning, fimbria-fornix, hippocampus, odor tracking, path integration, spatial learning, spatial navigation, piloting, rat, Canadian Centre for Behavioural Neuroscience
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T-maze Forced Alternation and Left-right Discrimination Tasks for Assessing Working and Reference Memory in Mice
Authors: Hirotaka Shoji, Hideo Hagihara, Keizo Takao, Satoko Hattori, Tsuyoshi Miyakawa.
Institutions: Fujita Health University, Japan Science and Technology Agency, Core Research for Evolutionary Science and Technology (CREST), National Institutes of Natural Sciences.
Forced alternation and left-right discrimination tasks using the T-maze have been widely used to assess working and reference memory, respectively, in rodents. In our laboratory, we evaluated the two types of memory in more than 30 strains of genetically engineered mice using the automated version of this apparatus. Here, we present the modified T-maze apparatus operated by a computer with a video-tracking system and our protocols in a movie format. The T-maze apparatus consists of runways partitioned off by sliding doors that can automatically open downward, each with a start box, a T-shaped alley, two boxes with automatic pellet dispensers at one side of the box, and two L-shaped alleys. Each L-shaped alley is connected to the start box so that mice can return to the start box, which excludes the effects of experimenter handling on mouse behavior. This apparatus also has an advantage that in vivo microdialysis, in vivo electrophysiology, and optogenetics techniques can be performed during T-maze performance because the doors are designed to go down into the floor. In this movie article, we describe T-maze tasks using the automated apparatus and the T-maze performance of α-CaMKII+/- mice, which are reported to show working memory deficits in the eight-arm radial maze task. Our data indicated that α-CaMKII+/- mice showed a working memory deficit, but no impairment of reference memory, and are consistent with previous findings using the eight-arm radial maze task, which supports the validity of our protocol. In addition, our data indicate that mutants tended to exhibit reversal learning deficits, suggesting that α-CaMKII deficiency causes reduced behavioral flexibility. Thus, the T-maze test using the modified automatic apparatus is useful for assessing working and reference memory and behavioral flexibility in mice.
Neuroscience, Issue 60, T-maze, learning, memory, behavioral flexibility, behavior, mouse
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Light/dark Transition Test for Mice
Authors: Keizo Takao, Tsuyoshi Miyakawa.
Institutions: Graduate School of Medicine, Kyoto University.
Although all of the mouse genome sequences have been determined, we do not yet know the functions of most of these genes. Gene-targeting techniques, however, can be used to delete or manipulate a specific gene in mice. The influence of a given gene on a specific behavior can then be determined by conducting behavioral analyses of the mutant mice. As a test for behavioral phenotyping of mutant mice, the light/dark transition test is one of the most widely used tests to measure anxiety-like behavior in mice. The test is based on the natural aversion of mice to brightly illuminated areas and on their spontaneous exploratory behavior in novel environments. The test is sensitive to anxiolytic drug treatment. The apparatus consists of a dark chamber and a brightly illuminated chamber. Mice are allowed to move freely between the two chambers. The number of entries into the bright chamber and the duration of time spent there are indices of bright-space anxiety in mice. To obtain phenotyping results of a strain of mutant mice that can be readily reproduced and compared with those of other mutants, the behavioral test methods should be as identical as possible between laboratories. The procedural differences that exist between laboratories, however, make it difficult to replicate or compare the results among laboratories. Here, we present our protocol for the light/dark transition test as a movie so that the details of the protocol can be demonstrated. In our laboratory, we have assessed more than 60 strains of mutant mice using the protocol shown in the movie. Those data will be disclosed as a part of a public database that we are now constructing. Visualization of the protocol will facilitate understanding of the details of the entire experimental procedure, allowing for standardization of the protocols used across laboratories and comparisons of the behavioral phenotypes of various strains of mutant mice assessed using this test.
Neuroscience, Issue 1, knockout mice, transgenic mice, behavioral test, phenotyping
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Morris Water Maze Experiment
Authors: Joseph Nunez.
Institutions: Michigan State University (MSU).
The Morris water maze is widely used to study spatial memory and learning. Animals are placed in a pool of water that is colored opaque with powdered non-fat milk or non-toxic tempera paint, where they must swim to a hidden escape platform. Because they are in opaque water, the animals cannot see the platform, and cannot rely on scent to find the escape route. Instead, they must rely on external/extra-maze cues. As the animals become more familiar with the task, they are able to find the platform more quickly. Developed by Richard G. Morris in 1984, this paradigm has become one of the "gold standards" of behavioral neuroscience.
Behavior, Issue 19, Declarative, Hippocampus, Memory, Procedural, Rodent, Spatial Learning
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Quantifying Cognitive Decrements Caused by Cranial Radiotherapy
Authors: Lori- Ann Christie, Munjal M. Acharya, Charles L. Limoli.
Institutions: University of California Irvine .
With the exception of survival, cognitive impairment stemming from the clinical management of cancer is a major factor dictating therapeutic outcome. For many patients afflicted with CNS and non-CNS malignancies, radiotherapy and chemotherapy offer the best options for disease control. These treatments however come at a cost, and nearly all cancer survivors (~11 million in the US alone as of 2006) incur some risk for developing cognitive dysfunction, with the most severe cases found in patients subjected to cranial radiotherapy (~200,000/yr) for the control of primary and metastatic brain tumors1. Particularly problematic are pediatric cases, whose long-term survival plagued with marked cognitive decrements results in significant socioeconomic burdens2. To date, there are still no satisfactory solutions to this significant clinical problem. We have addressed this serious health concern using transplanted stem cells to combat radiation-induced cognitive decline in athymic rats subjected to cranial irradiation3. Details of the stereotaxic irradiation and the in vitro culturing and transplantation of human neural stem cells (hNSCs) can be found in our companion paper (Acharya et al., JoVE reference). Following irradiation and transplantation surgery, rats are then assessed for changes in cognition, grafted cell survival and expression of differentiation-specific markers 1 and 4-months after irradiation. To critically evaluate the success or failure of any potential intervention designed to ameliorate radiation-induced cognitive sequelae, a rigorous series of quantitative cognitive tasks must be performed. To accomplish this, we subject our animals to a suite of cognitive testing paradigms including novel place recognition, water maze, elevated plus maze and fear conditioning, in order to quantify hippocampal and non-hippocampal learning and memory. We have demonstrated the utility of these tests for quantifying specific types of cognitive decrements in irradiated animals, and used them to show that animals engrafted with hNSCs exhibit significant improvements in cognitive function3. The cognitive benefits derived from engrafted human stem cells suggest that similar strategies may one day provide much needed clinical recourse to cancer survivors suffering from impaired cognition. Accordingly, we have provided written and visual documentation of the critical steps used in our cognitive testing paradigms to facilitate the translation of our promising results into the clinic.
Medicine, Issue 56, neuroscience, radiotherapy, cognitive dysfunction, hippocampus, novel place recognition, elevated plus maze, fear conditioning, water maze
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What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

How does it work?

We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

Video X seems to be unrelated to Abstract Y...

In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.