Following stroke, animals subsequently exhibit a variety of neurological deficits. The Bederson scale is a global neurological assessment that was developed to measure neurological impairments following stroke 7. Tests include forelimb flexion, resistance to lateral push and circling behavior. A grading scale of 0-3 is used to assess behavioral deficits after stroke. This scoring scale is a simple way to reveal basic neurological deficits. Ischemic animals will have significantly more neurological deficits than non-ischemic animals, resulting in a higher score 389. Many iteration scales have been developed and modified since the Bederson scale, all of them providing simple ways to detect impairments 91011. Although easy to perform, neurological ratings on this scale are limited because of their subjective nature. In addition, deficits on the Bederson scale resolve quickly in many common stroke models, rendering it less useful for the detection of long-term deficits after stroke.

One of the most common neurological scales used in animal studies of stroke is the modified neurological severity scores (mNSS). The mNSS rates neurological functioning on a scale of 14 or 18, depending on mice or rat, respectively. The mNSS includes a composite of motor (muscle status and abnormal movement), sensory (visual, tactile and proprioceptive), reflex and balance tests 312. One point is given for the inability to perform each test while one point is deducted for the lack of a tested reflex, and an overall composite score is given to determine impairment. Neurological rating scores have the ability to assess multiple deficits and can be good for testing over periods of 30-60 days 13. Despite the simplicity of administering the tasks, deficits may be specific to a certain modality or function and could be masked by the composite score. In addition, the reflexes tested in the mNSS are the pinna and startle reflexes, which are unlikely related to damage within the MCA territory.

Exploratory behavior in the rat provides a possibility to investigate the neural basis of spatial and motor behavior, which can be used as an assay of brain function 14. The cylinder test (Figure 1) provides a way to evaluate a rodent's spontaneous forelimb use and has been used in a number of motor system injury models of stroke 1516171819. To evaluate forelimb deficits, the animal is placed in a transparent Plexiglas cylinder and observed. Rats will actively explore vertical surfaces by rearing up on their hindlimbs and exploring the surface with their forelimbs and vibrissae. When assessing behavior in the cylinder, the number of independent wall placements observed for the right forelimb, left forelimb and both forelimbs simultaneously are recorded. Animals with unilateral brain damage will display an asymmetry in forelimb use during vertical exploration 6. The cylinder task has been found to be objective, easy to use and score, sensitive to chronic deficits that others fail to detect and have high inter-rater reliability 19. In addition, no pre-training is required, although it is best to obtain baseline data to test for pre-operative bias because, on occasions, some animals display independent use of one limb 17. It is best to use this assessment during the animal's dark cycle and under red lighting conditions because rodents are more apt to explore in a dark environment 19. This test has also been found to have the ability to detect even mild neurological impairments 6.

The grid walking task, often referred to as the foot fault task, is a relatively simple way to assess motor impairments of limb functioning (most commonly hindlimbs, but forelimbs have been evaluated as well) and placing deficits during locomotion in rodents. This task has been found to objectively demonstrate motor coordination deficits 2 and rehabilitation effects after stroke 512. An animal is placed on an elevated, leveled grid with openings. Animals without brain damage will typically place their paws precisely on the wire frame to hold themselves while moving along the grid. Each time a paw slips through an open grid, a "foot fault" is recorded. The number of both contra- and ipsilateral faults for each limb is compared to the total number of steps taken and then scored using a foot fault index 15. Intact animals will generally demonstrate few to no foot faults 220, and when faults occur, they do so symmetrically 20. Ischemic animals typically make significantly more contralateral foot faults than intact animals 2131621. The foot fault test has been shown to be a sensitive indicator for detecting impairments of sensorimotor function after ischemia in rodents 22 and requires very little pre-training 220. However, the foot fault has not been shown to provide a good model for testing long-term deficits beyond 30 to 60 days 13. This time-dependent test appears more appropriate for exploring early treatment effects due to spontaneous recovery 21.

Hindlimb testing in rats can be difficult because the hindlimbs in rodents are not generally used for complex movement 19. The ledged tapered beam-walking test, however, has been used as a reliable measure of hindlimb functioning 19. This test is a locomotor assessment that detects placement dysfunction of the hindlimbs after unilateral brain damage. In this task, animals must walk across an elevated balance beam that tapers at one end and has an under-hanging ledge. Foot faults made with hindlimbs are viewed as deficits in hindlimb function. After a stroke, an animal's foot faults will increase on the contralateral side as the ledge tapers and the difficulty of the task increases. For this task, pre-training is required until the animal successfully walks across the beam without turning around and without making any faults. It is also important to allow each animal a short break between trials so the animal does not habituate to the task 19. Although hindlimb testing has been found to be rather difficult to assess in the animal model, the ledged tapered beam task reveals deficits that rats might normally make compensatory adjustments that they can conceal 19. This assessment may be useful not only for determining the extent to which experimental therapies promote brain repair in but also may be helpful to show motor learning 4. The ledged beam task might be more sensitive to detect hindlimb placing deficits while the grid walking test might be more sensitive to impairments of forelimb functioning 20.

