Animal research for ME

[Jonathan Edwards]

showing that IgG from people with LongCovid, injected into mice caused the mice to exhibit symptoms

Now that I remember, @PeterW, we used to call it the Yak Dung Effect in the lab in 1980. Give a mouse or a rat anything and the graph will shift with p<0.01. Or at least one of your graphs will and that is the one to publish.

 

[forestglip]

It's true that if you expose mice immediately to high concentration CO2 they will exhibit signs of distress but if you increase the flow rate more slowly that doesn't happen

Can you provide a source that distress doesn't happen at slowly increased flow rates?

The effect of carbon dioxide flow rate
on the euthanasia of laboratory mice, 2014

Abstract
Laboratory rodents are commonly euthanized by exposure to gradually increasing concentrations of carbon
dioxide (CO2 ). Current recommended flow rates range between 10 and 30% chamber vol/min and result in
insensibility before exposure to painful concentrations (<40%). However, this method causes dyspnea, indi-
cated by deep, rapid breathing. In humans dyspnea is associated with a negative affective experience.
Sensations of dyspnea may explain why rodents find CO 2 concentrations >3% aversive. This study aimed to
assess the effect of CO 2 flow rates on time between the onset of dyspnea and various measures of insensibility
(recumbency, loss of the righting reflex and loss of the pedal withdrawal reflex) to identify flow rates that
minimize the potential experience of dyspnea. The results of this study indicate that a flow rate of 50%
chamber vol/min, while holding the CO 2 cage concentration just below 40%, minimizes the interval between
the onset of labored breathing and recumbency. Using a 50% flow rate this interval averaged (SE)
30.3 2.9 s versus 49.7 2.9 s at 20% chamber vol/min (F 3 ,22 ¼ 7.83, P ¼ 0.0013). Similarily, the interval
between the onset of labored breathing and loss of the righting reflex averaged 38.2 2.4 s at a flow rate
of 50% versus 59.2 2.4 s at 20% chamber vol/min of CO 2 (F 3, 22 ¼ 13.62, P < 0.0001). We conclude that higher
flow rates reduce the duration of dyspnea, but even at the highest flow rate mice experience more than 30 s
between the onset of dyspnea and the most conservative estimate of insensibility

Nitrogen gas produces less behavioural and neurophysiological excitation than carbon dioxide in mice undergoing euthanasia, 2019

Carbon dioxide (CO2) is one of the most commonly used gas euthanasia agents in mice, despite reports of aversion and nociception. Inert gases such as nitrogen (N2) may be a viable alternative to carbon dioxide. Here we compared behavioural and electrophysiological reactions to CO2 or N2 at either slow fill or rapid fill in C57Bl/6 mice undergoing gas euthanasia. We found that mice euthanised with CO2 increased locomotor activity compared to baseline, whereas mice exposed to N2 decreased locomotion. Furthermore, mice exposed to CO2 showed significantly more vertical jumps and freezing episodes than mice exposed to N2. We further found that CO2 exposure resulted in increased theta:delta of the EEG, a measure of excitation, whereas the N2 decreased theta:delta. Differences in responses were not oxygen-concentration dependent. Taken together, these results demonstrate that CO2 increases both behavioural and electrophysiological excitation as well as producing a fear response, whereas N2 reduces behavioural activity and central neurological depression and may be less aversive although still produces a fear response. Further studies are required to evaluate N2 as a suitable euthanasia agent for mice.

One interesting thing - while humans and many other animals can not directly detect low oxygen environments directly and are actually detecting high CO2, some animals apparently can sense low oxygen without high CO2:

Wikipedia: Inert gas asphyxiation

Diving animals such as rats and minks and burrowing animals are sensitive to low-oxygen atmospheres and will avoid them. For this reason, the use of inert gas (hypoxic) atmospheres (without CO2) for euthanasia is also species-specific.

differences in price between nitrogen and carbon dioxide will probably be trivial.

You might be right about price. Though on this Reddit thread, a few people who work with lab mice say the difference is a time cost. Nitrogen takes much longer to kill a mouse.

