D2/D3 dopamine receptor binding with - Schizophrenia Research

Schizophrenia Research 85 (2006) 232 – 244 www.elsevier.com/locate/schres

D2/D3 dopamine receptor binding with [F-18]fallypride in thalamus and cortex of patients with schizophrenia Monte S. Buchsbaum a,c,e,*, Bradley T. Christian b,d, Douglas S. Lehrer b,c, Tanjore K. Narayanan b, Bingzhi Shi b, Joseph Mantil b, Eileen Kemether a, Terrence R. Oakes d, Jogeshwar Mukherjee b,e a

Department of Psychiatry, Mt. Sinai School of Medicine, 1 Gustave Levy Place, Box 1505, New York, NY 10029, USA b Boonshoft Schizophrenia Center/Wallace–Kettering Neuroscience Institute, USA c Wright State University Department of Psychiatry, USA d Waisman Laboratory for Brain Imaging, University of Wisconsin-Madison, USA e Department of Psychiatry and Human Behavior/University of California-Irvine (Dr. Buchsbaum emeritus), USA Received 27 December 2005; received in revised form 11 March 2006; accepted 17 March 2006 Available online 19 May 2006

Abstract Background: Abnormalities in the dopaminergic system are implicated in schizophrenia. [F-18]fallypride is a highly selective, high affinity PET ligand well suited for measuring D2/D3 receptor availability in the extrastriatal regions of the brain including thalamus, prefrontal, cingulate, and temporal cortex, brain regions implicated in schizophrenia with other imaging modalities. Methods: Resting [F-18]fallypride PET studies were acquired together with anatomical MRI for accurate coregistration and image analysis on 15 drug naı¨ve schizophrenics (10 men, 5 women, mean age 28.5 years) and 15 matched controls (9 men, 6 women, mean age 27.4 years). Dopamine D2/D3 receptor levels were measured as binding potential (BP). The fallypride BP images of each subject were spatially normalized and subsequently smoothed for group comparison. Measures of significance between the schizophrenic and control groups were determined using statistical parametric mapping (SPM). The medial dorsal nucleus and pulvinar were also traced on coregistered MRI for detailed assessment of BP in these regions. Results: The thalamus of patients with schizophrenia had lower [F-18]fallypride BP than normal controls and this was the brain area with the greatest difference (range 8.5% to 27.2%). Left medial dorsal nucleus and left pulvinar showed the greatest decreases (21.6% and 27.2% respectively). The patients with schizophrenia also demonstrated D2/D3 BP reduction in the amygdala region, cingulate gyrus, and the temporal cortices. Conclusions: These findings suggest that drug naı¨ve patients with schizophrenia have significant reductions in extrastratial D2/ D3 receptor availability. The reductions were most prominent in regions of the thalamus, replicating other studies both with

* Corresponding author. Mount Sinai School of Medicine, Box 1505, 1 Gustave Levy Place, New York, NY 10029, USA. Tel.: +1 212 241 5294; fax: +1 212 423 0819. E-mail address: [email protected] (M.S. Buchsbaum). 0920-9964/$ - see front matter D 2006 Elsevier B.V. All rights reserved. doi:10.1016/j.schres.2006.03.042

M.S. Buchsbaum et al. / Schizophrenia Research 85 (2006) 232–244

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high affinity D2/D3 ligands and consistent with FDG-PET studies, further supporting the hypothesis of thalamic abnormalities in this patient population. D 2006 Elsevier B.V. All rights reserved. Keywords: Dopamine receptor; Thalamic association nuclei; Laterality; Limbic system

