so i'm gonna give a tiny bit of pharmacology because i don't know how many people here are pharmacologists
maybe a lot maybe not a lot
how did we study psychedelics without using humans
when I was in a purude university
worked with receptor targets, most of you know that, but I talk about it briefly
and when I was doing this talk I had my talk finished
i thought, you know there is nobody in this conference that talk about research chemicals
and large number of people these days are getting their first exposure to a psychedelic or an enctatogen
through a research chemical
so what I did was I changed the whole back in my presentation to talk a little bit about research chemicals
because lot of people don't know what they are
sadly a large number of them was pilfered from my publications
so i feel a little responsible for the fact they're out there
and then I should talk a little bit about if you want to be a researcher, what should you do and that's part of the CME requirement
so inform patients about new research, refer patients to clinical studies, develop strategies for conducting research and evaluating research
I'm gonna touch on some of these, but obviously this talk like that would be a 2 hour talk in depth
so I won't spend a lot of time, i do want to point out that the sort of revolution in neuroscience
was catalysed by the discovery of LSD and then few years later the finding that serotonin was in the brain]
and I think somebody earlier pointed that out, maybe it was Rick, that
prior to discovery of LSD and serotonin in the brain, psychiatric disorders were thought to be due to poor parenting
so we had mothers that were called refrigerator mothers, because they had a child with schizophrenia and they would say that's the mothers fault she didn't nurture the child properly
so there's no connection, so whe they discovered that
LSD had effects and serotonin was in the brain and the recognition that
there was a similarity in the skeletal fragment of LSD and serotonin
a number of people put together the idea well maybe behaviour and mental function is somehow related to neurochemistry
and that seem sort of silly today because it's obvious thing but back then they thought that brain was sort of an electrical box
nothing about neurochemistry had anything to do with behaviour
kind of weird thinking, but that's the way it was
and another thing to point out about psychedelics, and I used to used this definition a lot
because you try to tell people what psychedelics were
and Jaffe wrote a chapter in Goodman and Gilman which was a pharmacologist Bible
and I like this definition because it says that psychedelic
the feature that distinguishes them from other classes of drugs is their capacity reliably to induce or compel
states of altered perception thought and feeling that are not or can not
be experienced otherwise except in dreams or at times of religious exultation
I often give the seminar to pharmacology departments at medical schools
where i give this definition and I say
do you know of any other pharmacological class that has a definition like that?
a drug that produces religious experience? and so, very unique class
and i'm preaching to the choir you all know that but just to mention that
before we start
this is a list of what I'll call recently published
clinical studies, that have come out, and i probably dont have everything
listed here that's been published but the point
of the slide is, what i want you to notice is the colouring
so these purple ones ate the 1990s
these red ones are 2000-2010
and then after 2010, what you see is exponential explosion
of published studies that relate to clinical research
on psychedelics. And i think this trend is likely to continue
so this is really exciting trend and it really obvious when you look at these numbers
the literature in this field is exploding, so we're having an impact
the key brain receptors
for psychedelic are the serotonin 2a and serotonin 2c receptors
and for tryptamines they also activate the 1a receptor
LSD is a little bit different
somebody have a question?
LSDis a little bit different because it hits lots of different receptors
and people used to say that LSD was a promiscuous drug pharmacologicaly
and then they switched over
and they started sayin it had rich pharmacology
butfrom the point of view of understanding how they work if all these drugs
these types of compounds are considered to have a similar action in the brain
it pretty obvious that the serotonin 2a and 2c
receptors are probably the ones that are the most important
things like DOM, DOB, 2C-B, the kind of things you hear about
a lot on the streetm activate principally the serotonin
2a receptor, they may not fully activate it but they have a high
efficacy so they produce a good activation of the receptor
we dont know much about the serotonin 1a receptor
it may be important in distinguishing the effects of the tryptamines from phenethylamines
and with lsd its uniquely potent so its doing more than just activating these 2 receptots
and that's the quest i've been on for a long time
and will continue work on, to try to understand why LSDhas such profound effects
when if you look at its receptor effects at the 2a 2c or even 1a receptor
there is no evidence that it should be as potent in humans as it actually is
so it has unique pharmacology
so what happens inside the brain cell when we activate these receptors?
