Behavioral Tests for Rodents PDF

Okay, here's a comprehensive breakdown of the behavioral tests, brain imaging techniques, and brain manipulation methods discussed in the sources, organized for easy review. Behavioral Tests for Rodents Motor Function and Activity Gait Test ◦ Detailed Descri...

Okay, here's a comprehensive breakdown of the behavioral tests, brain imaging techniques, and brain manipulation methods discussed in the sources, organized for easy review. Behavioral Tests for Rodents Motor Function and Activity Gait Test ◦ Detailed Description: This test assesses how an animal walks. In humans, it's a simple observation, but in rodents, researchers may use video analysis to track their movements. ◦ Domains of Consideration: This test looks at broad motor function and can be affected by the animal's sensory system, including how they take in information from their environment. ◦ Test Selection: This test may not be very useful if your intervention does not affect gate or ne motor function. It is a very broad test and may not be sensitive to subtle differences or changes. ◦ Step-by-Step Procedures: There are no speci c step by step procedures detailed in the source, but it can be carried out via observational and video methods. ◦ Professor's Emphasis: This test highlights the complexity of motor function, which involves many brain regions, and the need to consider the sensory system. ◦ Data Representation: Data can be represented as observational notes, video analysis, and potentially metrics of gait. ◦ Test Applications: This test looks at broad motor function. ◦ Brain Regions: This test relates to activity in the motor cortex, basal ganglia, and cerebellum. ◦ Animal Experience: This test is fairly low stress as it just analyzes their normal movement. ◦ Critical Analysis: Rodents rely primarily on their sense of smell and not vision, so if an intervention affects their vision, gait might not be the best method of measurement. ◦ Examples: Not speci cally detailed in the source. Learning and Memory Novel Object Recognition (NOR) ◦ Detailed Description: This test evaluates recognition memory. Animals are habituated to an open eld arena, then exposed to two identical objects. After a delay (hour, day, or two days), one object is replaced with a novel one. Intact memory is indicated by more interaction with the novel object. ◦ Domains of Consideration: General activity, motivation to explore. ◦ Test Selection: This task is useful when you are primarily interested in object recognition, without any spatial component. ◦ Step-by-Step Procedures: ▪ Habituate animal to open eld. ▪ Training trial: Expose animal to two identical objects. fi fi fi fi fi  ▪Intertrial interval: 1 hour to 2 days. ▪Testing trial: Replace one object with a novel object; measure time/ interactions with each. ◦ Professor's Emphasis: This is a non-spatial memory test that measures an animal's preference for novel stimuli, and is reliant on areas of the temporal lobe. ◦ Data Representation: Data includes time spent or number of interactions with each object. A discrimination index (percentage of time with novel object) is often used. Control animals will spend more time with the novel object, while models of Alzheimer's will not discriminate, with their discrimination index near chance (50%). ◦ Test Applications: Used to assess recognition memory and the ability to discriminate between familiar and novel objects. ◦ Brain Regions: Hippocampus and cortical areas in the temporal lobe are important for this test. ◦ Animal Experience: Animals are motivated to explore new things, which is why they spend more time with the novel object, and the habituation process should reduce fear of the arena. ◦ Critical Analysis: This test does not measure spatial memory and relies on the animal's motivation to explore new objects. ◦ Examples: Control animals will spend more time exploring the novel object, indicating intact memory, while a model of Alzheimer's will show no preference, indicating impaired memory. Barnes Maze ◦ Detailed Description: This test evaluates spatial learning and memory using a circular platform with holes around the circumference. One hole leads to a dark escape box. Animals use cues around the room to navigate. ◦ Domains of Consideration: Agoraphobia, motor function. ◦ Test Selection: This test is good for spatial learning but may not be best if anxiety is a major confound. ◦ Step-by-Step Procedures: ▪ Place animal in center of the platform. ▪ Animal navigates to a target hole with escape box underneath. ▪ Measure time or path length to nd the box, as well as errors (visits to incorrect holes). ◦ Professor's Emphasis: The