1. Introduction The clinical efficacy of neuroleptics and their main action of blocking D2 receptors suggested that PET studies of dopamine receptor ligands would reveal differences between patients with schizophrenia and normal volunteers. The first study by Comar et al. (1979) with [C-11]chlorpromazine revealed a widespread pattern of cortical and subcortical uptake due to nonspecific binding. Subsequent studies with more specific ligands such as [C-11]methylspiperone (Wong et al., 1986) and [C-11]raclopride (Farde et al., 1990) focused exclusively on the striatum and had variable results (see review (Tune et al., 1993)). The development of high-affinity ligands such as [C11]FLB 457 (Olsson et al., 1999) and [F-18]fallypride (Mukherjee et al., 1999) opened the possibility of examining other brain regions identified as abnormal in functional activation imaging with PET, SPECT, EEG, and fMRI (see recent reviews) — the prefrontal cortex (Molina et al., 2005) (Andreasen et al., 1997; Buchsbaum and Hazlett, 1998) (Eyler et al., 2004; Suzuki et al., 2005), thalamus (Andreasen, 1997; Buchsbaum et al., 1996; Hazlett et al., 2004), cingulate gyrus (Haznedar et al., 2004; Quintana et al., 2004; Yasuno et al., 2005), and temporal lobe (Eyler et al., 2004) (Eyler et al., 2004). The medial dorsal nucleus (MDN), the largest association nucleus in the thalamus, has its major reciprocal connections with the prefrontal cortex, and is thus a prime candidate region for a schizophrenia diathesis. When thalamic nuclei were traced on coregistered MRI, decreased metabolic rates were found in the MDN in patients with schizophrenia in comparison to normal controls (Hazlett et al., 2004). This region is also known to have cell loss and volume reduction in patients with schizophrenia in postmortem studies (Byne et al., 2002; Danos et al., 2005; Pakkenberg, 1992; Popken et al., 2000; Young et al., 2000). In an early postmortem study, Oke

suggested elevated thalamic dopamine as important in schizophrenia (Oke et al., 1992). Autoradiographic and PET studies confirm moderate density D2/D3 sites in the MDN. Epidepride binding was about twice as high in the MDN than lateral dorsal or geniculate nuclei in autoradiographic studies (Rieck et al., 2004). PET studies have also confirmed thalamic D2/D3 binding (Farde et al., 1997; Mukherjee et al., 2002; Okubo et al., 1999; Rieck et al., 2004; Sedvall and Farde, 1995; Suhara et al., 2002; Talvik et al., 2003). There was markedly higher D2/D3 binding in the MDN and anterior nuclei of the thalamus than other thalamic areas (Okubo et al., 1999) and nearly twice as high in medial than in lateral thalamus in normal controls in PET measurement with [C-11]FLB 457 (Talvik et al., 2003). Several studies in patients with schizophrenia with high affinity ligands have found low D2 binding in the medial regions of the thalamus. After dividing the thalamus into lateral and medial segments, low D2 binding was found in previously untreated patients with schizophrenia (Talvik et al., 2003). Using a thalamic map derived from our earlier report (Buchsbaum et al., 1996), Yasuno found diminished [C-11]FLB 457 binding in the ventral medial and posterior subregions of the thalamus in 10 never previously medicated schizophrenics in comparison to 19 normal controls (Yasuno et al., 2004). Another PET study with [C-11]FLB 457 (Suhara et al., 2002) found significantly decreased binding potential in the anterior cingulate with a smaller effect in the whole thalamus (3.31 in never previously medicated patients and 3.58 in normals was p = 0.06, approximate effect size 0.79). The goal of this work was to use the high affinity, D2/D3 dopamine selective PET radioligand, [F18]fallypride to confirm and extend these earlier findings in never-medicated patients with significance probability mapping and anatomical tracing of the major nuclei of the thalamus.

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2. Methods 2.1. Subjects 2.1.1. Schizophrenia and related conditions Fifteen psychotic patients (10 men; 5 women; mean age, 28.5; SD, 8.9; 15 right-handed) were recruited from the greater Dayton, Ohio, area, and were evenly divided between inpatients and outpatients. After complete description of the study, all subjects completed a verbal binformed consent posttest.Q All participants passed this test and gave written informed consent. Subjects underwent evaluation using the Comprehensive Assessment of Symptoms and History (CASH) (Andreasen et al., 1992) Brief Psychiatric Rating Scale (BPRS, 18-item version) and

