The relevance of D2-AKT1 signalling for the pathophysiology

of cannabis-induced psychosis

Edinburgh, 14 April 2016 Dr Marco Colizzi

marco.v.colizzi@kcl.ac.uk

Dopamine dysregulation

-Prenatal and perinatal events -Genes:** NRG1, DTNBP1, COMT, DISC1, DRD2, RM3, ZNF, CNVs

Neurotransmiters’ genes: e.g. **COMT; AKT1

-Drug abuse e.g. cannabis, stimulants

P S Y C H O S I S

Family history of mental illness disorders

Adulthood adversity Migration Urbanicity

HPA-axis dysfunction

Birth events

Subtle motor, cognitive and social deficits

Chronic social adversity in childhood

Social anxiety,

Quasi-psychotic ideas, depression

Developmental cascade towards psychosis

An integrated model of schizophrenia

Howes & Murray, Lancet 2014

Cannabis and psychosis

Questions • Does cannabis trigger an earlier onset of psychosis in predisposed people? • Does cannabis use worsen the psychotic symptoms generally? • Does the cannabis use worsen the outcome and progress of the illness? • Is there a causal link?

Cannabis plant

• A complex plant with over 60 compounds and 400 chemicals.

There are 4 major compounds:

• Delta-9-tetrahydrocannabinol

(THC) * • Cannabidiol (CBD) • Delta-8-tetrahydrocannabinol * • Cannabinol *

*= has psychotogenic effect

THC (Gul et al., 2008)

CBD (Gul et al., 2008)

Cannabis and endocannabinoid system

Tetrahydrocannabinol (THC) Name Part of plant THC %

Marijuana Leaves, small stems 1.0-3-0

Sinsemilla Sterile female flowers 3.0-6.0

Ganja Compressed sterile f.f. 4.0-8.0

Hashish Cannabis resin 10.0-15-0

Skunk Flowers (grown) 5.0-20.0

Cannabis oil Alcoholic extract of resin 20.0-60.0

Cannabidiol (CBD) • No psychotomimetic effects in man

• Anti-epileptic, anti-anxiety and anti-psychotic

effects in men and animals

• No significant side-effects. Even large doses have

been tolerated Hollister, 1975; Zuardi et al., 1982; Pellow et al., 1985; Agurell et al, 1986; Zuardi & Guimaraes, 1997; Crippa et al., 2004

Sinsemilla potency comparison 1996/8 v 2005 UK (Potter et al., 2008)

Potency levels signifantly higher in 2005 (p<0.001)

Endocannabinoid system • 1988: A specific cannabinoid receptor was found in the rat brain (Devane-Howlett, 1988) •1990: The discovery of a complementary DNA encoding for the first G protein-coupled cannabinoid receptor, now known as CB1 (Matsuda et al., 1990) •1992: The discovery of the neurotransmitter, a brain molecule called “anandamide” was identified (Devane-Mechoulam, 1992) •1993: The discovery of the second neurotransmitter, arachidonoyglycerol (2-AG, Munro et al., 1993)

Existing evidence

Studies • Epidemiological studies • Clinical studies • Biochemical studies • Genetic studies • Imaging studies

Epidemiological studies

- A causal link: Swedish study (1969) of 50,000 Swedish conscripts followed up for 15 years showed an increased risk amongst those who smoked cannabis; 1. those who were heavy users at age 18 were 6 times more likely to develop schizophrenia over the next 15 years; 2. increased risk in those who have genetic vulnerability (i.e.; someone else in family suffers from severe mental illness (Andreasson et al., 1987).

- The reanalysis of the Swedish study showed that there is an associated risk of developing schizophrenia consistent with a causal relation. This association is not explained by use of other drugs or personality traits (Zammit et al., 2002). - A Dutch study: 7500 people followed for 3 years. Regular consumers were more likely to develop schizophrenia (Van Os et al., 2002).

- The Prospective Dunedin (New Zealand) Multidisciplinary Health and Development Study: 1037 individuals followed up from birth in New Zealand (Arseneault et al., 2002) 1. At age 11: information on psychotic symptoms 2. At ages 15, 18: their drug use assessed 3. At age 26: their psychotic symptoms assessed The study found that people who used cannabis by the age of 15 were four times as likely to show an increase in SCZ symptoms at the age of 26.

