Tuning of motor outputs produced by spinal stimulation during voluntary control of torque directions in monkeys

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1. 

   Version published to 10.7554/elife.78346 on eLife

   Dec 13, 2022
2. 

   eLife

   Nov 1, 2022

   Author Response

   Reviewer #2 (Public Review):

   Strengths:

   This is potentially a very large and robust dataset of spinal stimulation while the animal performs a wrist torque task. However, the authors do not detail the number of trials obtained for each combination of conditions - stimulation location, current intensity, movement direction, number of repetitions, etc.

   We have provided an additional table to present the summary of collected data (Table 1 and 2 in Supplementary File 1). Each experiment consisted of 63-1004 successful trials that were evenly distributed to 8 task targets. We described this in the text on line 823-824. However, we indicated the averaged evoked muscle responses or the averaged evoked torques using the stimulus triggered average throughout the manuscript, we believe that it is more important to show the …

   Author Response

   Reviewer #2 (Public Review):

   Strengths:

   This is potentially a very large and robust dataset of spinal stimulation while the animal performs a wrist torque task. However, the authors do not detail the number of trials obtained for each combination of conditions - stimulation location, current intensity, movement direction, number of repetitions, etc.

   We have provided an additional table to present the summary of collected data (Table 1 and 2 in Supplementary File 1). Each experiment consisted of 63-1004 successful trials that were evenly distributed to 8 task targets. We described this in the text on line 823-824. However, we indicated the averaged evoked muscle responses or the averaged evoked torques using the stimulus triggered average throughout the manuscript, we believe that it is more important to show the number of stimuli for averaging. Thus, we have kept the description of the number of stimuli in the typical examples of Figures 2A-C, 5A-C, 7B-D and 8A-C.

   Lines: 823-824 “Each experiment consisted of 63-1004 successful trials (Table 2 in Supplementary File 1).”

   Weaknesses:

   The authors' primary conclusion is that spinal stimulation at moderate current intensities facilitates the effects of descending inputs of the motor command. However, the authors need to expand on:

   i. The effect of these intensities of spinal stimulation on their own; without voluntary movement.

   ii. The robustness of the interactions observed.

   We added the results of stimulus-induced muscle responses (Figure 2A-C, 5A-C and 6A-D) and stimulus-induced torques (Figure 7B-D) during the hold period for the center target (i.e., during awake rest). These data allowed us to quantify the PStEs and the evoked torques without the effect of intended torque production. We could observe clearly the PStEs for Facilitation and the evoked torque. However, it was difficult to observe PStEs for Suppression because it required the substantial voluntary muscle activation to be inhibited. The robust interaction was demonstrated by the modulations of PStEs and the evoked torque from the awake rest to the voluntary torque production. We added further discussion on this point as follows:

   Results Lines 126-131 “The PStEs during the entire period of the task (insets on Figure 2A-C) showed either post-stimulus facilitative (Facilitation, insets on Figure 2A and C) or suppressive effect (Suppression, inset on Figure 2B). Spinal stimulation occasionally produced small magnitude of Facilitation during the hold period for the center target where the voluntary wrist torque production was not intended (center panels on Figure 2A). However, different magnitudes and/or types of PStEs were observed among the directions of voluntary torques (Figure 2A-C).”

   Lines 143-146 “Especially in PStEs of Facilitation, the magnitude of PStEs in the peripheral target close to the PD of background EMG (Figure 2A, 270° and 315°) was generally larger compared with that in the center target and smaller in the peripheral target opposite to the PD (Figure 2A, 90° and 135°).”

   Legend of Figure 2A-C Lines 170-171 “Muscle responses to spinal stimulation during the hold period for the 8-peripheral (peripheral panels) and the center targets (center of peripheral panels).”

   Results Lines 356-357 “Left insets and gray dots in right panels (Figure 5A-C) show the PStEs and background EMGs during hold period for the center target.”