 

[JemPD]

I find the idea of animal models at this stage, to be grossly unethical and at best a waste of time.

there is no such thing as an 'animal model' of ME. It simply cannot currently exist. We dont know what causes ME so its impossible... IMPOSSIBLE to accurately simulate one.

Exercise to exhaustion plus psychological stress does not = ME/CFS. If only it were! But no, that is just a common but woefully inaccurate, misconception.

So any experimentation on torture of, animals based on such ideas will not lead to any insight into ME/CFS. Any more than injecting a animals in the leg with lidocaine to create intermittent numbness, would lead to insight into MS. That wouldnt be an 'animal model' of MS, and scientists would laugh at the idea it could yield anything useful.

The whole thing is absurd and vile. Its one thing doing experiments to save lives, when it might actually save lives, (I mean i not sure of the ethics of doing even that but its a more difficult & nuanced decision).... but doing it for this?!?

I feel angry and outraged about it.

Not to mention the tragic waste of time and money that could be spent on something useful.

 

[PeterW]

I find the idea of animal models at this stage, to be grossly unethical and at best a waste of time.

there is no such thing as an 'animal model' of ME. It simply cannot currently exist. We dont know what causes ME so its impossible... IMPOSSIBLE to accurately simulate one.

Exercise to exhaustion plus psychological stress does not = ME/CFS. If only it were! But no, that is just a common but woefully inaccurate, misconception.

So any experimentation on torture of, animals based on such ideas will not lead to any insight into ME/CFS. Any more than injecting a animals in the leg with lidocaine to create intermittent numbness, would lead to insight into MS. That wouldnt be an 'animal model' of MS, and scientists would laugh at the idea it could yield anything useful.

The whole thing is absurd and vile. Its one thing doing experiments to save lives, when it might actually save lives, (I mean i not sure of the ethics of doing even that but its a more difficult & nuanced decision).... but doing it for this?!?

I feel angry and outraged about it.

Not to mention the tragic waste of time and money that could be spent on something useful.

The study I am referring to was not an animal model of ME/CFS, they injected mice with extracts of blood from ppl with LongCovid, and those mice then exhibited symptoms.

Having used mouse poison in the past, I don’t have an issue with undertaking studies which result in the death of mice.

Perhaps having seen animals being slaughtered the old fashioned way (knife to throat and bleeding out), I don’t have such a horrible opinion of it. I think there are worse ways to go than these.

 

[JemPD]

ah, well, as you can tell from my comment i was referring to studies using an 'animal model' of ME/CFS - of which there are quite a few & seem to be growing in number. Its those studies i object to.
The one you mention where they injected IgG... Is that the one you mean? I feel agnostic about that, but the poll just says 'research uisng animals', and there is no 'on the fence'/not sure, option.

 

[Jonathan Edwards]

The study I am referring to was not an animal model of ME/CFS, they injected mice with extracts of blood from ppl with LongCovid, and those mice then exhibited symptoms.

That's what we call an animal model. LC maybe, but I think the assumption is that they are trying to model something like ME/CFS.

 

[PeterW]

And when the experiment is done...as far as I know, almost all mice are killed with CO2 asphyxiation, likely a horribly agonizing death. Even though nitrogen asphyxiation is a painless option, CO2 is cheaper and easier, so that's what normally gets used.

many pigs in the UK are slaughtered using CO2 asphyxiation. That doesn’t disprove your point (about which I simply have no insight) but I do eat pork which will have been slaughtered that way.

 

[forestglip]

many pigs in the UK are slaughtered using CO2 asphyxiation. That doesn’t disprove your point (about which I simply have no insight) but I do eat pork which will have been slaughtered that way.

Yeah, I'm not a fan of food animals being treated inhumanely either. One organization I think is doing great work in this field is the Humane Slaughter Association, and I believe has had some big wins in changing industry in Europe. Though I think they only deal with farm animal slaughter, not lab animals.

https://www.hsa.org.uk/news-events/...lternatives-must-now-be-urgently-implementeda

The Humane Slaughter Association’s position regarding the slaughter of all animals is that they should be killed as humanely as possible with every reasonable effort made to reduce pain, suffering and distress to an absolute minimum.