It has recently been shown that rodent skilled paw movements are more similar to primate hand movements than once thought 23. Reaching chambers have been useful for the investigation of skilled forepaw use and motor functioning deficits after unilateral brain damage in rodents 52425. The chamber consists of a Plexiglas apparatus with a tray located on the outside of the wall that holds food targets. The single pellet tray with two indentions can be used to examine an animal's success at retrieving an individual pellet. A pellet is placed in the indention contralateral to the reaching limb in order to evaluate unilateral brain impairments (Figure 2). A reaching tray can also be used to analyze the overall number of pellets retrieved in a given amount of time. A removable barrier, designed to inhibit pellet reaching from one side of the body, can be used for forced retrieval from either side. An animal must reach through a window, grasp and retract the pellet. Attempts, successes, failures and drops are recorded in order to determine impairments. In addition, time from reaching to eating and paw preference for reaching can also be measured. Damage to the motor cortex region from stroke results in impairments of skilled reaching 2627.

The skilled reaching task has also been used to investigate the effects of intact forelimb usage on the recovery of the impaired forelimb after stroke 5. Rats have been found to compensate for motor deficits displayed in skilled reaching by relying on the unimpaired limb 28. By restraining the use of the unimpaired limb, a researcher can evaluate the effects of forced use of the impaired forelimb. In addition, altering the height of the single pellet tray has been shown to detect more subtle deficits such as postural adjustments 24. Grasping and digit movements can also be observed after motor cortex damage from stroke 23. Although paw dexterity, which is commonly impaired in human stroke patients, can be evaluated in this task, the rat paw is small and digit movements can be difficult to document.

Training for the skilled reaching task generally requires a long period of pre-training by first shaping the animals to acquire the skills of the task. In order for animals to be sufficiently motivated to participate in the task, animals are typically food deprived and kept at 85-90% of their normal body weight. Despite long periods of training, paw reaching is a valuable way to assess motor functioning because it is sensitive to movements that seem to be unimpaired in other voluntary behaviors 29, and rats' skilled forelimb movements are similar to those of humans 30. Additionally, deficits can be detectable for up to three months 31.

The staircase test (Figure 3) was developed to assess the independent use of forelimbs of rats 32 and was later adopted to assess skilled reaching in mice 33. This test allows for the bilateral measurement of an animal's forelimb extension, grasping skills and side bias by observing its behavior in a task requiring it to reach for food pellets. The apparatus is designed to encourage the animal to gain access to food by entering a narrow space, a natural rodent behavior 32. After the animal climbs on a platform, it must reach to either side to retrieve food from a double set of staircases. Pellets are placed on each step on both sides. The animal cannot simply scoop the pellet; it must make a coordinated reach and grasp to retrieve it. A normal animal will typically collect the pellets rapidly 34. Latency and the number of pellets from each side and location at increasing distances are then calculated to determine impairments. Animals will often attempt to reach and retrieve pellets but instead knock them to lower levels. To account for this, Adkins-Muir and Jones 35 have provided a useful improvement that allows the experimenter to know which pellets were grasped and retrieved and which were knocked to lower levels by staining the pellets different colors for specific staircase levels.

The staircase test allows for the assessment of forelimb reaching capacities, sensory capacities, dexterity and motor coordination 8. It has been shown by some to be sensitive to detecting long-lasting deficits from ischemia 18. A key advantage of this task is the ability to evaluate skilled use of the limbs independently following impairment. Although food deprivation and daily pre-training of a few weeks is generally required to teach the animals to retrieve the pellets 834, the test is easy to score 34. One potential drawback, however, is that the test may not detect detailed behavior and it lacks sensitivity to forepaw preference compared to other tests 32. Since rehabilitation efforts for unilateral brain damage are more sensitive when evaluating independent limb use of the contralateral limb as opposed to examining limb bias, the ability of the staircase test to evaluate independent limb functioning is a key feature 32. This task has been found to be an effective way to examine functional motor capabilities and goal-directed paw use after ischemia 3132.

