Naltrexone, NK cells and the calcium ion channel research

[Marylib]

What do you guys think about this? Seems the full study will be published soon.
@Gingergrrl @necessary8

I guess it links up the biomarker they are using with NK cells and the use of naltrexone as an immune-modulator.


https://www.frontiersin.org/article..._M2XjJ_oAJ0qYl1K9mRVltIpw6SENdhEMrvXFQ3-8wFEg

 

[pattismith]

Naltrexone restores impaired transient receptor potential melastatin 3 ion channel function in natural killer cells from myalgic encephalomyelitis/chronic fatigue syndrome patients

Helene Cabanas1, 2, 3, 4*, Katsuhiko Muraki4, 5, Donald Staines1, 2, 3, 4 and Sonya Marshall-Gradisnik1, 2, 3, 4

• 1School of Medical Sciences, Griffith Health, Griffith University, Australia
• 2National Centre for Neuroimmunology and Emerging Diseases, Menzies Health Institute, Griffith University, Australia
• 3Menzies Health Institute, Griffith University, Australia
• 4National Centre for Neuroimmunology and Emerging Diseases, Griffith University, Independent researcher, Australia
• 5School of Pharmacy, Aichi Gakuin University, Japan

Myalgic Encephalomyelitis/ Chronic Fatigue Syndrome (ME/CFS) is a seriously long-term and debilitating illness of unknown cause hallmarked by chronic pain and fatigue, memory and concentration impairment, and inflammation.

ME/CFS hypothesis involves ion channel disorders and more specifically impaired Transient receptor potential melastatin 3 (TRPM3) ion channel function, affecting calcium signalling and Natural killer (NK) cell functions.

Currently, substances called opioids, agonists of mu (µ)-opioid receptors (µOR), are the strongest painkillers clinically available for people suffering from strong or long-lasting pain characteristic of CFS/ME.

µOR have been reported to specifically inhibit TRPM3 and to be expressed in immune cells where they play an immunomodulatory and immunosuppressive role.

Naltrexone hydrochloride (NTX) acts as an antagonist to the μOR thus negating the inhibitory function of this opioid receptor on TRPM3.

Therefore, understanding the mechanism of action for NTX in regulating and modulating TRPM3 channel function in NK cells will provide important information for the development of effective therapeutic interventions for ME/CFS.


Whole-cell patch-clamp technique was used to measure TRPM3 activity in Interleukin-2 (IL-2) stimulated and NTX-treated NK cells for 24 hours on sixteen age- and sex-matched healthy controls and CFS/ME patients, after modulation with pregnenolone sulfate (PregS), NTX and ononetin.

We confirmed impaired TRPM3 function in ME/CFS patients through electrophysiological investigations in Interleukin-2 (IL-2) stimulated NK cells after modulation with pregnenolone sulfate and ononetin.

Importantly, TRPM3 channel activity was restored in IL-2 stimulated NK cells isolated from ME/CFS patients after incubation for 24 hours with NTX.
Moreover, we demonstrated that NTX does not act as an agonist by directly coupling on the TRPM3 ion channel gating.

The opioid antagonist NTX has the potential to negate the inhibitory function of opioid receptors on TRPM3 in NK cells from CFS/ME patients, resulting in calcium signals remodelling, which will in turn affect cell functions, supporting the hypothesis that NTX may have potential for use as a treatment for CFS/ME.

Our results demonstrate, for the first time, and based on novel patch clamp electrophysiology, potential pharmaco-therapeutic interventions in ME/CFS.

https://www.frontiersin.org/article...nzBRZTFJM7JBfvwspMuGxJyovwRNFukAwvlXlAF1mWmEc

 

[Marylib]

@pattismith Maybe this is nothing new and at the same time something new ? I'll just hang out until all you geniuses explain it all to me.

 

[pattismith]

@pattismith Maybe this is nothing new and at the same time something new ? I'll just hang out until all you geniuses explain it all to me.

