NeuroImage 52 (2010) 1505–1513

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Quantification of cerebral cannabinoid receptors subtype 1 (CB1) in healthy subjectsand schizophrenia by the novel PET radioligand [11C]OMAR

Dean F. Wong a,c,⁎, Hiroto Kuwabara a, Andrew G. Horti a, Vanessa Raymont a, James Brasic a,Maria Guevara a, Weiguo Ye a, Robert F. Dannals a, Hayden T. Ravert a, Ayon Nandi a, Arman Rahmim a,Jeffrey E. Ming e, Igor Grachev d,1, Christine Roy f, Nicola Cascella b

a Radiology, Johns Hopkins University, Baltimore, MD, USAb Psychiatry, Johns Hopkins University, Baltimore, MD, USAc Neuroscience, Johns Hopkins University, Baltimore, MD, USAd Clinical and Exploratory Pharmacology, Sanofi-aventis Research and Development, Bridgewater, NJ, USAe Clinical and Exploratory Pharmacology, Sanofi-aventis Research and Development, Malvern, PA, USAf Clinical and Exploratory Pharmacology, Sanofi-aventis Research and Development, Paris, France

⁎ Corresponding author. Radiology, Johns Hopkins Mroom 3245, 601 N. Caroline St., Baltimore MD 21287, U

E-mail address: dfwong@jhmi.edu (D.F. Wong).1 Current address: Merck/Schering-Plough, Kenilwor

1053-8119/$ – see front matter © 2010 Elsevier Inc. Adoi:10.1016/j.neuroimage.2010.04.034

a b s t r a c t

a r t i c l e i n f o

Article history:Received 23 December 2009Revised 7 April 2010Accepted 12 April 2010Available online 18 April 2010

Several studies have examined the link between the cannabinoid CB1 receptor and several neuropsychiatricillnesses, including schizophrenia. As such, there is a need for in vivo imaging tracers so that the relationshipbetween CB1 and schizophrenia (SZ) can be further studied. In this paper, we present our first human studies inboth healthy control patients and patients with schizophrenia using the novel PET tracer, [11C]OMAR (JHU75528),we have shown its utility as a tracer for imaging human CB1 receptors and to investigate normal aging and thedifferences in the cannabinoid system of healthy controls versus patientswith schizophrenia. A total of ten healthycontrols andninepatientswith schizophreniawere includedandstudiedwithhighspecific activity [11C]OMAR. TheCB1 binding (expressed as the distribution volume; VT) was highest in the globus pallidus and the cortex in bothcontrols and patients with schizophrenia. Controls showed a correlation with the known distribution of CB1 anddecline of [11C]OMAR binding with age, most significantly in the globus pallidus. Overall, we observed elevatedmeanbinding inpatientswith schizophreniaacross all regions studied, and this increasewas statistically significantin thepons (pb0.05), by theStudents t-test.Whenwerana regressionof the control subjectsVT valueswithageandthen compared the patient data to 95% prediction limits of the linear regression, three patients fell completelyoutside for the globus pallidus, and in all other regions there were at least 1–3 patients outside of the predictionintervals. There was no statistically significant correlations between PET measures and the individual BriefPsychiatryRatingScore (BPRS) subscores (r=0.49), but therewasa significant correlationbetweenVT and the ratioof the BPRS psychosis towithdrawal score in the frontal lobe (r=0.60), andmiddle andposterior cingulate regions(r=0.71 and r=0.79 respectively). In conclusion, we found that [11C] OMAR can image human CB1 receptors innormal aging and schizophrenia. In addition, our initial data in subjects with schizophrenia seem to suggest anassociation of elevated binding specific brain regions and symptoms of the disease.

edical Institutions, JHOC BldgSA. Fax: +1 410 955 0696.

th, NJ, USA.

2 SZ = schizophrenBPRS = brief psychiaSAS = Simpson Anginterest; TAC = timvolume; CB1 = cannPa=parietal cortex,Oparahippocampus, HpGP= globus pallidusaAC=anterior anteripCg = posterior cing

ll rights reserved.

© 2010 Elsevier Inc. All rights reserved.

ia; PET = positron-emission tomography; BMI = body mass index;tric rating scale; CDSS = Calgary Depression Scale for Schizophrenia;us Scale; MRI = magnetic resonance imaging; VOI = volume ofe–activity curves; BPND=binding potential; VT=total distribution

Introduction

Cannabis sativa, that is known asmarijuana, has been used by peopleas a drug of abuse and medication throughout history (Mechoulam,1986). Δ9-Tetrahydrocannabinol (Δ9-THC), the principal component ofmarijuana, is known to activate cannabinoid receptors (Pertwee, 2008).The cannabinoid receptors belong to GPCR-superfamily and currentlytwo subtypes of cannabinoid receptors have been identified (Howlettet al., 2002): CB1-subtype that is primarily found in brain and neuronal

tissue and CB2-subtype that is mainly found in immune tissue and, inlower concentration, in normal nervous tissue (Onaivi et al., 2008).Central CB1 receptors are involved in various brain functions anddisorders including schizophrenia2 and depression, drug addiction and

abinoid receptor type 1; Fr = frontal cortex, Tp = temporal cortex,c=occipital cortex, Fs=fusiformgyrus, Cg=cingulate cortex, PH== hippocamus, In = insula, Pu = putamen, CN= caudate nucleus,

, Th = thalamus, Cb = cerebellum; vAC= ventral anterior cingulate,or cingulate, dAC=dorsal anterior cingulate,mCg=middle cingulate,ulate.

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alcoholism (Mouslech and Valla, 2009). Even though the chemistry andpharmacology research of cannabinoids has reached enormous propor-tions (Hanus, 2009) the exact role of cannabinoid receptors in normalstate and disease remains elusive.

Multiple lines of evidence show that cannabis and CB1 receptorsplay a role in psychotic illness and particularly in SZ (Andreassonet al., 1987; Arseneault et al., 2002; Chavarria-Siles et al., 2008;D'Souza, 2007; D'Souza et al., 2005, 2009; Giuffrida et al., 2004;Henquet et al., 2005; Leweke et al., 1999; Ujike et al., 2002; van Oset al., 2002; Zammit et al., 2002).