   Legend of Figure 5A-C Lines 368-372 “The leftmost insets show PStEs during the hold period for the center target. The rightmost panels for each muscular condition show two-sided Pearson’s correlation coefficients between the magnitudes of background EMGs and PStEs. Gray dots in right panels indicate the result during the hold period for the center target that were not included for the correlation analyses.”

   Results Lines 394-402 “PStEs during the hold period for the center target increased as current intensity increased, showing a simple input-output property of stimulus-indued muscle responses (“Center target”, insets on Figure 6A-D). In general, including the hold period for the center target, the magnitudes of PStEs at low stimulus currents was linearly increased depending on the magnitudes of background EMGs (Figures 5A-C and 6A). However, the magnitudes of PStEs of Facilitation at medium currents were often larger during hold period for the center target (Figure 6B and C insets) compared to that during voluntary torque production even though the magnitude of background EMG was identical between them (Figure 6B and C, rightmost panels).”

   Legend of Figure 6A-D Lines 419-423 “The leftmost insets show PStEs during hold period for the center target intended to relax the wrist. The rightmost panels indicate two-sided Pearson’s correlation coefficients between the magnitudes of background EMGs and PStEs. Gray dots in right panels indicate the result during hold period for the center target that were not included for the correlation analyses.”

   Results Lines 452-460 “In another case, spinal stimulation at 300 μA mainly induced Facilitation effects on muscles with higher background EMG (outer peripheral panels in Figure 7C and Figure 7-figure supplement 1B), and the directions of the Evoked Torque were similar to the directions of voluntary torque independent of the direction of the Evoked Torque at the center target (center and inner peripheral panels in Figure 7C). Stimulation at 1700 μA exhibited large magnitudes of Facilitation in all muscles for all targets (outer peripheral panels in Figure 7D and Figure 7-figure supplement 1), and the Evoked Torques displayed ulnar-flexion directions regardless of the presence/absence or the direction of voluntary torque (center and inner peripheral panels in Figure 7D).”

   Legend of Figure 7B-D Lines 487-489 “StTAs of rectified EMGs (outer peripheral panels and center-bottom panel) and StTAs of wrist torque trajectories (inner peripheral panels and center-top panel).”

   Discussion Lines 669-680 “Compared with the hold period for the center target, the stimulus-induced muscle responses and torques at low to medium currents were generally more pronounced during the hold period for the peripheral targets (Figure 2A-C, Figure 7B and C, and Figure 7-figure supplement 1), indicating that the descending commands augmented activation in the spinal motoneurons and interneurons driven by spinal stimulation. Interestingly, at medium currents, the stimulus-induced facilitatory responses were sometimes smaller when the responses were recorded in the antagonistic muscles against the wrist torque direction regardless of the background EMG activity (Figure 2A and Figure 7-figure supplement 1B), suggesting that spinal reciprocal inhibitory function was evolved by the descending commands (Meunier and Pierrot-Deseilligny, 1998). Together, our findings indicate that voluntary commands amplify the functions of spinal circuits, including excitatory and inhibitory synaptic connections to motoneurons activated by spinal stimulation.”

   Specific comments:

   1. Interpretation of the main result - The authors state that they investigated the "effect of descending inputs on the stim-evoked EMG and torque output". But, their experimental design which compares post-stim EMG to pre-stim EMG provides a somewhat different result, i.e., the effect of spinal stimulation on voluntarily-evoked EMG and torque output. In other words, the voluntary output is held constant (independent variable) and the spinal stimulation parameters are varied (dependent variable).

   To get what the authors state, the design would have to be modified wherein the comparison would have to be between post-stim muscle activity recorded in the wrist neutral vs one of the holding state; Or comparison of post-stim muscle activity when the arm is passively torqued vs when voluntarily torqued.