No, I did some search and couldn't find it. It's a new and promising paper, thank you for posting!

"Received: 07 Aug 2019; Accepted: 14 Oct 2019. "

 

[Marylib]

My pleasure.

But "nothing new" I meant that fact that people have been using low dose naltrexone for awhile. By "something new," I meant that this is linking things together in terms of the calcium ion channel biomarker stuff. I think my comment was a bit confusing because I am more than a bit confused!

Yeah, it is definitely new.

Too bad low dose naltrexone has never been one of those things that is consistently helpful for me. But yes, they are going full steam ahead at that research group. And a biomarker for early diagnosis is so important. Maybe we will finally be rid of the term 'chronic fatigue syndrome.' That would be nice.

 

[Sad Dad]

Currently, substances called opioids, agonists of mu (µ)-opioid receptors (µOR), are the strongest painkillers clinically available for people suffering from strong or long-lasting pain characteristic of CFS/ME.

The reasearch may be good, but I'm a bit leery when they mischaracterize the symptoms of ME/CFS.

 

[Wishful]

The reasearch may be good, but I'm a bit leery when they mischaracterize the symptoms of ME/CFS.

Yah, "hallmarked by chronic pain and fatigue" is incorrect, at least for me. I did suffer from chronic pain for a period of time (few years?), and LDN did block it, but at some point the muscle aches stopped and I no longer qualify as suffering chronic pain. I'd say that pain, chronic or otherwise, is just one potential symptom associated with ME. I wouldn't describe my lethargy as fatigue either. When I do feel genuine fatigue, it's quite different from my ME symptoms.

Too bad low dose naltrexone has never been one of those things that is consistently helpful for me.

It was only temporarily helpful for me too. So, is 'modulating TRPM3 channel function in NK cells' an inherent part of ME, or just one of those secondary effects that shows up in some people some of the time, and thus not likely to lead to a treatment for ME?

 

[Gingergrrl]

What do you guys think about this? Seems the full study will be published soon. @Gingergrrl @necessary8

Thanks for tagging me and I just glanced through the study but am not sure if I understand it

ME/CFS hypothesis involves ion channel disorders and more specifically impaired Transient receptor potential melastatin 3 (TRPM3) ion channel function, affecting calcium signalling and Natural killer (NK) cell functions.

This part is very interesting to me especially after seeing that "ion transport issues" are part of the ICC Criteria (which I started a whole other thread about a few weeks ago). I am not sure what came first though, did they find ion transport issues like this study did and then put it into the ICC Criteria or was it somehow in the ICC Criteria first and then they worked backwards with these studies to try to prove it?

Naltrexone hydrochloride (NTX) acts as an antagonist to the μOR thus negating the inhibitory function of this opioid receptor on TRPM3.

I tried LDN back in 2014 and had a horrible reaction to it and it was not helpful for me at all (vs. I do very well with opiates for pain).

Whole-cell patch-clamp technique

What is "whole-cell patch clamp technique"?

Our results demonstrate, for the first time, and based on novel patch clamp electrophysiology, potential pharmaco-therapeutic interventions in ME/CFS.

I wish I knew what this meant!

 

[Marylib]

Well, @Gingergrrl , I wish I understood it too. I think the whole patch clamp technique refers to their device that somehow collects serum and measures calcium ion channel things. If I can think of it, I will look it up again and maybe have something to post that is actually helpful! Yes, opiates are amazing. Not being someone with alot of pain anymore, I can just count myself lucky.

About the criteria, I think this group has been pretty well focused on more strict criteria for some years now, but I am not good at history at the moment. I do know that chronic fatigue syndrome as a name (and the old Fukuda definition) seems to have pervaded the whole globe, and the ICC authors may have gotten pretty miffed about it.