Abnormal Involuntary Movement Scale (AIMS) (Psychopharmacology-Research-Branch, 1976), and were diagnosed according to DSM-IV (AmericanPsychiatric-Association, 1994) by a staff psychiatrist (D.S.L.). Patients were neuroleptic naı¨ve (n = 12) or almost neuroleptic naı¨ve (n = 3, defined in Table 1). All patients were negative for drugs of abuse on a urine screen at the time of the scan and smoking was prohibited on the scan day. Subject characteristics are summarized in Table 1. Following study evaluation, all subjects were immediately referred for psychiatric treatment. 2.1.2. Controls Fifteen normal control subjects (9 men; 6 women; mean age, 27.4; SD, 7.9; 14 right-handed) were age-

Table 1 Demographic and psychopathological data of healthy control subjects and patients with schizophrenia Parameter

Controlsa (n = 15)

Patientsa (n = 15)

Sex, no. M/F Race, no. white/African American/Afro-Caribbean/ Mixed Age, y ( p = 71) Education, y ( p = 11) Social classb p = 66 Handedness, no. right/left-handed Primary Diagnosis, DSM-IV (no. by diagnosis) Schizophrenia, paranoid type Schizophrenia, undifferentiated typec Schizoaffective disorder Schizophreniform disorderd Secondary Diagnosis, DSM-IV (no. by diagnosis) Alcohol abuse Schizotypal personality disorder Duration of illness, median/mean, wk Locus of care at time of study, no. inpatient/outpatient BPRS total score GAF score (past month) AIMSe, no. with total scores N 0 (mean score of positive subjects) Medication status Neuroleptic naı¨ve, number Almost neuroleptic naı¨vef

9/6 12/0/2/1 27.4 (7.9) 14.7 (2.1) 33.1(7.8) 14/1 NA

10/5 12/3/0/0 28.5 (8.9) 13.3 (2.4) 35.4 (18.0) 15/0

a

7 4 2 2

NA NA NA NA 2 NA

1 1 26/166 (257.3) 7/8 52.7 (9.1) 30.3 (9.7) (1.5) 12 3

All figures represent mean values (standard deviation) unless otherwise specified. Hollingshead Two Factor Index of Social Position (Hollingshead and Redlich, 1958); data for one ill subject was unobtainable; mean represents data for 11 remaining ill subjects. c Two subjects initially diagnosed as schizophreniform were determined at follow-up to be schizophrenic. d In both patients, symptoms fully remitted within six months; one patient has remained continuously on antipsychotic medication without return of psychotic symptoms; the other patient relapsed almost two years later after having been off of antipsychotics for one year. e Symptom scores are rated on a 0–5 scale, with 0 = none, 1 = questionable, 2 = mild, 3 = moderate, 4 = marked, and 5 = severe symptoms. f Three previously-medicated subjects reported lifetime neuroleptic exposure of: (1) no more than five doses, several years prior to study evaluation; (2) one intramuscular haloperidol hydrochloride injection one week prior to study; and, (3) 28 lifetime doses of risperidone, never more than 10 in a single year, most recent dosage three weeks before study. All of these doses would have negligible pharmacological action at the time of the study. b


and sex-matched to experimental subjects. Controls underwent psychiatric assessment performed by a staff psychiatrist (D.S.L.). Participating subjects had no history of psychiatric illness, substance use disorder, clinically significant head trauma, or neurological disease. Controls did not significantly differ from ill subjects with respect to sex, age ( p = 0.71), race, handedness, education ( p = 0.11), or family-oforigin socioeconomic status ( p = 0.66). Control subject characteristics are summarized in Table 1. The results of PET scans measuring [F-18]fluorodeoxyglucose (FDG) uptake in the MDN of a subset of these patients has recently been reported (Lehrer et al., 2005).

ligand-receptor binding) for the data analysis. Multiple, circular regions of interest were placed over outer lobules of the cerebellum, covering two 3.5 mm thick transaxial planes. 2.3. Data analysis 2.3.1. Binding potential parametric image analysis Parametric images of [F-18]fallypride binding potential were generated to permit group comparison over the entire volume of the brain using the SPM software. For this work, apparent binding potential is described as: BP ¼

