Side Effects of Opioids Are Ameliorated by Regulating TRPV1 Receptors

Round 1

Reviewer 1 Report

In the manuscript “Side effects of opioids are ameliorated by regulating TRPV1 receptors”, Wang et al review evidence in the literature that support functional interactions between the mu opioid receptor and TRPV1 channel and propose that modulation of TRPV1 activity may improve opioid analgesia and reduce side effects.

Although the topic is of interest for a large audience, I have some serious concerns regarding the content. On many occasions, the literature is improperly quoted leading to erroneous or mispresented concepts. In addition, several references are not appropriate or irrelevant. The manuscript should also be carefully edited as several sentences are not understandable.

Here follows some of the main concerns

• Antagonizing TRPV1 activity is a well-established strategy. However, it is associated with several strong side effects such as hyperthermia, which is never mentioned in the manuscript. However, these side effects were at the root of further pharmacological developments to improve the profile of the antagonists with clinical perspectives.
• Concepts such as opioid induced hyperalgesia and opioid tolerance are not clearly presented, and the text is often misleading. Opioid induced hyperalgesia results from opioid intake but hyperalgesia results from a pathological condition such as neuropathy
• Altogether, the content of section 2 is largely incorrect and very confusing. The authors indicate that activation of TRPV1 inhibits mu opioid phosphorylation preventing beta arrestin 2 recruitment and receptor internalization thereby preserving the pool of functional mu opioid receptors at the cell surface. They propose that it could be a therapeutic strategy. However, the data they refer to show decreased mu opioid phosphorylation by GRK5 in transfected HEK293 cells. Although this can take place in selected cell types in vivo, GRK5 is not the only GRK able to phosphorylate the mu opioid receptors (e.g. PMID 33147802) and the physiological relevance of the mechanism remains to be established.
• Also, the authors do not take into account that morphine is poorly promoting GRK phosphorylation of the mu opioid receptor and hence is poorly or not at all internalizing the receptor (PMID 23239825, 18612077). The same applies for herkinorin (PMID17090705) and ARM (PMID 19412545). In addition, GRK phosphorylated receptors at the cell surface get desensitized. It is commonly accepted that dephosphorylation the receptors enables resensitization.
• Finally, the impact on the activation by the endogenous opioid peptides is never envisaged
• The type of ligand envisaged is vague and constraints that apply on the choice or development of the TRPV1 agonist not mentioned.
• The expression of TRPV1 channels is increased in chronic pain conditions but the manuscript mixes data collected on naïve animals treated with morphine together with data in pathological models. It is very likely that the functional interactions between the mu opioid receptor and the TRPV1 channel are quite different in the two situations and therefore clear distinction should be made in the text between data collected in painful and non painful conditions.
• A section presenting and discussing all currently known molecular mechanisms underlying the mu opioid receptor-TRPV1 channel functional interactions should be included instead of providing dispersed elements of information throughout the manuscript
• Neuropathic pain is only one form of chronic pain. TRPV1 channels have been mostly studied in the context of inflammatory pain. The manuscript should clearly state the pain model or condition in which data were collected.
• The concept of biased agonist at the mu opioid receptor is currently highly debated which does not support the therapeutic option proposed by the authors (see for example (PMID 32234959) the text should take this into account
• Line 62-65 The authors mention brain areas with expression of the TRPV1 receptor but the cited references do not provide the information or are unrelated
• Line 39 mu, delta and kappa opioid receptors are receptor types and not subtypes
• the statement in lines 50 is incorrect. B-arrestin does not phosphorylate receptors but binds to GRK phosphorylated receptors
• line 59 TRPV1 also responds to chemical stimuli
• line 67 ref 12 refers to studies in rat
• reference 27 is not relevant
• the content of references 24 and 28 are improperly quoted. In particular, ref 24 focuses on GRK5 impact on mu opioid internalization in transfected cells
• line 139 morphine is by no mean a b-arrestin2 biased ligand
• lines 150-152 the sentence is not understandable
• lines 154-164 the content of this paragraph should be entirely revised
• lines 174-175 appropriate references for TRPV1 distribution are missing
• references 21 and 46 are by the same group and the mechanism they proposed should be discussed.
• line 196. The sentence is partly incorrect as it does not include physical dependence
• ref 49 is irrelevant. Several adenylate cyclases are inhibited downstream of mu opioid receptors
• The content of ref 50 is improperly quoted. It investigated the role of Gs in an operant paradigm and not changes in cAMP levels following repeated opioid stimulation
• Lines 224 the sentence is partly incorrect as it does not include decreased withdrawal
• Line 224 the Nac is part of the striatum. The authors surely mean dorsal striatum
• Lines 229-230 ref 44 refers to systemic injection and ref 54 to VTA. appropriate references are missing
• The content of ref 60 is improperly quoted. The study is on opioid induced hyperalgesia
• Lines 239-247 the content is very vague
• Line244 The authors indicate that there is a lack of functional mu opioid receptors in the spinal cord following sciatic nerve injury. Although debatable (see for example PMID 30876975), it is possibly model dependent and a result from a decrease of mu opioid receptors in primary afferents, which is not at all discussed in the manuscript
• Lines 258-271 upregulation and activity are two different concepts
• Line 272 this not a novel concept
• The content of ref 82 is improperly quoted
• Line 289 neuropathic pain is not more complex than other chronic pain conditions
• Line 310 the content of the sentence is incorrect
• Line 312 the sentence is not understandable
• Line 313 it is unclear whether the authors refer to opioid induced hyperalgesia or neuropathy induced hyperalgesia
• Line 321 the meaning of the sentence is unclear
• Line 327 appropriate references for changes in receptor binding are missing
• Line 332 current first line strategies to treat neuropathic pain rely on antidepressants and anticonvulsants. Strategies targeting TRPV1 is one of several others under development
• Lines 333 this conclusion is partly incorrect
• There are several recent articles extensively characterizing the signaling pathways activated by oliceridine together with the side effects. This part of the manuscript should be modified

