Name: the treadmill fatigue test: a simple, high-throughput assay of fatigue-like behavior for the mouse
Uploaded: 2016-05-31T15:00:00
Description: Watch this Scientific Journal Video about The Treadmill Fatigue Test: A Simple, High-throughput Assay of Fatigue-like Behavior for the Mouse at JoVE.com

This research was supported by the Intramural Research Program of the NIH, The National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK), Grant 1Z01 DK011006. We wish to thank Michele Allen for providing technical assistance, Eleni Solomos for editorial assistance, and the NIH veterinary and animal care staff for providing care for the mice used in developing this method.

This procedure was approved by the National Institute of Diabetes and Digestive and Kidney Diseases Animal Care and Use Committee.
1. Preparation
<ol>
	<li>To allow for rapid identification of each mouse prior to testing, tattoo the tails of all mice to be trained and tested with identifying marks. 
	NOTE: This step is optional. Permanent marker or other methods of identification can be used as an alternative to tattooing.</li>
	<li>Prior to training and testing mice, ensure that the tr...

<ol>
	<li>Hofman, M., Ryan, J. L., Figueroa-Moseley, C. D., Jean-Pierre, P., Morrow, G. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cancer-related+fatigue:+the+scale+of+the+problem.">Cancer-related fatigue: the scale of the problem.</a> Oncologist. 12 Suppl 1, 4-10 (2007).</li><li>Schwartz, A. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Daily+fatigue+patterns+and+effect+of+exercise+in+women+with+breast+cancer.">Daily fatigue patterns and effect of exercise in women with breast cancer.</a> Cancer Pract. 8 (1), 16-24 (2000).</li><li>Schwartz, A. L., Mori, M., Gao, R., Nail, L. M., King, M. 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Phys. 69 (5), 1496-1501 (2007).</li><li>Jean-Pierre, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+phase+3+randomized,+placebo-controlled,+double-blind,+clinical+trial+of+the+effect+of+modafinil+on+cancer-related+fatigue+among+631+patients+receiving+chemotherapy:+a+University+of+Rochester+Cancer+Center+Community+Clinical+Oncology+Program+Research+base+study.">A phase 3 randomized, placebo-controlled, double-blind, clinical trial of the effect of modafinil on cancer-related fatigue among 631 patients receiving chemotherapy: a University of Rochester Cancer Center Community Clinical Oncology Program Research base study.</a> Cancer. 116 (14), 3513-3520 (2010).</li><li>Mar Fan, H. 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E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+lonely+mouse:+verification+of+a+separation-induced+model+of+depression+in+female+mice.">The lonely mouse: verification of a separation-induced model of depression in female mice.</a> Behav. Brain Res. 207 (1), 196-207 (2010).</li><li>V&#245;ikar, V., Polus, A., Vasar, E., Rauvala, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Long-term+individual+housing+in+C57BL/6J+and+DBA/2+mice:+assessment+of+behavioral+consequences.">Long-term individual housing in C57BL/6J and DBA/2 mice: assessment of behavioral consequences.</a> Genes Brain Behav. 4 (4), 240-252 (2005).</li><li>Salem, G. 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The current protocol describes how to use a mouse treadmill to measure fatigue-like behavior. This method has several advantages over VWRA, a common preclinical assay of fatigue-like behavior. VWRA requires that mice choose to interact with the test apparatus. As a result, some inbred strains of mice rarely interact with the wheel16 and run so little that it may be difficult or impossible to identify a fatigue-induced decrease in activity. In contrast, the treadmill fatigue test eliminates that choice and ther...

