Hypothalamic Control of Forelimb Motor Adaptation

Abstract

The ability to perform skilled arm movements is central to everyday life, as limb impairments in common neurologic disorders such as stroke demonstrate. Skilled arm movements require adaptation of motor commands based on discrepancies between desired and actual movements, called sensory errors. Studies in humans show that this involves predictive and reactive movement adaptations to the errors, and also requires a general motivation to move. How these distinct aspects map onto defined neural signals remains unclear, because of a shortage of equivalent studies in experimental animal models that permit neural-level insights. Therefore, we adapted robotic technology used in human studies to mice, enabling insights into the neural underpinnings of motivational, reactive, and predictive aspects of motor adaptation. Here, we show that forelimb motor adaptation is regulated by neurons previously implicated in motivation and arousal, but not in forelimb motor control: the hypothalamic orexin/hypocretin neurons (HONs). By studying goal-oriented mouse-robot interactions in male mice, we found distinct HON signals occur during forelimb movements and motor adaptation. Temporally-delimited optosilencing of these movement-associated HON signals impaired sensory error-based motor adaptation. Unexpectedly, optosilencing affected neither task reward or execution rates, nor motor performance in tasks that did not require adaptation, indicating that the temporally-defined HON signals studied here were distinct from signals governing general task engagement or sensorimotor control. Collectively, these results reveal a hypothalamic neural substrate regulating forelimb motor adaptation.SIGNIFICANCE STATEMENT The ability to perform skilled, adaptable movements is a fundamental part of daily life, and is impaired in common neurologic diseases such as stroke. Maintaining motor adaptation is thus of great interest, but the necessary brain components remain incompletely identified. We found that impaired motor adaptation results from disruption of cells not previously implicated in this pathology: hypothalamic orexin/hypocretin neurons (HONs). We show that temporally confined HON signals are associated with skilled movements. Without these newly-identified signals, a resistance to movement that is normally rapidly overcome leads to prolonged movement impairment. These results identify natural brain signals that enable rapid and effective motor adaptation.

Keywords: hypothalamus; motor adaptation; motor learning; movement; orexin/hypocretin; upper limb.

Copyright © 2022 the authors.

Figures

Figure 1.

Mouse-robot interactions reveal forelimb motor adaptations to defined force fields. A, Left, Diagram of the ETH Pattus robotic manipulandum with top-down view of a head-fixed mouse, pulling the robot handle. Right, Diagram of the skilled motor task. Mice had to pull 9 mm in the y-direction within a ±5 mm x “tunnel” to obtain a reward. On trials 51–200, a velocity-dependent force (cyan arrows) pushed the animal's forelimb in the x direction. B, Top row, Diagram of the sequence of a single trial. Bottom row, Diagram of a single force field session. The mice performed 50 trials with no force field to get a baseline (black bar), then 150 trials of force field exposure (cyan to magenta bar), followed by 50 trials of no force field to determine the washout (red bar). C, Graphical explanation of end x-position and steering angle. D, Left panel, Typical example of trajectory evolution across a force field session. The force field was present in trials 51–200, individual lines represent average of 25 blocks. Right panel, Magnification of the first 0.5 mm of the left trajectories, demonstrating preemptive steering adaptation. E, Averages of five trajectories taken from preforce field (black), force field (blue, purple, magenta), and postforce field (washout, red) trial blocks. Data from 33 sessions from 12 mice. A, Left panel, Lateral displacement at the end of pulls (end x-position), expressed as mean ± SEM, versus trials (n = 33 sessions from 12 mice). The shared x-axis for panels F, H is given in panel H. Right panel, End x-position within indicated trials (mean ± SEM). RM ANOVA F(4,128) = 86.28, p < 0.0001; Sidak's multiple comparison tests ****p < 0.0001, ***p < 0.001. G, Left panel, Same as in F, left panel, for predictive steering angle. Right panel, Same as in F, right panel, for predictive steering angle. RM ANOVA F(4,128) = 18.07, p < 0.0001; Sidak's multiple comparison tests: ****p < 0.0001, ns = p > 0.05. H, Left panel, Same as in F, left panel, showing max y-velocity, ns = p > 0.05. Right panel, Same as in F, right panel, for max y-velocity. RM ANOVA F(4,128) = 3.578, p < 0.01; Sidak's multiple comparison tests: ns = p > 0.05. I, Left panel, Same as in F, left panel, for percentage of successful trials. Right panel, Same as in F, right panel, for percentage of successful trials. RM ANOVA F(4,128) = 5.532, p < 0.001; Sidak's multiple comparison tests: ns = p > 0.05, ****p < 0.0001.

