Endocannabinoids

Question 1

What are endocannabinioids?

Answer 1

• By 1990 researchers had cloned the gene for the
  brain cannabinoid receptor (termed CB1 receptor)
• Endogenous ligands for the cannabinoid
  receptors (endocannabinoids) were first
  identified in 1992

Question 2

Describe CB1 and CB2

Answer 2

CB1 is expressed widely in the CNS
» CB2 is expressed primarily on immune cells
‒ Only microglia in the CNS

Putative CB3 receptor for CBD recently

Question 3

Describe CB1 receptors

Answer 3

• Inhibitory through effects on adenylate
  cyclase (↓ cAMP), GIRK (↑ K+ efflux),
  and Ca2+-channels (↓ Ca2+ influx)
• CB1 receptors are exclusively
  expressed presynaptically

Question 4

Deescribe endocannabinoids

Answer 4

THC is a specific partial agonist of the CB receptors.

Anandamide was first identified in 1992 and named
from the Sanskrit word ananda, meaning ‘bliss’.

2-AG was identified in 1994-5. These two
endocannabinoids are the best described in the
CNS.

Question 5

Describe endocannbinoid synthesis

Answer 5

Endocannabinoids are formed from
arachidonoyl-containing phospholipids
(arachidonic acid is an essential ω6 fatty acid
found in peanut oil).

2-AG is synthesized in activity dependent
processes (such as Gq signalling to
phospholipase C - PLC) from the hydrolysis of
diacylglycerol (DAG) by DAG lipase (DAGL).

Question 5

Describe retrograde signalling

Answer 5

• Endocannabinoids do not function as classic
  neurotransmitters
• Endocannabinoids are capable of free diffusion
  through membranes
• They cannot be packaged into secretory vesicles
• Primary function is retrograde signalling to
  modulate presynaptic neurotransmitter release
• Release from postsynaptic compartments is
  through activity-dependent synthesis
  mechanisms

Question 6

Describe depolarization induced suppression of inhibition

Answer 6

• Post-synaptic depolarization can
  activate endocannabinoid synthesis
• Endocannabinoids are released and
  supress presynaptic GABA release

Question 7

Describe metabolism of endocannabinioids

Answer 7

• Two key enzymes are responsible for
  the breakdown of endocannabinoids
• Fatty-acid amide hydrolase (FAAH)
  metabolizes anandamide
• Monoacylglycerol lipase (MAGL)
  metabolizes 2-AG

Question 8

Describe endocannabinoid breakdown

Answer 8

• FAAH
• A common genetic variant in FAAH (P129T) decreases the rate of breakdown of anandamide (~20% of
  North Americans)
• Increased levels of anandamide throughout the brain
• FAAH P129T variant is associated with a greatly reduced risk of anxiety disorder and PTSD
• FAAH P129T mice show decreased anxiety in tests such as the elevated plus maze and more rapid
  extinction of conditioned fear responses
• Anxiety ↓ and fear extinction ↑ in humans carrying the mutation
• FAAH P129T is also overrepresented in problem users of street drugs, suggesting increased risk of
  drug and alcohol abuse
• BUT associates with decreased craving and withdrawal in cannabis users

Question 9

Describe stress and anxiety

Answer 9

• Endocannabinoids are highly involved in regulation of the physiological response to
  psychological stressors
• Acute stress activates two distinct pathways:
• Neuronal response – activation of the sympathetic nervous system via descending pathways from
  hypothalamus
• Neuroendocrine response – activation of the hypothalamic-pituitary-adrenal axis (HPA axis) leading
  to release of glucocorticoid hormones
• In many systems anandamide functions as a tonic (steady-state) regulator of activity
  while 2-AG functions as a phasic (on demand) regulator

Question 10

Describe HPA axis regulation

Answer 10

Hypothalamic release of CRH leads to pituitary
release of ACTH into circulation, and stimulates
adrenal release of cortisol.

Negative feedback is provided by cortisol to
the hypothalamus to attenuate stress signalling
– this feedback process may be impaired in
depression and anxiety disorders.

The limbic system provides a number of
modulatory inputs to the hypothalamus that are
critical in HPA response to psychological
stressors.

Question 11

describe limbic control of HPA axis

Answer 11

Under steady-state conditions
CB1 antagonism results in an
increase in glucocorticoid release

Direct microinjection into the
basolateral amygdala (BLA)
increases glucocorticoid release

Question 12

Describe anandamine’s role in stress response

Answer 12

• Under conditions of stress
  anandamide levels in the BLA drop as
  a result of FAAH induction.
• Decreased anandamide levels release
  the inhibitory tone leading to
  excitatory output to the PVN.
• CB1 agonists administered to the BLA
  supress stress-induced activation of
  the HPA axis.