Epidemiological studies

Systematic review of studies, cannabis use and psychosis risk (Moore et al., 2007) •Systematic review of evidence related to cannabis use and occurrence of mental health outcomes •Longitudinal and population based studies •Cannabis use increases the risk of development of a psychotic illness in a dose dependent manner •No strong evidence as yet that earlier cannabis use is particularly harmful

Systematic meta-analysis, cannabis use and earlier onset of psychosis (Large et al., 2011) • 443 articles: 83 studies met the inclusion criteria, involving: 1. 8167 substance-using patients 2. 14352 non-substance-using patients • Meta-analysis of age at onset of psychosis revealed that the age at onset was 2.70 years earlier among samples of cannabis users (p<.001) A further meta-analysis of other confounding factors (Large et al., 2012) •Possible confounding factors including: - Tobacco use - Diagnosis - Male gender - Study quality • The association between cannabis use and earlier onset of psychosis is robust • It is not the result of tobacco smoking by cannabis using patients or the other potentially confounding factors

Clinical studies

• Poor social functioning • Increased rates of violence • Increased rates of suicide • Increased rates of victimization • Homelessness • Criminal behaviour • Poorer physical health • Heavy burden on health services

• Early psychotic breakdown • Exacerbation or precipitation of symptoms • Poor adherence to treatment • Increased rates of hospitalization • Increased duration of hospitalization • Increased duration of psychotic episode

Review of clinical studies (Atakan et al., 2008) Cannabis use in those with serious mental illness has been associated with:

The adverse effect of cannabis use on outcome of First Episode

Psychosis is mediated by poor medication adherence

Colizzi et al., 2015 Schizophrenia Research

Gene environment interaction studies have mainly focussed on genetic variants involved in the regulation of the dopaminergic system: - COMT (Caspi et al., 2005) - AKT1 (van Winkel et al., 2011; Di Forti et al., 2012) - DAT1 (Bhattacharyya et al., 2012) - BDNF Val66Met (Decoster et al., 2011) - CNR1 and NRG1 (Pelayo-Teran et al., 2012) Some genes seem to explain susceptibility to cannabis related psychosis

Imaging studies - Long-term heavy cannabis users: reduction in bilateral hippocampal and amygdala volumes (Yucel et al., 2008) - First Episode Psychosis (FEP) cannabis users: decrease in GM density in the right posterior cingulate cortex, when compared with FEP with no cannabis use (Bangalore et al., 2008) - CNR1 rs2023239 by cannabis use interaction on bilateral hippocampus volumes (Schacht et al., 2012) Cannabis use alters the brain structure

Biochemical studies - The main psychotomimetic compound of the cannabis plant, delta-9-THC has an agonist/partial agonist effect on CB1 receptors (Pertwee, 2008) - Endocannabinoid and dopaminergic systems are co-localized in the CNS - Dopamine alterations have been reported in patients with schizophrenia (Laurelle, 1998) - Delta-9-THC increases dopamine release in the human striatum (Murray et al., 2014) - CBD reduces anxiety and has a profile similar to “atypical” antipsychotics (Atakan, 2008)

Genetic studies

Not every cannabis user develops psychosis..

Atakan et al., 2012

DRD2 gene (11q22-q23 )

Cannabis and psychosis: the role of DRD2 gene

D2L

D2S

Interaction between functional genetic variation of DRD2 and cannabis

use on risk of psychosis and on schizotypy and working memory Interaction between DRD2 rs1076560 and cannabis use on healthy subjects’ Working Memory Test (2-back) accuracy

Interaction between DRD2 rs1076560 and cannabis use on healthy subjects’ Schizotypy (SPQ questionnaire)

Interaction between DRD2 rs1076560 and cannabis use (lifetime and frequency of use) on psychosis risk

Colizzi et al., 2015 Schizophrenia Bulletin

*Adjusted for gender, age, ethnicity, alcohol,

stimulants and tobacco use

N = 252 N = 252

N = 439 N = 506

Beyond cAMP: the regulation of AKT and GSK3 by dopamine receptors

Dopamine-related behaviors

The effect of given genotype(s) on the risk of a disease can change in magnitude and direction (qualitative interaction) depending on the degree of environmental exposure

Interaction between DRD2 and AKT1 genetic variation on risk of psychosis in cannabis users

Colizzi et al., 2015 Nature Schizophrenia

Interaction between DRD2 rs1076560/AKT1 rs2494732 and cannabis use (lifetime and frequency of use) on psychosis risk

N = 402

N = 450

What do we know now?

