You know that I adressed this in the first part of my ponderings, right? I literally referenced the same two studies. I said:In your ponderings you're thinking about pannexin-1 (Panx1) in terms of its relationship with P2X7.
But it was discovered in 2008/2009 by at least two different groups (Qiu et al and Ma et al) that Panx1 actually has its own receptor for external ATP (eATP) that operates independently of P2X7. External ATP is able to inhibit Panx1 via this receptor.
So what about other inhibitory mechanisms of purinergic signaling, other than ecto-nucleotidases? The only other thing that I found, were some observations that P2X7 activation can inhibit Panx1-mediated ATP release.[source] Yes, this is the reverse of what I said at the beginning. There were two studies that reported this, one on murine cells, other on human HEK293 cells. This is a cell line cultured from embryonic kidney cells, often used in research because it's easy to modify them to do what you want, so they have different properties based on how you "bake them". In this case, Panx1 was artificially introduced to those cells through transfection.
Those observations raise two possibilities. First, that depending on the cell type, P2X7 activation might open or close Panx1 channels. The cell types in which P2X7 activation has been observed as leading to Panx1 channel opening are macrophages[source], astrocytes[source], and enteric neurons[source]. (And maybe more, I just haven't found the papers about it.) The P2X7-mediated Panx1 activation seems to be widely accepted in literature, but citations about it are often not present. It is very possible that in some cells P2X7 activation opens Panx1 and in other types closes them.
The second, more interesting possibility, is that there is some other factor, which decides if P2X7 activation in a cell opens or closes Panx1 channels. A distinction on P2X7 splice variants might be it.[source] Or it might be some yet undiscovered thing, possibly our mysterious blood factor, or something tied to it biochemically. We don't really know.
I consider the results of Qiu and Ma preliminary, because they are not on natural human cells. Qiu uses Xenopus, while Ma artificially transfects Panx1 into cells that do not have them normally. The research that shows ATP causing activation and opening of Panx1 is much higher quality, and larger. For me those two studies are indications that maybe sometimes eATP can close Panx1 channels, and we don't know what are the factors which decide if it opens or closes them.
Now, you propose, that we do know what the factor is, that it is solely the concentration. And you quote this as a source:
The problem here is that this sentence, in the original review that you linked, does not have a citation. It is nothing more than author's speculation. It's a very intetesting speculation, but a speculation nontheless. The Qiu and Ma studies do not prove this, because they induce Panx1 activation via electrical stimulation, not by administering low concentrations of eATP.At low ATP concentrations, the purinergic receptors [e.g., P2X7] activate Panx1 resulting in amplified ATP release. As the ATP concentration builds up in the vicinity or within the vestibulum of the Panx1 channel, the self inhibition will limit further ATP release.
What would actually prove this is the case, is if someone took the same human cell type that was used in previous studies to show that eATP opens Panx1, like macrophages or astrocytes, and demonstrated that when you apply higher amounts of eATP, the Panx1 channels close. If it was done on macrophages, they would also need to apply the appropriate cytokines to ensure the macrophage stays in the M1 polarization, because a shift to M2 can occur spontaneously, and it changes how they respond to eATP.
So if you find a study like that, I will immediately admit that you're right on this, and I was wrong. But for now I'm gonna say this again - We. Do. Not. Know. why sometimes eATP makes Panx1 close. We do not even know if such an effect occurs in actual, natural human cells. The action of Panx1 in humans might be dependent on dozens of factors that we don not yet understand, and which Qiu and Ma failed to reproduce in their studies.
I gave the values/sources I'm using previously:
Ah, yes. My bad. I missed it. Yes, it would seem that you're right here, at least in your math. But since this directly contradicts what Naviaux has said in his presentation, I think it's time we ask him directly about it, so I've sent an email to him. I'll let you know once he responds.
The one possibility I see, for you to be wrong on your conclusion, is that the initial concentration is 100uM, and only falls to 12uM after 2 days. It might be that it is the initial concentration that breaks the eATP release feedback loop, and the upregulation of P2X7 isnt enough to undo it later when the concentration falls. But we'll see what Naviaux says.
I get that, but what you're effectively suggesting here is that the mechanism of ME/CFS involves an underactivation of P2X7, Panx1, and ATP being released to a lesser degree than in healthy people. This is not only directly opposite to what Naviaux said about ME/CFS, it is also not consistent with symptoms, and many research findings. My "ponderings" thread is basically two long essays explaining in detail, how OVERactivation of Panx1, P2X7 and INCREASED release of eATP is consistent with pretty much all the symptoms, as well as many of the key research findings. So this is why I'm extremaly skeptical of this hypothesis of yours - there is a mountain of arguments on the opposite side. I can't say for sure that you're wrong, but as it currently stands, your version seems incredibly unlikely to me.So the idea with clemastine is just to try to capture what is outwardly an apparent net effect of an up-regulation at P2X7 and see what happens.
See, but in the same study, they say clemastine potentiates the formation of large pores, as assesed by large molecule dye uptake. So it means that clemastine does potentiate eATP release from cells. Now, they say Panx1 is not involved here, because the potentiation of dye uptake did not change when they added carbenoxolone, which is a Panx1 inhibitor. Further, they state that carbenoxolone by itself caused dye uptake! This is the reverse of a multitude of other studies, which show carbenoxolone decreasing dye uptake. Something very weird is going on here. Now, it's interesting to note that in this study they used HEK cells, the same as the Ma et all study we mentioned earlier. To me, this is a suggestion that HEK cells are not representative of how P2X7 and Panx1 behave in natural human cells.How a biochemical result is achieved can obviously be just as important as the result itself but note that clemastine effectively up-regulates P2X7 by making it more sensitive to eATP while apparently not affecting Panx1: