Cholecystokinin (CCK) is an essential modulator for neuroplasticity in sensory and emotional domains. Here, we investigated the role of CCK in motor learning using a single pellet reaching task in mice. Mice with a knockout of cck gene (CCK^-/-) or blockade of CCK-B receptor (CCKBR) showed defective motor learning ability; the success rate of retrieving reward remained at the baseline level compared to the wildtype mice with significantly increased success rate. We observed no long-term potentiation (LTP) upon high-frequency stimulation (HFS) in the motor cortex of CCK^-/- mice, indicating a possible association between motor learning deficiency and neuronal plasticity in the motor cortex. In vivo calcium imaging demonstrated that the deficiency of CCK signalling disrupted the refinement of population neuronal activity in the motor cortex during motor skill training. Anatomical tracing revealed direct projections from CCK-expressing neurons in the rhinal cortex to the motor cortex. Inactivating the CCK neurons in the rhinal cortex using chemogenetic methods significantly suppressed motor learning, and intraperitoneal application of CCK4, a tetrapeptide CCK agonist, rescued the motor learning deficits of CCK^-/- mice. In summary, our results suggest that CCK, which could be provided from the rhinal cortex, enables neuroplasticity in the motor cortex leading to motor skill learning.

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#Title: Cholecystokinin facilitates motor skill learning by modulating neuroplasticity in the motor cortex

[doi:10.5061/dryad.9ghx3ffms]

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Success rate WT-CKO.xlsx: Success rate of WT (wildtype) and CCK-/- (cholecystokinin gene knockout) mice. W represents wildtype mice and KO represents CCK (cholecystokinin) gene knockout mice. Every single mouse was identified with different serial numbers, like W-1, W-2, KO-1,KO-2, and so on (Figure. 1C and Figure 1-figure supplement 1);

WT-CKO hausdorff distance.xlsx: Hausdorff distance of trajectories from WT and CCK-/- mice at Day 1 and Day 3. C57 represents wildtype mice and CCK-/- represents CCK gene knockout mice. (Figure 1E);

Detail performance_WT_CKO.xlsx: Detailed performance of WT and CCK-/- mice. W represents wildtype mice and KO represents CCK gene knockout mice. Every single mouse was identified with different serial numbers, like W-1, W-2, KO-1,KO-2, and so on. M, N, D, S represent miss, no-grasp, drop and success, respectively (Figure 1G);

WT-CKO electrophysiology.xlsx: Normalized EPSP of WT mice and CCK knockout mice. WT represents wildtype mice and CCK-/- represents CCK gene knockout mice, Ch1, Ch2... represent different recording channels (Figure 1H);

WT-CKO electrophysiology before and after.xlsx: Normalized EPSP of WT and CCK knockout mice before HFS (-10-0) and after HFS (50-60) (Figure1I);

Basic movement.xlsx: Basic movement ability of the CCK-/-. Both CKO and KO represent CCK gene knockout mice, WT represents wildtype mice, M1,M2, M3...represent mean time or stride-length of different mice (Figure 1- figure supplement 1C,D,E,F);

Success rate Antagonist.xlsx: Success rate of mice injected with angtagonist or vehicle. A1, A2, A3,... represent every single mice injected with antagonist and V1, V2, V3,... represent every single mice injected with vehicle (Figure 2B, Figure 2-figure supplement 1);

Detail performance_Antagonist.xlsx: Detailed performance of mice injected with CCKBR antagonist or vehicle in the motor cortex. A1, A2, A3,... represent every single mice injected with antagonist and V1, V2, V3,... represent every single mice injected with vehicle (Figure 2C);

C57 Calcium population activity.xlsx: Population activity of C57BL/6 mice during reaching trials at day 1 and day 6. C57-1, C57-2, ... represent every single C57BL/6 mouse (Figure 3E and figure 3-figure supplement 1A, B);

CKO Calcium populational neuronal activity.xlsx: Population activity of CCK-/- mice during reaching trials at day 1 and day 6. K1, K2,... represent every single CCK gene knockout mouse (Figure 3E and figure 3-figure supplement 1C, D);

L365,260 Calcium populational neuronal activity.xlsx: Population activity of C57BL/6 mice injected with antagonist, L365,260, during reaching trials at day 1 and day 6. A1, A2, A3,... represent every single mouse injected with antagonist (Figure 3E and figure 3-figure supplement 1E, F);

Activated population activity.xlsx: Activated population activity of C57BL/6, CCK-/- and C57BL/6 injected with L365,260 (ANT) (Figure 3F);

Trial-to-trial population activity correlation.xlsx: Trial-to-trial population activity correlation of C57BL/6 mice, CCK-/- mice and C57BL/6 mice injected with L365,260 (Figure 3G);

Calcium imaging Hausdorff distance.xlsx: Hausdorff distance of trajectories from C57BL/6, CCK-/- and C57BL/6 injected with L365,26 at Day 1 and Day 6 (Figure 3H);

Success rate Chemogentics.xlsx: Success rate of CCK-Cre mice injected with hM4Di virus + Clozapine (hM4Di-CLZ), hM4Di virus+saline (hM4Di-saline), Control virus + Clozapine (control-CLZ). Serial numbers were used to identify every single mouse (Figure 5B, C, figure 5-figure supplement 1);

Success rate from RC to MC.xlsx: Success rate of C57 mice injected with retro-Cre virus in the motor cortex and the hM4Di or control virus in the rhinal cortex, and the manipulation after virus expression, including injection with saline, Clozapine. mCherry+CLZ represents control virus with clozapine, hM4Di+Saline represents hM4Di virus with saline, hM4Di+CLZ represents hM4Di virus with clozapine and hM4Di(SMC)+CLZ represents hM4Di injected only in the single site of the motor cortex contralateral to the dominant forelimb together with clozapine injection (Figure 6D, E, figure 6-figure supplement 3A, B, C);

Detailed performance _from RC to MC.xlsx: Detailed performance of C57 mice specifically inhbited the projections from the RC to the MC and the controls (Figure 6F);

Effect of manipulation on learned skill.xlsx: Effect of antagonist injection or chemogenetic manipulation on the learned motor skill. A-M means miss performance of the antagonist injected mouse, A-N means no-grasp performance of the antagonist injected mouse, A-D means drop performance of the antagonist injected mouse, A-S means success performance of the antagonist injected mouse, C-M means miss performance of the hM4Di with clozapine mouse, C-N means no-grasp performance of the hM4Di with clozapine mouse, C-D means drop performance of the hM4Di with clozapine mouse, C-S means success performance of the hM4Di with clozapine mouse. Serial numbers were used to identify different single mouse (Figure 2-figure supplement 1D, Figure 6-figure supplement 3D);

CCK4 electrophysiology.xlsx: Normalized EPSP of CCK knockout mice administrated with CCK4 or vehicle. Ch-serial nember means different recording channels (Figure 7A);

CCK4 electrophysiology before and after.xlsx: Normalized EPSP of CCK knockout mice administrated with CCK4 or vehicle before HFS (-10-0) and after HFS (50-60). (Figure 7B);

Success rate CCK4.xlsx: Success rate of CCK-/-mice injected with CCK4 or vehicle (Figuer 7D and Figure 7-figure supplement 1A,B);

Detail performance_CCK4_Vehicle.xlsx: Detailed performance of CCK-/- mice injected (i.p.) with CCK4 or vehicle. V1, V2, V3,... represent every single mouse injected with vehicle before training; C1, C2, C3,... represent every single mouse injected with CCK4 before training (Figure 7E);

Cholecystokinin facilitates motor skill learning by modulating neuroplasticity in the motor cortex

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Abstract

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