Carbon Dioxide (CO2) is widely used to stun pigs prior to slaughter. However, robust scientific evidence shows that it causes significant animal welfare issues. As early as 2003, the then UK Farm Animal Welfare Council (FAWC) called for CO2 to be phased out within five years1, yet two decades later this method continues to be widely used (for example, in the UK, over 80% of pigs are stunned with CO2). The HSA view is that a more humane alternative to CO2 should be adopted as soon as possible.

In support of this aim, in 2018 the HSA co-funded research alongside the UK Department for Environment, Food and Rural Affairs (DEFRA) to test one potentially humane alternative stunning method – Low Atmospheric Pressure Stunning (LAPS). Unfortunately, the research showed that LAPS was not a humane alternative for stunning pigs, although it had previously been shown to be a more humane method of poultry slaughter. The researchers conclude that “Collectively, our results show that both LAPS and CO2 stunning are associated with several indicators of poor welfare and are equally aversive to pigs.” The final report2 detailing the project findings has just been published and can be downloaded at: https://sciencesearch.defra.gov.uk/ProjectDetails?ProjectId=19805

It was important to test a method which could potentially have provided a humane and sustainable solution for pig stunning. However, now that LAPS has been shown not to be the alternative to CO2 that many had hoped, it is essential that other alternatives are adopted without undue delay.

 

[PeterW]

there is no such thing as an 'animal model' of ME. It simply cannot currently exist. We dont know what causes ME so its impossible... IMPOSSIBLE to accurately simulate one.

A load of our current medications were developped before we had any understanding of the mechanism underpinning them.

While the idea of building-up a formal mechanistic understanding to support drug development is very attractive, is that how many major drugs have been developed, or is there a need for serendipidy here?

 

[JemPD]

A load of our current medications were developped before we had any understanding of the mechanism underpinning them.

While the idea of building-up a formal mechanistic understanding to support drug development is very attractive, is that how many major drugs have been developed, or is there a need for serendipidy here?

I'm confused... I cant follow your reasoning... What i'm talking about is researchers exercising animals to exhaustion & then testing things on them, and assuming their results tell us something about ME. Whatever the results are they will only be results on what works (or doesnt), for treating a mouse who has exercised to exhaustion, & therefroe possibly a human.

Since ME does not = having exercised to exhaustion, i dont see how it can serendipitously tell us anything about it. It can only tell us something about what happens in a mouse body when they have swam until the point of collapse (Or sometimes they stress them out psychologically as well or instead of, the exhaustion).

Its like giving a bunch of mice very nasty psychological shocks, until they start trembling & shaking with it, & then expecting experimentation on them in that state to tell us something about parkinsons, just because it looks superficially similar. Its illogical.

I eat meat, with guilt but i do eat it, but i do not find it acceptable to kill more animals than we need for food (i know that happens but the question was what we find acceptable). Neither do i find it acceptable to do all kinds of experiments on animals hoping to discover something out of the experiments 'serendipitously'.

Yes we are going to need a pretty big stroke of luck to get anywhere, but i dont see it as necessary to toy with extreme cruelty, with animals, until we have a damn good reason to do so. I'm not certain i find it ethical even then, but that for me would be a different question.

Again, to be clear i am talking ONLY about studies on animals where researchers try to simulate ME/CFS - i dont believe that is possible, so to my mind research on animals with simulated ME/CFS is wrong and a waste. Playing around with them hoping for a stroke of luck is appalling.

The example you gave of injecting an infected person's blood to see if it will give them symptoms... well that is slightly different, i dont know about that, but i defer to Jonathan's comments both here and on the thread about that study, since it is all a bit scientifically beyond me.

All i'm saying is : No experiments on animals that are exercised to exhaustion , and or stressed to collapse, that are being performed because the researchers are too ignorant to know that that is simply not what ME/CFS is. I wish it were that but it isnt, it only looks a little like that to people who confuse it with CF or TATT.


LOL I honestly cant eat my ham sandwich now, i didnt think about that pig's death when i bought it. I dont find it acceptable to treat animals inhumanely during life or slaughter, and try to buy my meat accordingly, but i dont always, so i need to face that and stop it.
I only started eating meat for ease when i got too ill to cook for myself & was cutting out all kinds of other things trying to 'nutrition' my way out of this hell. But i carried on eating it even when i resumed bread etc, because it tastes good and it is easier.

