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Stizolobic and stizolobinic acid-action in mammals?

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Limpet_Chicken

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Oct 13, 2005
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I have been looking into psychoactive Amanita species recently, and want to know more about two compounds found therein in particular, stizolobic acid , and stizolobinic acid, but, seemingly excitatory amnino acid compounds.

The few references I can find suggest that they are competitive antagonists at the AMPA receptor in crayfish, and kainate receptor antagonists in frogs, but last time I checked, I am neither.

http://grande.nal.usda.gov/ibids/index.php?mode2=detail&origin=ibids_references&therow=230866

This seems to suggest, given wikipedias data on kynurenate, which I know little about (yet) seems to suggest that these might be antagonists at the glycine binding site on NMDA, and probably also a convulsant excitotoxin at high doses.

Care to discuss, or shed more light on this one people?

4-(2-amino-2-carboxy-ethyl)-6-oxo-pyran-2-carboxylic acid structure of stizolobic acid (I am unable to find the structure for stizolobinic acid)

Seemingly present in certain varieties of A.Muscaria at low, low levels, and at higher quantities by far in A.Pantherina, which might explain why that one has a reputation for higher toxicity, including memory problems for a while post use?
 
The structure you should know by now young padawan ;)

I'll see tomorrow what else I can find out about these 2 buggers...

YO! Murphy
 
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These molecules do not cross the BBB, beacause of the negative charge on the carboxylic acid... no???
How could they influence our neurochemistry? I don't think it's the cause of the higher toxicity of A.Pantherina.
 
@King Kong:
Please don't forget that ibotenic acid looks quite polar too (it IS in fact quite polar!) but crosses the BBB anyway. So, the structure (and therefore polarity) doesn't tell you always the whole story... Of course, in general I would agree with you.

Murphy
 
Amino acids can be tactively transported via amino-acid transporters!
 
"Excitatory action of a plant extract, stizolobic acid, in the isolated spinal cord of the rat."
Ishida, M.; Shinozaki, H.
Brain Research 1988, 473(1), p.193

Abstract

Actions of stizolobic acid, stizolobinic acid, and their derivs. were examd. on the isolated spinal cord of the newborn rat. The responses were recorded from the ventral root. Stizolobic acid and its bromo derivs. caused a depolarizing response in a concn.-dependent manner. Stizolobinic acid was considerably less potent than stizolobic acid. Depolarizing responses to stizolobic acid and its bromo derivs. were not affected by the existence of Mg2+ and specific N-methyl-D-aspartate (NMDA) antagonists. Kynurenate depressed responses to stizolobic acid. These results suggest that stizolobic acid acts as an excitatory amino acid in mammalian central neurons which binds preferably to other receptors than the NMDA-type receptor.
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"Stizolobic acid on frog spinal cord; Possible species dependent activation of excitatory amino acid receptors."
Maruyama, M.; Takeda, K.
Comparative Biochemistry and Physiology, Part C: Pharmacology, Toxicology & Endocrinology 1993, 104C(3), p.439

Abstract

The authors examd. the effects of stizolobic acid, an amino acid isolated from a plant, Stizolobium hassjoo, on the binding of [3H]glutamic acid and [3H]kainic acid to synaptosomes from frog (Rana catesbeiana) spinal cords and on the depolarization at the ventral roots of frog spinal cords. Stizolobic acid inhibited the binding of [3H]kainic acid more potently than that of [3H]glutamic acid. Among stizolobic acid derivs., 3-Br-stizolobic acid was the most potent inhibitor of the binding of [3H]kainic acid, but the inhibitory potency was 100-fold weaker than that of kainic acid. Stizolobic acid and its derivs. could cause depolarization of the ventral root of frog spinal cord in a dose-dependent manner, and 3-Br-stizolobic acid was a more potent inducer of depolarization than kainic acid, but the dose dependency of 3-Br-stizolobic acid was a little different from that of kainic acid. Evidently, stizolobic acid and its analogs act as kainic acid agonists in frog spinal cord. Stizolobic acid may interact with the different types of excitatory amino acid receptors depending on the species of animals.
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Makes me wonder if it wouldn't have effects similar to acromelic acids, some highly potent kainate type excitotoxins found in certain mushrooms of the genus Clitocybe.

http://www.ncbi.nlm.nih.gov/pubmed/1817797?dopt=Abstract

^makes me wonder just what they patch-clamped, looks like they discovered new kainate receptor subtypes.

http://www.nature.com/bjp/journal/v142/n4/abs/0705834a.html

More detail on acromelic acids, and DAMN are they potent 0.5 pg/kg causing symptoms, sheesh, remind me not to eat any of those mushrooms any time soon!