this is just a model of a brain synapse
but principaly these serotonin 2a receptors are located postsynaptically
so we have release of serotonin in a normal brain
it comes out of these vesicles and interacts with these receptors
so LSD or the other psychedelics will bind to these receptors and will induce a signalling in this receptor
or within the cell
and they can induce signalling in number of ways
they can either fully activate the receptor
that what serotonin does, lsd doesn't fully activate it
its actually fairly weak agonist, so if we look at LSD on this scheme it would probably be over here
about 30% of full activation
so not only does LSD not have the kind of receptor affinity or stick-to-it-iveness
we might expect from such a potent compound but it also doesn't activate the receptor fully
so its maybe related to its mechanism of action
and an antagonist which dont do anything, BOL as you just heard in a previous talk
its currently thought to be either antagonist or inverse agonist, an inverse agonist
actually drops the basal level below this
so BOL would fit into the scheme there
what happends when a psychedelic binds to the receptor?
these receptors are bundles of alpha-helical protein pieces
that are fit together in the membrane of the cell
and when the drug interacts with the exterior part of this receptor,
it binds inside inside an area that's in between what is called helices 3, 4, 5, 6 and 7 to some extend
it causes the shape of this protein bundle to change
and when the shape of that bundle changes, there are pieces of the protein that stick down into the cell
and they also change, so as the receptor changes, the interor pieces of the protein of the receptor also change
and when those pieces change they allow it to couple to signalling molecules within the cell
Now years ago pharmacologists used to talk about a drug that was an agonist
that would activate the receptor
or not activate it. And its alwas thought that the receptor
was sort of like a switch, you turn it on and turn it off and signal is produced.
Now today there is a concept in pharmacology that is called functional selectivity
which means that all drugs don't do the same thing to the receptor
so imagine serotonin, which is the natural transmitter
binding down inside the receptor
it causes the shape of the receptor to change in a specific way
and now we have an ensemble, the serotonin inside the receptor
and that ensemble produces a change in a pieces of the receptor that are inside the cell that allow it to couple
to some set of signalling molecules
if we put LSD in there
or 2C-B or you name your favourite drug
when the receptor binds it doesn't go to the same shape
that it had when the serotonin bound, because it's a different molecule
this new receptor shape induces new changes
in the interior part of the receptor and it no longer binds
the same way to the same set of signalling molecules that serotonin bound to
So we have a possibility of coupling to a lot of different things
so we have what we call G proteins, those are what
normally couple to G protein coupled receptors (GCR)
G alpha I which generally inhibits cellular activity
G alpha s which typically stimulates cellular activity
theres another series of G proteins called G alpha q that I dont have shown here
that typically also activate the cell
it can also be phosphorylated with an enzyme
that put phosphate groups on to serines inside these pieces of the receptor
and that can allow them to interact with another signalling molecule
called beta arrestin, so there are number
of these signalling pathways that can be activated
so when a drug binds to the receptor and activates it, it just doesn't turn it on and off, like light switch
it turns it on and it induces certain amount of signals inside the cell
a different drug produces a different set of signals
and yet a third drug will produce even a different set of signals
so we have the complexity of understanding how psychedelics work, because
they don't just turn the receptor on and off
they induce a whole bunch of different kinds of changes
and these changes are related to
the structure of the drug itself
So how does receptor signal change consciousness?
so this is when the hand-waving comes in
Franz Vollenvaider will give a talk i think tomorrow
and Robin Carhart-Harris will also talk about
some of the newer technologies involved in looking at how the brain
actually responds to psilocybin or to other psychoactive drugs
the kinds of electrical changes as measured by EEG
or glucose utilization as measure by PET
and a number of other techniques
so what happens?
well basically, we don't know clearly
but one of the things we know that the serotonin
2a receptor, which is the esential receptor
for producing the effects of psychedelics, is highly expresed in
all of the structures in the brain that are involved in perception
and counsciousness.