Questions • Does cannabis trigger an earlier onset of psychosis in predisposed people? Yes • Does cannabis exacerbate the psychotic symptoms generally? Yes • Does the cannabis use worsen the outcome and progress of the illness? Yes • Is there a causal link? Yes, but only in those with a predisposition to psychosis

Cannabis and psychosis: is it just dopamine?

• Involvement of presynaptic dopamine dysfunction in psychosis is compelling

(Howes & Murray, Lancet 2014) • Vulnerability to the

psychotogenic effects of cannabis use involves genes that control dopamine signalling postsynaptically (Colizzi et al., 2015)

• What about direct effect of

THC on dopamine? Little evidence to suggest that cannabis use affected acute striatal dopamine release or chronic dopamine receptor status in humans (Sami et al., 2015)

To summarize..

Cannabis and psychosis: the role of CNR1 gene

CNR1 gene (6q14-q15 ) Endogenous ligands

Cannabis

Functional genetic variation of the CNR1 and cannabis use interact on prefrontal connectivity and related working memory behavior

Association between CNR1 rs1406977 and postmortem CNR1 prefrontal mRNA expression

Interaction between CNR1 rs1406977 and cannabis use on healthy subjects’ Working Memory Test (2-back) accuracy

Interaction between CNR1 rs1406977 and cannabis use on Left VLPFC during Working Memory Test (2-back)

Colizzi et al., 2014 Neuropsychopharmacology

N = 208

N = 208

-> it is thought that cannabis or Δ9-THC may not act on dopamine firing directly but indirectly by altering glutamate neurotransmission. •Limited research carried out in humans tends to support the evidence that chronic cannabis use reduces levels of glutamate-derived metabolites in both cortical and subcortical brain areas. • Research in animals tends to consistently suggest that 1. Δ9-THC depresses glutamate synaptic transmission via CB1 receptor activation, 2. disrupting glutamate synaptic plasticity after prolonged exposure, affecting glutamate release, inhibiting 3. receptors and 4. transporters function, 5. reducing enzyme activity

Cannabis and glutamate

Hypotheses under investigation

• 1. is to employ a placebo-controlled acute pharmacological challenge design to test the hypothesis that relative to placebo acute ∆9-THC administration will be associated with a reduction in glutamate levels as indexed by proton magnetic resonance spectroscopy (1H-MRS) in the hippocampus and striatum Hypofunction in glutamate neurotransmission: primary

deficit determining the dopamine abnormalities consistently reported in schizophrenia (Howes et al., 2015)

Hypotheses under investigation

• 2. acute administration of ∆9-THC will be associated with the induction of transient psychotic symptoms in these healthy volunteers and that these effects on symptoms will be directly related to the changes in glutamate in the proposed brain areas induced by it

∆9-THC and memory • ∆9-THC reduces hippocampal neuron

activation, relevant to memory formation • Brain imaging studies show regular ∆9-THC

users have a smaller hippocampus • ∆9-THC affects memory in healthy subjects

Hypotheses under investigation

• 3. effect of acute ∆9-THC on hippocampal activity while performing a verbal paired associate learning task, and test for a mediating effect of altered glutamate neurotransmission in the association between acute cannabis exposure and hippocampal dysfunction during verbal memory processing

The present results support the theory that some individuals carry genetic vulnerability to the psychotogenic effect of cannabis. The character of this vulnerability is likely to be a polygenic gene-environment interaction. Identifying those genes and organizing them into genetic pathways will facilitate understanding the mechanisms that modulate the relationship between cannabis exposure and psychosis.

Conclusions

Prof. Alessandro Bertolino Group of Neuroscience, Bari

Prof. Sir Robin M Murray Dr. Marta Di Forti GAP study, London Dr Sagnik Bhattacharyya Prof. Philip McGuire

Acknowledgments