Monoamine transporters/activation of multiple neurotransmitter systems - MDMA/LSD: Flashcards
Compare and contrast the serotonin receptor subtypes, their mechanisms of action, and clinical applicability (where possible)
• All 5-HTRs, except 5-HT3Rs, are metabotropic GPCRs
o 5-HT is an important transmitter in the gut and the brain
• 5-HT3Rs are ionotropic receptors
• In humans, 14 genes encode 5-HTRs
o 13 GPCR genes + 1 gene for 5-HT3Rs
o Splice variants massively increase the diversity of functional receptors
5-HTRs have 7 families:
o 5-HT1-7
• Metabotropic 5-HTRs can be grouped by 2nd messenger system; Gq, Gs, Gi/o
Gq/11-coupled receptors:
• This group is made up of the 5-HT2A-CRs
• Activation of Gq/11 –> hydrolysis of membrane phosphoinositides
o –> increased DAG and IP3
o –> activation of PKC and increased i[Ca2+]
• 5-HT2ARs are found mostly on dendrites in several cortical areas and have excitatory effects
• 5-HT2BRs are important during development and are only weakly expressed in the adult brain
• 5-HT2CRs are found in the mesolimbic dopaminergic pathway (VTA) and the amygdala
o Activation reduces dopaminergic neurotransmission
o Antagonists have been proposed to be useful as anxiolytics
Gs-coupled receptors:
• Comprises: 5-HT4,6,7Rs
o Activation of Gs stimulates adenylyl cyclase –> increased cAMP and activation of PKA
• 5-HT4Rs are found in the basal ganglia, cortex, hippocampus, and substantia nigra
o Specific agonists can enhance learning and memory in animal models
• 5-HT6Rs are almost exclusively found in the CNS:
o Expression level is highest in the striatum, nucleus accumbens, and cortex. Also found in the hippocampus, thalamus, and amygdala. Implicated in many cognitive functions
o Reduces cholinergic, glutamatergic, and dopaminergic transmission
• 5-HT7Rs are found in the hippocampus, thalamus, hypothalamus, and cortex
o Functions and effects on neurotransmission are mainly unknown
Gi/o-coupled receptors:
• This group contains the 5-HT1A,B,D-F, 5A-BRs
o Activation of Gi inhibits adenylyl cyclase
o –>decreased cAMP
• 5HT1A is found pre- and post-synaptically; hyperpolarises cells
o May have a role in anxiety and repression
o Increased ACh, NA, DA release
• 5HT1BRs are mostly presynaptic, found on axon terminals
o They modulate neurotransmitter release – decreasing release of: 5-HT, DA, ACh, Glu
• 5HT1ERs aren’t in mouse or rat – function is unknown
• 5HT5A+BRs are found throughout the brain
o No specific agonists/antagonists; function is unknown, but are likely involved in cognition
5-HT3Rs:
• Cation-selective ligand-gated ion channels
o Members of a cys-loop superfamily including nAChRs and GABAARs
o Mediate fast excitatory neurotransmission
• Found in the cortex, hippocampus, and amygdala
o Principally located on inhibitory interneurons
• Probably involved in cognition, but the only current therapeutic use (as a CNS target) is as an antiemetic (anti-sickness drug)
o Clinical trials suggest antagonists could be anxiolytic and useful for treating withdrawal in addiction
Compare and contrast the effects, pharmacology, and potential clinical applications of lysergic acid diethylamide (LSD) and psilocybin
Lysergic acid diethylamide (LSD):
• Originally derived from the cereal fungus, ergot
• Used as a recreational drug from the 1960s onwards
• Effects include:
o Perceptual changes/hallucinations
o Strong emotions (euphoria)
o Spirituality/’connectedness’
o Pupil dilation
o Loss of appetite
o Wakefulness
• Pharmacology:
o Agonist of 5-HT2ARs (Ki = 2.7nM)
o Also agonises (fully or partially) 5-HT1A,B,D,E; 2C; 5A; 6; 7Rs
o Half-life = 175 minutes; effects last 9-12 hours
• Unlike other serotonergic hallucinogens, LSD activates dopamine receptors – particularly D2
• Medical uses?
