The overall goal of these behavioral tasks is to allow researchers to assay drug-related behaviors such as reward and sensitization.These methods can help answer key questions in the drug addiction field, such as which molecules or pathways contribute to drug-induced plasticity.The main advantage of these techniques is that they are high throughput and can be conducted using either home made or purchased equipment.When we do perform conditioned place preference assays using different drug doses, this approach can uncover differences resulting from altered drug sensitivity and provide additional confidence in your findings.This experiment makes use of three chambered CPP boxes equipped with photo beams for automated data collection.Have four or more on hand.This apparatus has two large chambers, A and B, into which mice can be introduced through a lid.Within each chamber is a dimmable light source.The A and B chambers are made visually distinct, for example, white versus black, and also tactilely distinct, such as with different flooring textures.Between the two large chambers is a smaller, interconnecting neutral chamber, or C, which is accessible by sliding doors.For further details on the CPP boxes, including important information on how to set up a balanced design, consult the text protocol.For these experiments, use mice that have undergone regular handling to minimize the impact of variable animal handling on the results.Prepare the test room, clean the CPP boxes thoroughly.Turn on the equipment and dim the room lights.Then, move the mice to the test area 60 to 90 minutes before the trial.On the first testing day, perform a pre-test.Prepare the chambers by opening the inter-chamber doors.Next, open the computer program.Load the test program.Therein, enter the animal IDs and issue the start command.When ready, gently place each mouse into the middle chamber of their assigned apparatus.After the pre-trials, return the mice to their home cages and export the data.From the pre-test results, determine each animal's preference for chambers A and B.Subtract the time spent in each from the other, and use the difference as an index.Give special consideration to animals with strong pre-test preferences, shown here in red, as described in the text protocol.Select the drug paired chamber for each mouse based on this result.The sum of the index scores for each test group should be made equivalent and near zero.In other words, choose the preferred side for about half of the mice.Also, assign animals evenly to the A and B chambers within each group.If conditioning multiple groups simultaneously is desirable, attempt to assign animals that are using the same apparatus to opposite chambers.For more details, consult the text protocol.A critical step in conditioned place preference when using a balanced design is assignment of the drug paired chambers so that pre-test scores are nearly equal for all test groups and as close to zero or no preference as possible.To begin, always weigh each mouse before the conditioning phase.On the first conditioning day, prepare syringes with cocaine or the chosen test drug according to each animal's body weight.On alternate days, prepare syringes with saline or another appropriate vehicle.Prepare the boxes by closing the inter-chamber doors.Load the conditioning program on the computer, enter the IDs, and start the program.Next, inject the animals with their drug or vehicle bolus and load them into their assigned box.It is critical to ensure that conditioning occurs in the proper A or B chambers for each mouse, each day, as assigned, since errors in this step can force removal of animals from the study.After no more than 30 minutes of conditioning, return the mice to their home cages.When all the trials are complete, export the data and return the mice to the colony.This assay makes use of four by eight photo beam arrays that are 20 inches by 11.5 inches.Have between four and 16 on hand.House each array in an open top black Plexiglas chamber.In the testing room, have a red light prepared to illuminate the chambers.In the software, prepare the program sessions for daily habituation and injection trials.First, set the habituation trial length to be 30 to 60 minutes.An hour is a good starting point.Have the beam breaks recorded in five minute or shorter bins.Second, set the time of the trial in each chamber to begin when the first photo beam is tripped, which comes after the trial has been initiated with a start signal.Third, set the injection trial length to be 60 to 120 minutes.If possible, for injection trials, choose to initiate the start signals from the photo beam arrays individually and not in unison.Locomotor sensitization requires an initial testing run of 10 or 11 consecutive days.On each day, bring the mice into the behavioral anteroom 30 to 60 minutes before the test.Now, place a fresh translucent mouse cage into each chamber.Make sure that the photo beams are centered on the long axis of each cage.When ready, remove the mice in random order from their home cages.First, scruff and weigh them.Then place each into their assigned locomotor testing cage for the habituation trial and cover with a lid.Each day involves a habituation trial with no injection, followed by an injection trial.During the habituation trial, prepare syringes based on each mouse's current body weight.After all habituation trials have ended, load the injection trial program and quietly enter the testing room.After injecting a mouse, quickly issue the cage start signal in the software, and then return that mouse to its test cage.Over the first three to four test days, administer saline or another appropriate vehicle.Over the next seven days, administer cocaine or the chosen test drug.After all the injection trials have ended, return the mice to their housing room.In the CPP assay, wild type C57 black 6n mice were conditioned with saline only, or two different concentrations of cocaine.The saline only group showed no change in preference after conditioning, as expected, while enhanced preferences were observed in groups treated with cocaine.The time spent in different chambers pre-and post-test demonstrated that more time was spent in the cocaine-paired chamber by the cocaine treated mice than in either other chamber.Mice that received only saline showed no striking differences in chamber preference, as expected.Next, the locomotor assay was performed on C67 black 6n mice.Saline injections were given for three to four days, followed by cocaine injections at four different doses for seven days.As expected, more pronounced changes in behavior were observed with increasing doses of cocaine.For a challenge, after one to two weeks of withdrawal, each group was given the original dose of cocaine.Additional challenges were then given with variable withdrawal periods and doses.Although this video demonstrated these techniques using commercially purchased equipment, many labs successfully performed them using home-constructed devices.While attempting these procedures, it is important to ensure that all animals receive equal treatment in order to reduce stress, errors and inter-experiment variability.Variables to consider include handling time and time spent in the testing room and chamber.Extensions of these assays including extinction and reinstatement for CPP and long term or cross-sensitization challenges for locomotor sensitization may be desirable.In addition, its important to use drug na