Question 13

Describe negative feedback

Answer 13

Glucocorticoids can act directly on the
hypothalamus or pituitary to provide
negative feedback preventing excess
glucocorticoid release

1. Rapid feedback in the PVN can be
   blocked by microinjection of CB1
   antagonists

2-AG levels increase resulting in inhibition
of GABA interneurons
Cortical output to the BNST is disinhibited
Net increase in inhibitory output from the
BNST to the PVN attenuates stress response

Question 14

Describe endocannabinoids in stress response

Answer 14

• AEA inhibits amygdala activation
• FAAH induction releases AEA
  inhibitory tone on stress
• 2-AG synthesis in the PVN contributes to
  rapid negative feedback
• • 2-AG synthesis in the PFC and HC
    contribute to slow negative feedback

Question 15

Describe endocannabinoids in appetite

Answer 15

• CB-1 is proposed to function downstream of
  ghrelin to initiate orexigenic effects in the PVN
• GHSR signalling (via Gq, PLC, PKC) induces
  DAGL activity
• 2-AG synthesis increases and signals to
  presynaptic CB-1 receptors
• CB-1 antagonists can block the orexigenic effects
  of ghrelin
• CB-1 agonism notably increases appetite,
  increases craving for highly rewarding foods, and
  increases hedonic value of food
• ‘Munchies’

Question 16

Describe cannabidiol

Answer 16

• CBD has a very low affinity for CB receptor
• CBD is a weak CB1 antagonist and a CB2 inverse
  agonist but may act indirectly
• CBD paradoxically potentiates the effects of THC
  at the CB1 receptor by unknown mechanisms
• CBD is also a 5-HT1A receptor agonist
• Contribute to antidepressant and anxiolytic effects
• Possible inhibitor of FAAH

Question 17

Describe therapeutic effects of CBD

Answer 17

High CBD cannabis is being explored as an anti-convulsant for treatment resistant epilepsy
(such as Dravet syndrome)

• CBD has marked antipsychotic effects in animal models of Schizophrenia
• CBD is a potent antioxidant and has been shown to be neuroprotective in models of ischemic
  stroke
• Selective breeding of cannabis for recreational use has resulted in current strains being high-
  THC, low-CBD– this trend is being reversed by current medical marijuana producers

Question 18

Describe CB1 psychotherapeutics

Answer 18

• Rimonabant– synthetic CB1 antagonist – orexilytic
• Nabilone– synthetic THC analogue (PO) – antiemetic,
  adjunct analgesic for neuropathic pain
• Dronabinol (THC, PO) – antiemetic, orexigenic
• Naboximol/Sativex (THC/CBD ~ 1:1) – mouth spray –
  MS symptoms incl. spasticity, neuropathic pain

Question 19

Describe FAAH inhibitors

Answer 19

FAAH inhibitors (experimental)
* Bial– BIA 10-2474 – serious adverse events in a
2016 phase I trial left 1 participant dead and 4 with
permanent neurological impairments –
determined likely due to poor drug and trial
design (low affinity, multiple high doses, and
bioaccumulation of the drug)

• Janssen– JNJ-42165279 – phase II trials for
  anxiety/depression suspended after Bial’s adverse
  events
• Sanofi– SSR-411298 – failed in a trial for
  depression, ongoing trials for cancer pain
• Vernalis– V-158866 – phase I trials for neuropathic pain

Question 20

Describe cannabis tolerance and dependence

Answer 20

• In animals, THC administration results in
  decreased CB1 receptor expression
• Many suggest psychological, not physiological
  dependence drives addictions (e.g. behavioural
  tolerance rather than pharmacodynamic tolerance)

Question 21

Describe self-administration in animal models

Answer 21

• THC administration often produces an
  aversive stimulus initially in animals
• THC self-administration has been
  demonstrated in squirrel moneys
  first trained on the operant
  procedure with cocaine

Question 22

Describe mesolimbic dopamine

Answer 22

• The mechanisms may be dependent
  on opioid receptors (animal models)
• Naltrexone abolishes THC self-
  administration and blocks NAc
  dopamine release
• Aversive effects of THC
  administration are abolished in κ-
  opioid receptor knockout mice
• However, in human trials naltrexone
  administration increased the
  positive subjective effects of
  inhaled THC administration in
  regular heavy marijuana users

Question 23

Describe cannabis withdrawal

Answer 23

• THC withdrawal can be precipitated in animals
• Administration of CB1 antagonist after period of prolonged
  administration
• Wet-dog shakes, excessive self-grooming, hyperactivity

Question 24

Describe cannabis-related psychosis

Answer 24

* A major concern with the use of cannabis is the occurrence of psychosis * Cannabis use (particularly during adolescence) is associated with an increased risk of psychosis * Alcohol, cannabis, amphetamine, or cocaine use is often a precipitant for the first psychotic episode in schizophrenia * Cannabis use interacts with genetic risk factors for SCZ