So if we look at the top of the brainstem lower midbrain we have a locus coeruleus
which sends norepinephrine projections to the whole forebrain
we have the raphe nuclei, the dorsal rapahe is also in the same area as the brain stem
that sends serotonin projections to the whole forebrain,
we have the ventral tegmental area which also have serotonin 2a receptors
we have serotonin 2a receptors both in thalamus
but particularly in the reticular nucleus and the reticular nucleus wraps around the thalamus
so the thalamus has the nuclei that are involved in processing all the senses
so auditory, vision, sensing, touch
all of that is processed through the thalamus
and those signals are sent onto the cortex for final sort of procesing and integration
while the reticular nucleus wraps around the thalamus as a single layer
and sends inhibitory projections down into the thalamus
and tells the thalamus what kind of information it should let through
so we serotoning 2a receptors there, and we have serotonin 2a receptors located on the apical dendrites
of these cortical cells which are the main processing units in cortex
the cortex is where we put together our picture of the world
its where we make our executive decisions, where we do abstract thinking
so the cortex is critical to the funtion of higher conciousness
it's critical to defining who we are
it's critical to defining whether we're conscious, unconscious
and its activity is determined by all these other areas
so any sensory information that comes in, is filtered through
the thalamus, that's regulated by reticular nucleus
we have the Raphe sending signals to the cortex
to the apical dendrites, we have a locus corelueus producing norepinephrine
there are alpha receptors responding to norepinephine located on these pyramidal cells
and the ventral tegmental area produces dopamine and also produces activation
so the confluence of all these things onto these senitive processing units
will change the way we perceive things
changes the whole process of sensory, perception, and consciousness
so it's not surprising that something that activate serotonin 2a receptors
or interacts with 2a receptors will change the way we see
and experience, because thats a fundamental receptor in the brain in all these areas
the serotonin 2a receptors and serotonin 2 receptor, its progenitor
is probably one of the oldest receptors
there's known. Serotonin as a transmitter is thought to be phylogenetically
probably the oldest of the monoamine transmitters
we consider serotonin, dopamine and epinephrine
because you find serotonin 2 receptors even in single cell organisms like
paramecium, where is responsible for the movement of the cells
so you know when nature finds something that works, like serotonin 2 receptor
it continues to use it and exploit it in ways, and as it turns out exploited it in modulating our perception and conciousness
one of the problems I had in Purdue university
working in the pharmacy school in the midwest
i had a lot of students who would come and volunteer to be a subject in my experiments
and i would say well i can't really do that
i can't even talk about anything i may or may not have done
i plead the 5th amendment
so how did we actually do our studies?
we spend a lot of time looking at animal models
trying to find something that would parallel the activity in animals that we saw in humans
no animal can model what happens is humans though
theres no animal model that can tell you i had a mystical experience
or i remembered my childhood
or anything like that, it's a real problem.
turns out that in pharmacology
the whole subject of pharmacology is the study of the action of drugs
and if you want to understand how the system in the body works
you apply the drug that modifies that system
so for example we know that they are beta 1 adrenergic receptors in the heart
so if we want to know what beta 1 receptors are doing
we can give the organism a drug that stimulates beta 1 receptors
and if we do we find out that beta 1 receptors control the heart rate
give a beta 1 agonist to human their heart will start beating faster
in a fight or flight response if you hear
a noise at night,
you walking down a dark street
and you hear someone say: Hey hippie, what are you doing down here?