   In our study, we compared pre-stim EMG and post-stim EMG in order to determine the presence/absence and the polarity (facilitation/inhibition) of PStEs. Our main aim in this study was to investigate the effect of descending commands (voluntary output) on the stimulus-evoked responses, and we concluded that the descending commands influence the spinal interneuron activities elicited by spinal stimulation. The motor task requires the control of the direction and magnitude of wrist torque attained in order to manipulate the magnitude of descending commands that were expressed as the background EMG activity at each muscle. Then, the result showing that PStEs were modulated by the variation of background EMG certainly indicates that the descending commands influence PStEs.

   In the revised version of the manuscript, we present additional data of PStEs and evoked torque while the wrist remained in the neural position (i.e., during awake rest) to address your comment.

   1. Most of the studies that have demonstrated the benefits of spinal stimulation, esp. in humans, have used sub-threshold stimulation. The manuscript does not provide direct information regarding the threshold of stimulation. Only table 2 provides such information but the data collection paradigm is so different from the actual task that it makes it difficult to make a relevant connection.

   • Why was the stimulation protocol under sedation different from during the wrist torque task? It would be really useful to describe the kind of involuntary movements evoked at different current intensities at the different spinal levels in awake, behaving animals. For instance, the higher amplitudes appear to just lock the arm into a full ulnar deviation. Such current intensities would be unlikely to be effective in enhancing movement in spinal cord injury. Thus, all the results for these amplitudes are somewhat irrelevant to therapeutic intervention. Similarly, does the moderate amplitude generate movement or muscle contraction?

   The stimulus evoked muscle responses changed their size depending on many variables, such as stimulus intensity, torque direction (i. e, voluntary muscle pre-activation in combination with other muscles activities), and the recording muscle. The stimulus threshold for each facilitatory and inhibitory effect is changed depending on these variables. Therefore, we did not aim to measure stimulus threshold independently. However, it was essential to map spinal somatotopic representation in relation with the site of the stimulus electrode for the experiment in Figure 4. Therefore, we delivered spinal stimulation with each electrode channel under anesthesia in order to capture muscle representation without concomitant voluntary descending drives in the intact monkey.

   As the reviewer indicated the importance, we agree to obtain the information of stimulus-evoked torques at each stimulus intensity while the wrist torque was neutral in the awake monkeys. In addition, we presented data of stimulus-evoked muscle responses and torques at each low, moderate, and high stimulus intensity while the monkeys’ cursor was maintained on the center target in Figures 2 and 7 (see the responses to previous comments).

   1. Please explain the term Spinal PD.

   Does the PD of the background EMG remain the same irrespective of the current intensity and site of stimulation? There is a decrease in background EMG amplitude in Fig. 2A and B with increasing stim amplitude. Can the authors please discuss this observation and how it would affect the efficacy of the spinal stimulation in facilitating descending inputs?

   Spinal PD is the preferred direction (PD) of facilitative evoked muscle responses (Facilitation) or suppressive evoked muscle responses (Suppression) that was calculated separately by the data obtained during the hold period for the peripheral targets. We added this explanation in the text (lines 146-149) and the legend of Figure 2D (lines 183-185).

   The amplitude of the background EMG changed with increasing current intensities, as the reviewer pointed out. Hence, it might be possible that the large ulnar-flexor torques due to the high stimulus currents had somewhat direction-biased effects on the required voluntary effort (i.e., for ulnar-flexor targets, less voluntary commands for ulnar-flexor muscles might be required under the support of stimulus evoked torque whereas for radial-extensor targets, more voluntary commands for radial-extensor muscles might be required under the opposed stimulus evoked torque). Nevertheless, we confirmed that the PD of the background EMG was consistent irrespective of the current intensity and stimulus site as presented in Figures 3A, 3B, 4B, and 4C (green polar plots). In addition, we showed that Spinal PD at high current was even opposite to the PD of background EMG, indicating that the magnitude of background EMG hardly explains the differences in the results between low to medium and high stimulus currents.

   Results Lines 146-149 “Significant PDs were observed in the 603 muscular conditions in 16 muscles for Facilitation (Spinal PD of Facilitation), 333 muscular conditions in 16 muscles for Suppression (Spinal PD of Suppression), and 1006 muscular conditions in 16 muscles for background EMG.”