Figure 2.

HON signals associated with skilled forelimb movements and motor adaptation. A, Targeting schematic (left) and typical expression (right) of AAV1-hORX-GCaMP6s in HONs, and example fiber placements in two additional mice (bottom). Dashed yellow box indicates fiber placement. V3, third ventricle; VMH, ventromedial nucleus of the hypothalamus; LH, lateral hypothalamus. B, Typical example of raw Ca2+-dependent (470 = 470 nm-excited fluorescence) and Ca2+-independent (405 = 405 nm-excited fluorescence) components of HON-GCaMP6s signal during trials. Epochs of the single trial are labeled: light blue line indicates the cue onset, blue bar indicates the pull, and magenta line indicates reward delivery. C, Z-scored HON-GCaMP6s activity of left (black) and right hemispheres (blue) during nonforce field trials with an intertrial interval of >15 s. Cue bar (gray) indicates the range of trial cue onsets across animals. Blue bar indicates the range of pull durations across animals, magenta bar indicates the range of reward delivery times. Data are mean ± SEM for n = 50 trials from 4 mice. D, Top, Diagram explaining the window of analysis of premotor and motor-related HON-GCaMP6s signals of a single trial (green bar). Bottom, Evolution of pull-associated HON-GCaMP6s signals across control nonforce field trials (left), and force field trials (right), aligned to pull completion. Traces are baseline subtracted from −2 to −1 s before pull start and averaged across sessions (NForce field = 15 sessions, NNo Force field = 9 sessions, from 4 mice). E, Average GCaMP6s signal amplitude, expressed as Δ Z – score, within indicated trial blocks from D of no force field (black) and force field (green) sessions. Two-way mixed-effects model analysis, interaction F(4,56) = 8.082, p < 0.0001; Sidak's multiple comparison tests: ****p < 0.0001, ns = p > 0.05. Signal trends across the force field trials (cyan shading) and equivalent no force field trials (yellow shading) are compared in Results. F, Average GCaMP6s signal amplitude versus trial number across force field (green) and no force field sessions (black), Δ Z – score represents signal amplitude (see Materials and Methods; NForce field = 15 sessions, NNo Force field = 9 sessions, from 4 mice).

Figure 3.

HON optosilencing impairs motor adaptation. A, Top, Targeting schematic (left) and typical expression of ArchT (right) in HONs. Bottom, Single trial diagram depicting when laser stimulation was applied (magenta bar). B, Whole-cell patch-clamp brain slice recording confirming optosilencing of an HON-ArchT neuron by green laser (representative example of n = 20 cells). C, Average trajectories of baseline, force field, and washout trajectories in control and optosilencing sessions. Blocks of 10 trials were averaged within a session and then averaged across sessions. Means ± SEM of 19 control and 14 optosilencing sessions from 7 mice. D, Average end x-position value versus trial number. Means ± SEM of 19 control (black) and 14 optosilencing (magenta) sessions from 7 mice. Inset, Exponential fits to the data (thick lines) and their confidence intervals (thin lines) demonstrating reduced adaptation rate in control versus optosilenced sessions (extra sum-of-squares F test of whether k fit parameters are different between datasets, F(1,4801) = 14.08, p < 0.001). Smoothed means are shown for visualization only, the fit was performed using individual trial points of unsmoothed data. E, For data shown in D, initial adaptation rates from individual sessions, obtained using least-squares linear fits of the first 20 force field trials of each session. Violin plot of density, dashed line at median and dotted line at the quartiles, two-tailed Mann–Whitney test, U(Ncontrol = 19, Noptosilenced = 14) = 73, *p < 0.05. F, Average end x-position value versus trial number of light control mice. Means ± SEM of 10 control (black) and 13 light on (orange) sessions from 6 mice. Inset, Exponential fits to the data (thick lines) and their confidence intervals (thin lines), demonstrating no difference in adaptation rate because of brain heating (extra sum-of-squares F test of whether k fit parameters are different between datasets, F(1,3378) = 3.330, p > 0.05) G. Same as in E, for light controls. Violin plot of density, dashed line at median and dotted line at the quartiles, two-tailed Mann–Whitney test, U(NLightOff = 10, NLightOn = 13) = 58, p > 0.05.

Figure 4.