Studies with post-mortem brain in patients with SZ have showed a25% increase of the binding of [3H]CP55,940, a non-subtype selectiveCB1/CB2 agonist, in the superficial layers of the posterior cingulatecortex (Newell et al., 2006) and a 64% increase of the binding of CB1selective cannabinoid antagonist [3H]SR141716A in the anteriorcingulate cortex (Zavitsanou et al., 2004). Dean et al. (2001) foundincreased binding of a cannabinoid agonist [3H]CP55,940 in thedorsolateral prefrontal cortex (DLPFC) in SZ patients (independent ofcannabis use prior to their death), while the increased numbers in thecaudate–putamen was thought to be the result of cannabis use. In thisstudy no change was observed in the hippocampus compared tonormal controls. In contrast to these radiotracer in vitro bindingstudies, reduced cortical CB1 receptor messenger RNA and proteinexpression have been found post-mortem in DLPFC of subjects withSZ (Eggan et al., 2008), while another immunohistochemical study(Koethe et al., 2007) showed no alteration of density of CB1 receptorimmunopositive cells in the brains of subjects with SZ. A more recentstudy demonstrated that immunodensity of CB1 receptors in thefrontal cortex was significantly decreased (71±7%) in antipsychotic-treated subjectswith schizophrenia butnot in drug-free subjects (104±13%) (Uriguen et al., 2009).

The most advanced technique for studying central receptors in thelivinghumanand animal brain is PET.Until recently thequantitative PETimaging of CB1 receptors has been hampered by the lack of suitableradioligands. PET imaging of CB1 receptors will help to study the role ofthe CB1 in many CNS diseases, including schizophrenia. Betterunderstanding of the fundamental mechanisms of the cannabinoidreceptor system could direct the development of medications to treatthese disorders. A preliminary PET study of CB1 in a subject withschizophrenia with [124I]AM281 suggested a feasibility of suchinvestigation, but a poor signal-to-noise ratio of the radioligand wasan obstacle for a further research including comparison of CB1 receptorsin normal controls and subjects with SZ (Berding et al., 2006).

During the last 2 years, advances inmedicinal radiochemistry haveled to the development of radiolabeled CB1 PET probes with betterimaging characteristics than those of [124I]AM281. Quantitative PET ofthe CB1 receptor has now become possible (Burns et al., 2007; Hortiet al., 2006; Terry et al., 2009; Van Laere et al., 2008b, 2009; Yasunoet al., 2008) and studies with several classes of new CB1 radiotracersare in progress (see for review (Horti and Van Laere, 2008)). Our newradioligand, [11C]OMAR ([11C]JHU75528 (Fan et al., 2006)), exhibitedgood results for quantification of CB1 receptors in the mouse andbaboon brain (Horti et al., 2006) and control human subjects (Horti etal., 2007; Wong et al., 2008b). In this report we are presenting theresults of PET imaging of CB1 receptors with [11C]OMAR in healthyhuman controls and patients affected by SZ (Wong et al., 2008a),which demonstrate the utility of [11C]OMAR to image CB1 receptors.

Materials and methods

Radioligand for human studies

[11C]OMAR ([11C]JHU75528) is a CB1 selective PET radioligand(Fan et al., 2006) that was synthesized here for human studies via thepreviously published procedure (Horti et al., 2006). The radioligandwas synthesized in high radiochemical purity (N99%) and specific

radioactivity (273±73 GBq/mmol, n=20). The precursor for theradiosynthesis was purchased from IsoSciences (King of Prussia, PA).Toxicology and radiation dosimetry studies were carried out anddemonstrated safety and tolerability. These safety and toxicologystudies were consistent with local IRB approval (see AppendixDescription 1).

Study design

To examine the distribution of CB1 receptors in the human brain,in both healthy controls and patients affected by SZ, we conducted asingle-center PET study. In this study control human subjects wereinitially compared with a single cohort of 10 subjects with SZ.

Study participants

Healthy adult control subjects were recruited via advertisements inlocal newspapers, word of mouth, and announcements at localuniversities. Adult subjects with SZ were recruited from the outpatientclinic of the Department of Psychiatry at Johns Hopkins Hospital.

All subjects received thorough medical and psychiatric screenings,which specifically examined current and earlier medications. Allcontrol subjects were free of neuroleptic, cannabinoid, serotonergic,and dopaminergic drugs at the time of PET scanning, and had beentreatment-free for a period of at least 6 months before the start of thestudy.

Study procedures were conducted in accordance with theDeclaration of Helsinki Principle of 2004. The Institutional ReviewBoard of The Johns Hopkins University School of Medicine approvedthe studies and the informed consent documents. All participantsgave their written informed consent.

The control cohort consisted of ten healthymales, with a mean ageof 33±11 years, ranging from ages 21 to 51 and was comprised of8 African-Americans, 1 Asian and 1 Caucasian. The SZ cohort consistedof 9males and 1 female, with amean age of 42±9 years, ranging fromages 30 to 54 and comprised of 7 African-Americans, 1 Indian and 2Caucasians (see Appendix Tables 1 and 2).

Inclusion criteria for the controls included: BMI between 18 and28 kg/m2, smoking consumption of less than 5 cigarettes a day andalso the ability to stop smoking while participating in the study, andfinally; all controls had to be deemed to be healthy by a detailedmedical history and complete physical examination. Potentialsubjects were also excluded if they had a history or presence ofdrug or alcohol abuse (alcohol consumptionN40 g/day).

The subjects with schizophrenia all had a DSM-IV-TR (AmericanPsychiatric Association [DSM-IV-TR], 2000) diagnosis of schizophre-nia, and were treated with olanazapine or risperidone monotherapyfor at least 2 months prior to screening. They did not differsignificantly in terms of duration of illness.

We excluded any of the schizophrenia subjects if they had a 1)BPRS hallucinatory behavior, unusual thought content, suspiciousnessor conceptual disorganization item scores N4 (moderately severe) onany single item; 2) CDSS total score N10 (depression) or a score of N0on item 8 (suicidality); 3) SAS score N0.6 (6 total); and 4) BarnesAkathisia Scale global score N3.

Study procedures

Dynamic PET scans of 90 min followed a slow-bolus injection of[11C]OMAR (mean total dose=703 MBq; specific radioactivi-ty=222–296 GBq/µmol).

All PET scans were carried out on the second-generation High-Resolution Research Tomograph (ECAT HRRT); (CPS Innovations,Inc.), an LSO-based, 2.5 mm-resolution, dedicated brain PET scanner(Sossi et al., 2005).

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For co-registration with the PET data to enhance anatomicaldefinitions, each participant received a volumetric MRI scan acquiredusing a spoiled GRASS (gradient recalled acquisition in steady state)(SPGR) sequence on a GE 1.5 T Signa Camera (GE Healthcare, ChalfontSt. Giles, UK).

PET scan procedures

An intravenous catheter was inserted to the antecubital vein forligand injection and an arterial catheter was inserted in the radialartery at the wrist on the other hand to obtain blood samples. Athermoplastic maskwas custom-made for each subject to reduce headmotion during the scan. At 15 min before ligand injection, the subjectwas positioned in the scanner. A 6-min attenuation scan wasperformed using a rotating Cs-137 point source. The 90-min emissionscan started with a slow-bolus injection (over 1 min) of radioligandabout 20 mCi (range; 19.53 to 20.43 mCi) in a 3-D list mode. Smallamounts of arterial blood (∼2 ml) were sampled every 5 s initiallyand at prolonging intervals toward the end of the scan. Radioactivityin plasma samples were measured with a gamma counter andcorrected for physical decay to the injection time. Large amounts ofarterial blood (∼5 ml) were sampled at 0, 5, 10, 30, 60 and 90 min fordetermination of [11C]OMAR and its radioactive metabolites in plasmaas described elsewhere (Hilton et al., 2000; Horti et al., 2006).

Reconstruction of emission scanPET images were reconstructed using the iterative ordered-subset

expectation-maximization (OSEM) algorithm correcting for attenua-tion, normalization, scatter, randoms and dead-time (Jones et al.,2006; Rahmim et al., 2005). The following frame schedule was used:four 15-s, four 30-s, three 1-min, two 2-min, five 4-min, and twelve 5-min frames, or a total of 30 frames for the 90 min scan. Theradioactivity was corrected for physical decay to the injection time.Each PET frame consisted of 256 (left-to-right) by 256 (nasion-to-inion) by 207 (neck-to-cranium) voxels.

Data analysis

Volumes of interestVOIs were generated by spatially normalizing (Ashburner and

Friston, 2007a) a standard VOI template (Tzourio-Mazoyer et al.,2002) to each individual subject's MRI that was then edited manuallyfor errors associated with spatial normalization. The pons VOI waslimited to the anterior portionwhere graymatter components are lessprominent. The cingulate cortex VOI, the region of particular interest,was divided into anterior, middle, and posterior cingulate VOIs by theanterior and posterior commissure planes (Planes A and B). The U-shaped anterior cingulate VOI was further divided into dorsal, rostral,and sub-callosal subdivisions by a coronal plane that passed throughthe anterior tip of corpus callosum (Plane C) (Bush et al., 2000; Fornitoet al., 2006, 2008; Vogt et al., 2003; Vogt and Laureys, 2005). Theclassification process was performed in the Talairach-oriented MRIspace (i.e., anterior and posterior commissure points were on the Yaxis, and mid-sagittal plane was one the YZ plane) (Talairach andTornoux, 1988). Finally, the VOIs were transferred to PET space usingMRI-to-PET co-registration parameters (Ashburner and Friston,2007b) and applied to individual PET frames to obtain TAC.

Radioactive plasma and radiolabeled metabolite dataThe radioactivemetabolites in bloodwere analyzed by HPLC under

the same conditions that we used previously in baboon [11C]OMARstudies (Horti et al., 2006). The HPLC analysis demonstrated nosubstantial difference between radioactive metabolites in the baboonand human blood (data are not presented). The average parent ligandin plasma was 41% at 60 min by HPLC. The radioactive metaboliteswere assumed not to enter the brain because of weak lipophilicity, as

has been shown in our previous rodent and baboon studies (Horti etal., 2006). The time profiles of total metabolites were fitted by a sum ofexponential functions (zero derivative at time 0) to separate the totalradioactivity into [11C]OMAR and metabolites. Metabolite-correctedplasma TAC was used in the model parameter estimation, with theexception of the total plasma TAC, which was used for the calculationof the radioactivity in the tissue vasculature when applicable.

Derivation of outcome variablesThe distribution of volume of [11C]OMAR VT was obtained for each

region by plasma reference graphical analysis (PRGA (Logan et al.,1990)). In a preliminary study, PRGA showed more robust estimatesof VT than the two-compartment model, as measured with time-dependency (i.e., PET frames used for data analysis) of estimates andthe magnitude of inter-subject variability. Because of the lack of areliable reference region for [11C]OMAR (see Results and discussion),we concluded that the BPND (Innis et al., 2007) is not appropriate forquantification of this radioligand.

Head motion correctionHead motions during the emission scan were not corrected in this

analysis because apparent head motions were not detected on visualinspections of TACs in each subject. In addition, VT estimates withoutand with head motion correction (HMC) were essentially identical forPRGA (VT[HMC]=1.01VT[noHMC]−0.004; R2=0.989).

Behavioral analysisWe carried out a correlational analysis of these summary categories

and the regional VT. We particularly focused on the summary categoriesof psychotic and withdrawal symptoms, as these are representative ofthe most prominent symptoms in SZ.

Results and discussion

Based on safety and tolerability studies (see Appendix Description 1),we elected to inject a mass not to exceed 0.14 μg/kg during eachradiotracer injection. The radiation dosimetry estimates in micesuggested that an [11C]OMAR dose of 10–20 mCi (370–740 mBq) issafe for human studies. There were no significant adverse experiencesreported during the studies, in either the healthy control subjects orsubjects with SZ.

Outcome variable of the [11C]OMAR human studies

The pons showed the lowest accumulation of the radioactivityamong brain regions. However, inter-subject variations (as measuredby standard deviation across subjects) increased from a 10% range forVT to a 20% range for BP when the pons was used as a reference region.Therefore, it is likely that VT estimates within the pons may beuncertain because of its relatively small size (∼10 ml) compared totypical reference regions (e.g., ∼90 ml for cerebellum and ∼50 ml foroccipital cortex) as well as the low counts in this region. In addition, ithas been noted that one of the highest reductions in VT occurred in thepons in the monkey brain as measured with [11C]MePPEP, a separateCB1 receptor ligand, after pretreatment with rimonabant (3.0 mg/kgi.v.). Other researchers (Burns et al., 2007; Yasuno et al., 2008) havesuggested that white matter (centrum semiovale) may be used as areference region for [18F]MK-9470, another CB1 receptor radioligand.However, in our study, white matter TAC continued to increase for the90 min period for [11C]OMAR even when white matter VOIs werelimited to voxels that were less affected by the partial volume effect(i.e., pure white matter voxels). Thus, we suggest that [11C]OMARshowed different kinetics in the white matter than cortical regionsand/or partial volume effects may not be completely eliminated evenfor HRRT. For all the above reasons, we concluded that pons andwhitematter may not be used as a reference region for calculation of BPND,

Fig. 1. Time–(radio-)activity curves (TACs) of selected brain regions shown as meanstandard uptake values (SUV; g/ml) across 10 healthy subjects.

Fig. 3. Brain regional total volume of distributions (mean VT±SD, n=10) in the controlcohort in the PET studies with [11C]OMAR.

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and that BPND may not be accessible for [11C]OMAR. Therefore, weused VT as the main outcome variable instead of BPND.