2.2. PET Scans All subjects refrained from alcohol, smoking, and caffeine 4 h before the PET scanning session. The PET scans were acquired using an ECAT EXACT HR+ scanner (Brix et al., 1997) in 3D mode. The subjects were placed in the scanner in the supine position, with the brain centered in the axial field of view. A 5 minute transmission scan was first acquired using a 68Ge/68Ga rod source to correct for the attenuation of photons. The head was fixed with a piece of surgical tape. Dynamic acquisition of the PET dynamic data was initiated with the 30 s bolus injection of [F-18]fallypride. The radiopharmaceutical was produced according to previously reported methods (Mukherjee et al., 1995) at high specific activity (N 2800 Ci/mmole) with a dose of 0.7 mCi/10 kg (range of 3.7–7.4 mCi). In an effort to increase subject compliance and minimize subject discomfort, a split session PET imaging protocol was implemented during the uptake of radiotracer. A split session PET involved 60 min of scanning (5 oneminute frames, 5 two-minute frames, 9 five-minute frames), 10-min break, and 50 min of scanning (10 five-minute frames) for a total PET imaging session of 2 h. The data were reconstructed using the ECAT v7.2 OSEM (3 iterations, 16 subsets) following the corrections applied for attenuation, normalization, and scatter. Prior to further data processing the dynamic frames were spatially aligned using the AIR 3.08 software (Woods et al., 1992) to correct for patient motion. The cerebellum was used to represent the reference region (with negligible specific

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f2 Bmax V ; KD

where, f 2 is the unbound fraction of radioligand in the free cellular space, B max V is the available receptor density, and K D is the equilibrium dissociation constant. The BP images reported here were calculated using a multilinear variation (Ichise et al., 2002) of the Logan (Logan et al., 1996) distribution volume ratio (DVR). This multilinear method of BP calculation was shown to be least sensitive to noise in the dynamic PET data. This model employs the use of a tissue reference region (cerebellum) to represent the kinetics of unbound radioligand in the tissue (Cunningham et al., 1991; Logan et al., 1996). Reference tissue methods were also found suitable for mapping of the thalamus with fallypride by Siessmeier et al. (2005). The functional equation in the multilinear form is given as: Z 0

T

C ðt Þdt ¼ DVR

Z

T

0

Cref ðt Þdt þ

Cref ðT Þ k¯ 2

þ bWC ðT Þ; where C(t) is the PET measured time varying voxel concentration, C ref(t) is the reference (cerebellum) concentration, DVR is the distribution volume ratio, k¯ 2 is the average tissue-to-plasma efflux constant and bW is a constant of integration and T representing the time midpoint of each PET frame. The advantage of this method is that the integration of the tissue concentration (C(t)) tends to reduce the parameter bias introduced by the higher noise levels present in

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the voxel-by-voxel data. The apparent binding potential is then calculated as BP= DVR-1. 2.3.2. Statistical approach Since we hypothesized a thalamic decrease in BP we first evaluated the mean BP within targeted traced regions of interest with repeated measures MANOVA using the traditional p b 0.05 criterion. Then to document the exact shape of the thalamic area showing significance, we examined significance probability maps of the brain through the thalamic level with threshold at p b 0.005, uncorrected, using the SPM package. Lastly we explored the entire brain at a corrected p threshold level of 0.05 and with a color bar scale indicating the p value between p b 0.05 and the lowest p value. This allows readers to chose the appropriate p value for their own application. Other authors have published region of interest data outside the thalamus including the anterior cingulate cortex (Suhara et al., 2002) and temporal cortex (Tuppurainen et al., 2003). Any replication needs to be at p b 0.05; it would introduce Type II bias if we tested our own hypothesized region, the thalamus at p b 0.05 but required the anterior cingulate findings of Suhara (Suhara et al., 2002) to be tested at 0.005 or lower. Similarly, since we have already reported the traced medial dorsal nucleus to be low in volume and metabolic activity in two other samples (Byne et al., 2001; Kemether et al., 2003), it would provide Type II error bias to present only 0.005 rather than 0.05 significance probability mapping of the MDN. 2.3.3. ROI based analysis One of the authors with established reliability (E.K.) traced the whole thalamus, MDN and pulvinar on raw unresliced and coded MRI anatomical images exactly as previously reported (Byne et al., 2001; Kemether et al., 2003). These tracings were used to create binary mask images. Unsmoothed fallypride images were coregistered to these anatomical images by first coregistering the early integrated fallypride data (0–5 min) using the SPM registration algorithm (Friston, 1995) then applying this transformation matrix to the BP image as above. Binding potential within these areas and the whole thalamus not including the MDN or pulvinar were obtained by applying the binary mask images to the [F-18]fallypride BP images.