What measurement is ag ?
 
Limpet_Chicken said:
What measurement is ag ?
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In this context, it's "attogram", I assume. Holy crap! Those compounds are fucking toxic!!!
ACRO-B induced allodynia in a dose-dependent manner from 50 pg/kg to 50 ng/kg
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Geeeee...
The comparison with the "stizos" is not so far-fetched. Just look at the acromelic acids:


Murphy
 
Hehe, if one thing, that gives me something to quote the next time anybody tells me Amanita Phalloides/A.Virosa/A.Ocreata etc contain the most toxic substance in any mushroom=D

Attogram, thats what I thought, but part of me thought something along the lines of 'bugger me, that has to be wrong', that has to be tricky to even measure accurately, wonder how they did it.
 
Yo, I thought exactly the same thing... that is next to nothing! 50 ag/kg?! I must be wrong, seriously. This is hardly believable...
 
How WOULD one go about weighing a compound in that miniscule a quantity, sheesh, the merest dust particle in the air could foul up measurements big time.

Or perhaps using something like an antibody and protein immunofluorescence, or radioactive tracer methods of some kind?
 
The answer is: You can't. Most probably, you weigh out an amount that is easy to handle and then dilute in several steps...
 
But, if one cannot weigh to suitable accuracy, how can one be sure of the dilution one is using?

Riddle me that one =D
 
...ummm, I don't get your point, sorry.
What I described in short is the normal procedure for preparing extremely low concentrated solutions: You weigh out e.g. 1 g of your compound. That can be done with good accuracy, no problem.
Then you dilute, lets say 1:100, and this again 1:100 and so on... Of course, you have to take the error of your equipment into consideration but that's not problematic, too. The error is negligible. Use accurate pipettes and stuff and you're done...

Murphy

P.S. Or did I understand something in your post wrong?! Sometimes english confuses me...
 
Well the issues is that your measuring 1g could be way, way off when you dilute it down to that sort of level. There's no way to have it that accurate.
 
(my last post on that issue)

I have to insist: The correct term would be (I hope so): "Dilution series". That's how it's done.

Example: To my knowledge nobody weighs out his dosage of LSD, as there are hardly any scales for common folks that could handle µg-levels. Instead, one takes an amount that can be measured exact enough and dilutes this until the final concentration is reached. Done.
Diluting further will lead to down to nM, pM, fM and even aM levels.

Take for example an error limit of your scale of 100 µg: That is just 0,01% of your 1 gram! Therefore, the error is negligible. Diluting now in 1:1000-steps will yield the desired concentration. Perform everything in triple (minimum!) to ensure correctness of results.

There's no reason IMHO why this shouldn't work.
 
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My point was just that with an error limit of 100ug, you have an error limit of 10^14 attograms, which sort of precludes an accurate measurement, no? If I'm wrong- which should always be considered a strong possibility, just feel free to slap me.
 
I agree Hammy, I was going to say more or less the same thing, but I can't do fractions, at least, only (very) basic ones, so I couldn't post the math to back it up.

I don't suppose you could explain the '^' notation could ya? I didn't get the whole powers thing in school, fucking dyscalculia, its a pain in the bollocks.
 
Bah! You guys drive me crazy. ;) Therefore, one more post (although the previous one was supposed to be the last :D ).

- Weigh out 1 g. If the scale got an accurary of 0.1 mg (100 µg), the error will be only 0.01 %. That's nothing. Negligible.
- Dissolve this in 1000 ml of your solvent of choice.
- Take from this solution 1 ml and fill up to 1000 ml. This was the first dilution step (1:1000).
- Repeat this as long as you have reached your final concentration.

Every step got its own error, due the accuracy of used equipment (pipettes and volumetric flasks), which have to be taken into consideration. But these are relative errors and not absolute ones! Of course, one can not measure out weights in the nano-, pico-, femto- or even attogram-range. No way! Therefore: Dilution series!

How do you think are nM-concentration prepared? nM is a common for bioassays of all kind (enzyme- or receptor-studies). Nobody weighs out the amount needed and dissolves it in the appropriate volume. The error would be far too big and the results get irreproducible. Nope - they do a dilution series!
An even more extreme example: Homeopathic preparations. These are often far below nM and for this reason get produced by the same way: Dilution, step by step...

I have no other idea how to explain this. Any native english-speaker like to chime in?

Peace! Murphy

Edit: Even if they deal with bacteria, here are some pics that could help. Just look at the steps: They get from 1:100 to 1:1.000.000...
http://www.sigmaaldrich.com/img/assets/4261/micro_7.gif
http://www.physics.csbsju.edu/stats/serial_dilution.gif
 
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