Fatigue affects a wide range of people, can markedly reduce quality of life, and frequently has an unclear or unknown pathogenesis. Cancer-related fatigue (CRF), for example, is experienced by the majority of cancer patients undergoing treatment and can persist long after cancer treatment has been completed and in the absence of detectable cancer1. Moreover, fatigue is also a prominent symptom in numerous other diseases and disorders, including chronic fatigue syndrome, depression, diabetes, and fibromyalgia. Fortunately, there are non-pharmacological interventions that are capable of helping some people experiencing fatigue (e.g., exercise can reduce CRF for some breast cancer patients2,3), but many individuals still lack effective treatment. Furthermore, existing drug treatments for CRF have not been found to be broadly, if at all, efficacious4-7.
Despite the therapeutic need and lack of drug treatment options, preclinical assays of fatigue to aid in the discovery and development of novel fatigue treatments are lacking, especially in animal models. One of the only preclinical assays of fatigue for rodent studies is voluntary wheel running activity (VWRA)9-15, in which mice or other rodents are given free access to a running wheel and their daily running activity is recorded. In many studies, VWRA is the only measure of fatigue-like behavior, with fatigue-like behavior defined (in either VWRA or the current protocol) as a decrease in the measured physical activity in the experimental group. Although VWRA can provide a useful longitudinal measure of fatigue-like behavior, it is a relatively low-throughput assay, running varies considerably between inbred mouse strains16, and it requires subjects to be individually housed, which may cause changes in behavior and test performance17-19. Other assays, such as home cage behavioral monitoring and analysis, can also provide continuous data collection and some systems may allow for subjects to be housed in pairs20. These assays have utility, but may be less sensitive as a means of detecting fatigue-like behavior and, like wheel running, are also low-throughput.
In contrast to VWRA, mouse treadmill tests do not rely upon voluntary activity and can be completed in a short time frame, allowing for higher throughput. In comparison to VWRA, these tests employ external motivators. Specifically, there is usually an electrified metal grid located to the rear of the moving belt to provide mice with an electric shock should they cease to run. In addition to this shock grid, mice may be motivated to run on the treadmill via several other methods, including prodding, poking, or touching them with a hand, brush, or other tool and directing short puffs of air at them. Instead of fatigue, mouse treadmill tests are often used to measure aerobic and/or anaerobic exercise capacity21-25. Mice are motivated to run until they are incapable of or unwilling to continue running on the treadmill as a means of escaping further electric shocks. Testing then ends when mice meet the criterion for exhaustion. In these protocols, to ensure that mice reach true physiologic exhaustion, the criterion for exhaustion is often defined as spending five continuous seconds laying on top of the shock grid and failing to continue running in the face of repeated aversive stimuli. Thus, fatigue-like behavior may be masked in typical treadmill tests due to the strong aversive nature of the external motivation and criterion for ending the test. Interestingly, and in contrast to many other studies using rodent treadmills, a recent publication describes another version of a treadmill fatigue test, which was used as part of an examination of the effects of social stress in mice26. Although the method used by this group markedly differed from the current protocol (i.e., they employed a single-lane treadmill and required 10 sec of electric shock as the criterion for ending their test), their study highlights the utility of and interest in developing a quick, simple fatigue test using the mouse treadmill.
Fatigue is likely to be detectable by means other than wheel running and alterations in routine behaviors. CRF makes patients feel exhausted by a lesser amount of muscle fatigue, as determined by electromyographic analysis, than people without CRF27. Additionally, reduced motivation has been noted in and is measured by several scales measuring human fatigue28,29. Thus, a useful preclinical assay of fatigue-like behavior should distinguish between healthy and fatigued mice on the basis of a measure other than physiological capability and should not obscure decreases in motivation. To achieve that end while avoiding limitations of VWRA and other assays, the current method was developed by adapting the mouse treadmill test. This method uses a shock grid as the sole external motivator to make mice run on the treadmill. Mice quickly learn that the grid provides an aversive stimulus and will promptly move away from it when placed on the treadmill and maintain some distance from it when running.
When mice fatigue, they spend progressively more time toward the back of the treadmill instead of maintaining speed toward the front end. Therefore, the criterion for test completion in this protocol is spending five continuous seconds in the designated fatigue zone (i.e., the rear of the treadmill, ranging from approximately one body length from the shock grid to, and including, the shock grid). This takes advantage of the aversive nature of the grid without requiring mice to receive many or any actual shocks after training. By allowing mice to complete testing using the current criterion rather than exhaustion (as defined above), this method provides a means of using the treadmill to measure fatigue-like behavior rather than its maximal (or near-maximal) physiological capability. Thus, this method can provide a simple, high-throughput assay of fatigue-like behavior in mice and can serve either as an independent or complementary measure to other assays of fatigue-like behavior.