HON optosilencing does not impair reward rates or motivational metrics. A, Average percentage of successful trials versus trial number of control and HON optosilencing sessions. Means ± SEM of 19 control and 14 optosilencing sessions from 7 mice. B, For data shown in A, initial adaptation rates from individual sessions, obtained using least-squares linear fits of the first 20 force field trials of each session. Violin plot of density, dashed line at median and dotted line at the quartiles, Mann–Whitney test, U(Ncontrol = 19, Noptosilenced = 14) = 131, ns = p > 0.05. C, As in A for reaction time, calculated as the time between cue onset and pull onset. D, As in B for data in C, Mann–Whitney test, U(Ncontrol = 19, Noptosilenced = 14) = 105, ns = p > 0.05.

Figure 5.

Forelimb adaptation and effects of HON optosilencing follow a sensory prediction error-based model. A, Orange line shows the average end x-position value versus trial number for control force field data from 19 sessions. The shaded orange area depicts the SEM. Black line depicts a sensory prediction error-based model that was fitted to the force field data of each session. The blue line depicts the reward prediction error-based model. The average of 50 realizations of the model with different random initial parameters is visualized. The sensory prediction error-based model shows population-level root-mean-square error of the fit is 1.12 cm, and the reward prediction error-based model shows population-level root-mean-square error of the fit is 1.98 cm. B, Normalized command strength contributions of the reward prediction error-based model (blue) and the sensory prediction error-based model (black) of control sessions. The command strength of the sensory and reward-based models were normalized with respect to the sensory and reward prediction error, respectively. Violin plot of density, dashed line at median and dotted line at the quartiles, two-tailed Mann–Whitney test, U(Ncontrol = 152, NOptosilenced = 156) = 737, ****p < 0.0001. C, As in A for only the sensory prediction error-based model for both control and optosilencing sessions. D, As in B for the normalized contributions of sensory prediction error-based model for both control and optosilencing sessions. Violin plot of density, dashed line at median and dotted line at the quartiles, two-tailed Mann–Whitney test, U(Ncontrol = 152, NOptosilenced = 156) = 6295, ****p < 0.0001.

Figure 6.

HON optosilencing disrupts sensory prediction error-based model through impaired sensorimotor integration. A, Left panel, Average max y-velocity versus trial number. Means ± SEM of n = 19 force field sessions and 14 optosilencing sessions from 7 mice. Right panel, Initial adaptation rates from individual sessions for data shown in left panel, obtained using least-squares linear fits of the first 20 force field trials of each session. Violin plot of density, dashed line at median and dotted line at the quartiles, two-tailed Mann–Whitney test, U(NControl = 19, NOptosilenced = 14) = 129, ns = p > 0.05. B, Average probability of a trial having a counterforce feedback correction versus trial number. Inset describes an example trial with counterforce feedback correction (for calculation, see Materials and Methods). C, Left panel, Same as A, but for steering angle. Inset, Exponential fits to the data (thick lines) and confidence intervals (thin lines), illustrating reduced adaptation rate (extra sum-of-squares F test of whether k fit parameters are different between datasets, F(1,4797) = 13.49, p < 0.001). Smoothed means are shown for visualization only, the fit was performed using individual trial points of unsmoothed data. Right panel, Initial adaptation rates from individual sessions for data shown in A, obtained using least-squares linear fits of the first 20 force field trials of each session. Violin plot of density, dashed line at median and dotted line at the quartiles, two-tailed Mann–Whitney test, U(Ncontrol = 19, Noptosilenced = 14) = 70, *p < 0.05. D, Left panel, Same as in A, left panel, for light control mice, demonstrating no difference in adaptation rate because of tissue heating (extra sum-of-squares F test of whether k fit parameters are different between datasets, F(1,3372) = 0.9682, p > 0.05). Right panel, Same as (A, right panel) for data shown in left panel, two-tailed Mann–Whitney test, U(NLightOff = 10, NLightOff = 13) = 61, ns = p > 0.05. E, Left panel, Average steering angle value versus trial number, without the force field. Means ± SEM of 11 control and 13 optosilencing sessions from 7 mice. Right panel, As in A, right panel, for data in left panel, two-tailed Mann–Whitney test, U(NControl = 11, NOptosilenced = 13) = 64, ns = p > 0.05. F, Left panel, Average end x-position value versus trial number, without the force field. Means ± SEM of 11 control and 13 optosilencing sessions from 7 mice. Right panel, As in A, right panel, for data in left panel, two-tailed Mann–Whitney test, U(NControl = 11, NOptosilenced = 13) = 53, ns = p > 0.05.