The overall goal of this procedure is to assess the behavioral effects of compensatory limb use and a mouse model of stroke. This is accomplished by first determining the preferred reaching limb by allowing mice to reach with either limb during shaping procedures. The second step is to establish the motor skill by training only the preferred reaching limb on the pasta matrix reaching task.
Next compensatory limb use is taught by training mice to use the less affected limb on the pasta matrix. Reaching task beginning four days after unilateral ischemic injury contralateral to the preferred reaching limb. The final step is to assess the effect of less affected limb training on the functional outcome of the stroke affected limb.
Ultimately, impaired functional outcome of the affected limb is demonstrated by a decrease in the number of successful reaches over several days of testing. This method can help answer key questions in basic stroke research, including what are the implications of behavioral interventions on functional outcome. Generally, individuals new to this method will struggle because of the time and patients taken to teach mice this highly skilled task.
The benefit is that the sensitivity of this test exceeds that of other sensory motor tests. In mice. Begin the procedure by cutting uncooked CAPI pasta pieces using a razor blade.
Next place the pasta pieces in the pasta matrix. Habituate a mouse to the testing chamber for three sessions of five to 10 minutes each with a full pasta matrix placed in front and several pieces of pasta at the bottom for eating to shape the reach. Place the mouse in the testing chamber with a full pasta matrix in front of it, as in habitation training.
Record the number of reaches with each limb and denote the number of reaches with the right and left limb.Separately. Define a reach as the extension of a limb through the reaching aperture, such that the wrist breaks the plane of the chamber, making the four paw visible outside of the chamber. It is not necessary for the reach to be directed toward or make contact with the pasta during the shaping sessions.
Limit the mouse to a maximum of 10 reaches or 10 minutes of time in the chamber, whichever occurs. First, to encourage the mouse to reach. Hold several pieces of pasta just inside the chamber through the reaching aperture.
Draw the pasta pieces out when the mouse shows interest, such as sniffing or biting, thus encouraging them to reach after the retreating pasta pieces. If the mice do not show interest in the dry pasta, what the tips of the pasta pieces in a small cup of warm water to make them more palatable. Shaping sessions are also used to determine limb preference.
When a mouse exhibits a minimum of 70%of their daily reaches with a single limb, that four limb is defined as the preferred limb. Place a mouse in the testing chamber with a half full matrix in front. Train the mouse to reach only with its preferred limb by filling only the sign of the matrix contralateral to the preferred limb.
During this phase of training, leave the portion of the matrix corresponding to the less affected limb unfilled in order to discourage reach attempts with this limb. Administer once daily, 15 minute, or 100 reach training sessions to help the mouse successfully learn to target and break the pasta pieces. During early training sessions, place a small amount of pressure on the pasta pieces while they are in the matrix by holding another pasta piece lightly, behind and perpendicular to the intended reaching target.
This small amount of support help stabilize the thin pasta and is easier for the mouse to grasp. Successful reaches require the mouse to reach through the aperture. Grab a piece of pasta, break the pasta piece and remove it from the matrix.
Record the total number of reaches up to a maximum of 100, and the number and location of the successfully broken pasta pieces. In this procedure. Train the less affected limb beginning four days after ischemic insult to the motor cortex contralateral to the preferred reaching limb.
Just as with preoperative reaching training, administer once daily, 15 minute, or 100 reach training sessions. Remove the first column of pasta for the first week of less affected limb training. In order to ensure that early reach attempts with the affected limb are not successful, then place the mouse in the reaching chamber with the half filled matrix in front.
Ensure that the filled half of the matrix is opposite to the preoperative filled half, which will force the mouse to now reach with a less affected limb. Encourage reaching with the less affected limb by reinforcing extension of this limb through the reaching aperture to do so. When the mouse extends the less affected limb in the first few days of training, place a small piece of pasta on the floor of the reaching chamber.
Regardless of whether or not the reach is successful, count only those reaches made with the less affected limb toward the 100 reach total. Do not count furtive attempts with the affected limb towards the total number of reaches. Then return the most medial vertical column of pasta to the matrix after the first seven days of less affected limb training record the total number of reaches and the number and location of successfully broken pasta pieces to determine the impact of compensatory use of the less affected limb on functional recovery of the stroke affected limb.
Test the performance of the affected limb on the PMRT. Following less affected limb training. Place the matrix in front of the chamber oriented identically to preoperative training procedures.
This will force the mice to now reach with their stroke affected limb record, the total number of reaches and the number and location of successfully broken pasta pieces to compare with pre-stroke performance levels. Shown here are the representative reaching patterns with the affected limb of mice before and after unilateral ischemic insult of the sensory motor cortex opposite the preferred limb. Anterior denotes pasta pieces located increasingly further in front of the mouse while lateral denotes pasta pieces located increasingly to the side of the mouse.
The representative pattern changes of the affected limb over days of postoperative training are shown here. This graph shows a representative example of how less affected limb training affects performance of the affected limb on the PMRT training of the less affected limb for 15 days following unilateral ischemic insult of the sensory motor cortex impairs functional outcome of the affected limb and impairs successful recovery While attempting this procedure. It's important to remember to avoid over-training the mice during the shaping phase by letting them reach too many times.
Otherwise, the learning curve will not accurately reflect the early stage of learning on this test. Following this procedure. Other methods, including immunohistochemistry, cortical mapping, and in vivo imaging can be used to investigate other questions concerning the impact of behavioral interventions on neuroanatomy.
