This protocol integrates an automated opera and strategy shifting task with a flexible stress paradigm, enabling experimenters to efficiently assay the impact of stress on cognition. The advantages of this technique or its efficiency with the automation of the cognitive task and its flexibility because it can be paired with many other stress paradigms. This method provides insight to the interplay between the stress and cognition.
It could also be extended to assess other variables such as the effect of drug exposure on cognition. Visual demonstration ensures that newcomers to this technique know how to properly test the function of the operant chambers before running subjects to avoid any glitches. Use operant chambers for behavior training and testing.
Ensure that the chambers contain at least two retractable levers with two stimulus lights above. A house light and a dispenser for reinforcement of these tasks. Make sure that the levers are on either side of the central reinforcement delivery area with one stimulus light above each lever.
Then use the house light to eliminate the chamber without interfering with detection of the light stimulus. Use dustless food pellets for reinforcement in food restricted rats. Making sure that the pellets are not high in sucrose or fat, control the presentation of stimuli, lever operation and data collection with software capable of operating the chamber.
Fill the bottom tray of each operant box with fresh bedding to collect waste. After each session dump each tray, clean it with alcohol wipes and add fresh bedding before placing a new animal in the chamber. Decide whether the stress procedure should be performed before, during or after a training on the operant strategy shifting paradigm and execute the stress procedure at the same time daily with respect to operant training.
In a separate room that is neither the colony room or the strategy shifting paradigm room, place the rat in a broom style transparent restraint tube and seal the opening. Taking care not to pinch the limbs or tail stagger the stress procedure for subjects depending on how many operant chambers are available. Before placing the rat in the chamber, ensure that there are enough food pellets in the dispenser and that the operant boxes are properly functioning.
Do this by loading and initiating a program in an empty chamber and manually testing that the correct lever delivers one reward per lever press. Before placing the rat in the box for the first day training, manually set one food pellet reward on the correct lever as designated upon loading the training procedure within each chamber. Train the rat using a fixed ratio schedule such that each correct lever press is rewarded with one reinforcement.
Counterbalance the correct lever per day across subjects or experimental conditions. Allow the rat to press the lever until it reaches the criterion by pressing the correct lever at 50 times which usually takes between 30 and 45 minutes. On the following day, force the rat to perform this task on the opposite lever using the same program and designating the opposite lever as the correct one.
On the test day, place the rat in the operant chamber following stress procedures and test it in side discrimination, side reversal and light discrimination tasks serially. Ensure that the light discrimination task only illuminates the light above the correct lever. For the side discrimination task, reward the rat for pressing the lever on its least preferred side as determined from the third day of training.
Regardless of the light cue, the task ends upon pressing the correct lever eight times consecutively. For the side reversal test, use the side discrimination program again but designate the opposite lever to be the correct one. The rat should be rewarded for pressing this lever regardless of the light cue.
It must press the lever eight times consecutively to end the task. To perform the light discrimination task, reward the rat for pressing the lever with the light illuminated above. Again, each operant testing is complete upon pressing the correct lever eight times consecutively.
This adapted automated operant strategy shifting paradigm was used to determine if repeated restraint stress affects cognition in male and female Sprague Dawley rats. A reduced number of trials or criterion indicated a better performance on each task. Following acute restraint, males completed the side reversal task insignificantly fewer trials than unstressed control males.
Conversely, stressed females required a greater number of trials to complete the task. The total number of errors made for each attention task is shown here consistent with the number of trials to criterion. Stressed males made significantly fewer errors than control males in the side reversal task.
Whereas stressed females made more errors. Furthermore, in the light discrimination task, females also made significantly more errors. In some, these data suggest that repeated stress improves cognitive performance in males but impairs cognitive performance in females.
Total errors were further categorized into perseverative or regressive errors. Stressed males made fewer perseverative errors in the side reversal task than control males. While stressed females made a greater number of perseverative errors than control females in both the side reversal and light discrimination tasks.
The brain areas underlying the side reversal task were examined to determine whether they displayed a similar sex differences in neural activity. Stress induced a significant increase to neuronal activation in the orbital frontal cortex of males compared to controls. However, the opposite was observed in females.
When attempting this protocol pay attention to which subject is in which task of the strategy shifting paradigm. After finishing that task, manually ensure that it moves to the next task with correct settings. Along with this procedure, experimenters can perform other behavioral studies on the effects of restraint or proceed directly to perfusion and immunohisticchemical techniques to determine which brain regions were important in strategy shifting.
In addition to increasing the efficiency and throughput of the paradigm, this technique allows us to differentiate types of errors made by subjects into perseverative, regressive, and random errors.