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			<td>Fisher Scientific</td>
			<td>03-572-5</td>
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			<td height="21" style="height:21px;">Compressed air</td>
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			<td height="21" style="height:21px;">Alcohol (70%)</td>
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the treadmill fatigue test: a simple, high-throughput assay of fatigue-like behavior for the mouse

Fatigue is a prominent symptom in many diseases and disorders and reduces quality of life for many people. The lack of clear pathogenesis and failure of current interventions to adequately treat fatigue in all patients leaves a need for new treatment options. Despite the therapeutic need and importance of preclinical research in helping identify promising novel treatments, few preclinical assays of fatigue are available. Moreover, the most common preclinical assay used to assess fatigue-like behavior, voluntary wheel running, is not suitable for use with some strains of mice, may not be sensitive to drugs that reduce fatigue, and has relatively low throughput. The current protocol describes a novel, non-voluntary preclinical assay of fatigue-like behavior, the treadmill fatigue test, and provides evidence of its efficacy in detecting fatigue-like behavior in mice treated with a chemotherapy drug known to cause fatigue in humans and fatigue-like behavior in animals. This assay may be a beneficial alternative to wheel running, as fatigue-like behavior and potential interventions can be assessed in a greater number of mice over a shorter time frame, thus permitting faster discovery of new therapeutic options.

This protocol allows fatigue-like behavior to be measured in mice using a treadmill. The data presented in this section was obtained by training and testing 3 separate groups of mice using the current protocol (excluding Figure 1A and 1C). To induce fatigue, 5-fluorouracil (5-FU), a cytotoxic chemotherapy drug known to cause fatigue in humans30 and fatigue-like behavior in mice10,13, was administered.&#160;All data presented are from...

Watch this Scientific Journal Video about The Treadmill Fatigue Test: A Simple, High-throughput Assay of Fatigue-like Behavior for the Mouse at JoVE.com

The Treadmill Fatigue Test: A Simple, High-throughput Assay of Fatigue-like Behavior for the Mouse

Fatigue is a common, undertreated and frequently poorly-understood symptom in many diseases and disorders. New preclinical assays of fatigue may help to improve current understanding and future treatment of fatigue. To that end, the current protocol provides a novel means of measuring fatigue-like behavior in the mouse.

Fatigue is a prominent symptom in many diseases and disorders and reduces quality of life for many people. The lack of clear pathogenesis and failure of ...

