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Little_Ghost_again
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[*] posted on 17-10-2015 at 06:06
Nicotine and Lobeline


Seeing as both are of the same group with Nicotine seen as the main representative, is it likely that both have almost identical properties on the body?

For example Nicotine is highly addictive, Lobeline seems highly potent but I cant dig up much about it being used in modern medicine. So I guess I am asking if it could replace nicotine and would it be as addictive?

Also what would be the best approach to isolate it?

I have done experiments with Drosophila and alcohol tolerance and would like to see if Lobeline is as addictive as nicotine using fruit flies




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[*] posted on 17-10-2015 at 09:25


That would be an interesting experiment. Nicotine and lobeline have somewhat different pharmacology, so it is difficult to make a direct comparison between the two. I would imagine that lobeline would be addictive to some extent, due to its ability to release dopamine via binding to VMAT2, as well as its activity as a dopamine reuptake inhibitor. Nicotine, I think, increases dopamine levels via a downstream signaling process initiated by the agonistic binding to nicotinic acetylcholine receptors. Lobeline is a mixed agonist/antagonist at these receptors, so maybe this is why it is somewhat comparable to nicotine.

As for isolation of lobeline, all I have to suggest would be to try an acid-base extraction on Lobelia inflata or other lobeline containing plant. Of course, other amines would come through, but maybe the crude extract would be enough for your purposes. Chromatography may help you get a more lobeline-rich fraction.
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Little_Ghost_again
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[*] posted on 17-10-2015 at 11:41


Quote: Originally posted by Crowfjord  
That would be an interesting experiment. Nicotine and lobeline have somewhat different pharmacology, so it is difficult to make a direct comparison between the two. I would imagine that lobeline would be addictive to some extent, due to its ability to release dopamine via binding to VMAT2, as well as its activity as a dopamine reuptake inhibitor. Nicotine, I think, increases dopamine levels via a downstream signaling process initiated by the agonistic binding to nicotinic acetylcholine receptors. Lobeline is a mixed agonist/antagonist at these receptors, so maybe this is why it is somewhat comparable to nicotine.

As for isolation of lobeline, all I have to suggest would be to try an acid-base extraction on Lobelia inflata or other lobeline containing plant. Of course, other amines would come through, but maybe the crude extract would be enough for your purposes. Chromatography may help you get a more lobeline-rich fraction.


Thanks thats really useful, I was hoping Chromatography might be able to separate this. I know it used to be used in pharmacy but not too sure yet for what.

I am actually interested in seeing which of the two is the more addictive or which develops dependence the quickest.
I have the wrong Lobelia but it did give a positive.




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[*] posted on 17-10-2015 at 13:19


How would you test addiction in fruit flies ?



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Little_Ghost_again
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[*] posted on 17-10-2015 at 13:23


with food aga, you put three trays in and see which the feed from most. One has nicotine one has the other and one has just normal fly food. while you breed them and grow then to adult hood in a single container you just feed them one type of the foods, if they are addicted then when released they should mainly feed and lay eggs in the one they were fed on most of the time if that makes sense




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[*] posted on 17-10-2015 at 13:24


badly explained I will try again



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[*] posted on 17-10-2015 at 13:27


you have 4 sets of flies in different containers, you breed several gens in each container and only feed them one type of food, you add the thing like nicotine to only one.
Then after say 3 gens you release adult flies into a main container with 3 food dishes in, then you wait and see which flies feed from which container and which they lay eggs in

How do you tell which flies are which?

simples you use 3 different eye colours ;)




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[*] posted on 17-10-2015 at 14:05


That would require either a really good shooter with a micro paintball gun, or Genetic Differences between the test subjects.

Introducing genetic differences would invailidate the results - they could easily be more attracted to the Lobeline due to their genetic difference.

Eye colour would mean a few Controls need running before the Main Event, so a micro sharp-shooter would take less time.




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[*] posted on 17-10-2015 at 16:40


Quote: Originally posted by aga  
That would require either a really good shooter with a micro paintball gun, or Genetic Differences between the test subjects.

Introducing genetic differences would invailidate the results - they could easily be more attracted to the Lobeline due to their genetic difference.

Eye colour would mean a few Controls need running before the Main Event, so a micro sharp-shooter would take less time.


Eye colour is a genetic difference but should have no effect on the results due the locus of the gene. HOWEVER...

I have been SLATED for extremely poor design! He who knows all was not in a good mood and dismissed my plan as crap, I did however get a few clues.

For a start the eye colour was brought up and the genetic difference which is what aga also said.
If this was the only problem then two simple solutions to it.

do the experiment 3 times and each time swap which each colour gets which food. If the results correlate then this would show I am correct and the genetic difference has no affect on the results, YAY.