In addition to reaching, manual dexterity is often impaired after suffering damage to the central nervous system. Rats live in environments that require them to manipulate objects and use a wide array of motor skills to gain access to food 19; thus dexterous manipulation of items are part of their repertoire. While there are only a few tests that have the ability to measure skillful forepaw use, the pasta test has been shown to reveal manipulative skilled impairments. When eating a piece of pasta, a rat's fine motor skills are often asymmetrical at first (Figure 4). It will use one paw to perform supportive functions and the other paw to move the piece of pasta. One paw, referred to as the "grasp paw", is used in a whole paw grasp and is typically positioned further away from the mouth. The other paw, referred to as the "guide paw", holds the pasta between one or two digits and the thumb, is positioned closer to the mouth and is used to guide the pasta into the mouth. Adjustments are made with both paws as the pasta is eaten. As the pasta becomes shorter, the paws move together into a symmetrical holding pattern. As symmetrical holding is acquired, one paw becomes directly placed on top of the other, and the digits become interposed. The number of adjustments for each paw is recorded as well as the number and type of atypical behaviors. After stroke, a rat will show differences in the number of adjustments made with each paw as well as an increase in atypical behaviors. This test is quantitatively measureable, relatively simple to administer and is sensitive to sensory, motor and lateralized impairments 36. While other dexterous tasks can be labor intensive, the pasta test requires no shaping and little pre-training.

The ladder rung test, initially developed for rats 37 and later adapted for mice 38, is another assessment used to assess skilled walking for both fore- and hindlimb placing. This test is designed to measure placing, stepping and inter-limb coordination 37. The apparatus consists of a horizontal ladder which an animal spontaneously walks across. The spacing of the ladder rungs can be varied to prevent a compensatory reliance for impairments through learning the spacing and location of the rungs 37. For analysis, video recordings of foot faults are generally scored on a rating scale according to quality of limb placement. Animals with motor system injury have been shown to display impairments in the ladder rung test after stroke 373839. This task requires minimal training and is useful for measuring chronic deficits and long-term treatments 37.

A very basic assessment used to detect neurological deficits is a test of forelimb flexion. This test requires an experimenter to suspend a rat by its tail. The posture of the forelimb is then observed and rated. Typically, an animal will extend its forelimbs toward the ground. A rat that has undergone MCAO will flex the contralateral forelimb and twist its body towards the contralateral side of damage 1621. Deficits will tend to decrease with time as recovery occurs. This measure has been found to be particularly sensitive to detecting deficits for several weeks following MCAO 16.

A rat's vibrissae play a very important role in its sensory environment 19. Forelimb placing can be assessed by stimulating a rat's vibrissae to trigger a response. To test forelimb function, an animal is held by its torso, forelimbs hanging free, while brushing its vibrissae on the corner edge of a table. Damage to the motor system will elicit paw placing impairments. Non-brain damaged animals will typically elicit a response of placing the forelimb ipsilateral to the side of vibrissae stimulation on the table 6840. This task can be scored as either calculating the percentage of placing responses for contralateral and ipsilateral responses 1516 or on a scoring scale 8. Animals with unilateral brain damage have been found to have difficulty eliciting a placing response on the contralateral side 61740. Intact rats will generally have a high success rate with this task 19. Although the forelimb placing test has been found to detect even mild neurological impairments 616, a well-handled rat is required for accurate scoring. It is also important to have an experienced examiner in order to prevent abrupt movements resulting in an elicited response 4.

The corner test is a sensorimotor functional assessment that has been shown to be reliable for identifying and quantifying sensorimotor and postural asymmetries. It has also been shown to provide a simple way of detecting contralateral deficits and ipsilateral turning biases. This test was first described while investigating unilateral nigrostriatal damage in the rat 18 and was later used to investigate focal cerebral ischemia in the mouse 13. The apparatus consists of two boards placed closely together at a 30 degree angle to form a narrow alley. An animal is then placed in between the boards facing the corner. As the animal approaches the corner, both sides of the vibrissae are simultaneously stimulated which leads the animal to rear and turn 180 degrees. Intact animals will usually turn around to the right or left randomly while animals with unilateral brain damage will preferentially turn around in the ipsilateral direction, leading with the non-impaired limb and displaying an asymmetry in corner turning 61213. Baseline data is recommended for the reduction of variability and identification of preferential side. In addition to identifying sensorimotor deficits, the corner test has been shown to be an objective assessment of long-term functional outcome (up to 90 days) after stroke in both the rat and the mouse 4. The corner test might be more sensitive in detecting deficits than the other symmetry tests because it reflects multiple asymmetries, including forelimb, hindlimb, postural and turning bias 6. Some feel that the corner test is a measurement that is equated with neglect in stroke patients.