   Legend of figure 2D Lines 183-185 “ Spinal PD (top panels) and Background EMG PD (middle panels) show the PDs calculated by the magnitudes of Facilitation or Suppression of PStEs and by the magnitudes of background EMG activity, respectively, during the hold period for the peripheral targets.”

   1. Line 546 - The authors speculate that higher current intensities resulted in direct activation of motoneurons. While this is certainly possible, It seems somewhat do the authors see proof of this in their data? Latency measurements?

   We newly analyzed the results for onset latency of PStEs as Figure 8, and added the relating descriptions in the Results, Discussion, and Materials and Methods of the revised manuscript. Please refer the responses to the 2nd comment from Reviewer 1. The results showing the latency shortening at the high currents support our statement that higher current intensities result in direct activation of ventral root axons.

   1. Line 589 - "However, in the rostrally-innervated muscles, the PDs for facilitation effects from caudal sites were opposite to those for background EMGs (Figure 4G, bottom-left panel), suggesting the direct activation of motor nerves." Can the authors clarify how they infer direct activation of motoneurons from the discrepancy between spinal PD and background EMG PD?

   We revised the Discussion as follows:

   Lines 702-710 “However, an exception was observed in some cases of rostrally-innervated muscles that showed facilitation effects. The Spinal PDs for facilitation in the rostrally-innervated muscles from caudal sites were opposite to those for background EMGs (Figure 4G, bottom-left panel). The magnitude of these responses was quite small (Figure 4E, left panel), but this feature of responses was similar to the response at higher current (Figure 3F, lower panel). These results suggest that some motoneurons of rostrally-innervated muscles may not receive excitatory ascending inputs from afferents of the caudal part of the spinal site. Although there is a considerable distance between them, current targeting to the caudal site might spread to ventral roots of rostrally-innervated muscles.”

   • I wonder why the authors did not look at the effect of spinal stimulation-evoked EMG and torque during the movement of the cursor? This could be used to determine the parameters that improve the performance of the task, by either increasing the speed or decreasing the effort required to perform the task.

   As the aim of this study was to reveal fundamental characteristics of descending commands on stimulus effects, we systematically and quantitatively explored evoked motor outputs, but did not directly investigate how the spinal stimulation improves the motor task to suggest a therapeutic interventional approach.

   For the analyses shown in Figure 7, we have shown the data of evoked torques, instead of the movement of the cursor, and concluded that the magnitudes and directions of evoked torque change depending on the current intensity and direction of voluntary torque production.

   • I wonder if the current dataset allows the generation of a map that shows the lower and upper limits of current intensity that result in facilitation of descending inputs for each muscle, at each stimulation location. Additionally, is this map stable across days/sessions.

   In the present study, we showed that descending commands amplified the functions of intraspinal neural elements regardless of stimulus sites (Figures 4G and H). In addition, we revealed that the current of 150-1350 μA boosted torque production in a direction corresponding with the direction of voluntary torque production (Figure 7C and F).

   Since it took many days to get these data with various stimulus conditions (stimulus current and site), we could not compare motor outputs to spinal stimulation in the same stimulus condition across days/sessions. Future studies will be needed to investigate the stability of motor outputs. We add this issue in discussion as follows:

   Lines: 750-752 “However, the effectiveness of subdural stimulation in controlling dexterous hand movements and the long-term stability of motor output need to be determined in future studies.”

   Reviewer #3 (Public Review):

   1. To characterize the effects of stimulation, stimulation was first delivered during an anesthetized experiment to map the evoked responses from each electrode. A major result of the paper is that the level of background activity affects the response to stimulation. It would be interesting to see these baseline responses to stimulation in awake monkeys while they were sitting quietly and not attempting a task to see if these align well with the anesthetized responses.

