Dear Dr Grill,

Thank you very much for submitting your manuscript, 'Kilohertz waveforms optimized to produce closed-state Na+ channel inactivation eliminate onset response in nerve conduction block', to PLOS Computational Biology. As with all papers submitted to the journal, yours was fully evaluated by the PLOS Computational Biology editorial team, and in this case, by independent peer reviewers. The reviewers appreciated the attention to an important topic but identified some aspects of the manuscript that should be improved.  Indeed, you will see that in some cases the reviewers ask for clarification or more details, and they have also raised a variety of open-ended questions, which reflect their interest in the work.  We hope that you will find these useful and thought-provoking and look forward to reading your responses.

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Kim Blackwell

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Reviewer's Responses to Questions

Comments to the Authors:

Please note here if the review is uploaded as an attachment.

Reviewer #1: An interesting, drug-free method for blocking nerve conduction relies on inactivation of voltage-gated sodium (Nav) channels via externally applied, high-frequency electrical stimuli. This technique can potentially be used to treat pain and spasticity. However, one caveat is that the nerve responds to the onset of the stimulus, which can cause transient muscle contractions and pain.

Yi and Grill propose here a procedure for optimizing the stimulus waveform in such a way that the onset response is eliminated. To generate action potentials, Nav channels first activate, open and generate current, then inactivate and remain inactivated for a period of time that makes the nerve refractory to stimuli. However, Nav channels can also inactivate without opening, a process termed closed state inactivation (CSI). The kHz waveform designed by Yi and Grill exploits CSI.

The problem addressed is important and interesting, the modeling work is good, and the manuscript is clear and well written. Overall, I really enjoyed reading the manuscript and I didn't see any major issue. I only have a number of minor comments and suggestions.

Fig. 2: I think it might be good to show what happens when a standard voltage step is applied, for readers who are less familiar with Nav channels, and also to put the magnitude of the residual INa in context. Otherwise, the current graph in A is uninterpretable.

The states could be directly called C11, I11, etc., instead of SC11, SI11, etc.

Do I understand correctly that the voltage was clamped only at one node (10), and the rest of the axon was completely free? If so, I am still a little surprised that no APs were generated (escaped) farther away from node 10. I think it's really important to show a waveform that is not fully optimized and permits APs to escape, side by side with the optimized waveform, so readers understand what has been achieved here. The authors might want to cite a paper where this phenomenon of escaped APs is actually exploited to selectively silence axonal Nav channels (Milescu at al, 2010).

The optimal waveform was obtained here in the absence of an external stimulus, but would it be different if excitation were present?

The authors might want to emphasize/explain that the goal was not to block conduction in the entire axon by CSI, but only at one node (or a few).

The study convinced me that, in general, a waveform can be designed to produce conduction block without onset response. The waveform discussed here works with a computational compartmental model that describes a certain type of axon, and the authors admit that it may not work with other types. Is there a strategy that would allow a clinical device to generate an optimal waveform for a specific patient? That is, without triggerring an onset response during the optimization process, or at least reduce that. For example, wouldn't it be safe to just ramp the voltage up so very slowly, in hundreds of ms, say? Could you show a simulation with a much shallower ramp? If the optimization was started with a square waveform, I could see how the optimizer would stop when it was just shallow enough. Of course, I could be entirely wrong, and this waveform could really be the only one that works. Please explain and illustrate.

Could you give estimates on how long would optimization take on a desktop computer?

Reviewer #2: review is uploaded

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Have all data underlying the figures and results presented in the manuscript been provided?

Large-scale datasets should be made available via a public repository as described in the PLOS Computational Biology data availability policy, and numerical data that underlies graphs or summary statistics should be provided in spreadsheet form as supporting information.

Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: No

Reviewer #2: No

Attachments

Attachment

Submitted filename: Comments V2.pdf

Dear Dr Grill,

We are pleased to inform you that your manuscript 'Kilohertz waveforms optimized to produce closed-state Na+ channel inactivation eliminate onset response in nerve conduction block' has been provisionally accepted for publication in PLOS Computational Biology.

Before your manuscript can be formally accepted you will need to complete some formatting changes, which you will receive in a follow up email. A member of our team will be in touch with a set of requests.

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Thank you again for supporting Open Access publishing; we are looking forward to publishing your work in PLOS Computational Biology. 

Best regards,

Jonathan Rubin

Associate Editor

PLOS Computational Biology

Kim Blackwell

Deputy Editor

PLOS Computational Biology

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Reviewer's Responses to Questions

Comments to the Authors:

Please note here if the review is uploaded as an attachment.

Reviewer #1: My comments and suggestions have been fully addressed.

Reviewer #2: Great job addressing the comments!

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Have all data underlying the figures and results presented in the manuscript been provided?

Large-scale datasets should be made available via a public repository as described in the PLOS Computational Biology data availability policy, and numerical data that underlies graphs or summary statistics should be provided in spreadsheet form as supporting information.

Reviewer #1: Yes

Reviewer #2: None

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PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: No