The rotarod test is used to assess motor coordination and balance alterations in rodents 1. It was first introduced by Dunham and Miya 41 to study neurological functioning and was later improved to investigate motor deficits of naïve mice and the effects of drug administration 42. Since then, many have used it to explore brain injury such as stroke 10. The rotarod apparatus is a rod which rotates at an adjustable speed. The speed of the rod increases with time, and the amount of time the animal remains on the device is recorded. Animals with ischemia have been shown to have significantly shorter times of staying on the spinning rod 12. For this test, a researcher only needs to devote a minimal amount of time to train each intact animal to perform this task before brain injury 2. Human survivors of stroke often suffer from lack of coordination 2, and this behavioral assessment offers an approach to investigate coordination deficits in the animal model. However, since stroke patients are not challenged with similar devices such as rotarods to test their balance, the translational significance of the rotarod is unclear. Despite this, its sensitivity to detect motor impairments in other ischemic models has been well-established 142.

First developed to investigate stimulus-directed movement asymmetries resulting from unilateral nigrostriatal damage 18, the adhesive removal test is now commonly used to investigate stroke related impairments of tactile extinction 3821. Two adhesive tapes of equal size are applied as bilateral tactile stimuli on the dorsal side of the paws, which rats will naturally remove from their body by grooming. Tactile responses are measured by recording the time of initial contact with both the ipsilateral and contralateral paws and how much time after each contact it takes the animal to remove the adhesive from each side. Time to contact and time to remove separate out sensory vs. motor deficits. Rats with unilateral brain damage typically develop a bias for both contacting and removing the adhesive from the unimpaired, ipsilateral limb first 418192021. For this assessment, pre-training and baseline data are recommended to obtain optimal level of performance as well as identifying any pre-operative asymmetries 819. It is also essential to maintain consistency in the animal's testing environment, usually in the home cage, because small changes may impact the functional outcome 19.

The adhesive removal test can also be used to measure an animal's sensory asymmetries. The magnitude of sensory asymmetry is measured by adjusting the ratio of the size of the adhesive tapes on each limb. The adhesive on the impaired limb is increased while the adhesive on the unimpaired limb is decreased. This process is repeated until it has been determined between which two levels a bias exists. The magnitude of asymmetry test is highly sensitive to even minor deficits and treatment effects 18. Changes can be quantified over time to measure behavioral recovery 20 and reinstatement of symmetrical tactile sensation 17. This test can reveal asymmetrical biases in stimulus-directed activity in brain damage of sensorimotor areas and long-lasting deficits after focal ischemia 8 and MCAO 21.

The Morris water maze 43 is a commonly used cognitive and behavioral assessment tool that assesses spatial learning and various memory systems. The apparatus consists of a round pool filled with opaque water. Animals are required to swim to a submerged platform to escape the water. Initial heading angle, latency to reach the platform and path length can be analyzed to study swimming behaviors and determine memory impairments. It has been used to evaluate learning for both reference and working memory impairments after stroke 4916. Damage to particular regions of the forebrain can be detected by using various procedural variables of the water maze 4. MCAO has been found by some to cause prolonged disturbance of spatial memories in rats using this task 91621. Additionally, the water maze has been shown to reveal long lasting impairments in rats 16 and detect memory impairments in mice 44. Other researchers, however, have been unable to detect spatial deficits after stroke in both the rat 1545 and mouse models 1. Some have suggested that ischemic rats only show slight impairments in cognitive functioning of navigation because they use a non-spatial strategy to find the platform 46 while others state the discrepancy may be due to variability in protocols 1.

Modifications of the traditional use of the maze have been developed by moving the platform's location within a quadrant to assess spatial navigation memory and flexibility of learning 47. An advantage to this method is the ability to detect subtle search strategy deficits by probing spatial memory early and continuously as the animals learn 4. Using the platform moving procedure, stroke animals have been found to display deficits after learning 4. The water maze is also known for producing highly quantifiable data. A primary advantage of using the water maze over other common behavioral mazes to test memory is that there are no olfactory trails for animals to use scent tracking to find the target. In addition, food deprivation is not required for motivational purposes.

The radial arm maze is another method to study learning behavior and memory impairments in an animal stroke model. This task is also sensitive to hippocampal damage. The apparatus consists of a central platform with eight or twelve radial arms projecting from the center. Food targets are placed at the end of the arms. Generally, animals must be at least partially food deprived to initiate motivation. Animals are provided with ample spatial cues for spatial orientation. When testing for reference memory, an animal is required to retain a memory for the procedures of the task. The stimulus-response association remains constant for all trials 48. Working memory assessment, however, requires the animal to remember which arms (target arms change from trial to trial) previously contained the food targets for one trial. Ischemic animals have been found to make more errors than non-ischemic animals when searching for food 4546. Some studies have shown impairments in both reference and working memory 4546 while others have found that ischemic rats showed more impairments of working memory than reference memory 4849. These findings suggest that spatial cognitive functioning is impaired after ischemia, but it is difficult to fully explain the increase of working memory errors over reference memory errors 46.