[11C]OMAR binding and brain regional distribution in control subjects

The brain time–activity curves of [11C]OMAR peaked at approxi-mately 20 min post injection (% SUV ranged from 136 to 207% in theputamen) and decreased gradually thereafter to reach a SUV between80 and 117% at 90 min. (Fig. 1). In all 10 control subjects the VT valuesranged from 0.79 to 1.82 across multiple cortical and subcorticalregions. Because of the high resolution of the HRRT PET scanner wewere able to assess relatively small regions including the globuspallidus and subdivisions of cingulate cortex.

Within the controls, the globus pallidus showed the highest VT, witha mean of 1.47±0.25 (SD), with a range of 0.95 to 1.82 in individualsubjects (Figs. 2 and 3). The cingulate cortex (mean 1.23±0.16(SD),range of 1.01–1.49) and the putamen (mean 1.32±0.20(SD), range of1.05–1.63) also showed high binding. The intermediate uptake was inthe hippocampus, cerebellum and cortex. The lowest accumulation of

Fig. 2. Trans-axial images of distribution volume (VT) of [11C}OMAR, mean of 10 healthy subjVolumes of interest of selected structures in the standard space are shown on the VT imagcaudate nucleus (C).

the [11C]OMAR radioactivity was seen in thewhitematter, the pons andthalamus. It is noteworthy that the previous autoradiography studywith cannabinoid agonist [3H]CP55,940 (Glass et al., 1997; Herkenhamet al., 1990) demonstrated the highest binding in the hippocampus andfrontal cortex and intermediate binding in the globus pallidus andputamen, whereas [11C]OMAR manifests the highest accumulation ofradioactivity in the basal ganglia, including the globus pallidus andputamen (Fig. 3). However, the in vivo regional distribution of [11C]OMAR in thehuman brain (Fig. 4) is consistentwith the PEThumandatafrom other CB1 selective PET radioligands including [18F]MK-9470(Burns et al., 2007; Van Laere et al., 2008a), [11C]MePPEP (Terry et al.,2009) and [124I]AM281 (Berding et al., 2006). The differences betweenin vivo and in vitro CB1 binding have been discussed by otherresearchers (Burns et al., 2007; Van Laere et al., 2008a) who concludedthat there could be a number of reasons for the differences, including alimited availability of the hippocampusCB1 receptors for in vivo bindinggiven that ∼85% of these receptors are located in intracellular vesicles(Burns et al., 2007; Coutts et al., 2001).

Effect of aging on the [11C] OMAR binding in controls and subjects withschizophrenia

We also correlated the [11C]OMAR regional volume of distribution(VT) with age. Within the control group we found a significant age-associated decline of [11C]OMAR VT in the globus pallidus (r=−0.66,

ects in a standard space (left panel), a standardMRI (right), andmerged image (middle).e and MRI. Regions are insula (I), putamen (P), glubus pallidus (G), thalamus (T), and

Fig. 4. Correlation of PET regional volumes of distribution of [11C]OMAR in controls(VT,mean±SD, n=10) with post-mortem autoradiographic CB1 density of binding [3H]CP55,940 in the adult human brain (Bmax). The regional Bmax values were calculatedhere as means of all sub-regions that are presented in the autoradiography paper (Glasset al., 1997).

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pb0.05), the region with the highest uptake of [11C] OMAR. Otherregions of intermediate to high [11C] OMAR uptake, such as theputamen and cortex also showed a decline with age, but this trendwas not statistically significant. This supports results from previousstudies that found no significant trend of the CB1 radioligand [18F]MK-9470 uptake in the hippocampus formation with aging within a groupof normal male controls (range=18–70 years old) (Van Laere et al.,2008a), although an age related increase of [18F]MK-9470 uptake inthe entorhinal cortex/amygdala was observed in female controls.

Fig. 5. Effect of aging on the binding of [11C]OMAR (VT) in the various brain regions. Amonglines, dotted lines indicated 95% prediction interval). Data from subjects with schizophreni

Within the cohort with SZ, there was no significant associationwith age in any region. Furthermore, when the patients with SZ wereplotted against the regression of the controls with age, at least one toseveral of the schizophrenia data points fell outside of the 95%prediction interval of the regression line. This was the case in theglobus pallidus, where we saw a significant decline with age incontrols, as well as in all other regions. Fig. 5 shows the regressionwith age in several regions, including globus pallidus.

These PET results are consistent with an autoradiography study thatdemonstrated a substantially larger binding of [3H]CP55,940 in thebasalganglia of the neonatal brain (Bmax

Putamen=86 fmol/mg, BmaxGlobus pallidus=

118 fmol/mg) versus the human adult brain (BmaxPutamen=44 fmol/

mg, BmaxGlobus pallidus=82 fmol/mg) (Glass et al., 1997), but the PET

data contradict to another [3H]CP55,940 autoradiography study thatfound that the CB1 binding in a gender-mixed set of human brainsincreases progressively from fetus and early prenatal stages toadulthood (Mato et al., 2003). We are not aware of the post-mortemCB1 binding studies within the aging human brain.

In a separate study occupancy-CB1 antagonist drug concentrationdata from schizophrenic subjects were indistinguishable from those ofhealthy volunteers. In calculation of occupancy, distribution volumeof non-displaceable ligand (VND) was also estimated from a scatterplot of observed distribution volume of baseline blocking. The resultsclearly indicate that VT likely represents specific binding to the CB1 inthe human brain. (unpublished data)

[11C]OMAR PET imaging of subjects with schizophrenia

A comparison of [11C]OMAR binding in the control subjects andsubjects with SZ demonstrated a higher VT values in all brain regions

the controls only, regression analysis of[11C]OMAR VT with aging was done (solid blacka was then plotted against this regression (red triangles).

Fig. 6. Regional total volume of distributions (mean VT±SEM) of [11C]OMAR are greater in SZ subjects than in controls. VT was signficantly higher in SZ in the Pons (PO), pb0.05.

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of subjects with SZ (Fig. 6), and this increase was significant in thepons by t-test (pb0.05). The data did contain one outlier withextremely low CB1 binding (potentially a plasma input functionproblem), but all other psychological and demographic variables werewithin the group means. There was indeed an increase (15–22%) inbinding in most brain regions of the subjects with SZ, compared tocontrols, but the difference was not significant. However, in the ponsthe increase (23%) of VT in subjects with SZ compared to controls wassignificant (SZ VT

GP=0.93±0.27; controls VTGP=0.72±0.07). Previ-

ous autoradiography data in schizophrenia demonstrated an increaseof the [3H]SR141716 binding in the anterior cingulate cortex(Zavitsanou et al., 2004) and [3H]CP55,940 binding in the posteriorcingulate cortex (Newell et al., 2006). We measured the VT values of[11C]OMAR in sub-regions of the cingulate cortex of SZ and controlsubjects (Fig. 7), and found a non-significant increase of [11C]OMARbinding in all subdivisions of the cingulate cortex in subjects with SZ.