2.3.4. Spatial normalization for exploratory significance probability mapping For many radiotracer studies, such as [O-15]water and FDG, it is possible to directly transform an image of integrated radiotracer uptake into a normalized coordinate system. However, for [F-18]fallypride BP images there are several preprocessing steps that need to be performed. The highly selective D2/D3 binding of [F-18]fallypride results in an image dominated with striatal information and only limited information in the cortical regions. As a result the current algorithms for image coregistration and normalization fail when trying to match [F-18]fallypride with MRI, FDG PET or water PET. We have previously reported a comparison of various methods to spatially normalize the data (Christian et al., 2004). For the work reported here, the following steps were undertaken to spatially normalize the [F-18]fallypride BP images: 1. Create an image of the first several minutes of [F-18]fallypride uptake (following the bolus), this image represents primarily the delivery (i.e. unbound) of fallypride throughout the brain, also mixed with significant bound fallypride in the high D2/D3 density regions such as the striatum. 2. Spatially normalize the data from 1) to the Montreal Neurological Institute (MNI) FDG template, using 7 9 7 basis functions, 16 nonlinear iterations. 3. Apply the transformation matrices from 2) to the BP parametric images. Sum the transformed BP images. 4. Spatially normalize the individual BP parametric images to the image created in 3). 5. Sum the transformed images from 4) to create the fallypride BP template. This template can then be used for all spatial normalizations. The MNI image dimensions were chosen for the size of the bounding box of the output image. 6. Smooth all of the normalized images (8 mm filter) before the application of SPM statistical analysis but not for the application of the anatomical tracings. For many radioligands, the first several minutes of tracer uptake in the brain represents primarily ligand delivery, i.e. flow, thus a template representing blood flow such as [O-15]water would be the most suitable.


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Table 2 Mean thalamic nuclei binding potential Structure

Normals Left

Medial dorsal Pulvinar Residual thalamus a b

2.18 1.14 2.57

Right 2.15 1.16 2.36

Patients with schizophrenia

% change

Left

Right

Left

Right

1.84 1.04 2.16

21.6% 27.2% 12.5%

14.4% 10.3% 8.5%

a

1.7 0.83b 2.25

t = 2.66, df = 24, p = 0.014. t = 3.54, df = 24, p = 0.0016.

However, with [F-18]fallypride, the binding in the high D2/D3 receptor density regions such as the striatum is so rapid, that the first several minutes of PET data is actually dominated by radioligand in the bound state (Christian et al., 2004). For this reason, we have chosen to use the FDG template for the spatial transformation of step 2) into standardized space. 2.3.5. SPM analysis Following spatial normalization and spatial smoothing (8 mm) of the [F-18]fallypride BP images, statistical parametric maps of the groups was performed using SPM2 (http://www.fil.ion.ucl.ac.uk/spm/ spm2.html). A schizophrenic and control group comparison was made using the two-sample t-test criterion. There was no global scaling applied in the analysis, as the BP images are directly comparable. The t-test results were then displayed on the MNI template with a color bar to indicate level of significance.

