Behavioral tests.

Motor and cognitive functions were assessed by the same operator blinded to the treatment. A group of animals was tested at 6, 10, and 12 weeks of age for the assessment of the motor behavior by the RotaRod test. Another group of animals was tested at 9 and 13 weeks of age for the assessment of cognitive deficits by the fear conditioning test. For RotaRod, the test was performed using the accelerating RotaRod apparatus (Ugo Basile, catalog 47600). Mice were moved to the RotaRod room at least 30 minutes before starting the test. Mice were tested 2 days a week. On each testing day, mice were first trained at a constant speed of 4 rpm for 5 minutes on the apparatus. Mice were then tested in 3 consecutive sessions with acceleration from 4 to 40 rpm in 5 minutes. The time at which mice fell from the rotating bar was recorded, for a maximum of 5 minutes. The best performance of each day was recorded, and the average between the 2 trials was reported in the graph. Contextual and cued fear conditioning was performed using the Fear Conditioning System (Ugo Basile, series 46000). WT-TAT-Ala-ADAM10^709–729, R6/2-TAT-Ala-ADAM10^709–729, and R6/2-TAT-Ala-ADAM10^709–729 mice were tested by cued fear conditioning according to the following protocol: on day 1, mice were transferred to the behavioral room at least 30 minutes before starting the test. The conditioned stimulus (CS) was a 10-second 3.5 kHz tone delivered 150 seconds after the beginning of the test. The unconditioned stimulus (US) was a 2-second 0.5 mA footshock that coterminated with the CS. The conditioning was repeated 3 times, every 150 seconds. After the last conditioning, animals were let in the same chamber for 150 seconds, and then they were placed in their cage. During day 2, animals were left in their housing room to rest. On day 3, cued fear conditioning was assessed using a modified conditioning chamber obtained by attaching patterned contexts on the walls and floor. Once mice were placed in this modified chamber, a protocol similar to that of day 1 was applied, except for the electrical footshock that was not administered. Freezing of animals was assessed and recorded during the tone activation in order to focus the analysis during the CS. Latency to freeze and time of freezing were recorded. WT, R6/2, R6/2-A10cKO, and A10cKO mice were tested by contextual and cued fear conditioning according to the following protocol: on day 1, mice were trained as described above for WT and R6/2 mice treated with TAT peptides. On day 2, contextual memory was assessed by placing animals in the same conditioning chamber for 5 minutes and measuring freezing response in terms of time of freezing and latency to freeze. On day 3, cued fear conditioning was assessed as described above for WT and R6/2 mice treated with TAT peptides.