professor emphasized that anti-anxiety effects of interventions can confound results. This is because the motivation for the animal to nd the dark box may change. For example, if an intervention reduces anxiety, they will explore the platform more and therefore take longer to nd the goal box. ◦ Data Representation: Data includes the time to nd the escape box, path length, and number of errors. Control animals will take a shorter time to nd the box and make less errors, while transgenic models will take longer and make more errors. ◦ Test Applications: Used to assess spatial learning and memory. ◦ Brain Regions: Hippocampus is important for this test. ◦ Animal Experience: Animals are motivated by agoraphobia and the desire to nd a dark, safe space. fi fi fi fi fi fi  ◦ Critical Analysis: Changes in anxiety levels can alter the results, as can motor impairments. Examples: The professor's study found that exercise, which reduces anxiety, caused mice to take longer to nd the goal box due to increased exploratory behavior in the open space, even though their spatial learning was intact. Morris Water Maze (MWM) ◦ Detailed Description: This test assesses spatial learning and memory using a pool of water with a hidden platform. Animals swim to nd the platform, motivated by the desire to escape the water. ◦ Domains of Consideration: Anxiety, motor function, motivation. ◦ Test Selection: This test is a good option when anxiety is a concern, because the motivation to escape water is a strong motivator and can overcome changes in agoraphobia. ◦ Step-by-Step Procedures: ▪ Hidden trials (spatial learning): Animal swims to nd hidden platform over multiple days. ▪ Probe trial (spatial memory): Platform is removed, and time spent in the target quadrant is measured. ◦ Professor's Emphasis: This test provides a strong motivator (escape from water), which reduces the confound of differences in anxiety. Path length may be a better measure than time if motor function is affected. ◦ Data Representation: Data includes latency to nd the platform, path length, and percentage of time in the target quadrant during the probe trial. Control animals will take a shorter time to nd the platform and spend more time in the correct quadrant. Transgenic animals may have impaired performance on these tasks. ◦ Test Applications: Assesses spatial learning (hidden trials) and memory (probe trial). ◦ Brain Regions: Hippocampus is critical for this test. ◦ Animal Experience: Animals are motivated to escape the water, which can be a stressor, but this can be a stronger motivator than the Barnes maze. ◦ Critical Analysis: Swim speed can confound results if motor function is impaired. Path length may be more appropriate. ◦ Examples: A study using nitrous oxide showed that higher doses of nitrous oxide impaired spatial learning and memory in the water maze, with animals taking longer to nd the platform and spending less time in the target quadrant on the probe trial. Radial Arm Maze ◦ Detailed Description: This test uses a maze with multiple arms to assess spatial learning and memory. Motivation may be food rewards, where food is placed in some of the arms. Researchers measure the time it takes to nd all food, the number of errors, and how often the animal goes to the same arms on different trials. This can be used to evaluate reference and working memory. ◦ Domains of Consideration: Motivation, motor function fi fi fi fi fi fi fi  ◦ Test Selection: This test is helpful if you want to look at a multitude of things, including how long it takes to nd food, how often they make errors, and their reference and working memory. ◦ Step-by-Step Procedures: ▪ Place food at the end of multiple arms, sometimes all of them. ▪ Allow the animal to explore and collect the food. ▪ Measure the time to collect all food, the errors they make, and the number of times they revisit old arms. ◦ Professor's Emphasis: There are different types of memory this test can measure, including reference memory (across days) and working memory (within a single trial). ◦ Data Representation: Data can include, time to collect all food, the number of errors (visiting arms that don't have food), and the number of repeated visits to the same arms. ◦ Test Applications: Measures spatial memory and learning ◦ Brain Regions: Not speci cally detailed in the source, but similar to the other spatial memory tests, is related to the hippocampus ◦ Animal Experience: Animals are motivated by their desire for food. ◦ Critical Analysis: If the animal is not properly food deprived it may not be properly motivated. This may also be compounded if any intervention causes taste aversion or impairs their motivation