 

[JES]

Boy is this confusing. The common wisdom rationale for taking naltrexone in form of LDN was the idea that it blocks opioid receptors temporarily. With a dosage of around 4 mg, this blocking effect will last a few hours, which tricks the body into a rebound stimulation of endorphins after this short-term blockade.

But this article suggest that blocking opioid receptors is actually what you would want to do to restore TRPM3 channel activity. In the experiment they used a 24 hour incubation period to restore TRPM3 channel activity.

So could it be that in order for this effect to materialize, you would ideally want to take standard, not low dosage naltrexone, which in 50 mg form will block the opioid receptors pretty much continuously. This is of course the mechanism by which naltrexone is used to treat alcoholism and it would completely negate the rebound effect, but I think it's possible that the TRPM3 restoring effect would require this sort of dosage.

 

[Wishful]

With a dosage of around 4 mg, this blocking effect will last a few hours,

Confusing indeed, as to exactly how it's working. For me, 3 mg would block pain for between 36 and 48 hrs. I can't remember how long it took to start blocking pain.

Also, I wouldn't call it a treatment for ME/CFS. I'd call it a treatment for one specific symptom that some PWME have, and it's a treatment that works only for some, and causes bad side effects for others. I think there are areas of research with higher potential benefits for more PWME.

 

[skandar]

What is "whole-cell patch clamp technique"?


I wish I knew what this meant!

It is a common experimental approach used to measure ion flux in single cells. Usually it is done on excitable cells (e.g., neurons). So one can assess the effect of a drug on Ca flux through a calcium channel, or Na flux through a sodium channel, Cl flux, K flux, etc.

 

[Gingergrrl]

Well, @Gingergrrl , I wish I understood it too.

Glad I am not the only one

So could it be that in order for this effect to materialize, you would ideally want to take standard, not low dosage naltrexone, which in 50 mg form will block the opioid receptors pretty much continuously.

@JES Do you mean that this study was using the full dose of naltrexone and not LDN?!

It is a common experimental approach used to measure ion flux in single cells. Usually it is done on excitable cells (e.g., neurons). So one can assess the effect of a drug on Ca flux through a calcium channel, or Na flux through a sodium channel, Cl flux, K flux, etc.

Thank you for explaining, and if I understand, then the patch clamp is some kind of device that allows the researchers to measure how much calcium is flowing into and out of an individual cell? How does this relate to the naltrexone?

 

[JES]

@JES Do you mean that this study was using the full dose of naltrexone and not LDN?!

Actually it's difficult to say what dosage in humans the study dosage would correspond to, since the study was done on isolated cells, not humans ingesting naltrexone:

Importantly, TRPM3 channel activity was restored in IL-2 stimulated NK cells isolated from ME/CFS patients after incubation for 24 hours with NTX.

So they demonstrated they were able to restore TRPM3 channel activity by blocking opioid receptors for 24 hours. But from what I have read, naltrexone in form of LDN only blocks opioid receptors for a couple of hours after each dosage. So it's a bit speculation from my side, but using naltrexone instead of LDN would seem to make more sense if the desired effect is to block opioid receptors and restore TRPM3 channel activity.

 

[Marylib]

Confusing indeed, as to exactly how it's working. For me, 3 mg would block pain for between 36 and 48 hrs. I can't remember how long it took to start blocking pain.

Also, I wouldn't call it a treatment for ME/CFS. I'd call it a treatment for one specific symptom that some PWME have, and it's a treatment that works only for some, and causes bad side effects for others. I think there are areas of research with higher potential benefits for more PWME.

Wow - you guys are really amazing with these posts.

I think this is what does my head in when I try to put this finding in context, because the mind quickly goes to the use of low-dose naltrexone for symptom relief. This has been quite popular with some people.

I am thinking along the lines of @Wishful at the moment. It seems to me that this paper is not yet proposing treatment with naltrexone. Actually, I think some clinicians associated with this research are trialling an actual calcium channel blocker. (@Gingergrrl was in on this discussion as not being able to take this kind of drug in her case.)

