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Direct Application of MDMA to the Brain is not Neurotoxic

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BilZ0r

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I often read that the "direct application of MDMA to the brain is not neurotoxic*" or some such things. Examples can be seen on BlueLight in zorn's MDMA/neurotoxicity misconceptions thread, an in Monks et al., The role of Metabolism in MDA and MDMA toxicity.

Now I wanted to look into this a bit more closely, as in the next couple of months, I'm hopefully going to do some in vitro work using primary cell culture, to see if BZP is neurotoxic (crosses fingers and prays for grant money)... Because if as I've read, MDMA isn't neurotoxic if applied directly to the brain, or into the cerebral ventricles, then it would be hard to believe that MDMA is neurotoxic in cultured neurons.

First I did a pubmed search. The two articles I found straight away were Schmidt and Taylor and Paris and Cunningham. These two papers are odd "proofs" that MDMA isn't neurotoxic when applied directly. Schmidt and Taylor in fact DO show MDMA is neurotoxic when MDMA is applied ICV at a cumulative dose of 2 or 1mg/kg for 1 hour in rats, as defined by TPH activity (but not 5-HT at 1mg/kg). Paris and Cunningham on the other hand injected MDMA into the Raphe, which is hurt when MDMA is systemically applied, so that doesn't matter in my opinion.

So I looked what that Monks et al., paper cited as their evidence. Well they cited one paper by O'Shea which I could see had anything to do with anything. The other paper though, was VERY interesting though. I have it here so everyone who is interested can look at it too.

This extremely well executed paper has one major flaw, at least from my calculations. But lets start at the beginning. Firstly, the measure the brain concentrations of MDMA following peripheral administration of a neurotoxic dose (fig 1). Then they make sure that when they directly apply MDMA, it both produces the necessary concentrations, distances (fig 2) and maintained it (fig 3). They made sure the MDMA released serotonin (fig 4) and dopamine (fig 6) like it should. Then they looked at neurotoxicity.

Table 1 and table 2 basically sum it up. On first look everything looks pretty normal, like they say in the results: No differences. But look at little closer here folks. First thing you notice is that there is a big difference between the right and left. The Right is where they injected the MDMA, and left is the control. Okay, so maybe I can believe that serotonin is naturally that asymmetrical (I'm still checking that one out, but initial papers I've read say no, they are normally symmetrical)... I did my own two way ANOVA on the results, and yes, there is a significant difference between the sides, which they don't report, but no difference caused by the treatment (p=0.877).

But table 2! They so no change. Look at it yourself, there is blatantly a difference between the sides. But according to the two way ANOVA I did, there is JUST a significant difference between the normothermic and blanket group (p=0.0407) (Even a two tailed non-paired t-test of right-normothermic vs right-blanket gives a p=0.0012).

Now I don't know what to make of all of this. But it seems to me this is a paper reporting that direct application of MDMA IS neurotoxic, so long as the animal is made hyperthermic. Either way, I've seen papers reporting neurotoxicity to cultured neurons from MDMA, at similar concentrations too.

Basically what I'm getting as is, does anyone know what's going on with that paper, and does anyone know any better evidence on the "toxic metabolite theory"


* When I say neurotoxic, I'm generally talking about lowered 5-HT, 5-HIAA, and SERT. I apologize if you object to this usage.
 
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* When I say neurotoxic, I'm generally talking about lowered 5-HT, 5-HIAA, and SERT. I apologize if you object to this usage.
good that you mention it, when "I" say neurotoxic I mean when a certain drug does irreversible damage to the brain that produce noticeable physical/mental sideeffects


from what I can understand, your clonclusion is that with all the research you have done... someone who has just done mdma has less serotonin, 5-HT & 5-HIAA than someone who hasn't done mdma? if that's the case -> old news brother :\

but how does the brain of a person from someone that has done mdma a week to 3 or 6 weeks ago compare to someone's who hasn't done any mdma? that's the question


(i probably sound like an idiot now I know :p )
 
MDMA isn't neurotoxic upon direct application (as I suspect MDA is not as well) becuase it's (in my opinion) 3,4,dihydroxyamphetamine that is the problem. This is (again in my opinion) the single reason that MDA is more toxic than MDMA.

Good luck, and be sure to keep us posted...


X
 
Well then why is MDMA neurotoxic in culture? And as I say, it looks to me like these guys showed that MDMA IS neurotoxic when applied directly, and so do Schmidt and Taylor.