to eat. ◦ Examples: Not speci cally detailed in the source. Y Maze ◦ Detailed Description: This test, shaped like a Y, is used to assess spatial working memory by evaluating spontaneous alternation. Animals are expected to explore new arms and keep alternating through the three arms rather than repeatedly visiting the same one or two. ◦ Domains of Consideration: General activity levels, motor function. ◦ Test Selection: This is a good test if you are interested in working memory, and is simpler than a radial arm maze. ◦ Step-by-Step Procedures: ▪ Place animal in one arm of the maze ▪ Measure how often they alternate between the three arms ▪ Measure the number of arm entries ◦ Professor's Emphasis: This test relies on natural curiosity. A high percentage of alternation (consistently visiting different arms) indicates intact spatial working memory. ◦ Data Representation: Data includes the percentage of alternation and the number of arm entries. Control animals should have a high alternation percentage, while models of cognitive impairment may display a low alternation percentage. ◦ Test Applications: This test is used to assess spatial working memory ◦ Brain Regions: This test is dependent on the hippocampus and the prefrontal cortex, as the latter is important for keeping the information in mind for short periods of time. ◦ Animal Experience: Animals are motivated by their natural curiosity to explore. fi fi fi  ◦Critical Analysis: Differences in motivation or general activity can confound results. ◦ Examples: Not speci cally detailed in the source. T-Maze ◦ Detailed Description: The T-maze is similar to the Y-maze but uses two arms (T- shape). In this test, one of the arms is blocked on the rst trial to force the animal to turn either left or right to obtain a reward (e.g. food). Then a short interval later, both arms are available. The animal must remember the direction that they previously chose, and then choose the opposite arm. This tests spatial working memory. The duration it takes to make a decision can measure motivation. Domains of Consideration: Motivation. Test Selection: This is a simpler task that still measures spatial working memory. It can also measure motivation by evaluating the amount of time it takes to make a decision. Step-by-Step Procedures: ◦ Block one arm and place food at the end of the available arm ◦ After a short interval (e.g. 30 seconds) allow the animal to choose between the two arms ◦ Measure the percentage of time they chose the correct arm. ◦ Measure the duration it takes them to make the decision Professor's Emphasis: This test uses short intervals to measure working memory. The motivating factor is important and it can show how changes in motivation can affect memory performance. Data Representation: Data includes the percentage of correct arm choices and the trial duration. Control animals may choose the correct arm over chance (50% or greater), while models of cognitive impairment may choose incorrectly more often than chance. Trial duration may be reduced in control animals due to proper motivation, while those that have impaired motivation will take longer to make decisions. Test Applications: This test is used to assess spatial working memory and the in uence of motivation. Brain Regions: This test relies on spatial learning and memory, and is associated with activity in the hippocampus and prefrontal cortex. Animal Experience: Animals are motivated by food rewards and will be hungry if properly food deprived before the test. Critical Analysis: If an intervention affects taste or motivation, it may impair performance. Examples: Not speci cally detailed in the source. Drug Abuse Liability and Addiction Locomotor Sensitization ◦ Detailed Description: Measures the increased motor response to a drug after repeated exposures. Animals are given a drug over a series of days and their locomotor activity is measured in an open eld arena. ◦ Domains of Consideration: General activity, stereotypy. ◦ Test Selection: This is appropriate to see how repeated exposure to drugs affects their motor response, and is an indicator of the likelihood of experiencing dependence and withdrawal. fi fi fi fi fl  ◦ Step-by-Step Procedures: ▪ Administer a drug over multiple days. ▪ Measure distance traveled in an open eld each day. ▪ Observe for stereotypy, which are non-rewarding behaviors such as rocking back and forth. ◦ Professor's Emphasis: This test uses the principle of sensitization which is increased response to a drug after repeated exposure. This is thought to be correlated to neuroplasticity and increased likelihood of dependence. ◦ Data Representation: Data includes the distance