   As we had similar comments from Reviewer 2, we presented additional data of the evoked muscle responses and evoked torques during the hold period for the center target where the wrist torque production was not intended in awake monkeys (Figure 2A-C, Figure 5A-C, Figure 6A-D and Figure 7B-D). These data support our results that descending commands amplify the function of intraspinal elements. Please refer the responses to the 2nd comment from Reviewer 2 for the revisions to the text.

   On the other hand, the currents and frequencies of subdural spinal stimulation used in the anesthetized monkeys were different from those in awake monkeys. Thus, we could not compare the evoked motor outputs between anesthetized and awake conditions in present study.

   1. To understand the coordinated effects of stimulation across muscles, the authors present wrist torque data in Figure 7. These data are certainly important from a functional perspective and provide some information about coordination, but additional detail about coordination across muscles would be helpful throughout the paper. Currently, most of the results are presented on a per-muscle basis but don't describe whether there were (un)coordinated responses across muscles. For example, was there co-contraction of agonists or antagonists during stimulation? Increased activity of multiple antagonists could potentially lead to increased joint stiffness or fatigue without resulting in an increase in joint torque at the wrist.

   As you suggested, the inter-muscular relationship is another aspect of important information to comprehend the coordination of forearm muscles. Based on our data, the monkeys properly engaged each muscle as agonist with following anatomical constraint. We found antagonistic voluntary contraction to be quite rare or mostly non-dominant even during high intensity electrical stimulation, suggesting that the stimulus evoked responses of each muscle were independent of the voluntary activation (i.e., background EMG) of antagonistic muscles. We added these results in Figure 7-figure supplement 1 and the relating descriptions in the text as follows:

   Results Lines 460-467 “During the 8-directional torque task, the monkeys properly engaged each muscle as agonist (Figure 7-figure supplement 1). We found the antagonistic voluntary contraction were quite rare or mostly non-dominant even during high intensity electrical stimulation. There was a tendency that the magnitude of PStEs was stronger in agonists and weaker in antagonists at low and medium currents (Figure 7-figure supplement 1A and B). On the other hand, stimulation at high currents tended to induce large magnitudes of facilitation effects for all targets irrespective of agonist and antagonists (Figure 7-figure supplement 1C).”

   Legend of Figure 7-figure supplement 1 Lines 1179-1189 “Figure 7-figure supplement 1. Subdural spinal stimulation simultaneously evoked facilitative and suppressive effects in multiple muscles and activated synergistic muscle groups. (A-C) StTAs of rectified EMGs in five wrist muscles during the hold period for the center and the 4 peripheral targets. Each polar plot was normalized by the maximum value of each muscle. Each example in (A-C) corresponds to the cases of Figure 7B-D, respectively. At low and medium currents of stimulations, large magnitudes of PStEs were observed in the muscles with high background EMG. For instance, stimulation given at the flexion directed target in (B) strongly facilitated wrist flexor muscles (e.g., FCR, PL and FCU), while stimuli at the extension directed target strongly facilitated wrist extensor muscles (e.g., ECR and ECU). On the other hand, at high current of stimulation, the magnitudes of PStEs hardly changed regardless of the magnitudes of background EMGs and the directions of voluntary torque.”

   Discussion Lines 644-650 “The inter-muscular relationship characterized by the PDs of background EMGs in the wrist muscles (Figure 7-figure supplement 1) demonstrate that the monkeys consistently engaged each muscle as agonist, and that antagonistic voluntary contractions were rare irrespective of stimulus currents (see polar plots of background EMGs of Figure 7-figure supplement 1A-C). This result indicates that the presumed different activation in the spinal excitatory and inhibitory interneurons at different current intensity is not supported by the change of wrist torque production strategy.”

   1. Authors infer from the consistent ulnar wrist torque during high amplitude stimulation that these responses are likely to direct activation of the ventral motor pathway rather than activation through the dorsal sensory pathway and spinal circuitry. Is there any evidence in the EMG data (e.g. decreased latency, more consistent pulse-to-pulse amplitude of evoked EMG responses) to further support this finding?