what happens: your heart start beating, you start going [quick breathing]; that's epinephrine produced by adrenals
and it stimulates beta 1 receptors in the heart
so similarly we can argue to study consciousnesses
we can use agents that perturbs consciousnesses, so, psychedelics
they're the best drugs to look at consciousness and how we can perturb it
I like to use this slide, but I don't hear the sound that goes with it
so if you could hear this you would hear Grace Slick singing The White Rabbit, right
anyway, the ones that mother gives you don't do anything at all, but the ones Sasha gives you would probably have some effect
we use the techinque called drug discrimination, the two-lever drug discrimination
and it's actually a pretty good model for what these drugs do
we have an operant chamber that has two levers in the front
and we can train the rats to press one lever or another
so if we give a rat a drug, LSD in this case, what we can do is turn on the right lever
and this is a trough that delivers 50mg sucrose pellets, so it's rat candy
so we put the rat in the box, we turn this level on, and he'll explore the box, and learn that if he presses this lever he gets a food pellet
if he presses the left lever, nothing happens, we turn if off
the next day we put the rat in, we turn on the left lever, and we give him nothing, placebo
and he presses the right lever, nothing happens, he presses the left lever he gets food pellets
the next day we give him LSD, we put him back in, we turn on the right lever, we give him food pellets if he presses the right lever
it takes 2-3 months to train rats to do this, but once you do, the rat will recognise very reliably the effects of a drug like LSD
and when you train rats and they're really well trained they'll press to get a good pellet anywhere from 1500 to 2500 times in a 15 minute training session
so they're really pressing that lever
and the way it works is, there's something unique about this called the third state hypothesis
if you now give the rat some other drug, like amphetamine or MDMA, and he's been trained with LSD, he won't respond on the LSD lever, he'll respond on the non-drug lever
so they uniquely recognise the drug they were trained with
so if we give this rat a drug that we've made that we think might have activity like LSD, and we've trained them on the right lever
he presses the right lever, he's telling us "I think you gave me LSD"
and if he doesn't press it, he presses the left lever, "I don't think you gave it to me"
so we always tested our compounds after we started using this in about 1984
we've tested our compounds with drug discrimination, and it actually is a fairly good assay
this is just a correlation that I put together some years ago where we look at the relative potency compared with LSD in rats
and relative potency compared to LSD in humans
and the numbers aren't exact but you see here the ETH-LAD compound we made which is more potent than LSD in humans
it's also more potent in rats
we have a relative potency of about 185 which is exaggerated compared to its actualy potency which is around 140 times LSD
LSD is 100, DOB 2.37, DOI 9.26, psilocin 2.61, MDMA 2.4, mescaline .06
so there's a good general correlation
this assay works well, but will sometimes give you false positive, so it will sometimes tell you something is active when in humans it may not be active
but it was a mainstay of all the biological work in animals that we did and it served us well
now I'm gonna talk about a compound that we worked with some time ago that some of you may of heard of called 25I-NBOMe
that's an extremely potent compound, there have actually been a couple of deaths from overdose
this stuff the dose of it is about 500 micrograms, 200-500 micrograms
people have gotten hold of a pure powder, they insufflate it up their nose and die
and I'm just going to tell you how we did a little bit of work with this before it actually came out as a so-called research chemical
a German fellow called Rolf Heim (?) had published that these compounds with an N-benzyl were very potent
and we were interested into whether they would actually have psychedelic-type effects
so here's a compound called 2,5-dimethoxy phenethylamine, it's basically inactive in humans but we used it as a model for early studies
this is the affinity, which is how tightly it sticks to the receptor, so 300nM is not a particularly potent compound
if you put an N-methyl on this compound, you see it gets even worse, you see it gets up to 1900
it's even less effective in binding
add an N-propyl, which is even larger, there's about comparable decreased affinity
so if you put something onto the nitrogen of these phenethylamines, generally it ruins their activity
if you take DOB and you put a methyl on it, almost kills the activity
2C-B, you put a methyl on it, almost kills the activity
by contrast, when you put an N-benzyl on, the affinity goes from 380 to 68 nM
and if you put a methoxy over here, it goes to 2.8nM