3. Results 3.1. MDN and pulvinar Patients with schizophrenia had lower BP in the MDN (1.78 F 0.53) than normal volunteers (2.16 F 0.36; t = 2.15, df = 1,24, p = 0.041, mean of left and right sided ROI). This was also true for the pulvinar (0.94 F 0.27 vs. 1.15 F 0.18, t = 2.27, p = 0.03). However the remainder of the thalamus after removal of the areas of the medial dorsal and pulvinar showed no significant group difference (2.21 F 0.92 vs. 2.47 F0.46, t = 0.89, p = 0.37). The entire thalamus did differ (1.34 F .36 vs. 1.58 F 21, t = 2.12, p = 0.04). When the medial dorsal and pulvinar were entered into a diagnostic group (normal, patient) structure (MDN, pulvinar) hemisphere (right, left) there was a main effect of group ( F = 5.17, df = 1,24, p = 0.032) and a group hemisphere interaction ( F = 5.58,

Fig. 1. [F-18]Fallypride binding potential in the brain, p b 0.005 in yellow on MNI anatomical brain background. Yellow indicates normals N patients with schizophrenia. Significant area is most prominent in medial dorsal and pulvinar with a third area in the posterior portion of the striatum. MNI z level from SPM program. Images shown in standard orientation (image left is brain right hemisphere).

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Fig. 2. 3D (orthogonal) view of [F-18]fallypride binding differences between normals and patients with schizophrenia. The color yellow indicates normals N patients with schizophrenia. The second row with the p b 0.05 probability level reveals areas in the cingulate (indicated by an arrow in the first column) also found significant by Suhara et al., 2002). The SPM maps are overlaid on spatially normalized images of summed data (0–5 min fallypride representing early radioligand delivery and binding for anatomical reference).

df = 1,24, p = 0.027) indicating a larger left hemisphere than right hemisphere difference between normals and schizophrenics but higher order interactions with diagnostic group were not significant (see Table 2). Since low BP in the centromedian and posterior subregions of the thalamus was found associated with high BPRS positive symptoms earlier (Yasuno et al., 2004), we examined the BPRS subscale scores vs. right + left BP value correlations in our own data. There were significant negative correlations (r b 0.55, p = 0.05, two tailed, 0.47, 1 tailed in replication) between whole thalamus and hostility and suspiciousness ( 0.59) and with positive symptoms ( 0.49) and between MDN and pulvinar and hostility and suspiciousness ( 0.81, 0.86 respectively). However since this earlier report (Yasuno et al., 2004) examined the centromedian nucleus, which is outside our two traced nuclei, we also examined the residual thalamus (outside the

two traced nuclei) and found correlations with BPRS Total ( 0.51) positive symptoms ( 0.73), hostility and suspiciousness ( 0.69), and general symptoms ( 0.57). Thus, our data, associating significantly more marked symptoms with low BP in areas outside the medial dorsal, is quite consistent with the previous report (Yasuno et al., 2004) and appears to be more strongly contributed to by more lateral thalamic structures. There were no significant correlations with age for normals or patients matching earlier results (Talvik et al., 2003). 3.2. Exploratory mapping We present consecutive 2 mm slices through the thalamus (Fig. 1) with p b 0.005 threshold to explore the spatial extent of the thalamic BP differences. Areas of difference closely follow the traced areas in the MDN and pulvinar with the area in the thalamus

Fig. 3. [F-18]Fallypride binding differences between normals and patients with schizophrenia. The color yellow indicates normals N patients with schizophrenia. In each row, three consecutive slices are presented with the threshold set at p b 0.05 probability level and the color bar extending from t = 0 to the smallest value obtained in a map anywhere in the row. Thus in the second row, left column, z = 12, t values between 3.5 and 4.0 are revealed in the region of the left medial dorsal nucleus of the thalamus. The SPM maps are overlaid on spatially normalized anatomical MRI images from the SPM package. This allows an exploratory survey of the brain from the viewpoint of a confirmatory analysis examining consistency with postmortem binding studies, SPECT studies or other [F-18]fallypride studies to exploration of not otherwise considered areas at a protected p value of the reader’s choice.


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outside of these areas not reaching the threshold. Fig. 2 presents the 3D orthogonal view with the two statistical thresholds to reveal areas in the cingulate, frontal lobe and temporal lobe which also differ in showing lower BP and Fig. 3 presents a brain survey from z = 24 down to z = 24. Areas in the cingulate gyrus (rows 1–3, z = 26 through z = 0) and temporal lobe (rows 4–5, z = 12 to 24) show binding differences.