Fig. 7. Correlation of [11C]OMAR VT in the brain regi

In order to examine evenmore rigorous subject matching, we useda t-test to compare the VT values of [11C]OMAR in the two SZ andcontrol subjects matched by age, educational level and also parentalSES, and found a significantly higher mean value for some regionssuch as the occipital cortex (mean VT=1.21(sd=0.05) c.f. meanVT=0.92(sd=0.05); p=0.029) and the putamen (mean VT=1.42(sd=0.06) c.f. mean VT=1.08(sd=0.04); p=0.015). However, asthis subsample contained only a very small number of subjects, thesefindings are supportive butmust be replicated.Wewould suggest thatfuture studies should consider utilizing matching criteria that includeracial group and educational level (Dickinson et al., 2007; Resnick,1992) in larger samples.

All patients with SZ that we studied were taking the antipsychoticdrugs olanzapine or risperidone, so there is a chance that these drugsmay have affected the CB1 binding, as there have been a number ofrecent reports on this matter. Thus, it was found that risperidone

ons versus BPRS withdrawal scores (SZ group).

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treatment increased CB1 binding in the rat caudate nucleus,hippocampus and amygdala (Secher et al., 2010). In addition, exvivo experiments in Sprague rats have demonstrated that olanzapinesignificantly decreases CB1 binding in the dorsal vagal complex of thebrainstem in rats (Weston-Green et al., 2008). However, no evidenceof binding of antipsychotic drugs clozapine, olanzapine and haloper-idol at CB1 receptor was found in vitro (Theisen et al., 2007). Otherantipsychotic drugs such as aripriprazole and haloperidol have beenshown to have little effect on the CB1 binding ex vivo (Weston-Greenet al., 2008). Experiments with rats that were chronically treated withvarious antipsychotic drugs demonstrated (Sundram et al., 2005) thatclozapine decreases CB1 binding ([3H]CP55,940) in the nucleusaccumbens while other regions (cortex, hippocampus and striatum)showed no change. However, the same study (Sundram et al., 2005)showed no effect with haloperidol, chlorpromazine or olanzapine.Overall, the existing body of research suggests that most antipsychoticdrugs do not bind with CB1 receptor in vitro and do not change theCB1 radiotracer binding in the cortex and striatum (the regions withthe highest density of the CB1 receptor), but may decrease the CB1radiotracer binding in the brainstem and amygdala, or, in case ofrisperidone, may increase the CB1 binding in the hypothalamus,hippocampus and amygdala. Risperidone and olanzapine seem toshow the opposite effect on the CB1 binding of radiotracers in rodents.Because all of the SZ patients were prescribed olanzapine orriperidone we compared the [11C]OMAR binding in both subgroups,but found no significant difference (Appendix Fig. 1).

Fig. 8. Correlation of [11C]OMAR VT in the brain regions of SZ subjects

[11C]OMAR PET binding and relationship of behavioral subscores (BPRSfor subjects with schizophrenia)

The elevated binding of [11C]OMAR in subjects with SZ (Figs. 6 and7) suggests that CB1 binding might correlate with type and severity ofclinical symptoms of schizophrenia. However, we found no correlationbetween total score on the brief psychiatric rating scale (BPRS) and[11C]OMAR VT. In addition, there was no obvious association betweenthe VT values and BPRS subscores, although there was a trend for theBPRS withdrawal symptom scores, whose severity showed a trendlevelwith a decline of VT in the cortical brain regions (Fig. 7). However,when we examined the difference and ratio between BPRS subscoresfor positive symptoms (psychosis-type symptoms) to negativesymptoms (withdrawal-type symptoms), we found some significantresults. The SZ subjects (when examining 9 of the patients with SZ,excluding the outlier mentioned above) with the highest psychosis towithdrawal scores ratio had the highest elevated CB1 receptors,suggesting a possible interaction between positive and negativesymptoms and CB1 receptors (Fig. 8). We found a significantcorrelation between the psychosis to withdrawal scores ratio and VT

values in a number of different brain regions, including the frontal lobe(r=0.49, p=0.05), and the middle and posterior cingulate (r=0.71p=0.03, and r=0.79, p=0.004 respectively). These results suggestthat CB1 receptor binding in the subjects with SZ increases withseverity of the positive symptoms and decreases with severity ofnegative symptoms.

and severity of symptoms: VT versus positive/negative symptoms.

1512 D.F. Wong et al. / NeuroImage 52 (2010) 1505–1513

Conclusions

[11C]OMAR readily enters human brain and shows a regional braindistribution that is consistent with that of cannabinoid receptorssubtype 1 (CB1). The imaging properties of [11C]OMAR are sufficientfor studying CB1 receptors in the human brain, including in thosewithneuropsychiatric disorders such as schizophrenia. It has reversiblekinetics during the 90 minute PET scan. There were no significantsafety issues with [11C] OMAR at high specific activity.

The total volume of distribution VT values of [11C]OMAR in themale control subjects show little age dependence in most brainregions, with low to intermediate binding that is consistent with theobservations of other researchers that have utilized the CB1 tracer[18F]MK-9470. However, [11C]OMAR binding does appear to declinewith increasing age in the healthy controls in the globus pallidus andputamen, the regions with highest values of VT.

Having demonstrated the feasibility and safety of [11C] OMAR inhumans, our initial studies applied to a patient group, demonstratedelevated VT values in a small cohort of subjects with schizophreniacompared to normal controls. This VT difference was significant in thepons when examining all controls versus all included SZ patients, and inthe globus pallidus when the subjects with SZ were compared with the95% prediction limits of the linear regression of the controls’ VT vs. age.The age-matched cohort was a small group of 6 controls and 6 patients.Lastly, there were significant differences in the occipital lobe andputamen when additional matching criteria including racial group andeducation levelwere incorporated.Withinpatientswith SZ, theVT valuesin certain brain regions correlated with positive and inversely withnegative symptoms, suggesting that it may be possible to characterizethe CB1 VT as it relates to type and severity of clinical symptoms in SZ. Itmay well be that accounting for the negative, as well as the positivesymptomsof thedisordermayhelp our furtherunderstandingof the roleof the CB1 cannabinoid receptors in SZ. We recognize that the ratio anddifferences of BPRS subscores have not been widely cited, and that thisstudy involved a small sample. However this preliminary findingsuggests a potential relationship between behavioral characteristicsand CB1 binding, and is provocative. If supported with large numbersthis could provide neurochemical clues to CB1 involvement inschizophrenia. It is possible that subjects with SZ who have an inherenttendency to a particular symptomatology have abnormal VT values. Itmay be even possible that VT values may help predict those that are atgreater risk of early decline into the negative syndrome of SZ, and thusthese important initial findings warrant greater investigation.

Acknowledgments

This study was supported by Sanofi-Aventis and NIH grantsDA000412, MH079017, S10RR023623-01 and S10RR017219 to DeanF. Wong.

