Differential participation of the corticospinal and corticorubral neurons during motor execution in the rat

Abstract

The sensorimotor cortex is crucial for learning and executing new movements with precision. It selectively modulates sensory information flow and represents motor information in a spatially organized manner. The pyramidal system is made up of layer 5 pyramidal tract neurons (PTNs), which are organized into populations with distinct morphological, genetic and functional properties. These subpopulations project to different subcortical structures in a segregated manner. To understand whether PTNs projecting to different structures play distinct functional roles in motor control, we characterized two types of layer 5 neurons in the motor cortex: corticorubral (CR) neurons, which project to the red nucleus, and corticospinal (CS) neurons, which project to the spinal cord. To analyze movement performance in rats, we compared the selective optogenetic inhibition of motor cortex CS or CR neurons during lever movement execution in response to a light stimulus. As the animals progressed through the training sessions, the variability of lever trajectories decreased, and the movements became more stereotyped. Photoinhibition of CS or CR neurons increased the performance variability of learned movements but differentially affected kinematic parameters. CR neuron inhibition affected amplitude, duration, reaction times, speed, and acceleration of the movement. In contrast, the inhibition of CS neurons mainly altered the duration and acceleration of the movement. The results indicate that the same motor order would be expected to have different causal effects when sent to different brain regions.

Keywords: Corticorubral; Corticospinal; Layer 5; Pyramidal tract neurons; Sensorimotor cortex.

Fig. 1

Anatomical distribution of CS and CR neurons in the cortex. Retrograde tracer injections (A); at the top, the site of spinal cord injection (BDA), and at the bottom, the site of injection in the red nucleus (FluoroGold). In (B), a coronal section of the cortex is shown (AP: +1 mm); in green, the soma of neurons labeled with BDA, with projections to the spinal cord; in red, the somas of neurons labeled with FluoroGold, with projections to the red nucleus; and in light blue, the somas of neurons labeled with Neun. In (C); in green, the distribution of CS neurons; in red, the distribution of CR neurons; in gray, the distribution of neurons marked with Neun; and in purple, double-marked neurons. Shadows indicate standard error, and dashed vertical lines denote the different layers of the cortex. Venn diagrams show the percentage of CR and CS neurons in each cortex. In (D), density profiles show the relative density of CR neurons (above) and CS neurons (below). The gray dashed line at bregma. The magenta dashed line indicates the coordinate where depth distribution profiles were performed. S1FL (anterior limb somatosensory cortex); M2 (secondary motor cortex); M1 (primary motor cortex); S1HL (posterior limb somatosensory cortex).

Fig. 2

Behavioral paradigm. In (A), light stimulus or delay (700–1300 ms), response time (2000 ms), and inter-trial interval (2000–4000 ms) are illustrated. Learning curve, n = 20 (B), with vertical lines indicating standard error. In (C), comparison of hits between beginner (sessions 1–3) and expert rats (sessions 26–28) (n = 20), showing lever presses for one session per condition on the right, individual trajectories in gray lines, and the average trajectory in shaded line. In (D), experimental design timeline.

Fig. 3

Expression of the virus pAAV-hsyn-Jaws-KGC-GFP-ER2 and photoinhibition protocol. In (A), injection sites in the spinal cord at C5 level (left) and in the red nucleus (right). In (B), virus expression in cortices (Bregma 2.20) M1 and M2 for CS and CR neurons. In (C), amplification of the expression site (red box in B) of CS and CR neurons. In (D), Coronal section showing the location of the optic fiber (white dotted line) and location of the fiber implantation area for each experiment are shown in green (CS group) and red (CR group) dots. S1FL (anterior limb somatosensory cortex); M2 (secondary motor cortex); M1 (primary motor cortex); S1HL (posterior limb somatosensory cortex). (E), inhibition protocol during early phase and late phase; shaded region indicates the time of inhibition.

Fig. 4

Effect of CS neuron inhibition on lever pressing trajectories. (A), lever trajectories of the CS neuron group upon inhibition. Averaged control lever press trajectories are shown in the solid green line under control conditions (left), during early inhibition (middle), and during late inhibition (right) is presented. Individual lever presses are shown in gray. Different kinematic parameters of lever pressing movement were evaluated: hits (B), amplitude (C), duration (D), reaction time (E). Comparisons were made between early and late 465 nm light control lever presses versus early inhibition and late inhibition. (One way ANOVA, Tukey’s multiple comparisons test).

Fig. 5

Effect of CR neuron inhibition on lever pressing trajectories. (A), lever trajectories of the CR neuron group upon inhibition. Averaged control lever press trajectories are shown in the solid red line under control conditions (left), during early inhibition (middle), and during late inhibition (right). Individual lever presses are shown in gray. Different kinematic parameters of lever pressing movement were evaluated: hits (B), amplitude (C), duration (D), reaction time (E). Comparisons were made between early and late 465 nm light control lever presses versus early inhibition and late inhibition. (One way ANOVA, Tukey’s multiple comparisons test).

Fig. 6

Kinematic parameters of lever pressing trajectory between inhibited and control trials. (A), averaged lever press trajectories of CS inhibited animals (Left graphs). The solid green line under control conditions (upper), during early inhibition (middle), and during late inhibition (lower). The averaged p-value (± SE) for all rats with early (middle graph) and late (right graph) inhibitions in CS neurons. Significant values, highlighted in orange, were identified in the ROC curve analysis throughout the temporal window (p-value < 0.05). The gray horizontal dotted line indicates the threshold of 0.05, while the magenta vertical line marks the beginning of the leverage movement. (B), the same as A but for CR inhibited rats.