PONE-D-23-30728Behavioral sensitization and tolerance induced by repeated treatment with ketamine enantiomers in male Wistar ratsPLOS ONE

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Reviewers' comments:

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Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Partly

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2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

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3. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

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4. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: Yes

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5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: The results of this study showed that repeated treatment with S-ketamine cause locmotor sensitization and tolerance to the ataxic effects, although R-ketamine also has simiar behavior but it is not observed before repeated treatment. The study could be accepted after major revision:

1) racemic ketamine could be included in the study, I think addtional experiemnts of racemic ketamine will strengthen the conclusion of the study and make it more convincing.

2) The authors need to show the evidence that 15 mg/kg is a suitable dosage choice. So this study could add a dose-dependent prelimimnary study comparing 5, 10 and 15 mg/kg.

3) why the authors did not adopt two administration of ketamine per week, instead of every 3 days, this protocol has been accepted for depression treatment by ketamine clinically.

4) I could not find the figure legends for the study.

Reviewer #2: In this study, authors compared abuse potential and psychotomimetic potential of two ketamine enantiomers, S-ketamine and R-ketamine by means of locomotor sensitization and stereotyped behavior. Based on the results, authors concluded that S-ketamine has higher abuse potential and psychotomimetic potential than R-ketamine. Given that there are many reports demonstrating that R-ketamine has more potent antidepressant-like effects than S-ketamine, while that R-ketamine is much safer than S-ketamine, the objective of the study is important and timely. Although there are several studies which compared side effect liabilities of S-ketamine and R-ketamine, this is a nice addition to show differences of these enantiomers. However, there are several concerns on interpretation of the results, and experimental procedures. The followings are comments to be considered.

1. Authors used locomotor activity after acute treatment and potential to induce locomotor sensitization after repeated treatment. Behavioral sensitization is partly regarded as abuse potential, but this phenomenon is not equal to abuse potential. For example, reference #17 measured molecular changes relating abuse potential in the nucleus accumbens which are accompanied with behavioral sensitization to relate this behavioral changes with abuse potential. Please be very careful to clearly mention abuse potential by only the results of behavioral sensitization.

Moreover, it is premature to say that increase in locomotor activity after acute administration is indicative of abuse potential.

Likewise, both ataxia and stereotyped behavior do not necessarily reflect psychotomimetic/dissociative symptoms, because these behavioral abnormalities may be induced by several factors including impaired motor coordination.

2. Please justify 15 mpk SC dose that authors used for this study. Although authors claimed that doses were selected based on the previous reports (and cited references accordingly), intraperitoneal administration was used in the previous reports. In contrast, authors used subcutaneous administration, which is expected to provide higher exposure of both enantiomers, because ketamine is rapidly and extensively metabolized before entering circulation in case of IP administration but not SC administration. Indeed, S-ketamine induced ataxia at 15 mpk SC, a condition which cannot be used to evaluate antidepressant-like effects in FST. Please mention possible differences in exposure between the present study and the previous ones. Moreover, please discuss differences in exposure of ketamine enantiomer between IP administration and SC administration. Without discussion on exposure level, it is difficult to compare doses for abuse potential with ones for antidepressant-like effects.

3. Accordingly, authors’ conclusion that R-ketamine has abuse potential is not appropriate. Zanos et al. suggested abuse potential at higher dose than dose showing antidepressant-like effects. Therefore, authors need to clarify that dose used in the present study may be higher than one showing antidepressant effects.

4. Figure 2. Please show time course of locomotion as well. Therefore, authors may incorporate Figure 4A into Figure 2. Instead, it is not necessary to show both A and B in Figure 1, and individual data in Figure 4 can be transferred to supplementary data. Because ataxia and behavioral abnormalities were observed in S-ketamine dose, it is not meaningful to compare peak time in locomotor increase between R-ketamine and S-ketamine.

5. Regarding description on more robust antidepressant-like effects of R-ketamine than S-ketamine, the followings are representative reports on this finding. Authors should cite these reports.

Zhang et al., Pharmacol Biochem Behav. 2014 Jan;116:137-41.

Yang et al., Transl Psychiatry. 2015 Sep 1;5(9):e632.

6. There are many citations which are not appropriate. For example, abuse potential of ketamine is not described in reference #10, and reference #12 did not mention differential side effects between S-ketamine and R-ketamine. Please carefully confirm contents of all of citations, and correct them accordingly.

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Reviewer #1: No

Reviewer #2: No

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Behavioral sensitization and tolerance induced by repeated treatment with ketamine enantiomers in male Wistar rats

PONE-D-23-30728R1

Dear Dr. Elersič,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

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Kind regards,

Kenji Hashimoto, PhD

Section Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: All comments have been addressed

Reviewer #2: All comments have been addressed

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2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Yes

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3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

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4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

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5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: Yes

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6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: This is a timely study comparing the ketamine enantiomers. The authors have addressed all the comments. No addtional comments. This revised manuscript could be accepted in Plos One.

Reviewer #2: Authors responded adequately to all of my comments, conducted additional experiments as requested, and revised the manuscript accordingly. I appreciate all of authors' efforts to improve the manuscript. I have no further comments.

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7. 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

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