The fact that naltrexone is altering the NK cell problem, through its action on the faulty TRPM3 receptor - that must be the big deal. Because the NK cell problems came up ages ago in immunological research - Mary Fletcher was trying to propose the low NK thing as a biomarker ages ago.

So for years, people have been trying to figure out how to find a fix for the NK cell problem (ampligen, isoprinosine, equilibrant, etc.)

So I am thinking that this paper from the Australian group is building on the already solid work in immunology. And this latest finding would further confirm the biomarker - the clamp device to measure the flux of intracellular calcium. In other words, it is "proof" that using an immune modulator can fix the faulty receptor in these isolated human cells. And the agent they have used is naltrexone. So this would make naltrexone a candidate for actual treatment.

Open Medicine Foundation is perhaps still working on the metabolic trap hypothesis, which comes from a different angle, I think.

But Open Medicine Foundation is now recruiting for patients with connective tissue problems, as this something that keeps popping up in the patient population - along with the structural defects like cervical cranial instability.

This narrowing of this structural passage would seem to link up in terms of the newly recognized glymphatic system??

https://www.sciencedirect.com/science/article/abs/pii/S0306987718305486

So if this is the case, we are now looking at several disease states or syndromes, one of which seems to be fixed through surgery and the other through correcting the defect in moving calcium through the cells.

https://www.ncbi.nlm.nih.gov/pubmed/29391028

I have no idea if all this makes sense to anyone else, so let me know what you think. I feel like I am operating far above my own abilities to put all these recent findings in context.

 

[skandar]

Thank you for explaining, and if I understand, then the patch clamp is some kind of device that allows the researchers to measure how much calcium is flowing into and out of an individual cell? How does this relate to the naltrexone?

Hi @Gingergrrl . Good questions. I'll try: the bona fide electrophysiologists use patch clamp because it is a powerful, but very specialized skill (it is difficult and requires a good deal of training). They are measuring ion flux through the area of the plasma membrane that is covered by the patch (i.e., in the microelectrode). So depending on which channels are in the patch, this is the area of ion flux being measured.

TRPM3 are not well-characterized among the TRP superfamily (~38 different genes), but they are thought to form a pore for calcium entry. How I understand this to be related to naltrexone is that the u (mu) opioid receptors are G-coupled protein receptors, and when naltrexone binds to them, this causes changes in phospholipid bilayer membrane signaling that in turn causes TRPM3 to alter the amount of calcium that gets into the cell.
There is a good schematic diagram of this in Figure 2 here: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3698859/

 

[Gingergrrl]

Actually it's difficult to say what dosage in humans the study dosage would correspond to, since the study was done on isolated cells, not humans ingesting naltrexone:

because the mind quickly goes to the use of low-dose naltrexone for symptom relief.

Thank you both for explaining (@JES and @Marylib) and I had also incorrectly assumed that this study was talking about LDN and not full-dose Naltrexone.

Actually, I think some clinicians associated with this research are trialling an actual calcium channel blocker. (@Gingergrrl was in on this discussion as not being able to take this kind of drug in her case.)

Good memory and I was talking about this in another thread re: the ICC Criteria having a section re: ion transport issues. I can't personally take a calcium channel blocker but most (many?) people can and I find this entire research very interesting that there is now a whole line of thinking that the issue or mechanism of action relates to the calcium channels (like in my case).

And this latest finding would further confirm the biomarker - the clamp device to measure the flux of intracellular calcium. In other words, it is "proof" that using an immune modulator can fix the faulty receptor in these isolated human cells.

Are they saying that calcium channel issues is a potential biomarker for ME/CFS?

But Open Medicine Foundation is now recruiting for patients with connective tissue problems, as this something that keeps popping up in the patient population - along with the structural defects like cervical cranial instability.