Sorry if that was very waffly i not good at succinct even before the fog descended

 

[PeterW]

I would agree with you that animals exercised to exhaustion are not in any way a proxy for ME/CFS.

The question is, if insights can be gained from animal testing, should we be doing it?

that is significant as the UK ME/CFS charities all refuse to fund animal research.

This would include injecting the blood of people with LongCovid into mice to see if they exhibit symptoms.

I feel that is legitimate animal research, and I am disappointed it has not been done before.

 

[forestglip]

This would include injecting the blood of people with LongCovid into mice to see if they exhibit symptoms.

I feel that is legitimate animal research, and I am disappointed it has not been done before.

I tend to ask myself for anything involving animals, would this be acceptable to do to a human? Would society be okay with injecting healthy humans with LC blood, trying to induce LC symptoms? If not, what's the reason it feels okay to do to an animal?

 

[PeterW]

I tend to ask myself for anything involving animals, would this be acceptable to do to a human? Would society be okay with injecting healthy humans with LC blood, trying to induce LC symptoms? If not, what's the reason it feels okay to do to an animal?

Well, on a practical level, many of us do eat meat, poison and trap mice, kill slugs etc. All things which our society does not accept being done to other humans.

So many of us do differentiate between human and non-human animals on a daily basis.

 

[forestglip]

Well, on a practical level, many of us do eat meat, poison and trap mice, kill slugs etc. All things which our society does not accept being done to other humans.

So many of us do differentiate between human and non-human animals on a daily basis.

That's true. So I'd go to the question, why does it seem okay? Is it because there's actually some difference that matters morally in all these cases, or does it just feel that way because animals are so different?

To me, it mostly seems like the latter, as well as because that's how it's always been done.

 

[PeterW]

That's true. So I'd go to the question, why does it seem okay? Is it because there's actually some difference that matters morally in all these cases, or does it just feel that way because animals are so different?

To me, it mostly seems like the latter, as well as because that's how it's always been done.

Well many people have thought about that.

So the question of morality of testing on animals (and whether it should be done in ME/CFS) essentially boils down to the simple question of “are you vegan”?

 

[Jonathan Edwards]

The question is, if insights can be gained from animal testing, should we be doing it?

that is significant as the UK ME/CFS charities all refuse to fund animal research.

This would include injecting the blood of people with LongCovid into mice to see if they exhibit symptoms.

I feel that is legitimate animal research, and I am disappointed it has not been done before.

I think the situation is more nuanced.

All immunology, from which we have to build theories of disease, has involved animal experimentation to establish how the immune system normally works and responds to stimuli.

ME/CFS charities are happy to fund research that makes use of that knowledge. They fund research that involves use of lab reagents generated in animals.

What they are not happy to do by and large is fund studies where animals are deliberately caused distress. How strict they are and how open to change in policy they might be I do not know but that would be a reasonable position if history has told us that such studies are by and large useless.

And that is the case.

Research is not legitimate if there isn't snowballs chance in hell of it telling us something useful. Giving a mouse a slug of human IgG is a bit like giving a human two pints of mouse IgG. Since all of it (not just some extra mysterious 'thing in the blood') is foreign protein there will inevitably be a reaction. So testing mice for symptoms vaguely compatible with ME/CFS or fibromyalgia after injection of human IgG is going to be pretty hard to interpret. Moreover, people did hundreds of experiments like this in the 1970s and 1980s and ended up none the wiser.

Small molecule drug development is still very dependent on being able to simulate things like inflammation in animals for early efficacy testing. For biologic drugs this doesn't necessarily apply. But in ME/CFS and fibromyalgia we have no inflammation to simulate and anyway the drug companies would do that sort of study - and only once a pathway had been identified. We don't have any identified pathways.

 

[PeterW]

I think the situation is more nuanced.

All immunology, from which we have to build theories of disease, has involved animal experimentation to establish how the immune system normally works and responds to stimuli.