Appendix A. Supplementary data

Supplementary data associated with this article can be found, inthe online version, at doi:10.1016/j.neuroimage.2010.04.034.

References

American Psychiatric Association, 2000. Diagnostic and statistical manual of mentaldisorders (Revised 4th ed.). Washington, DC.

Andreasson, S., Allebeck, P., Engstrom, A., Rydberg, U., 1987. Cannabis and schizophre-nia. A longitudinal study of Swedish conscripts. Lancet 2, 1483–1486.

Arseneault, L., Cannon, M., Poulton, R., Murray, R., Caspi, A., Moffitt, T.E., 2002. Cannabisuse in adolescence and risk for adult psychosis: longitudinal prospective study. BMJ325, 1212–1213.

Ashburner, J.T., Friston, K.J., 2007a. Non-linear registration. In: Friston, K.J., Ashburner, J.T.,Kiebel, S.J., Nichols, T.E. (Eds.), Statistical Parametric Mapping: The Analysis ofFunctional Brain Images. Academic Press, pp. 63–80.

Ashburner, J.T., Friston, K.J., 2007b. Rigid body registration. In: Friston, K.J., Ashburner, J.T.,Kiebel, S.J., Nichols, T.E. (Eds.), Statistical Parametric Mapping: The Analysis ofFunctional Brain Images. Academic Press, pp. 49–62.

Berding, G., Schneider, U., Gielow, P., Buchert, R., Donnerstag, F., Brandau,W., Knapp,W.H.,Emrich, H.M., Muller-Vahl, K., 2006. Feasibility of central cannabinoid CB1 receptorimaging with [124I]AM281 PET demonstrated in a schizophrenic patient. PsychiatryRes. 147, 249–256.

Burns, H.D., Van Laere, K., Sanabria-Bohorquez, S., Hamill, T.G., Bormans, G., Eng, W.S.,Gibson, R., Ryan, C., Connolly, B., Patel, S., Krause, S., Vanko, A., Van Hecken, A.,Dupont, P., De Lepeleire, I., Rothenberg, P., Stoch, S.A., Cote, J., Hagmann, W.K.,Jewell, J.P., Lin, L.S., Liu, P., Goulet, M.T., Gottesdiener, K., Wagner, J.A., de Hoon, J.,Mortelmans, L., Fong, T.M., Hargreaves, R.J., 2007. [18F]MK-9470, a positronemission tomography (PET) tracer for in vivo human PET brain imaging of thecannabinoid-1 receptor. Proc. Natl. Acad. Sci. U.S.A. 104, 9800–9805.

Bush, G., Luu, P., Posner, M.I., 2000. Cognitive and emotional influences in anteriorcingulate cortex. Trends Cogn. Sci. 4, 215–222.

Chavarria-Siles, I., Contreras-Rojas, J., Hare, E., Walss-Bass, C., Quezada, P., Dassori, A.,Contreras, S., Medina, R., Ramirez, M., Salazar, R., Raventos, H., Escamilla, M.A.,2008. Cannabinoid receptor 1 gene (CNR1) and susceptibility to a quantitativephenotype for hebephrenic schizophrenia. Am. J. Med. Genet. B Neuropsychiatr.Genet. 147, 279–284.

Coutts, A.A., Anavi-Goffer, S., Ross, R.A., MacEwan, D.J., Mackie, K., Pertwee, R.G., Irving,A.J., 2001. Agonist-induced internalization and trafficking of cannabinoid CB1receptors in hippocampal neurons. J. Neurosci. 21, 2425–2433.

D'Souza, D.C., 2007. Cannabinoids and psychosis. Int. Rev. Neurobiol. 78, 289–326.D'Souza, D.C., Abi-Saab, W.M., Madonick, S., Forselius-Bielen, K., Doersch, A., Braley, G.,

Gueorguieva, R., Cooper, T.B., Krystal, J.H., 2005. Delta-9-tetrahydrocannabinoleffects in schizophrenia: implications for cognition, psychosis, and addiction. Biol.Psychiatry 57, 594–608.

D'Souza, D.C., Sewell, R.A., Ranganathan, M., 2009. Cannabis and psychosis/schizo-phrenia: human studies. Eur. Arch. Psychiatry Clin. Neurosci. 259, 413–431.

Dean, B., Sundram, S., Bradbury, R., Scarr, E., Copolov, D., 2001. Studies on [3H]CP-55940binding in the human central nervous system: regional specific changes in densityof cannabinoid-1 receptors associated with schizophrenia and cannabis use.Neuroscience 103, 9–15.

Dickinson, D., Ramsey, M.E., Gold, J.M., 2007. Overlooking the obvious: a meta-analyticcomparison of digit symbol coding tasks and other cognitive measures inschizophrenia. Arch. Gen. Psychiatry 64, 532–542.

Eggan, S.M., Hashimoto, T., Lewis, D.A., 2008. Reduced cortical cannabinoid 1 receptormessenger RNA and protein expression in schizophrenia. Arch. Gen. Psychiatry 65,772–784.

Fan, H., Ravert, H.T., Holt, D.P., Dannals, R.F., Horti, A.G., 2006. Synthesis of 1-(2, 4-dichlorophenyl)-4-cyano-5-(4-[11C]methoxyphenyl)-N-(piperidin-1-yl)-1H-pyr-azole-3-carboxamide ([11C]JHU75528) and 1-(2-bromophenyl)-4-cyano-5-(4-[11C]methoxyphenyl)-N-(piperidin-1-yl)-1H-pyrazole-3-carboxamide ([11C]JHU75575) as potential radioligands for PET imaging of cerebral cannabinoidreceptor. J. Labelled Comp. Radiopharm. 49, 1021–1036.

Fornito, A., Whittle, S., Wood, S.J., Velakoulis, D., Pantelis, C., Yucel, M., 2006. Theinfluence of sulcal variability on morphometry of the human anterior cingulate andparacingulate cortex. Neuroimage 33, 843–854.

Fornito, A., Yucel, M., Wood, S.J., Adamson, C., Velakoulis, D., Saling, M.M., McGorry, P.D.,Pantelis, C., 2008. Surface-based morphometry of the anterior cingulate cortex infirst episode schizophrenia. Hum. Brain Mapp. 29, 478–489.

Giuffrida, A., Leweke, F.M., Gerth, C.W., Schreiber, D., Koethe, D., Faulhaber, J.,Klosterkotter, J., Piomelli, D., 2004. Cerebrospinal anandamide levels are elevatedin acute schizophrenia and are inversely correlated with psychotic symptoms.Neuropsychopharmacology 29, 2108–2114.