o Not currently licensed
o Meta-analysis of 6 clinical trials found LSD significantly improved outcomes in alcoholism (Krebs and Johansen, 2012)
Psilocybin: • Found in ‘magic mushrooms’ from the psilocybe family • Similar effects to LSD: o Euphoria o Hallucinations o Altered time perception o ‘connectedness’ • Side effects: o Nausea o Panic attacks • Pharmacology: o Converted to psilocin in first-pass hepatic metabolism o Agonist at 5-HT2ARs o High affinity for 5-HT2B,CRs o Lower affinity for 5-HT1A,BRs o Half-life: ~2 hours, effects last 4-5 hours • Potential clinical use
Clinical trials:
• Ongoing clinical trials:
o Depression – psilocybin
o Depression & anxiety in advanced cancer patients (single dose psilocybin)
o Alcohol addiction (LSD, psilocybin)
o Cocaine and nicotine addiction (psilocybin)
o ADHD (psilocybin)
• LSD and psilocybin share a common mechanism: action at 5-HT receptors
o The 5-HT2AR is necessary for the effects of ‘classic’ hallucinogens – such as LSD, psilocybin, or mescaline (found in the peyote cactus)
Briefly outline the monoamine neurotransmitter family and their transporters
• Include serotonin, dopamine, and norepinephrine (noradrenaline)
• All have similar synthesis pathways
• All have unique transporters
o Transporters are not entirely specific – they will preferentially transport their ‘target cargo’, but will also transport some other monoamines
E.g. – DAT, the dopamine transporter, will preferentially transport dopamine but will also transport serotonin and noradrenaline too
Compare and contrast the monoamine transporters (including vesicular transporters), and how they relate to the action of psychostimulants
• 3 are located presynaptically:
o SERT – serotonin transporter
o DAT – dopamine transporter
o NET – norepinephrine transporter
• Monoamine transporters are not hugely selective for their substrates:
o Dopamine has a higher affinity for NET than for DAT
• Vesicular monoamine transporters load monoamines into synaptic vesicles:
o VMAT1 is found only peripherally
o VMAT2 is found in central and peripheral regions
• Monoamine transporters are involved in the mechanism of action of many psychostimulants:
o MDMA
o Cocaine
o Amphetamine
o Ritalin
Outline the pharmacology, effects, and potential clinical applications of MDMA
• 3,4-methylenedioxymethamphetamine (MDMA)
• AKA ecstasy
• 1912 – first synthesised; not derived from natural substances
• Recreationally used since the 1980s
• Effects include:
o Increased wakefulness and energy
o Euphoria
o Increased empathy, sociability
o Heightened senses
o Bruxism (teeth clenching and grinding)
o Elevated heart rate and blood pressure
o Increased temperature (hyperthermia)
o Addiction
o Anxiety, hyperactivity, insomnia
• Pharmacology:
o Enters neurons via monoamine transporters (affinity: SERT~=NET>DAT); then inactivates VMAT2, and inactivates/reverses SERT/NET/DAT
o MDMA has the lowest efficacy at DAT; roughly equivalent efficacy at SERT and NET
o Weak agonist at 5-HT1+2Rs
o Half life = ~8 hours
• Clinical use?
o Currently not licenced
o Ongoing phase 3 clinical trial for severe PTSD
o Imperial college is launching a study to use MDMA in alcoholism
Compare and contrast the selectivity of cocaine, ritalin, MDMA, amphetamine, and methamphetamine on the SERT, DAT, and NET
These vary in their selectivity for different monoamine transporters:
o Cocaine has roughly equal affinity for SERT, NET, and DAT; but most for DAT
o Ritalin has the most affinity for DAT, and least for SERT
o MDMA is roughly equal for all; most for SERT and least for DAT
o Amphetamine has highest for NET, middling for DAT, lowest for SERT
o Methamphetamine has about equal for NET and DAT, least for SERT
Briefly outline the dopamine receptor families and subtypes
• All dopamine receptors are GPCRs
• 5 receptors in total
o D1-like: D1+5Rs – Gs-coupled; stimulates adenylyl cyclase to increase cAMP and activate PKA. Mainly inhibits postsynaptic activity
o D2-like: D2-4Rs – Gi-coupled; inhibits adenylyl cyclase decrease cAMP; an important target for antipsychotics. Causes pre- and postsynaptic inhibition; will stimulate/inhibit hormone release
Why are monoamine transporters therapeutic targets? Which disorder(s) do they target? Which theory was this based on? What did this lead to?