Method 2 involves marking with simple dye put on the backs while the flies are put out. PITA and requires filming.

But this apparently is not the only problem/mistake, so having been given a few half hearted tips I will be back with a new plan. I will still do the eye colour because if nothing else this will show if the gene responsible for eye colour also has any effect on addiction, to me thats worth trying.

What I have been told is actually not to feed the flies when ready and leave males and vergin females separate, place normal food in the experiment chamber and the food they have been used to behind a mesh screen they cant reach.

Now I am told to watch then redesign, so I am thinking maybe if they are addicted then they wont lay anywhere near the same numbers of eggs. Apparently if flies are extremely agitated they wont breed.

There is a whole list of other stuff so I need to sleep on this and think it through, I am sure the first couple of runs will be crap but I will get to see how they behave etc. From there I can work out how to make the experiment better.

I am also told that studies using other drugs have been done, so last resort will be to track those down and see what techniques were used. But first I want to design my own experiment for a change.
Yeah ok to some this isnt science, but I want to work it out myself and not copy someones work. I will update this with pics (flies are quick to breed ) until this gets dropped into in the bin.

So as it stands there are some large flaws to iron out, but fuck it I will work this out.

[Edited on 18-10-2015 by Little_Ghost_again]




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[*] posted on 17-10-2015 at 16:50


I also found this

.
Pharmacological Action. Lobeline belongs to a pharmacologically
similar group, which includes nicotine, cytisine, gelsemine, coniine and
sparteine. These alkaloids act chiefly on the central nervous system, the
sympathetic ganglia and the myoneural junctions in voluntary muscle.
Nicotine may be regarded as the typical alkaloid of the group. Lobeline
is a potent, respiratory stimulant and is used in medical practice as an
analeptic,, e.g., in cases of collapse due to poisoning by noxious gases,
narcotics, etc., and there is now an extensive pharmacological and clinical
literature on the drug.19

So there should be papers around on it, apparently it was also used to determine blood circulation times.

I think it also gives me a clue on one of the flaws I was berated over lol.

[Edited on 18-10-2015 by Little_Ghost_again]




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[*] posted on 17-10-2015 at 23:07


Quote: Originally posted by Little_Ghost_again  
For example Nicotine is highly addictive...


Nicotine is not nearly as addicting as you may think. By itself, nicotine actually produces pretty weak reinforcing effects, especially when compared to true "highly-addictive" drugs. What makes smoking cigarettes so addicting is the fact that tobacco smoke contains not only nicotine, but various monoamine oxidase inhibitors (MAOIs), too. These inhibit monoamine oxidase enzymes and prolong the life of monoamines like dopamine, norepinephrine, serotonin etc, significantly increasing the recreational (and thus addictive) effects of nicotine. It's a synergy between nicotine and MAOIs that make smoking as addictive as it is, not so much nicotine itself. This is one of the main reasons that nicotine patches and e-cigs just don't quite "do it" for smokers. No matter how much nicotine they consume, if it's not from an actual cigarette, it is never truly satisfying.

If you're interested, here's a reference demonstrating just how significant the differences are between rats who have been administered nicotine alone, and rats who have been pretreated with MAOIs first:

http://www.jneurosci.org/content/25/38/8593.full

[Edited on 10-18-2015 by Darkstar]
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[*] posted on 18-10-2015 at 00:15


Quote:
do the experiment 3 times and each time swap which each colour gets which food

Sounds like a good plan.

How long is the lifecycle of these flies ?




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[*] posted on 18-10-2015 at 01:53


Quote: Originally posted by aga  
Quote:
do the experiment 3 times and each time swap which each colour gets which food

Sounds like a good plan.

How long is the lifecycle of these flies ?


You can go from egg to ready to breed in around 7-10 days, I normally aim for around 8-9 days and some last 30 days (wild type) I tend to not use flies over around 21 days old.

Still huge flaws to sort out.

If nicotine isnt addictive on its own why do they offer nicotine replacement patches? they contain nicotine and little else.
I know other things are more addictive but my interest is plant extracts and keeping things legal.

Nicotine I can buy pure and I can grow the right Lobelia, for now I will use the L.Officianalis I have and use that.

Flies get addicted quickly to things, making them tolerant is also pretty easy. With alcohol you can get flies that can tolerate alot of alcohol in around 8 generations.




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[*] posted on 18-10-2015 at 02:04


OOO

new info, apparently what might make it hard is the flies that normally eat nicotine mix might be affected if they eat the Lobelia mix. I think this has something to do with the bit I posted above about it being used as an analeptic.

I need to check that out, I thought this would be easy like the alcohol one I did but apparently not! I still have the alcohol tolerant strain but they havnt had a drink in generations lol, I wonder if any of them still have any tolerance?