 

Author Response

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Author Response File: Author Response.pdf

Reviewer 2 Report

In this review article by Xia-qing Wang et al., the authors studied the
relationships between opioid side effects and the function and
regulation of TRPV1 receptors. The choice of topic is topical and timely,
as otherwise highly effective opiate painkillers have serious and harmful
side effects. The authors' review of the literature provided important
data on the interactions between TRPV1 and opioid receptors and
signaling pathways and their ligands.  The article reviews the side effects of opioid agonists and the possibilities of reducing these side effects in four chapters (Introduction, Tolerance and opioid-induced hyperalgesia, Addiction and Neurophatic pain). The article discusses in detail the interactions, regulation and role of different signaling pathways in the most effective reduction of pain. The authors' conclusions are scientifically sound and sufficiently modest. I suggest the authors justify and define the terms ‘tolerance’, ‘hyperalgesia’ and especially ‘addiction’. The latter includes tolerance, dependence and withdrawal symptoms together. To the best of my belief I recommend the acceptance of this paper for publication.

                             

            

 

Author Response

We thank the Reviewer for the summary of our results and positive comments. We have defined the terms ‘tolerance’, ‘hyperalgesia’, and ‘addiction’ in lines 91-92, 92-94, and 177-179, respectively.

 

Reviewer 3 Report

See attached

Comments for author File: Comments.pdf

Author Response

Please see the attachment.