ME/CFS charities are happy to fund research that makes use of that knowledge. They fund research that involves use of lab reagents generated in animals.

What they are not happy to do by and large is fund studies where animals are deliberately caused distress. How strict they are and how open to change in policy they might be I do not know but that would be a reasonable position if history has told us that such studies are by and large useless.

And that is the case.

Research is not legitimate if there isn't snowballs chance in hell of it telling us something useful. Giving a mouse a slug of human IgG is a bit like giving a human two pints of mouse IgG. Since all of it (not just some extra mysterious 'thing in the blood') is foreign protein there will inevitably be a reaction. So testing mice for symptoms vaguely compatible with ME/CFS or fibromyalgia after injection of human IgG is going to be pretty hard to interpret. Moreover, people did hundreds of experiments like this in the 1970s and 1980s and ended up none the wiser.

Small molecule drug development is still very dependent on being able to simulate things like inflammation in animals for early efficacy testing. For biologic drugs this doesn't necessarily apply. But in ME/CFS and fibromyalgia we have no inflammation to simulate and anyway the drug companies would do that sort of study - and only once a pathway had been identified. We don't have any identified pathways.

Not quite right - this thread was first started after 2 UK ME/CFS charities stated to me, quote “our position on using animals in research is unequivocal - we don’t fund it”. Another said similar.

Regarding the limitations of different research methodologies - you will know these better than me but, I would note that many research institutions and larger research funders do have a less restrictive approach:

Oxford Uni: https://www.ox.ac.uk/news-and-events/animal-research/research-using-animals-an-overview

Parkinson’s UK: https://www.parkinsons.org.uk/about-us/animal-research-at-parkinsons-UK

Vs Arthritis: https://versusarthritis.org/researc...r-approach-to-research/our-research-policies/

When we look at research funding in the UK, we see that ME/CFS funding is insufficient both from government agencies (see Prof Danny Altmann’s recent comments that funders are “not interested”) but also from the UK charity sector (which funds even less research than government does).

It is worth querying whether the UK charity sector is being too dogmatic around the research that it will fund. It is also worth querying whether fixed ideas about methodologies including animal research has, or in the future could, get in the way of understanding this disease better or testing potential therapeutics.

 

[Jonathan Edwards]

Not quite right - this thread was first started after 2 UK ME/CFS charities stated to me, quote “our position on using animals in research is unequivocal - we don’t fund it”.

Well, having been on the steering committee of a research group funded by such a charity that had to use reagents derived from laboratory animals I can say that things are not so unequivocal. You cannot do any immunology without doing that. And there wouldn't even be any immunology if there hadn't been background animal studies. The charities mayn't realise that but I suspect they do and I don't actually think they would object. Charles Shepherd would be aware of the situation for instance, as would the people from MERUK I have met.

 

[Jonathan Edwards]

It is worth querying whether the UK charity sector is being too dogmatic around the research that it will fund.

It is worth querying but I can assure you that as someone who once had a specific interest in animal models I think the ME charity policy is exactly right. There are not experiments that involve direct harm to animals as disease modelling exercises worth doing in ME/CFS and I doubt there ever will be. Oxford has had a reputation for having a pretty borderline ethical policy, hence the animal rights episodes around 1990. Versus Arthritis has a long history of funding animal model studies that I consider totally unjustifiable and a complete waste of time.

Worth querying, yes, but I wouldn't get too bothered about it. It is not where the problems lie. The last thing we want is for people's money to go into the sort of pointless me-too research that dominates the scene these days. DecodeME is a rock solid project. There have been other good searching exercises in immunology that have come out negative. The metabolic studies make sense but it seems unlikely they will turn up much when we know so much of the metabolism is normal. Otherwise our main problem is that we do not have leads. Setting up animal models with no leads is futile because you can always set up a model that does what you manipulate it to do and it still has nothing whatever to do with the disease in hand.

 

[poetinsf]

Animal research is often more for exploring than for proving. In diseases with unknown aethiology like ME/CFS, just reliably reproducing the disease would be a huge step towards understanding and solving the problem. And you can't do that with humans for obvious reasons. If you have to justify by proving the significance first, no such exploration will never take place.