RCUK Academic Fellow Psychological Medicine & Psychiatry Institute of Psychiatry King’s College London Modelling the effects of cannabis in adolescent mice Dr Cathy Fernandes

cannabis is generally considered to be a safe drug –side-effects include cognitive impairment (Sitskoorn et al, 2004), anxiety attacks and paranoia (Ames, 1958) increased risk of developing schizophrenia (Arseneault et al, 2004) Introduction worrying trends.... – –increase in the concentration of the main active ingredient of cannabis, ∆9 -tetrahydrocannabinol ( δ-THC ) – –age of first-time cannabis users is rapidly decreasing with 40% of 15 year olds in the UK having experienced the drug (European School Survey Project on Alcohol and Other Drugs)

effects of cannabis during development –neuronal networks are still under development (Romeo, 2003) –role of the endocannabinoid system (Fernandez-Ruiz et al, 2000) Introduction focus on prenatal/perinatal periods (Viveros et al, 2005) – –impaired executive function (Fried et al, 1998), impaired social behaviours and emotional reactivity (Trezza et al, 2008) fewer studies on the effects of postnatal (adolescent) exposure – –long-lasting cognitive effects (Stiglick & Kalant, 1985), sensory motor gating deficits and anhedonia (Schneider and Koch, 2003) – –synthetic cannabinoid compounds used, incremental or irregular dosing regimens [pharmacokinetic analyses?]

What are we modelling? Model= ‘ small measure ’ a simplified representation used to explain the workings of a real system behavioural endophenotypes genetic perturbation environmental perturbation

Endophenotype concept Kas et al (2007) association with the psychiatric disorder in the population heritable and primarily state-independent endophenotype and disorder co-segregate within families occur more in non-affected family members than in general population (Gottesman & Gould, 2003)

What are we modelling? make use of (natural) variation multiple perturbations (inbreds, outbreds, crosses) knock them out (one) at a time ? one perturbation (knock-out/transgenics)

diverse range of genetic manipulations physiological & anatomical similarity numerous behavioural models available Mouse - a model organism

childhood adolescence adulthood (> pd 7-28) (pd 28-49) (> pd 56) critical postnatal ontogenetic stages neurophysiological and hormonal processes (Spear, 2000) Developmental stages puberty (pd 35-56)

∆9 -tetrahydrocannabinol ( δ-THC ) administration –20 mg/kg once daily by oral gavage for 14 days two different inbred strains of mice (males only) developmental stages –juvenile (7-21 days old) versus adolescent (puberty; days old) behavioural phenotypes ( drug-free, 10 wks old) –working/episodic-like memory (delayed matching to place task) molecular phenotypes –synaptic markers –epigenetic profiling of candidate genes –genomewide transcription profiling Modelling the effects of cannabis C57BL/6JDBA/2J

δ-THC exposure in early life but not during late adolescence causes a working/ episodic-like deficit in adulthood dependent on genetic background C57BL/6J DBA/2J juvenile adolescent DMP task saving time (Zeng et al) 8 trials/day, 8 platform locations Behavioural effects of δ-THC in mouse * STRAIN x DRUG x AGE: F(1,72)=13.9, p < *

immunohistochemical staining for synaptic marker synaptophysin Molecular effects of δ-THC in mouse prefrontal cortex (PFC) DBA/2J C57BL/6J hippocampus (CA3) synaptophysin increased synaptophysin in PFC and CA3 after δ-THC exposure in juvenile but not adolescent mice dependent on genetic background but synaptophysin is reduced in prefrontal cortex in schizophrenic brains * * (* p <0.05)

individual susceptibility to the harmful effects of cannabis genetic factors (Harrison & Weinberger, 2005) Genetic influence on the effects of δ-THC COMT (catechol-O-methyltransferase) – –Val158Met polymorphism associated with the increased risk of psychosis incurred by adolescent-onset cannabis use (Caspi et al, 2005) – –Val158 carriers appear to be more sensitive to the psychotic experiences and cognitive impairments following administration of δ-THC (Henquet et al, 2006)

mRNA expression level (current data confounded by the mutation) Genetic influence on the effects of δ-THC protein level and activity of COMT (in collaboration with Elizabeth Tunbridge, Oxford) DNA methylation of Comt promoter (in collaboration with Jon Mill, IoP) mutation in Comt between DBA/2J and C57BL/6J mice (Fernandes et al)

early life exposure to δ-THC in DBA/2J but not C57BL/6J mice resulted in a long-lasting, mild impairment in cognition and changes in synaptic density genetic background and developmental stage influence the effects of δ-THC evidence for a genetically mediated vulnerability (resilience) to the effects of cannabis exposure in early life is Comt mediating this gene by environment interaction? Summary

What next... extend the behavioural characterisation of the effects of δ-THC - -anxiety, social behaviours, cognitive abilities pharmacokinetic analyses to translate levels of δ-THC broaden the molecular characterisation of the effects of δ-THC - -genomewide gene expression profiling (hippocampus and PFC) - -genomewide DNA methylation profiling (PFC) - -DNA methylation of specific candidate genes (Comt and Nrg1 across several brain regions)

Acknowledgements Elke BinderElke Binder Rachel KemberRachel Kember Leo SchalkwykLeo Schalkwyk Maria Teresa OrtinMaria Teresa Ortin Paul MorrisonPaul Morrison Jon CooperJon Cooper Jelle MostertJelle Mostert Jon MillJon Mill David CollierDavid Collier Robin MurrayRobin Murray Funding Research Councils UK Institute of Psychiatry