   We added the results of the onset latency of PStEs as Figure 8, and the related description in the Results, Discussion, and Materials and Methods. The results showing the decreased latency at high stimulus current supports our argument that stimulus-evoked muscle response at the high currents resulted from the direct activation of ventral motor pathways. Please refer the response to Reviewer 1 for the revisions to the text.

   Read the original source
3. 

   eLife

   Jun 23, 2022

   Evaluation Summary:

   In this manuscript, the authors specifically look at the interaction between epidural stimulation of the spinal cord and the descending input evoked voluntarily in 2 intact monkeys. The results show that spinal stimulation could facilitate or suppress voluntarily evoked EMG and wrist torque, depending on voluntarily evoked activity as well as the stimulation parameters. This shows that spinal stimulation could enhance the descending inputs in cases of partial lesions. The conclusions of this paper are well supported by data, although they could be made stronger with additional analysis and clarification.

   (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #2 agreed to share their …

   Evaluation Summary:

   In this manuscript, the authors specifically look at the interaction between epidural stimulation of the spinal cord and the descending input evoked voluntarily in 2 intact monkeys. The results show that spinal stimulation could facilitate or suppress voluntarily evoked EMG and wrist torque, depending on voluntarily evoked activity as well as the stimulation parameters. This shows that spinal stimulation could enhance the descending inputs in cases of partial lesions. The conclusions of this paper are well supported by data, although they could be made stronger with additional analysis and clarification.

   (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #2 agreed to share their name with the authors.)

   Read the original source
4. 

   eLife

   Jun 23, 2022

   Reviewer #1 (Public Review):

   The authors explore the effects of spinal stimulation using subdural arrays in non-human primates (NHPs). The experiments are conducted in intact able-bodied NHPs first under anesthesia, and then awake and executing a voluntary 8 target center-out isometric motor task at the wrist. The data show that the stimulation effects in awake NHPs have directional tuning matching the task (enhance the task) and comprise both excitatory and inhibitory effects on muscles at appropriate intensities and sites. The data indicate that both types of pathways are recruited and contribute to the stimulation effects.

   The experiments focus on 2 NHPs. Stimulation effects are tested under sedation and then in the awake NHP during the voluntary task and the results are compared. The data are carefully explored using bootstrapping …

   Reviewer #1 (Public Review):

   The authors explore the effects of spinal stimulation using subdural arrays in non-human primates (NHPs). The experiments are conducted in intact able-bodied NHPs first under anesthesia, and then awake and executing a voluntary 8 target center-out isometric motor task at the wrist. The data show that the stimulation effects in awake NHPs have directional tuning matching the task (enhance the task) and comprise both excitatory and inhibitory effects on muscles at appropriate intensities and sites. The data indicate that both types of pathways are recruited and contribute to the stimulation effects.

   The experiments focus on 2 NHPs. Stimulation effects are tested under sedation and then in the awake NHP during the voluntary task and the results are compared. The data are carefully explored using bootstrapping and circular statistics, and various analyses of variance.

   The conclusion that stimulation at appropriate levels can induce both excitatory and inhibitory effects to enhance voluntary motor responses is strong. The lowest currents could show suppressive effects on voluntary activity. The balance of these opposing effects is modulated by current intensity, with inhibitory effects swamped by excitation at higher stimulation intensity. The two are balanced in ways that facilitate voluntary activity over a range of intensity from 175uA to 1300uA. The relative strengths of the two effects are thus variable, with excitation often dominant, possibly for much of what might be clinically chosen intensities for many purposes.

   The relative roles of the inhibitory and excitatory effects in therapeutic regimes remain to be determined. Data provided are consistent with the gating of voluntary controls with both excitatory and inhibitory effects. In injured CNS the required levels of background controls by descending systems may be absent or oddly biased. However, the data support possible enhanced inhibitory control as well as excitatory in at least some conditions. The role of intensity in regulating the balance of these may also matter in the future in the design of therapies.