traveled and the presence of stereotypies. Increased distance traveled over time indicates sensitization. Increased stereotypies are another indicator of sensitization. ◦ Test Applications: This test shows that repeated exposure to a drug changes its effects on motor function, which may be indicative of addiction. ◦ Brain Regions: Dopamine-related brain regions. ◦ Animal Experience: Animals experience increased motor activity, due to drug effects. ◦ Critical Analysis: This test is not an indicator of whether an animal will choose to take a drug again. It only measures the body's increased response to the drug. ◦ Examples: Repeated exposure to a psychostimulant leads to increased locomotion. Conditioned Place Preference (CPP) ◦ Detailed Description: This test utilizes the associative learning principle of classical conditioning. Animals learn to associate a drug with a speci c environment. The CPP apparatus has two distinct chambers. The drug is administered in one chamber, and saline in the other over a series of days. If the drug is rewarding, animals will start to prefer the chamber associated with the drug, indicated by more time spent on the drug-paired side during a test. ◦ Domains of Consideration: Drug reward. ◦ Test Selection: This test is an appropriate measure of whether an animal nds a drug rewarding, and therefore can indicate its potential for abuse. ◦ Step-by-Step Procedures: ▪ Conditioning: Pair one chamber with drug administration and the other chamber with saline. ▪ Test: Measure time spent in each chamber without any drug or saline. ◦ Professor's Emphasis: This test shows how animals learn to associate a drug with a speci c location, indicating its rewarding properties. ◦ Data Representation: The preference score is the time spent in the drug-paired chamber compared to the saline chamber. Animals that nd the drug rewarding will spend more time in the drug-paired chamber. ◦ Test Applications: This test is used to measure drug reward by evaluating an animal's preference for a drug associated context. ◦ Brain Regions: Reward circuits, including the mesolimbic dopamine pathway. ◦ Animal Experience: Animals experience the drug's effects in one context and the absence of it in the other and will spend more time in the context associated with the drug if it is rewarding. fi fi fi fi fi  ◦Critical Analysis: Animals are not choosing to take the drug and this may not correlate with self administration of a drug. ◦ Examples: Not speci cally detailed in the source. Self-Administration ◦ Detailed Description: This test measures drug-seeking behavior by allowing animals to self-administer a drug via an action (e.g., lever press). This test is a model of operant learning, where an animal has to take action to receive the reward. The test typically uses a xed ratio in which a set number of actions must be performed for a single infusion. The amount of times the animal chooses to perform that action for a reward is an indication of its motivation to obtain the drug. ◦ Domains of Consideration: Motivation, impulsivity. Test Selection: This test is the most direct measure of drug seeking and abuse liability. It allows you to examine motivation, and is better than condition place preference, where the animals do not have to choose to self-administer the drug. Step-by-Step Procedures: ◦ Train animal to press a lever to receive a drug infusion. ◦ Measure the number of lever presses and the breakpoint on a progressive ratio schedule. Professor's Emphasis: This is a very valid model of addiction as animals are choosing when and how much of a drug to take. Data Representation: Data includes number of lever presses, break point on a progressive ratio schedule, or continued lever pressing in spite of negative consequences (such as shock). Higher break points indicate higher motivation for the drug. Compulsive drug seeking is measured when animals will continue to work for the drug even with negative consequences such as foot shock. Test Applications: Used to evaluate drug-seeking, compulsive drug use, motivation, and the escalation of drug use. Brain Regions: Mesolimbic dopamine pathway, prefrontal cortex. Animal Experience: Animals experience the rewarding effects of the drug, are motivated to obtain the drug, and may experience withdrawal if the drug is removed. They may also experience foot shock if there are negative consequences involved. Critical Analysis: It can be dif cult to get rodents to self-administer some drugs, such as marijuana. Examples: Animals will work harder for a drug when they are hungry than when they are sated. Stimulating the prefrontal cortex has been shown to reduce drug seeking