 

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

The revised version of the manuscript is greatly improved and now reads easily. Here follow a few additional comments and suggestions

• Line 125: in ref 26 the work has been performed in transfected cells. this should be indicated
• Ref 56 refers to expression of TRPV1 in the cortex not the striatum and Ref 57 does not mention the striatum
• Lines 224-226: the reference to the role of the p38/NFkB signaling pathway in memory could be included in the previous paragraph to avoid redundancy.
• lines 225-226: the content is a repetition of lines 216-219 and can be deleted
• Lines 231 and 326: the dorsal striatum is critical for reward-guided and habitual behavior rather than mediating the reward response per se as is the case for the nucleus accumbens. This should be clarified especially in line 326
• Lines 317-323: the content is only a repetition and could be omitted to get straight to the conclusion
• Similarly lines 327-330 could be omitted

Minor

• Line 220 phospho protein kinase A
• Line 255 resistance to opioids
• Line 269 Similarly, Luo et al revealed that…
• Line 273 antinociceptive role
• Line 275 is predominantly mediated by

Author Response

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Author Response File: Author Response.pdf

Reviewer 3 Report

General comments

1. Still needs general English language editing for clarity
2. If TRPV1 interacts with B arrestin which is shared by multiple GPCRs, a discussion of the drawbacks/dangers of this approach is warranted.

 

Specific comments

1. Page 1, line 11 – Missing words makes the sentence unclear. Should read “combining nonopioid analgesics with opioid analgesics to target multiple…”
2. Page 1 lines 33-34 – opioid receptors have different sequences, classification based on ligand selectivity it outdated (especially as many ligands, such as endogenous neurotransmitters) are not very selective.
3. Page 1 Lines 33-34 – Be consistent in either spelling out Greek letters (mu) or using Greek letters (µ). Authors switch back and forth.
4. Page 1 line 42-43 and page 2 line 44 – Authors again claim that B arresting signaling is the main cause of adverse effects of opioids. No citations are given and this is a hotly debated claim as the original studies have not been replicated. This debate should be acknowledged and the language softened.
5. Page 2 line 49 – should read “combining non-opioid analgesics with opioid mediated analgesics to target multiple sites …” English language problem you combine something with something else and both items need to be specified.
6. Page 3 lines 115-116 – clarify language. Do the phosphorylation sites interface with G proteins or does the C-terminus interface with G proteins. Document generally needs English language editing.
7. Page 3 line 28 – clarify that the shuttling of B arrestin to the nucleus hinders its association with ALL GPCRs, not specifically MOR. This is not a MOR specific effect. Again, needs editing to clarify language.
8. Page 3 lines 125-132 – phosphorylation desensitizes MOR and reduces signaling. The lack of internalization (because of lack of B arrestin) may actually be counter-productive as internalization leads to resensitization and trafficking to the surface, INCREASING active MOR at the surface. In other words – trafficking of B arresting to the nucleus by TRPV1 prevents MOR from being internalized, leaving phosphorylated desensitized MOR at the surface. This is contrary to the author’s statement on line 132. Authors should consider this in their discussion or draw back on their claims.
9. Page 3 lines 143-149 – I still find the discussion of DOR agonists distracting and confusing. It does not strengthen the authors’ case that targeting both MOR and TRPV1 could be a viable pain management strategy. Either clarify why this is included or delete.
10. Page 4 lines 171-174 – needs English language editing. This paragraph is confusing.
11. Page 4 line 183 – “regions of the brain” needs English language editing.
12. Page 5 lines 244-247 – needs English language editing, missing verbs, odd word choice.
13. Page 6 line 255-256 – needs English language editing, sentence is confusing.
14. Page 7 lines 313 – 338 – these paragraphs are confusing. I think it is a wordy description of the information in Table 1. If so, this is redundant and can be removed. If not, it needs to be clarified. The numbering is confusing and there is no scaffolding/narrative for this section of the paper.
15. Page 7 lone 341 – the authors claim that compounds which target multiple receptors have lower affinity than high selectivity single target compounds. Why? Is this relevant? Is this always true. Unclear what the authors are attempting to convey.
16. Page 8 line 361-362 – the statement that B arrestin KO mice consistently show decreased morphine tolerance, resp depression and constipation is blatantly untrue. There has been difficulty in reproducing these results and the idea that B arresting biased MOR agonists have reduced side effects is hotly debated. The authors need to consider this throughout the paper and include appropriate references (as mentioned in the first set of reviews and that they incorporated earlier in the manuscript), instead of making cursory edits.

Author Response

Please see the attachment.

Author Response File: Author Response.pdf