The overall goal of this procedure is to measure fatigue-like behavior in mice to establish a fatigue model, and test whether intervention can reverse the fatigue. This method can help answer key questions relating to fatigue, such as what interventions can help reduce fatigue or reverse fatigue models. The main advantage of this technique is that it can quickly assess fatigue-like behavior and or anti-fatigue interventions with a single piece of hardware.Visual demonstration of this method is critical. It is important to see and understand how the mice behave in order to successfully perform this task. Prior to training and testing mice, ensure that the treadmill is on a flat surface, and set it to the desired angle of inclination, which is usually 10 degrees.Next, ensure that the electric shock frequency and intensity are set appropriately, which is usually to two hertz and 1.22 milliamps. Under the treadmill, place a piece of paper or an absorbent pad to collect feces and urine. Then, place a pad over one third of the treadmill housing furthest from the shock grid to create a darkened space and entice the mice to stay in that area.Now, proceed with training the mice. With the treadmill off and the speed set to zero, quickly load the mice, holding them by their tails, directly onto the treadmill belt. After placing each mouse, promptly turn on its corresponding shock grid.First, allow the mice to explore freely. Stop the exploring time once each mouse has received at least one shock, or after about three minutes. Then, turn on the treadmill and slowly increase the speed until it begins moving at about 1.5 to 3.0 meters per minute.Ensure that all the mice begin walking. If a mouse does not begin walking, or walks toward the shock grid, tap the mouse with the wire brush or use tail tickling to encourage it to walk on the treadmill. Now, with all the mice walking, slowly increase the treadmill speed to eight meters per minute.Try to ensure that mice are walking forward on the treadmill while increasing the speed during this first training session. If mice are exploring are turning to face the shock grid, cease increasing treadmill speed until they resume walking. Then start a timer and continue monitoring the behavior.After five minutes, increase the speed to nine meters per minute. After seven minutes, increase the speed to 10 meters per minute. After 10 minutes, end the training session by stopping the treadmill.Allow the mice to briefly explore the treadmill a little longer, and return them to their home cage. It is not necessary to have all mice from one cage running at the same time, but try to minimize the number of times mice are removed from any given cage. Between sessions, clean the treadmill and shock grids with alcohol wipes.Also, replace the paper or absorbent pad beneath the treadmill. On the second day of training, repeat the process used on the first day, but increase the treadmill speed to 10 meters per minute before starting the timer. After five minutes, increase the treadmill speed to 11 meters per minute.After 10 minutes, increase the speed to 12 meters per minute and let the treadmill run until 15 minutes have expired. Then, end the training run. Additional days of training can be performed if the mice are having trouble with the task.Once trained, allow at least one full day of rest before the fatigue test. A gap of up to seven days is possible without retraining. In this test, fatigue-like behavior is defined as spending five consecutive seconds in the fatigue zone.The proper technique for removing mice involves gripping near the base of the tail and promptly lifting them out of the treadmill. The fatigue zone is the region of the treadmill belt within a body length of the shock grid, as well as the grid itself. Prior to testing, apply marks to the top or side of the treadmill lanes to delineate the fatigue zone.Next, set the treadmill speed to 12 meters per minute, but do not start the treadmill. Also, make certain that the shock grids are turned off. Now, individually place the mice into their separate lanes and immediately turn on the corresponding shock grid after each mouse is loaded.To begin the test, simultaneously start the treadmill and a stopwatch. Increase the treadmill speed as indicated. This schedule is based on testing female C57 Black 6 mice and may need to be modified to accommodate differently abled mice.Carefully observe all mice throughout the test, but do not intervene to help or motivate mice that enter the fatigue zone. Do not use the wire brush or the tail tickling techniques. Only intervene when a mouse remains in the fatigue zone for five continuous seconds.When this occurs, promptly remove the mouse from the treadmill and record the duration and distance it ran. It is critical that the experimenter carefully observes all the mice on the treadmill, and quickly removes those that have spent five continuous seconds in the fatigue zone. Once no mice remain on the treadmill, prepare for the next run.Mice were trained for five days, and then, to induce fatigue, 5-fluorouracil was administered for five days. This drug is known to cause fatigue in humans and fatigue-like behavior in mice. In the exercise capacity test, which uses tail tickling and air puffs to motivate mice during testing, no difference was seen between groups.On the following day, however, the mice were tested using the treadmill fatigue test. This test revealed a significant difference in the distance run during testing between chemotherapy treated and control mice. To validate that the treadmill fatigue test was measuring fatigue-like behavior, mouse voluntary wheel running was measured in a separate experiment during the five days of 5-fluorouracil treatment, and for an additional night beyond the final day of treatment.Similar results were obtained. In a different experiment, the treadmill fatigue test was also able to detect fatigue-like behavior in mice that had been treated with a different dose of 5-fluorouracil, once weekly, for two weeks. When untreated mice were tested regularly, they became less willing to run upon repeated testing.This data suggests that testing with this protocol should be limited to a single test to avoid a high rate of non-compliant mice. While attempting this procedure, it is important to observe your mice closely during the training and testing. Do not intervene until the criterion for fatigue has been met.After watching this video, you should have a good understanding of how to measure fatigue-like behavior in mice using this protocol, how mice behave on the treadmill, and how to quickly remove mice from the treadmill.