4. Discussion We found decreased binding potential with [F-18] fallypride, a D2/D3 high-affinity ligand in the MDN and pulvinar regions of the thalamus in patients with schizophrenia in comparison to normal controls. These data are consistent with three other studies. First, the reduced binding potential in the medial half of the thalamus divided geometrically has been reported in 9 never-medicated patients and 8 controls using [C-11]FLB 457 (Talvik et al., 2003). These findings were more prominent in the right medial area. Second, in a study of 10 never-medicated patients with schizophrenia and 19 normal controls also using [C-11]FLB 457 (Yasuno et al., 2004), the central medial and posterior regions of the thalamus were found to have lower BP in patients; these regions were identified using a thalamic template we developed and are the regions identified as the MDN and pulvinar. Third our findings are similar to a study (Suhara et al., 2002) of 11 never-medicated patients and 18 normals with [C-11]FLB 457 who showed a trend level ( p = 0.06) effect for circular regions of interest placed on the thalamus. In that study, normals had BP of 3.58 F 38 and patients 3.31 F 0.30 which showed an F = 3.90, p = 0.06. The anterior cingulate was significantly lower ( p b 0.02) in patients and the temporal cortex was marginally lower ( p = 0.16). Their cingulate cortex and temporal lobe values are replicated in our p b 0.05 maps (Figs. 2 and 3). The anterior cingulate was also found significantly low in a second FLB-457 study (Yasuno et al., 2005). The consistency of these findings over the three studies with our American sample as well as with patients in Sweden and Japan is noteworthy. The striatum was not analyzed in any of these studies and was not considered here due to long equilibration times. However, taken together with frontal and thalamic

findings, recent results with ligand imaging in the caudate (Abi-Dargham et al., 2000; Hirvonen et al., 2003) suggest that dopamine abnormalities may be present in all parts of the fronto-stratio-thalamic circuit. Both the cingulate (Choi et al., 2005; Haznedar et al., 2004; Quintana et al., 2004; Yamasue et al., 2004) and the temporal lobe (Yamasue et al., 2004) (Whitford et al., 2005) (Loberg et al., 2004) are regions for which we and others have recently reported diminished functional activity and smaller volume in patients with schizophrenia (see also review (Shenton et al., 2001) and reviews in recent articles). These findings were stronger in the left hemisphere than the right (greater than 20% reduction in the left medial thalamus and pulvinar). This difference is greater than the test–retest variability of less than 10% reported in our previous study in healthy volunteers (Mukherjee et al., 2002). Earlier studies showed stronger findings for the right medial thalamus (Talvik et al., 2003) and two studies averaged right and left sides (Suhara et al., 2002; Yasuno et al., 2004). Our own MDN volumetric findings showed that the decrease in volume was greater for schizophrenics on the left than the right (Kemether et al., 2003), but the diagnostic group asymmetry difference did not reach statistical significance as in the current fallypride results. These same patients did not show a FDG activation asymmetry in the medial dorsal nucleus (Lehrer et al., 2005) suggesting that any asymmetry is not related to activation, although subjects rested during the fallypride uptake so a systematic behavioral bias in dopaminergic activity cannot be ruled out. It should be noted that the low values were not found for the remainder of the thalamus outside the medial dorsal and pulvinar, a larger volume containing other nuclei, suggesting some anatomical specificity for the findings. The use of a specific manually-traced coregistered MRI template for the nuclei and thalamus and the actual size of the medial dorsal nucleus and pulvinar (each about 0.5 cc) minimize partial volume effects, but the contribution of the third ventricle, typically about 1 mm wide at this point, cannot be fully excluded. Head motion between the initial and final fallypride images might blur or misposition the anatomical template and fallypride image, but the specific localization of the effect to the region of the medial dorsal nucleus with ROI tracing, and the medial dorsal-pulvinar contour of