No shakawkaw, my conclusion is (if I have one), that MDMA very well might be neurotoxic when applied directly to the brain.
 
Are you referring to Cell Biol Toxicol. 1999 Jun;15(3):137-42? Is MDMA metabolised in culture?


X
 
No, that paper is shithouse, and all it's really testing is amphetamine medaited internalization of transporters... I was talking about Stumm et al., FASEB J. 1999 13(9):1065-72.

Well 3,4-dihydroxyamphetamine, so thats CYP2D6 produced right? Yeah, it could very well be expressed in neurons in culture (1)... Interesting... Still, if it was, that would mean that MDMA should be neurotoxic if directly applied...

and that culture experiment above did leave the amphetamines in there for 96 hours...
 
I seem to remember reading that animals deficient in CYP2D6 are more susceptible to neurotoxicity?

Colado MI, Green AR "The spin trap reagent alpha-phenyl-N-tert-butyl nitrone prevents 'ecstasy'-induced neurodegeneration of 5-hydroxytryptamine neurones" Eur J Pharmacol, 1995; 280(3):343-6.
It's cited on TheDEA's neurotoxicity page at reference 21
 
Yeah, he old Dark Agouti CYP2D6 poor metaboliser model... that's a double edged sword, because if your 2D6 deficient you're going to end up with a higher plasma concentration.
 
Well 3,4-dihydroxyamphetamine, so thats CYP2D6 produced right?

Yes, I believe that 3,4,DHA is produced through the CYP2D6 pathway.

Yeah, it could very well be expressed in neurons in culture (1)... Interesting... Still, if it was, that would mean that MDMA should be neurotoxic if directly applied...

MDMA -[O-demethylation]-> 3,4,DHMA -[N-demethylation]-> 3,4,DHA

OR

MDMA -[N-demethylation]-> MDA -[O-demethylation]-> 3,4,DHA

There's (at least) 2 steps to 3,4,DHA from MDMA, I'm not sure if both are 2D6 dependant. If MDMA ---> DHMA is not 2D6 dependant but DHMA ---> is then there would be a discrepancy.

Since Deprenyl seems to block the neurotoxic effects, I'm guessing that it's actually not DHA, but a metabolite thereof (and/or hydroxl radicals produced during deamination) that is actually causing the problems.

(PERSONAL THEORY ALERT!) I don't think MDMA is taken up into the axon, I think it sticks to the transporter (and causes it to work backwards, transporters are capable of transporting in either direction depending on concentration gradient). That methylenedixoy bridge should be too bulky to get in to the terminal but remove it and you've got a very DA-like substance that should have very little problem getting in.

We all know that axon terminals can and do release more than one type of NT so it figures that SERT can transport DA into the terminal. So DHA, which looks just like DA except for that alpha-methyl group gets in, gets deaminated by MAO-B and produces a bunch of bad stuff (DOPAL?).

Ok, I've lost myself again. Where were we?


X
 
The N-demethylation is CYP3A4, the the demethylenation is CYP2D6...

But you theory on MDMA mediated release is probably a bit whack. Most theories on amphetamine mediated monoamine release center around phosphorylation of the transporters by PKC, which is a intracellular calcium dependent process... nothing really to do with the gradient as such.
 
I was inferring that the concentration gradient had anything to do with amphetamine or MDMA induced release of monoamines. Only that SERT is normally capable of transporting in either direction, depending on concentraton gradient.


X
 
BilZ0r said:
Yeah, he old Dark Agouti CYP2D6 poor metaboliser model... that's a double edged sword, because if your 2D6 deficient you're going to end up with a higher plasma concentration.

CYP2D6 deficiency reduces the production of ether-ring cleaved metabolites, making the side-effect of greater MDMA levels irrelevant if the question is whether or not a ring-cleaved metabolite is the source of toxicity.

At any rate. We have a pretty solid basis for believing that the mechanism of MDMA neurotoxicity is the production of free radicals during the breakdown of Mystery Chemical X by MAO.

So, what's the chemical? I like MDMA as the culprit; after all, it has a decent affinity for the enzyme and we know it's available in good quantity under almost any conditions. The search for a toxic metabolite is mostly based on the reduced (eliminated?) neurotoxicity of locally injected MDMA into the brain vs. systemic injections, but we now know that this was probably a red herring because local injections would also likely not produce the sort of hyperthermic response of a systemic injection.