the-treadmill-fatigue-test-simple-high-throughput-assay-fatigue-like

Research

JoVE Journal

Behavior

The Resident-intruder Paradigm: A Standardized Test for Aggression, Violence and Social Stress

This video publication explains in detail the experimental protocol of the resident-intruder paradigm in rats. This test is a standardized method to measure offensive aggression and defensive behavior in a semi natural setting. The most important behavioral elements performed by the resident and the intruder are demonstrated in the video and illustrated using artistic drawings. The use of the resident intruder paradigm for acute and chronic social stress experiments is explained as well. Finally, some brief tests and criteria are presented to distinguish aggression from its more violent and pathological forms.

This video shows the resident-intruder paradigm in rats. This test is a standardized method to measure offensive aggression, defensive behavior and violence in a semi-natural setting. The use of the paradigm for social stress experiments is explained as well.

This video publication explains in detail the experimental protocol of the resident-intruder paradigm in rats. This test is a standardized method to ...

Paw-Dragging: a Novel, Sensitive Analysis of the Mouse Cylinder Test

The cylinder test is routinely used to predict focal ischemic damage to the forelimb motor cortex in rodents. When placed in the cylinder, rodents explore by rearing and touching the walls of the cylinder with their forelimb paws for postural support. Following ischemic injury to the forelimb sensorimotor cortex, rats rely more heavily on their unaffected forelimb paw for postural support resulting in fewer touches with their affected paw which is termed forelimb asymmetry. In contrast, focal ischemic damage in the mouse brain fails to result in comparable consistent deficits in forelimb asymmetry. While forelimb asymmetry deficits are infrequently observed, mice do demonstrate a novel behaviour post stroke termed &#8220;paw-dragging&#8221;. Paw-dragging is the tendency for a mouse to drag its affected paw along the cylinder wall rather than directly push off from the wall when dismounting from a rear to a four-legged stance. We have previously demonstrated that paw-dragging behaviour is highly sensitive to small cortical ischemic injuries to the forelimb motor cortex. Here we provide a detailed protocol for paw-dragging analysis. We define what a paw-drag is and demonstrate how to quantify paw-dragging behaviour. The cylinder test is a simple and inexpensive test to administer and does not require pre-training or food deprivation strategies. In using paw-dragging analysis with the cylinder test, it fills a niche for predicting cortical ischemic injuries such as photothrombosis and Endothelin-1 (ET-1)-induced ischemia &#8211; two models that are ever-increasing in popularity and produce smaller focal injuries than middle cerebral artery occlusion. Finally, measuring paw-dragging behaviour in the cylinder test will allow studies of functional recovery after cortical injury using a wide cohort of transgenic mouse strains where previous forelimb asymmetry analysis has failed to detect consistent deficits.

Classical forelimb asymmetry analysis of the cylinder test is routinely used to assess behavioural deficits in rats following brain injury or stroke; however, it fails to detect consistent deficits in mice. This study demonstrates that quantifying paw-dragging behaviour is a more sensitive analysis of brain injury in mice.

The cylinder test is routinely used to predict focal ischemic damage to the forelimb motor cortex in rodents. When placed in the cylinder, rodents explore ...