Glass, M., Dragunow, M., Faull, R.L., 1997. Cannabinoid receptors in the human brain: adetailed anatomical and quantitative autoradiographic study in the fetal, neonataland adult human brain. Neuroscience 77, 299–318.

Hanus, L.O., 2009. Pharmacological and therapeutic secrets of plant and brain (endo)cannabinoids. Med. Res. Rev. 29, 213–271.

Henquet, C., Krabbendam, L., Spauwen, J., Kaplan, C., Lieb, R., Wittchen, H.U., van Os, J.,2005. Prospective cohort study of cannabis use, predisposition for psychosis, andpsychotic symptoms in young people. BMJ 330, 11.

Herkenham, M., Lynn, A.B., Little, M.D., Johnson,M.R., Melvin, L.S., de Costa, B.R., Rice, K.C.,1990. Cannabinoid receptor localization in brain. Proc. Natl. Acad. Sci. U.S.A. 87,1932–1936.

Hilton, J., Yokoi, F., Dannals, R.F., Ravert, H.T., Szabo, Z., Wong, D.F., 2000. Column-switching HPLC for the analysis of plasma in PET imaging studies. Nuclear Med.Biol. 27, 627–630.

Horti, A.G., Van Laere, K., 2008. Development of radioligands for in vivo imaging of type 1cannabinoid receptors (CB1) in human brain. Curr. Pharm. Des. 14, 3363–3383.

Horti, A.G., Fan, H., Kuwabara, H., Hilton, J., Ravert, H.T., Holt, D.P., Alexander,M., Kumar, A.,Rahmim, A., Scheffel, U., Wong, D.F., Dannals, R.F., 2006. 11C-JHU75528: a radiotracerfor PET imaging of CB1 cannabinoid receptors. J. Nucl. Med. 47, 1689–1696.

Horti, A.G., Fan, H., Ravert, H.T., Holt, D.P., Hilton, J., Kumar, A., Alexander, M., Rahman,A., Hoffman, A., Lupica, C., Kuwabara, H., Wong, D.F., Dannals, R.F., 2007. [11C]JHU75528, a PET radioligand for imaging of cerebral cannabinoid CB1 receptors. 39-th meeting of European Brain and Behaviour Society, Trieste, Italy.

Howlett, A.C., Barth, F., Bonner, T.I., Cabral, G., Casellas, P., Devane, W.A., Felder, C.C.,Herkenham, M., Mackie, K., Martin, B.R., Mechoulam, R., Pertwee, R.G., 2002.International Union of Pharmacology. XXVII. Classification of cannabinoidreceptors. Pharmacol. Rev. 54, 161–202.

Innis, R.B., Cunningham, V.J., Delforge, J., Fujita, M., Gjedde, A., Gunn, R.N., Holden, J.,Houle, S., Huang, S.C., Ichise,M., Iida, H., Ito, H., Kimura, Y., Koeppe, R.A., Knudsen, G.M.,

1513D.F. Wong et al. / NeuroImage 52 (2010) 1505–1513

Knuuti, J., Lammertsma, A.A., Laruelle,M., Logan, J.,Maguire, R.P.,Mintun,M.A.,Morris,E.D., Parsey, R., Price, J.C., Slifstein, M., Sossi, V., Suhara, T., Votaw, J.R., Wong, D.F.,Carson, R.E., 2007. Consensus nomenclature for in vivo imaging of reversibly bindingradioligands. J. Cereb. Blood Flow Metab. 27, 1533–1539.

Jones, J.P., Rahman, A., Sibomana, M., Crabb, A.H., Burbar, Z., Cavanaugh, C.B., Michel, C.,Wong Dean, F., 2006. Data processing methods for a high throughput brain imagingPET research center. IEEE Medical Imaging Conference.

Koethe, D., Llenos, I.C., Dulay, J.R., Hoyer, C., Torrey, E.F., Leweke, F.M., Weis, S., 2007.Expression of CB1 cannabinoid receptor in the anterior cingulate cortex inschizophrenia, bipolar disorder, and major depression. J. Neural Transm. 114,1055–1063.

Leweke, F.M., Giuffrida, A., Wurster, U., Emrich, H.M., Piomelli, D., 1999. Elevatedendogenous cannabinoids in schizophrenia. Neuroreport 10, 1665–1669.

Logan, J., Fowler, J.S., Volkow, N.D., Wolf, A.P., Dewey, S.L., Schlyer, D.J., Macgregor, R.R.,Hitzemann, R., Bendriem, B., Gatley, S.J., 1990. Graphical analysis of reversibleradioligand binding from time–activitymeasurements applied to [N- 11 C-methyl]-(−)-cocaine PET studies in human subjects. J. Cereb. Blood Flow Metab. 10, 740.

Mato, S., Del Olmo, E., Pazos, A., 2003. Ontogenetic development of cannabinoidreceptor expression and signal transduction functionality in the human brain. Eur.J. Neurosci. 17, 1747–1754.

Mechoulam, R., 1986. Interview with Prof. Raphael Mechoulam, codiscoverer of THC.Interview by Stanley Einstein. Int. J. Addict. 21, 579–587.

Mouslech, Z., Valla, V., 2009. Endocannabinoid system: an overview of its potential incurrent medical practice. Neuro Endocrinol. Lett. 30, 153–179.

Newell, K.A., Deng, C., Huang, X.F., 2006. Increased cannabinoid receptor density in theposterior cingulate cortex in schizophrenia. Exp. Brain Res. 172, 556–560.

Onaivi, E.S., Ishiguro, H., Gong, J.P., Patel, S., Meozzi, P.A., Myers, L., Perchuk, A., Mora, Z.,Tagliaferro, P.A., Gardner, E., Brusco, A., Akinshola, B.E., Hope, B., Lujilde, J., Inada, T.,Iwasaki, S., Macharia, D., Teasenfitz, L., Arinami, T., Uhl, G.R., 2008. Brain neuronalCB2 cannabinoid receptors in drug abuse and depression: from mice to humansubjects. PLoS ONE 3, e1640.

Pertwee, R.G., 2008. Ligands that target cannabinoid receptors in the brain: from THC toanandamide and beyond. Addict. Biol. 13, 147–159.

Rahmim, A., Cheng, J.C., Blinder, S., Camborde, M.L., Sossi, V., 2005. Statistical dynamicimage reconstruction in state-of-the-art high-resolution PET. Phys. Med. Biol. 50,4887–4912.

Resnick, S.M., 1992. Matching for education in studies of schizophrenia. Arch. Gen.Psychiatry 49, 246.

Secher, A., Husum, H., Holst, B., Egerod, K.L., Mellerup, E., 2010. Risperidone treatmentincreases CB1 receptor binding in rat brain. Neuroendocrinology 91, 155–168.