So if this is the case, we are now looking at several disease states or syndromes, one of which seems to be fixed through surgery and the other through correcting the defect in moving calcium through the cells.

That is my current take, too, that different groups of researchers are looking at different patho-mechanisms of the diseases or at different diseases completely (I am not yet sure which)?!

I have no idea if all this makes sense to anyone else, so let me know what you think. I feel like I am operating far above my own abilities to put all these recent findings in context.

Likewise and I have no ability to put this all together either.

I'll try: the bona fide electrophysiologists use patch clamp because it is a powerful, but very specialized skill (it is difficult and requires a good deal of training). They are measuring ion flux through the area of the plasma membrane that is covered by the patch (i.e., in the microelectrode). So depending on which channels are in the patch, this is the area of ion flux being measured.

Thank you for explaining @skandar and there seem to be several research studies re: calcium channels and NK cells but I am not sure if they are all looking at the same issue or different issues?

TRPM3 are not well-characterized among the TRP superfamily (~38 different genes), but they are thought to form a pore for calcium entry.

Do you know if this would relate to calcium channel autoantibodies (or would that be a totally separate issue)?

 

[skandar]

Thank you for explaining @skandar and there seem to be several research studies re: calcium channels and NK cells but I am not sure if they are all looking at the same issue or different issues?

I think the studies by this group (Marshall-Gradisnik et al) on NK cells are all on the TRPM3 channel.
What is complicated is that there are may different kinds of calcium channels and almost all cell types have calcium channels on the plasma membrane. I'm most familiar with calcium channels on excitable cells like muscle because they initiate excitation-contraction coupling. But calcium is an intracellular signaling molecule in most cells. For instance, this is a nice article on T lymphocytes, and they mention the type of calcium channel you have been referring to (TRPs).
"The repertoire of calcium-conducting proteins in T lymphocytes includes store-operated CRAC channels, transient receptor potential channels, P2X channels, and L-type voltage-gated calcium (Cav1) channels. In this paper, we will focus mainly on the role of the Cav1 channels found expressed by T lymphocytes, where these channels appear to operate in a T cell receptor stimulation-dependent and voltage sensor independent manner."
https://www.frontiersin.org/articles/10.3389/fimmu.2013.00243/full

Do you know if this would relate to calcium channel autoantibodies (or would that be a totally separate issue)?

I do not know the ME/CFS literature on calcium channel autoantibodies. I am aware of reports of a few autoantibodies such as specific adrenergic receptors and acetylcholine receptors, and ANA.

Taken together, I think some investigators are trying to understand the role of TRPM3 in certain immune cells because of the SNV's in the TRPM3 gene. Then, I believe there is an attempt to understand how opioid receptors and drugs that people take for pain (opioid receptor agonists) or drugs that block opioid receptors (naltrexone), might regulate immune cell function in ME via ion flux. But I'm not certain, it's not a part of the ME literature that really caught my attention at the moment.

 

[Marylib]

Here is a page about on the areas of research for that group. I don't know when it was last updated:

https://meaustralia.net/current-science/

@Gingergrrl I think that the research group in Australia are in the stages of proposing using their clamp device as a biomarker, in terms of to the TRMP3 genetic defect (SNP) - but I don't have all the literature around it at the moment. Their specialty is neuro-immune, I think. But they are looking at mitochondrial aspects too.

I guess this would be in the same category as the device Open Medicine Foundation is proposing- the little computer chip they use to measure the plasma with another technique. I don't really know what is happening with OMF in that regard at the moment, but I would like to know more. I need help understanding it all. LOL

I did recently see a video of Dan Peterson (a new working group) who said he feels there are several different disorders or diseases involved in what is currently known as M.E. and CFS.

 

[Dechi]

I was on LDN for more than 4 months and did not have conclusive benefits. It seems to have had an impact on my neuropathic pain, but it’s still not clear if it stopped for good or will come back again. Other than that, not much changed while I was on it.