The Social Dimension of Stress: Experimental Manipulations of Social Support and Social Identity in the Trier Social Stress Test

In many situations humans are influenced by the behavior of other people and their relationships with them. For example, in stressful situations supportive behavior of other people as well as positive social relationships can act as powerful resources to cope with stress. In order to study the interplay between these variables, this protocol describes two effective experimental manipulations of social relationships and supportive behavior in the laboratory. In the present article, these two manipulations are implemented in the Trier Social Stress Test (TSST)&#8212;a standard stress induction paradigm in which participants are subjected to a simulated job interview. More precisely, we propose (a) a manipulation of the relationship between different protagonists in the TSST by making a shared social identity salient and (b) a manipulation of the behavior of the TSST-selection committee, which acts either supportively or unsupportively. These two experimental manipulations are designed in a modular fashion and can be applied independently of each other but can also be combined. Moreover, these two manipulations can also be integrated into other stress protocols and into other standardized social interactions such as trust games, negotiation tasks, or other group tasks.

Previous research on the social dimension of stress has focused on two important variables: social identity and social support. This protocol introduces an effective experimental manipulation of these two social variables and describes their implementation in a standard stress induction paradigm (Trier Social Stress Test).

In many situations humans are influenced by the behavior of other people and their relationships with them. For example, in stressful situations ...

The 4 Mountains Test: A Short Test of Spatial Memory with High Sensitivity for the Diagnosis of Pre-dementia Alzheimer's Disease

This protocol describes the administration of the 4 Mountains Test (4MT), a short test of spatial memory, in which memory for the topographical layout of four mountains within a computer-generated landscape is tested using a delayed match-to-sample paradigm. Allocentric spatial memory is assessed by altering the viewpoint, colors and textures between the initially presented and target images.
Allocentric spatial memory is a key function of the hippocampus, one of the earliest brain regions to be affected in Alzheimer&#39;s disease (AD) and impairment of hippocampal function predates the onset of dementia. It was hypothesized that performance on the 4MT would aid the diagnosis of predementia AD, which manifests clinically as Mild Cognitive Impairment (MCI).
The 4MT was applied to patients with MCI, stratified further based on cerebrospinal fluid (CSF) AD biomarker status (10 MCI biomarker positive, 9 MCI biomarker negative), and with mild AD dementia, as well as healthy controls. Comparator tests included tests of episodic memory and attention widely accepted as sensitive measures of early AD. Behavioral data were correlated with quantitative MRI measures of the hippocampus, precuneus and posterior cingulate gyrus.
4MT scores were significantly different between the two MCI groups (p = 0.001), with a test score of &#8804;8/15 associated with 100% sensitivity and 78% specificity for the classification of MCI with positive AD biomarkers, i.e., predementia AD. 4MT test scores correlated with hippocampal volume (r = 0.42) and cortical thickness of the precuneus (r = 0.55).
In conclusion, the 4MT is effective in identifying the early stages of AD. The short duration, easy application and scoring, and favorable psychometric properties of the 4MT fulfil the need for a simple but accurate diagnostic test for predementia AD.

This article describes the 4 Mountains Test (4MT), a hippocampus-dependent test of working allocentric spatial memory. The hippocampus is affected early in Alzheimer's disease (AD) and this article outlines the 4MT methodology and results of patient testing, which demonstrates the value of the 4MT in the diagnosis of pre-dementia AD.

This protocol describes the administration of the 4 Mountains Test (4MT), a short test of spatial memory, in which memory for the topographical layout of ...

The Rodent Psychomotor Vigilance Test (rPVT): A Method for Assessing Neurobehavioral Performance in Rats and Mice

The human Psychomotor Vigilance Test (PVT) is a widely used procedure for measuring changes in fatigue and sustained attention. The present article describes a rodent version of the PVT&#8212;termed the &#34;rPVT&#34;&#8212;that measures similar aspects of attention (i.e., performance accuracy, motor speed, premature responding, and lapses in attention). Data are presented that demonstrate both the short- and long-term usefulness of the rPVT when employed with laboratory rats. Rats easily learn the rPVT, and learning to perform the basic procedure takes less than two weeks of training. Once acquired, rat performances in the rPVT show a high degree of similarity to these same performance measures in the human PVT, including similarities in, lapses in attention, reaction times, vigilance decrements across session time (i.e., the human &#34;time-on-task&#34; effects), and the response-stimulus interval (RSI) effect described for humans. Thus the rPVT can be an extremely valuable tool for assessing the effects of a wide range of variables on sustained attention quite similar to human PVT performances, and thus can be useful for developing novel treatments for neurobehavioral dysfunctions.

The Rodent Psychomotor Vigilance Test rPVT: A Method for Assessing Neurobehavioral Performance in Rats and Mice

A rat version of the human Psychomotor Vigilance Test (PVT) is described that measures aspects of attention similar to those measured with the human PVT, including aspects of human vigilance such as performance accuracy, motor speed, premature responding, and lapses in attention.

The human Psychomotor Vigilance Test (PVT) is a widely used procedure for measuring changes in fatigue and sustained attention. The present article ...

A Method to Test the Effect of Environmental Cues on Mating Behavior in Drosophila melanogaster

An individual's sexual drive is influenced by genotype, experience and environmental conditions. How these factors interact to modulate sexual behaviors remains poorly understood. In Drosophila melanogaster, environmental cues, such as food availability, affect mating activity offering a tractable system to investigate the mechanisms modulating sexual behavior. In D. melanogaster, environmental cues are often sensed via the chemosensory gustatory and olfactory systems. Here, we present a method to test the effect of environmental chemical cues on mating behavior. The assay consists of a small mating arena containing food medium and a mating couple. The mating frequency for each couple is continuously monitored for 24 h. Here we present the applicability of this assay to test environmental compounds from an external source through a pressurized air system as well as manipulation of the environmental components directly in the mating arena. The use of a pressurized air system is especially useful to test the effect of very volatile compounds, while manipulating components directly in the mating arena can be of value to ascertain a compound's presence. This assay can be adapted to answer questions about the influence of genetic and environmental cues on mating behavior and fecundity as well as other male and female reproductive behaviors.

A Method to Test the Effect of Environmental Cues on Mating Behavior in Drosophila melanogaster

We demonstrate an assay to analyze the environmental and genetic cues that influence mating behavior in the fruit fly Drosophila melanogaster.

An individual's sexual drive is influenced by genotype, experience and environmental conditions. How these factors interact to modulate sexual behaviors ...

A Simple and Low-cost Assay for Measuring Ambulation in Mouse Models of Muscular Dystrophy

Measuring functional outcomes in the treatment of muscular dystrophy is an essential aspect of preclinical testing. The assessment of voluntary ambulation in mouse models is a non-invasive and reproducible activity assay that is directly analogous to measures of patient ambulation such as the 6-minute walk test and related mobility scores. Many common methods for testing mouse ambulation speed and distance are based on the open field test, where an animal&#39;s free movement within an arena is measured over time. One major downside to this approach is that commercial software and equipment for high-resolution motion tracking is expensive and may require transferring mice to specialized facilities for testing. Here, we describe a low-cost, video-based system for measuring mouse ambulation that utilizes free and open-source software. Using this protocol, we demonstrate that voluntary ambulation in the dystrophin-null mdx mouse model for Duchenne muscular dystrophy (DMD) is decreased relative to wild-type mouse activity. In mdx mice expressing the utrophin transgene, these activity deficits are not observed and the total distance traveled is indistinguishable from wild-type mice. This method is effective for measuring changes in voluntary ambulation associated with dystrophic pathology, and provides a versatile platform that can be readily adapted to diverse research settings.

This protocol describes a flexible, low-cost system for measuring mouse ambulation in an open field activity assay. We show that a 6-minute ambulation assay based on this system detects a decrease in voluntary movement in mdx mice, and accurately distinguishes improvement in a muscle-specific rescue of these animals.

Measuring functional outcomes in the treatment of muscular dystrophy is an essential aspect of preclinical testing. The assessment of voluntary ambulation ...