Sossi, V., De Jong, M., Barker, W., Bloomfield, P., Burbar, Z., Camborde, M.L., Comtat, C.,Eriksson, L.A., Houle, S., Houle, S., Keator, D., Knob, C., Krais, R., Lammertsma, A.A.,Rahmim, A., Sibomana, M., Teras, M., Thompson, C.J., Trebossen, R., Votaw, J., Walker,M.J., Wienhard, K., Wong, D.F., 2005. The second generation HRRT: a multi-centrescanner performance investigation. IEEE Nucl. Sci. Symp. Conf, pp. 2195–2199.

Sundram, S., Copolov, D., Dean, B., 2005. Clozapine decreases [3H] CP 55940 binding tothe cannabinoid 1 receptor in the rat nucleus accumbens. Naunyn SchmiedebergsArch. Pharmacol. 371, 428–433.

Talairach, Tornoux, 1988. Co-Planar Stereotaxic Atlas of the Human Brain. ThiemeMedical Publishers, New York.

Terry, G.E., Liow, J.S., Zoghbi, S.S., Hirvonen, J., Farris, A.G., Lerner, A., Tauscher, J.T.,Schaus, J.M., Phebus, L., Felder, C.C., Morse, C.L., Hong, J.S., Pike, V.W., Halldin, C.,Innis, R.B., 2009. Quantitation of cannabinoid CB1 receptors in healthy human brainusing positron emission tomography and an inverse agonist radioligand. Neuro-image 48, 362–370.

Theisen, F.M., Haberhausen, M., Firnges, M.A., Gregory, P., Reinders, J.H., Remschmidt,H., Hebebrand, J., Antel, J., 2007. No evidence for binding of clozapine, olanzapine

and/or haloperidol to selected receptors involved in body weight regulation.Pharmacogenomics J. 7, 275–281.

Tzourio-Mazoyer, N., Landeau, B., Papathanassiou, D., Crivello, F., Etard, O., Delcroix, N.,Mazoyer, B., Joliot, M., 2002. Automated anatomical labeling of activations in SPMusing a macroscopic anatomical parcellation of the MNI MRI single-subject brain.Neuroimage 15, 273–289.

Ujike, H., Takaki, M., Nakata, K., Tanaka, Y., Takeda, T., Kodama, M., Fujiwara, Y., Sakai, A.,Kuroda, S., 2002. CNR1, central cannabinoid receptor gene, associated withsusceptibility to hebephrenic schizophrenia. Mol. Psychiatry 7, 515–518.

Uriguen, L., Garcia-Fuster, M.J., Callado, L.F., Morentin, B., La Harpe, R., Casado, V., Lluis,C., Franco, R., Garcia-Sevilla, J.A., Meana, J.J., 2009. Immunodensity and mRNAexpression of A2A adenosine, D2 dopamine, and CB1 cannabinoid receptors inpostmortem frontal cortex of subjects with schizophrenia: effect of antipsychotictreatment. Psychopharmacology 206, 313–324.

Van Laere, K., Goffin, K., Casteels, C., Dupont, P., Mortelmans, L., de Hoon, J., Bormans, G.,2008a. Gender-dependent increases with healthy aging of the human cerebralcannabinoid-type 1 receptor binding using [(18)F]MK-9470 PET. Neuroimage 39,1533–1541.

Van Laere, K., Koole, M., Sanabria Bohorquez, S.M., Goffin, K., Guenther, I., Belanger, M.J.,Cote, J., Rothenberg, P., De Lepeleire, I., Grachev, I.D., Hargreaves, R.J., Bormans, G.,Burns, H.D., 2008b. Whole-body biodistribution and radiation dosimetry of thehuman cannabinoid type-1 receptor ligand 18F-MK-9470 in healthy subjects.J. Nucl. Med. 49, 439–445.

Van Laere, K., Goffin, K., Bormans, G., Casteels, C., Mortelmans, L., de Hoon, J., Grachev, I.,Vandenbulcke, M., Pieters, G., 2009. Relationship of type 1 cannabinoid receptoravailability in the human brain to novelty-seeking temperament. Arch. Gen.Psychiatry 66, 196–204.

van Os, J., Bak, M., Hanssen, M., Bijl, R.V., de Graaf, R., Verdoux, H., 2002. Cannabis use andpsychosis: a longitudinal population-based study. Am. J. Epidemiol. 156, 319–327.

Vogt, B.A., Laureys, S., 2005. Posterior cingulate, precuneal and retrosplenial cortices:cytology and components of the neural network correlates of consciousness. Prog.Brain Res. 150, 205–217.

Vogt, B.A., Berger, G.R., Derbyshire, S.W., 2003. Structural and functional dichotomy ofhuman midcingulate cortex. Eur. J. Neurosci. 18, 3134–3144.

Weston-Green, K., Huang, X.F., Han, M., Deng, C., 2008. The effects of antipsychotics onthe density of cannabinoid receptors in the dorsal vagal complex of rats:implications for olanzapine-induced weight gain. Int. J. Neuropsychopharmacol.11, 827–835.

Wong, D.F., Kuwabara, H., Horti, A., Kumar, A., Brasic, J., Ye, W., Alexander, M., Raymont,J., Galecki, J., Charlotte, M., Cascella, N., 2008a. PET imaging of cannabinoid CB1 typereceptors in healthy humans and patients with schizophrenia using [11C]OMAR.NeuroReceptor Mapp. 41, T51.

Wong, D.F., Kuwabara, H., Horti, A.G., Kumar, A., Brasic, J., Ye, W., Alexander, M., Hilton,J., Williams, V., Ravert, H.T., Dannals, R.F., 2008b. Imaging of human cannaboid CB1type human receptors with [11C]OMAR. 55th Annual Meeting of the Society ofNuclear Medicine, New Orleans, Louisiana.

Yasuno, F., Brown, A.K., Zoghbi, S.S., Krushinski, J.H., Chernet, E., Tauscher, J., Schaus,J.M., Phebus, L.A., Chesterfield, A.K., Felder, C.C., Gladding, R.L., Hong, J., Halldin,C., Pike, V.W., Innis, R.B., 2008. The PET radioligand [11C]MePPEP binds reversibly andwith high specific signal to cannabinoid CB1 receptors in nonhuman primate brain.Neuropsychopharmacology 33, 259–269.

Zammit, S., Allebeck, P., Andreasson, S., Lundberg, I., Lewis, G., 2002. Self reportedcannabis use as a risk factor for schizophrenia in Swedish conscripts of 1969:historical cohort study. BMJ 325, 1199.

Zavitsanou, K., Garrick, T., Huang, X.F., 2004. Selective antagonist [3H]SR141716Abinding to cannabinoid CB1 receptors is increased in the anterior cingulate cortexin schizophrenia. Prog. Neuropsychopharmacol. Biol. Psychiatry 28, 355–360.