so this is really curious; you put stuff onto the nitrogen, the activity goes down, way down
but if you put on a benzyl or a methoxy-benzyl, it goes up
and this compound with an iodo is the 25-I-NBOME, and the affinity in this case 0.04 nM, so it's a super potent compound
so we did a lot of work trying to model the receptor, and our earliest work with an activated model of rhodopsin
which we converted into a serotonin 2a receptor
and this is something in March of 2013, the crystal structure of the human serotonin 2b receptor was published
this shows it with ergotamine bound, which is the way it was crystallised
and we've now converted that into a homology model, this is now the serotonin 2a receptor
and this is LSD bound
so we now have a very good receptor model to go in and look at what these things actually do
so early on, after doing some virtual computer docking with this N-benzyl compound, it appeared to us that the N-benzyl, which is right here
this thing that's in spheres is the N-benzyl compound
and these things that are labelled here are the residues that are in the active site, the binding site of the serotonin 2a receptor
and we docked it in virtually, we saw that this N-benzyl groups appeared to be interacting with phenylalanine 339
as a kind of edge-to-face interaction
so we thought, let's mutate all these residues and see if that assumption if that assumption or that hypothesis is true
so we mutated that phenylalanine to leucine
so phenylalanine is like a benzene ring, it's aromatic
leucine is just a series of aliphatic carbon atoms, you lose the aromatic character which is necessary for these rings to stack if it binds in the receptor
but it still keeps some hydrophobicity
so when we looked at the mutated receptor, where that phenylalanine was mutated, now we put those compounds in, we saw the affinity drop from 380 to 4200
so about a tenfold drop in affinity, about a fivefold drop in affinity and in this case a sixfold drop in affinity
so that didn't tell us anything, but when we looked at the N-benzyl, you see when we did the same mutation we got a 40-fold drop in affinity, and a 500-fold loss in affinity with the methoxy-phenyl
so this was strong was strong evidence that that residue, phenylalanine 339, was interacting with the N-benzyl group
now with our new model now, we're trying to dock these compounds in and get a better idea of how exactly they're fitting into the receptor
one of the other things we did though was to try to discover what shape that molecule might have when it bound to the serotonin 2a receptor
so we made a whole series of compounds, so here's a compound 2C-B, its affinity is about 6nM in a series of experiments, and here's the N-methoxy-benzyl-2C-B, about .2nM
so you can see the increase in potency, 6 down to .2
so we use this is a model because the bromine atom was easier to get on than the iodine, so we made most of our compounds with bromine
and I had a fairly large group of students so I assigned each student one of these molecules
I said let's make all the possible combinations of 2C-B with the N-benzyl group attached
so in each one of these although it's not immediately obvious, they all have the equivalent of an N-benzyl
so here's the 2C-B nitrogen, and here's the N-bencyl
here's the 2C-B nitrogen with the N-benzyl
here's the 2C-B nitrogen and the N-benzyl is over here
here's the N-benzyl down here
this one is a little harder to see, 2C-B is here, here's the N-benzyl
2C-B is here with the N-benzyl
and then this one substituted, this is this so they're on the same side of the ring, and this is trans on the opposite side of the ring
and the affinities for this compound is 2.6nM
we actually crysallised this into its stereoisomers, this is a pure stereoisomer, and this is the one that's most active
so though we don't strike the .2nM affinity, if you look at these, 70, 2000, 160, 170, 300, 800, 800, 2.8
we're pretty sure that that represents the shape that these N-benzyl compounds mainatin when they bind to the receptor
so we're doing docking studies now to see how that fits into the receptor
we get a better understanding of how this super potent compound can activate the receptor
so what I did when I changed my talk was to cut out the back end and talk about research chemicals
and I don't think anybody here at this conference is talking about research chemicals
so I thought that this we should really spend some time on because a lot of people are taking these
they don't know what they are , they don't know where they came from
and a lot of young people their first experience with a psychedelic-type compound is one of these research chemicals
and there are several kinds of research chemicals that we could talk about
the psychedelics, the entheogens, the psychostimulants that are like amphetamine, or entactogens or empathogens
depending on what you prefer
synthetic cannabinoids, and I'll briefly talk about those
and ketamine analogues, there are one or two of those out there, I'm not going to spend time on those