the matching SPM-identified regions tends to mitigate against group differences in head motion (which would tend to diminish normal-schizophrenia contrast) as a sole source of the effect. When using a composite parameter such as BP to serve as an index for receptor availability, B max, several interpretations of the results can be made. For example, the decrease in [F-18]fallypride binding in the schizophrenia cohort could be attributed to a decrease in D2/D3 receptor density or an increase in endogenous dopamine competition or changes in the apparent affinity (via K D). Following the traditional dopamine theory, we postulate that the decrease in [F-18]fallypride binding in the thalamus of the schizophrenics may be due, at least in part, to an increase in the endogenous dopamine concentration in the region of the synapse, thus competing with the [F-18]fallypride for the binding sites. This suggestion is consistent with findings in the striatum of patients with schizophrenia by Abi-Dargham and colleagues, reporting a baseline dopamine occupancy of the D2/D3 receptor sites of 19% vs. 9% in controls, using [I-123]IBZM before and after dopamine depletion with a-MPT (Abi-Dargham et al., 2000). However striatum-a-MPT depletion experiments may not be directly applicable to our thalamus findings and resolution of the question of 1) lower apparent B max (receptor downregulation) or 2) different radioligand or 3) increase in dopamine concentration must await further study. Our results in the thalamus appear to be largely attributable to D2 receptors since studies using quantitative autoradiography with [(125)I]7-OH-PIPAT, a relatively specific D3 agent, failed to demonstrate significant localization in thalamus, temporal lobe, or cingulate (Stanwood et al., 2000). Our current BP values are lower than those we previously reported (Mukherjee et al., 2002) because in that study we used circular regions of interest placed over regions of peak intensity vs. MRI traced over nuclei without reference to fallypride distribution. Comparing BPTs with [C-11]FLB 457 (Talvik et al., 2003) reveals somewhat higher BP for [C-11]FLB 457 than for fallypride due to the differing kinetics of the radioligands. While the D2 receptor is the most prominent of the dopamine receptors in the thalamus, it must be noted that low levels of D3 receptors are also found in the

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MDN (Gurevich and Joyce, 1999) and cannot be excluded as a partial source of our effects. Examination of slices at z = 8 (Fig. 3, right column, 5th row) suggests that differences in binding potential may also extend into the nucleus accumbens, an area relatively high in D3 receptors. The findings reported here of dopamine receptor change in the MDN of the thalamus in patients with schizophrenia are noteworthy in replicating several other receptor studies (Suhara et al., 2002; Talvik et al., 2003; Yasuno et al., 2004). They are also consistent with MRI volumetric reduction (Ananth et al., 2002; Andreasen et al., 1994; Byne et al., 2001; Kemether et al., 2003; Konick and Friedman, 2001), reduced FDG uptake (Buchsbaum et al., 1996; Hazlett et al., 2004; Lehrer et al., 2005) and postmortem studies (Byne et al., 2002; Pakkenberg, 1990, 1992; Young et al., 2000). These results, taken together with the strong and well-documented connections between the prefrontal regions and the MDN, implicate a fronto-thalamic circuitry deficit as important in the functional anatomy of schizophrenia.

Acknowledgements This work was supported by the Boonshoft Schizophrenia Center, the Wallace–Kettering Neuroscience Center, and by a grant to Dr. Buchsbaum, Anatomy and function of the thalamus in schizophrenia MH60023. The support of the United States Air Force, Air Force Research Laboratory (AFRL/HEOP), Air Force Materiel Command, under cooperative agreement F33615-98-2-6002, for use of imaging resources, is gratefully acknowledged. Marylin Brackney, Dr. King-Wai Chu, Kelly Dunigan, Kerry Kovacs, Candice Lee, Steve Mattmuller, Aaron Murray, Maruthi Narayanan, and Tonya Perkins, provided important technical support. Drs. Martin Satter (Kettering Medical Center), Nathaniel Alpert (Massachusetts General Hospital) and Chris Endres (Johns Hopkins) furnished valuable discussions on the analysis. MRI data were acquired by Dr. Mehdi Adenih. The project was approved by the IRBs of Kettering Medical Center, Wright State University and Mount Sinai School of Medicine. Gerald M. Szkotnicki, Executive Director, Wallace–Kettering Neuroscience Institute, Charles F. Kettering Memorial

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Hospital provided critical facilitating administrative and organizational support.

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