• Clinical depression:
o The second-largest cause of years lived with disability worldwide (8.2%, Ferrari et al 2013)
o 19.7% of people in the UK aged 16+ showed symptoms of anxiety or depression (Evans et al 2016)
o Monoamine theory of depression:
Review by David Nutt: Slattery et al 2004
First proposed in 1965; focus on norepinephrine then 5-HT
Argues that deficiencies in brain monoamine levels is the cause of clinical depression
• Led to development of SSRIs and SNRIs, as well as monoamine oxidase inhibitors
Compare and contrast SSRIs and SNRIs
SSRIs: • Common SSRIs include: o Citalopram – celexa o Escitalopram – Lexapro o Fluoxetine – Prozac, sarafem first launched in 1987, peak sales were $2.6billlion per annum half-life: 1-3 days (acute use); 4-6 days (chronic use) o Fluvoxamine - luvox o Paroxetine – brisdelle, paxil, pexeva o Sertraline - zoloft All of these have high SERT affinity SNRIs:
Serotonin-norepinephrine reuptake inhibitors
• Common SNRIs:
o Duloxetine – Cymbalta, irenka
o Venlafaxine – Effexor, Effexor XR
• Statistically better than SSRIs, but not clinically better than SSRIs due to SNRI side-effects and ‘adverse events’
What are the main limitations of the monoamine theory of depression?
• Side effects: o Headache and nausea o Sleep disturbances o Dry mouth o Appetite dysfunction o Sexual dysfunction o Hypertension (SNRIs only) o Tachycardia (SNRIs only) • Limitations of monoamine hypothesis: o Antidepressants don’t elevate the mood of non-depressed individuals o Pharmacological changes happen immediately, but antidepressant effects only begin 2-4 weeks after treatment onset
What else are monoamine transporters therapeutic targets for? Which disorders cause abnormal expression of receptors (name 6)
• Inhibitors: o Substance abuse • Treating: o Depression o ADHD o OCD o Anxiety o Chronic pain syndrome o Parkinson’s disease o Narcolepsy o Smoking cessation • Transporters are abnormally expressed in: o Substance abuse o Depression o ADHD o Parkinson’s o Schizophrenia o Bipolar disorder o Hypertension o Anorexia nervosa o Aggression o Alzheimer’s o Anger o Anxiety o Autism o Delirium o OCD o PTSD o Suicide o Panic o Unipolar disorder o TCI/TPQ HA o (Lin et al 2011)
How can we measure drug activity in vitro? What are the advantages and disadvantages of this?
• Can use cell-free extracts (such as liver microsome or free enzymes), or cell-based assays
For example, ligand-binding assay:
o Take a radiolabelled compound
o Wash it through a filter with a membrane-bound receptor
o Measure residual radioactivity
Or a competitive binding assay:
o Measure how much of radiolabelled compound A is displaced by non-labelled compound B
• Other methods:
o Plasma protein binding
o Cytochrome P450 turnover
o Electrophysiology of expressed channels
Advantages of in vitro assays: o Cheap and reproducible o High-throughput o Can study a single protein/receptor in isolation o Doesn’t require animals
Disadvantages of in vitro assays: o Recombinant receptors don’t always behave like native receptors o Can’t measure active metabolites o Limited pharmacokinetic data o Questionable physiological relevance
How can we measure drug activity in vivo? What are the advantages and disadvantages of this?
• Measure the effect of a compound in a whole animal setting
o Can use any measure, from behaviour to biochemical changes
• For example:
o In vivo microdialysis
Insert small probe (catheter) into tissue of interest
• Mimics capillary; consists of shaft with a semipermeable hollow fibre membrane at the tip
Small solutes can cross via diffusion
Advantages of in vivo assays:
o Allow the measurement of the effects of a compound without knowing the mechanism
o You’re able to measure the effects of the drug and it’s active metabolites (for example, psilocybin and psilocin)
o You can measure pharmacokinetics
o More physiological relevance than an in vitro assay
Disadvantages of in vivo assays:
o Uses the whole animal (costs money and has ethical implications lacked by in vitro work)
o Low throughput methods
o Can be difficult to determine the drug’s mechanism of action
o Negative results are difficult to interpret – efficacy vs bioavailability
Compare and contrast the serotonergic, dopaminergic, noradrenergic, and cholinergic projections; which functions/pathways are they involved in?
Dopaminergic projections: • Mesolimbic and nigrostriatal pathways: o From the ventral tegmental area or the substantia nigra, respectively o Involved in: Reward Motivation Executive function Motor control Noradrenergic projections: • Norepinephrine projections arise from the locus coeruleus • Important for vigilance and arousal Cholinergic projections: • Originate in the basal forebrain • Functions include: o Wakefulness o Attention o Learning and memory
Explain the why neurotransmitter co-release is possible, giving examples of the neurotransmitters released
• A single terminal can release more than 1 neurotransmitter
• Many neurons release both a ‘classic’ neurotransmitter, as well as neuropeptides
o E.g. neuropeptide Y (NPY), somatostatin, ATP
• Some neurons release more than 1 classical neurotransmitter:
o ACh and GABA can be co-released, both in retinal and in central neurons
o Some VTA neurons release both GABA and Glutamate