What i should have done was continue the experiment after I stopped giving alcohol and see how many generations until the tolerance went.
I also tried caffeine but some how I couldnt get it right and the caffeine killed the flies, at the time I did this I didnt have many pheno/geno types of D. melonogasta. I have a few now so might try again, I also have better ways to administer tiny amounts of caffeine, my flies never seemed to want to go near it and when then did 12 hours later it was curtains




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[*] posted on 18-10-2015 at 02:13


Quote: Originally posted by Darkstar  
Quote: Originally posted by Little_Ghost_again  
For example Nicotine is highly addictive...


Nicotine is not nearly as addicting as you may think. By itself, nicotine actually produces pretty weak reinforcing effects, especially when compared to true "highly-addictive" drugs. What makes smoking cigarettes so addicting is the fact that tobacco smoke contains not only nicotine, but various monoamine oxidase inhibitors (MAOIs), too. These inhibit monoamine oxidase enzymes and prolong the life of monoamines like dopamine, norepinephrine, serotonin etc, significantly increasing the recreational (and thus addictive) effects of nicotine. It's a synergy between nicotine and MAOIs that make smoking as addictive as it is, not so much nicotine itself. This is one of the main reasons that nicotine patches and e-cigs just don't quite "do it" for smokers. No matter how much nicotine they consume, if it's not from an actual cigarette, it is never truly satisfying.

If you're interested, here's a reference demonstrating just how significant the differences are between rats who have been administered nicotine alone, and rats who have been pretreated with MAOIs first:

http://www.jneurosci.org/content/25/38/8593.full

[Edited on 10-18-2015 by Darkstar]


I see what your saying but in the first part of the paper it say
"Tobacco addiction remains the most prevalent addiction in the world today, with significant associated pathology and costs to society. Nicotine is the neuroactive compound that is considered to be most responsible for the psychoactive properties of tobacco and the development and maintenance of tobacco addiction (Jaffe and Kanzler, 1979; Stolerman and Jarvis, 1995; Pontieri et al., 1996). "

Nicotine is so contentious with alot of studies contradicting each other, then you have to take into account who has paid for which study.

On the one hand you have Nicotine patches that are used for weaning people off nicotine. On the other hand Drugs like Zyban (sp?) are aimed at the uptake pathways you mentioned.
So to be totally honest its a case of pick a side and stick to it. Not that satisfactory.
For me I am not sure it matters, then maybe it matters alot.
Nicotine and Lobeline and both from the same family, although both see to have different effects at some levels.

I am keeping this as simple as I can, I want to compare how addictive they are compared to each other. i can breed many generations quickly so it wont be long before I know its a bust.




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[*] posted on 21-10-2015 at 05:35


First attempt was a disaster! I am using CG syringes to measure the nicotine and laboeline, I mistakenly used a 10ul syringe in a serial dilution of nicotine instead of the 1ul one I had intended to use! The numbers etc are a bit worn and I cant tell them apart easily!
Anyway the first lot of flies exposed to the nicotine food DIED! In fact they lasted just over an hour so I am going to have to drop the does considerably to what I had intended.




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[*] posted on 21-10-2015 at 09:52


Geee, I'm a bit late with the warning that nicotine was used as an insecticide too... I know you know this, but wanted to tell you that the dilution is very important.

And it is also very important with lobeline too.
How do you know that you have enough a.i. in their food to "catch their interest" but not too much to cause them "bad feelings" and shyness or refusal of the laced food?

Maybe if you used one hundredth (one somethingth) of the Drosophila LD50 dose? The best dose (if there's any) will be surely different with the different compounds.
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[*] posted on 21-10-2015 at 10:27


Quote: Originally posted by Pumukli  
Geee, I'm a bit late with the warning that nicotine was used as an insecticide too... I know you know this, but wanted to tell you that the dilution is very important.

And it is also very important with lobeline too.
How do you know that you have enough a.i. in their food to "catch their interest" but not too much to cause them "bad feelings" and shyness or refusal of the laced food?

Maybe if you used one hundredth (one somethingth) of the Drosophila LD50 dose? The best dose (if there's any) will be surely different with the different compounds.


Each batch of flies are raised on the laced food first, so they dont get a choice of meal. The dose I have no idea what to base it on but I was going to start with 100th of the amount I ended up using!

I have fly cultures going constantly so in another 7 days I will go again with a reduced amount of nicotine, i am using the dose to start with of both chemicals.

These are all things my dad picked up at the start, so its a bit of a learning curve at the moment. I have split my stock flies into more cultures so I can increase the number of breeding flies I have.
At least that way i can get going again quickly until I iron out the wrinkles.

The food is commercial fly food and the only difference being what I add into the laced ones.




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