That various amphetamine metabolites of MDMA can be demonstrated to be neurotoxic is meaningless; neurotoxicity is a pretty standard quality of amphetamines of any sort.

If a metabolite being attacked by MAO is the source of toxicity, then we must assume that either the attack is on the amine of the amphetamine (in which case MAO can be assumed to also attack the amine of MDA (at least, and probably MDMA as well)...or the MAO/metabolite interaction in question is something else, such as an interaction with the hydroxy/methoxy groups of ether-ring cleaved metabolites. But that still butts heads against the increased (not reduced) neurotoxicity in the CYP2D6 rats. Perhaps we can go for some sort of ring oxidation?

I remain of the opinion that the 'toxic metabolite' school has rather significant weaknesses.
 
I had typed up a huge reply but the more I think about this the more ambiguous everything gets.

Someone please fund a study to compare the effects of Deprenyl + massive dose of MDMA, Deprenyl + massive dose of MDA, massive dose of MDMA alone, massive dose of MDMA alone, large dose of 34HHMA +Deprenyl, large dose of 34HHA + Deprenyl, 34HHMA alone, and 34HHA alone (in non-human primates of course). Measure the metabolites. Take PETs before and after.


X
 
TheDEA.org said:
CYP2D6 deficiency reduces the production of ether-ring cleaved metabolites, making the side-effect of greater MDMA levels irrelevant if the question is whether or not a ring-cleaved metabolite is the source of toxicity.

Not necessarily in my mind. Lower active CYP2D6 --> Higher Peak MDMA, Higher MDMA AUC --> More hyperthermic, also probably lower peak CYP2D6 metabolite, but possibley higher metabolite AUC


The search for a toxic metabolite is mostly based on the reduced (eliminated?) neurotoxicity of locally injected MDMA into the brain vs. systemic injections, but we now know that this was probably a red herring because local injections would also likely not produce the sort of hyperthermic response of a systemic injection.

Yeah, but in the study I started this on, they used a blanket to get the animals temperature up by around 2ºC.

That various amphetamine metabolites of MDMA can be demonstrated to be neurotoxic is meaningless; neurotoxicity is a pretty standard quality of amphetamines of any sort.

Possibley, but some of the alpha-dopmaine-glucuronides are hardly amphetamines any more

If a metabolite being attacked by MAO is the source of toxicity, then we must assume that either the attack is on the amine of the amphetamine (in which case MAO can be assumed to also attack the amine of MDA (at least, and probably MDMA as well)...or the MAO/metabolite interaction in question is something else, such as an interaction with the hydroxy/methoxy groups of ether-ring cleaved metabolites. But that still butts heads against the increased (not reduced) neurotoxicity in the CYP2D6 rats. Perhaps we can go for some sort of ring oxidation?

I remain of the opinion that the 'toxic metabolite' school has rather significant weaknesses.

Well as I said before, possibley both the increased AUC as well as increased hyperthermia... and I don't see why it "butts heads", if there was less CYP2D6, there would be more substrate for MAO..
 
..also, do we know if dark aguoti rats are poor metabolizers because of low expression of a relatively normal cyp2D6, or becasue the CYP2D6 is actually bad? I know you'd think it was the later, but It might not be...

Hmmm... Look at:
http://www.ncbi.nlm.nih.gov/entrez/...ve&db=pubmed&dopt=Abstract&list_uids=10471068

So It might be low level of expression... which could mean that CYP2D6 might be rate limiting to a degree, at least in the brain. So if we were to hypothesize that MDMA toxicity was due to metabolism in the brain, then the rat being a poor metaboliser might not alter the rate of MDMA-->toxic stuff, all it would later is that amount of MDMA in the plasma and the amount of hyperthermia.
 
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Interestingly this paper found a decrease in neurotoxicity in Dark Agouti Rats...

and I went back and looked at the original female DA rats as a CYP2D poor metaboliser model paper
Colado MI, Williams JL, Green AR.
The hyperthermic and neurotoxic effects of 'Ecstasy' (MDMA) and 3,4 methylenedioxyamphetamine (MDA) in the Dark Agouti (DA) rat, a model of the CYP2D6 poor metabolizer phenotype.
Br J Pharmacol. 1995 Aug;115(7):1281-9.

and it doesn't show that CYP2D6 metabolism effects toxicity one way or the other. (Well it hints that low CYP2D6 make for more toxicity, but doesn't get significance)
 
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