behaviors in animals, and inhibiting that region increases drug seeking behaviors in those animals that had previously been resistant to addiction. Other Behavioral Domains Pain and Nociception ◦ Detailed Description: These tests measure sensitivity to pain, using both mechanical and thermal stimuli. Mechanical sensitivity is measured using Von Frey laments to assess withdrawal thresholds and thermal sensitivity can be measured using a hot plate test to look at withdrawal latency. ◦ Domains of Consideration: Sensory sensitivity, motor response fi fi fi fi  ◦ Test Selection: These tests are appropriate if an intervention causes or alleviates pain. ◦ Step-by-Step Procedures: ▪ Apply Von Frey laments of varying thicknesses to a paw to determine the mechanical threshold for withdrawal. ▪ Place an animal on a hot plate and measure how long it takes them to withdraw their paw. ◦ Professor's Emphasis: Pain is an adaptive mechanism. Data Representation: Data includes the threshold at which the animal withdraws from mechanical stimulation or the latency to withdraw the paw on a hot plate. Test Applications: These tests measure pain sensitivity. Brain Regions: Primarily sensory pathways. Animal Experience: Animals will experience noxious stimuli and will withdraw to avoid pain. Critical Analysis: These tests rely on their motor response to sensory stimulation. Examples: Not speci cally detailed in the source. Brain Imaging Techniques Structural Imaging ◦ CT Scans ▪ Uses X-rays to create 3D images of brain structure by combining sequential 2D images. Can show white and gray matter and ventricles. ◦ MRI ▪ Uses non-ionizing radiation and differences in water content in different types of tissue to visualize brain structure. Can show white and gray matter and ventricles. Functional Imaging ◦ fMRI ▪ Measures changes in cerebral blood ow to identify active brain regions. Active areas require more blood ow. ▪ Used to measure the activation of reward related areas in response to loved ones. ◦ PET Imaging ▪ Uses radioactively tagged molecules that bind to receptors or transporters, which are then picked up by the scanner. This allows measurement of neurotransmission, receptor levels, and activity levels by use of radio labeled glucose. ▪ Used to look at dopamine receptor levels and changes in activity in people with substance use disorders. Post-Mortem Analyses (only done on deceased animals or humans) ◦ Autoradiography ▪ Uses radioactive tags to label receptors or transporters in thin sections of brain tissue. ◦ Immunocytochemistry/Immuno uorescence fi fi fl fl fl  ▪ Uses staining or tagging to identify speci c proteins in thin sections of brain tissue. ▪ Used to look at new and mature cells in the hippocampus. ◦ In Situ Hybridization ▪ Uses tags to label mRNA for speci c genes in brain tissue, allowing quanti cation and localization of gene expression levels. ◦ Clarity ▪ Clears out the brain's lipids so it can be stained for a speci c protein in 3D. This allows the assessment of neural connectivity. Electrophysiology ◦ Uses electrodes to measure electrical activity in brain tissue or living animals. Can be done in tissue slices or in vivo, to record neural ring activity in real time in response to a stimulus. Microdialysis and Voltammetry ◦ Probes are inserted into a brain region to measure the levels of neurotransmitters, such as dopamine, while animals are awake. ◦ Can measure the abuse potential of drugs. Two-Photon Imaging ◦ Live imaging through a cranial window that looks at deeper structures in the brain, such as blood vessels, over longer periods of time. Brain Manipulation Methods Lesion Studies ◦ Purposefully damaging or removing a part of the brain to observe the resulting changes in behavior. Methods include chemical injection, radiofrequency waves, knife cuts, and aspiration. ▪ Very crude and can affect more areas than desired. Electrical Stimulation ◦ Stimulating neurons by emitting electrical currents through surgically placed electrodes. This method is used to assess a region's role in behavior. ▪ Not very speci c and will stimulate all cell types. Transcranial Magnetic Stimulation (TMS) ◦ Non-invasive method that uses magnetic pulses to stimulate brain regions, particularly the prefrontal cortex. ▪ Used to reduce cravings in those addicted to drugs. Optogenetics ◦ Combines light and genetic techniques to activate or inhibit speci c neurons using light-gated ion channels. ▪ Channelrhodopsin (ChR2): activated by blue light; causes excitation. ▪ Halorhodopsin (NPHR): activated by yellow light; causes inhibition. ▪ Can target speci c cell types and is used to assess drug seeking behavior. fi fi fi fi fi fi fi fi