   Read the original source
5. 

   eLife

   Jun 23, 2022

   Reviewer #2 (Public Review):

   This manuscript will be of great interest to neuroscientists and biomedical engineers in the field of neuromodulation. The results shed light on the possible interactions between epidural spinal cord stimulation and the descending inputs of the motor command. They have systematically explored the parameter space that could prove useful for boosting the descending inputs which could help restore movement after spinal cord injury. Most of the conclusions of this paper are well supported by data. However, the manuscript needs to expand on the justification of specific experimental choices made.

   Strengths:

   This is potentially a very large and robust dataset of spinal stimulation while the animal performs a wrist torque task. However, the authors do not detail the number of trials obtained for each combination of …

   Reviewer #2 (Public Review):

   This manuscript will be of great interest to neuroscientists and biomedical engineers in the field of neuromodulation. The results shed light on the possible interactions between epidural spinal cord stimulation and the descending inputs of the motor command. They have systematically explored the parameter space that could prove useful for boosting the descending inputs which could help restore movement after spinal cord injury. Most of the conclusions of this paper are well supported by data. However, the manuscript needs to expand on the justification of specific experimental choices made.

   Strengths:

   This is potentially a very large and robust dataset of spinal stimulation while the animal performs a wrist torque task. However, the authors do not detail the number of trials obtained for each combination of conditions - stimulation location, current intensity, movement direction, number of repetitions, etc.

   Weaknesses:

   The authors' primary conclusion is that spinal stimulation at moderate current intensities facilitates the effects of descending inputs of the motor command. However, the authors need to expand on:
   i. The effect of these intensities of spinal stimulation on their own; without voluntary movement.
   ii. The robustness of the interactions observed.

   Specific comments:

   1. Interpretation of the main result - The authors state that they investigated the "effect of descending inputs on the stim-evoked EMG and torque output". But, their experimental design which compares post-stim EMG to pre-stim EMG provides a somewhat different result, i.e., the effect of spinal stimulation on voluntarily-evoked EMG and torque output. In other words, the voluntary output is held constant (independent variable) and the spinal stimulation parameters are varied (dependent variable).
   To get what the authors state, the design would have to be modified wherein the comparison would have to be between post-stim muscle activity recorded in the wrist neutral vs one of the holding state; Or comparison of post-stim muscle activity when the arm is passively torqued vs when voluntarily torqued.

   2. Most of the studies that have demonstrated the benefits of spinal stimulation, esp. in humans, have used sub-threshold stimulation. The manuscript does not provide direct information regarding the threshold of stimulation. Only table 2 provides such information but the data collection paradigm is so different from the actual task that it makes it difficult to make a relevant connection.
   - Why was the stimulation protocol under sedation different from during the wrist torque task?
   It would be really useful to describe the kind of involuntary movements evoked at different current intensities at the different spinal levels in awake, behaving animals. For instance, the higher amplitudes appear to just lock the arm into a full ulnar deviation. Such current intensities would be unlikely to be effective in enhancing movement in spinal cord injury. Thus, all the results for these amplitudes are somewhat irrelevant to therapeutic intervention.
   Similarly, does the moderate amplitude generate movement or muscle contraction?

   3. Please explain the term Spinal PD.
   Does the PD of the background EMG remain the same irrespective of the current intensity and site of stimulation? There is a decrease in background EMG amplitude in Fig. 2A and B with increasing stim amplitude. Can the authors please discuss this observation and how it would affect the efficacy of the spinal stimulation in facilitating descending inputs?

   4. Line 546 - The authors speculate that higher current intensities resulted in direct activation of motoneurons. While this is certainly possible, It seems somewhat do the authors see proof of this in their data? Latency measurements?

   5. Line 589 - "However, in the rostrally-innervated muscles, the PDs for facilitation effects from caudal sites were opposite to those for background EMGs (Figure 4G, bottom-left panel), suggesting the direct activation of motor nerves." Can the authors clarify how they infer direct activation of motoneurons from the discrepancy between spinal PD and background EMG PD?