A Simple Non-invasive Method for Temporary Knockdown of Upper Limb Proprioception

Proprioception may be the least well measured of all contributors to the neural control of movement. New precise, reliable measures of proprioception are needed for clinical diagnosis of impairment, and to measure outcomes of proprioceptive training. The purpose of this simple, non-invasive method is to temporarily knockdown upper limb proprioception in healthy adults, to an extent that would be useful in the development and testing of upper limb proprioception measures. Knockdown models have two main advantages over studying humans with impaired proprioception: participant availability and the ability to control the extent of impairment across participants. Current published methods of temporary proprioception knockdown of the upper limb, such as ischemic nerve blocks and cryotherapy, are invasive, impractical, or uncomfortable for the participant. Here, vibration over the ulnar groove was used to reduce upper limb proprioception. High frequency vibration may reduce proprioceptive acuity by inhibiting pacinian corpuscle-induced input. The effect of vibration used in this protocol was confirmed using two quantitative measures. This method was simple to administer, comfortable for participants, and practical.

The goal of this protocol is to demonstrate a practical method to temporarily interfere with proprioception in the upper limb of healthy humans.

Proprioception may be the least well measured of all contributors to the neural control of movement. New precise, reliable measures of proprioception are ...

Measuring Active and Passive Tameness Separately in Mice

Domesticated animals such as dogs and laboratory mice show a high level of tameness, which is important for humans to handle them easily. Tameness has two behavioral components: a reluctance to avoid humans (passive tameness) and a motivation to approach humans (active tameness). To quantify these components in mice, we previously developed behavioral tests for active tameness, passive tameness, and the willingness to stay on a human hand, each designed to be completed within 3 min. The data obtained were used for selective breeding, with a large number of mice analyzed per generation. The active tameness test measures the movement of the mouse toward a human hand and the contact it engages in. The passive tameness test measures the duration of time that a mouse tolerates human touch. In the stay-on-hand test, a mouse is placed on a human hand and touched slowly using the thumb of that hand; the duration of time that the animal remains on the hand is measured. Here, we describe the test set-up and apparatus, explain the procedures, and discuss the appropriate data analysis. Finally, we explain how to interpret the results.

Tameness in animals includes a reduction of avoidance responses to humans (passive tameness) and an increase in active approaches to humans (active tameness). Here, we describe detailed protocols for three behavioral tests (active tameness, passive tameness, and stay-on-hand tests) to separately measure the active and passive tameness of mice.

Domesticated animals such as dogs and laboratory mice show a high level of tameness, which is important for humans to handle them easily. Tameness has two ...

An Instrumented Pull Test to Characterize Postural Responses

Impairment of postural reflexes, termed postural instability, is a common and disabling deficit in Parkinson's disease. To assess postural reflexes, clinicians typically employ the pull test to grade corrective responses to a backward perturbation at the shoulders. However, the pull test is prone to issues with reliability and scaling (score/4). Here, we present an instrumented version of the pull test to more precisely quantify postural responses. Akin to the clinical test, pulls are manually administered except pull force is also recorded. Displacements of the trunk and feet are captured by a semi-portable motion tracking system. Raw data represent distance traveled (in millimeter units), making subsequent interpretation and analysis intuitive. The instrumented pull test also detects variabilities influencing pull test administration, such as pull force, thereby identifying and quantifying potential confounds that can be accounted for by statistical techniques. The instrumented pull test could have application in studies seeking to capture early abnormalities in postural responses, track postural instability over time, and detect responses to therapy.

Impairment of postural reflexes, termed postural instability, is difficult to quantify. Clinical assessments such as the pull test suffer issues with reliability and scaling. Here, we present an instrumented version of the pull test to objectively characterize postural responses.

Impairment of postural reflexes, termed postural instability, is a common and disabling deficit in Parkinson's disease. To assess postural reflexes, ...