but there is a methoxy-substituted ketamine analogue out there that people seem to be using quite a bit
also I'd point out that this paper just came out, I just got it for review a week ago, showing that lifetime use of psychedelics in the US population in 2010 is over 30 million people
and that includes LSD, psilocybin, mescaline and a lot of other things were not classified
so there's a large number of people that are using these
so psychedlics, or hallucinogens, we know they're characterised by changes in perception, thought, feeling, distortion of space
feelings of portentiousness, something important is about to happen
high doses can elicit anxiety, or psychotic behaviour
generally they have low toxicity and non-addictive
now the peculiar thing about the 25I-NBOMe compounds is they have killed people
there are no overdose deaths from LSD, mescaline, psilocybin
some of these newer so-called research chemicals do have toxic properties that are not fully appreciated
the psychostimulants are related to amphetamine, they produce elation and euphoria, give you energy,
but their chronic use or high doses can produce depression and fatigue, dependence and continued can produce acute psychosis, amphetamine psychosis
and most psychostimulants have cardiovascular effects
they release norepinephrine into the circulation and that causes blood vessel constriction, and that's a major toxic problem of some of the research chemicals
and entactogens also produce elation and euphoria, depending on the dose and the pharmacology
large doses or chronic use can produce depression and fatigue
and high doses can also produce psychotic behaviour
people who repeatedly take large doses over short periods of time most these also have cardiovascular side effects because they release norepinephrine
they cause vasoconstriction
these are different from psychostimulants, the pure amphetamine, and in the beginning when MDMA was brought out and there was a lot of publicity
there were drug abuse experts who said well, MDMA is just another methamphetamine
we all know today that MDMA is not just another methamphetamine
we did a number of experiments in the early days to prove that MDMA was not just another amphetamine or methamphetamine
we also proved it wasn't another psychedelic amphetamine, that it had a specific category
and many of you know that I created this name entactogens so set them off as a specific class of drugs
so they wouldn't be lumped into the hallucinogenic amphetamine class
they are dfferent; I collaborated with a fellow in Germany named Wilfred Dimpfel who planted electrodes in the brains of rats
give them different drugs and then we'd characterise the power spectra
so frontal cortex, hippocampus, striatum and reticular formation
and then separated these into six frequency bins, and could do pattern recognition
now clearly here's amphetamine, LSD and MDA, and although there's some similarities, each one is unique
and so it's possible to show that electically in the rat brain they also were distinct and that ... psychostimulants were different from entactogens and also psychedelics
so most of the research chemicals target receptors or reuptake sites for monoamines
we talk about the phenethylamine type, typically they're interacting with targets for serotonin, dopamine or norepinephrine
and depends on the substituents on the aromatic ring what kind of pharmacology you will actually get
so we had the G-protein coupled recptor and this would include the serotonin 2a receptor showed you earlier
these seven transmembrane helices bound together, they couple to intracellular signalling
dopamine, serotonin, norepinephrine, cannabinoid receptors that are all members of this GPCR family
we have things that interact with reuptake transporters
so after transmitters are released from the nerve terminal, there are specific protein that pump the transmitter back in to conserve it, reuse it
we have reuptake transporters for dopamine, norepinephrine, and serotonin
and things like cocaine block these transporters so they can't transport at all
MDMA and amphetamine actually get inside the substrates for the transporter
once they get inside, they displace the stored dopamine, serotonin, norepinephrine
and it spills out in the reverse direction through the transporter
ketamine and some of the others interact with what is called an ionotropic receptor, a glutamate receptor
and I'm not going to spend any time on those but that's a third kind of target
so the reserch chemicals typically they hit either this or this
the psychostimulants and entactogens they're principally interacting with the reuptake transporters
and the psychedelics and cannabinoids they're principally interacting with a GPCR
a G-protein coupled receptor
in terms of preclinical characterisation, we work with a lot of these in the lab
hallucinogens have affinity for serotonin 2a receptors, and in drug discrimination in LSD-trained rats we get generalisation