   • I wonder why the authors did not look at the effect of spinal stimulation-evoked EMG and torque during the movement of the cursor? This could be used to determine the parameters that improve the performance of the task, by either increasing the speed or decreasing the effort required to perform the task.
   • I wonder if the current dataset allows the generation of a map that shows the lower and upper limits of current intensity that result in facilitation of descending inputs for each muscle, at each stimulation location. Additionally, is this map stable across days/sessions.

   Read the original source
6. 

   eLife

   Jun 23, 2022

   Reviewer #3 (Public Review):

   This manuscript describes results from a set of experiments to explore the effects of cervical spinal cord stimulation on motor control of the arm. The long-term clinical goal is to use spinal cord stimulation to improve motor function after neural injuries by facilitating volitional control of the limb. Towards that goal, they performed a set of experiments in two awake, behaving monkeys in which stimulation was delivered to the spinal cord via a set of electrodes implanted subdurally, and the monkeys were trained to perform a target tracking task using wrist flexion. Stimulation was delivered during the task and wrist torque and arm muscle EMG were recorded to quantify the effects of stimulation. The authors found that cervical stimulation can produce either facilitation or suppression of volitional muscle …

   Reviewer #3 (Public Review):

   This manuscript describes results from a set of experiments to explore the effects of cervical spinal cord stimulation on motor control of the arm. The long-term clinical goal is to use spinal cord stimulation to improve motor function after neural injuries by facilitating volitional control of the limb. Towards that goal, they performed a set of experiments in two awake, behaving monkeys in which stimulation was delivered to the spinal cord via a set of electrodes implanted subdurally, and the monkeys were trained to perform a target tracking task using wrist flexion. Stimulation was delivered during the task and wrist torque and arm muscle EMG were recorded to quantify the effects of stimulation. The authors found that cervical stimulation can produce either facilitation or suppression of volitional muscle activity and that the direction of that facilitation or suppression was typically aligned with the direction of volitional movement. Further, they report that the amplitude of stimulation and the amplitude of background muscle activity affected the degree of facilitation or suppression observed in the muscle activity, and that high amplitude stimulation was likely causing direct activation of the ventral motor pathway, rather than indirect muscle activation via the dorsal sensory pathway. These results suggest that tuning of stimulation amplitude will be important for achieving facilitatory responses in a motor neuroprosthesis.

   The conclusions of this paper are well supported by data, although they could be made stronger with additional analysis and clarification.

   1. To characterize the effects of stimulation, stimulation was first delivered during an anesthetized experiment to map the evoked responses from each electrode. A major result of the paper is that the level of background activity affects the response to stimulation. It would be interesting to see these baseline responses to stimulation in awake monkeys while they were sitting quietly and not attempting a task to see if these align well with the anesthetized responses.

   2. To understand the coordinated effects of stimulation across muscles, the authors present wrist torque data in Figure 7. These data are certainly important from a functional perspective and provide some information about coordination, but additional detail about coordination across muscles would be helpful throughout the paper. Currently, most of the results are presented on a per-muscle basis but don't describe whether there were (un)coordinated responses across muscles. For example, was there co-contraction of agonists or antagonists during stimulation? Increased activity of multiple antagonists could potentially lead to increased joint stiffness or fatigue without resulting in an increase in joint torque at the wrist.

   3. Authors infer from the consistent ulnar wrist torque during high amplitude stimulation that these responses are likely to direct activation of the ventral motor pathway rather than activation through the dorsal sensory pathway and spinal circuitry. Is there any evidence in the EMG data (e.g. decreased latency, more consistent pulse-to-pulse amplitude of evoked EMG responses) to further support this finding?

   Read the original source
7. 

   Version published to 10.1101/2022.03.16.484577v1 on bioRxiv

   Mar 18, 2022