Imaging Brain Mu-Opioid Receptors in Abstinent Cocaine Users: Time Course and Relation to Cocaine Craving

Imaging Brain Mu-Opioid Receptors in Abstinent Cocaine Users: Time Course and Relation to Cocaine Craving David A. Gorelick, Yu Kyeong Kim, Badreddine Bencherif, Susan J. Boyd, Richard Nelson, Marc Copersino, Christopher J. Endres, Robert F. Dannals, and J. James Frost Background: Cocaine treatment upregulates brain mu-opioid receptors (mOR) in animals. Human data regarding this phenomenon are limited. We previously used positron emission tomography (PET) with [11C]-carfentanil to show increased mOR binding in brain regions of 10 cocaine-dependent men after 1 and 28 days of abstinence. Methods: Regional brain mOR binding potential (BP) was measured with [11C]carfentanil PET scanning in 17 cocaine users over 12 weeks of abstinence on a research ward and in 16 healthy control subjects. Results: Mu-opioid receptor BP was increased in the frontal, anterior cingulate, and lateral temporal cortex after 1 day of abstinence. Mu-opioid receptor BP remained elevated in the first two regions after 1 week and in the anterior cingulate and anterior frontal cortex after 12 weeks. Increased binding in some regions at 1 day and 1 week was positively correlated with self-reported cocaine craving. Mu-opioid receptor BP was significantly correlated with percentage of days with cocaine use and amount of cocaine used per day of use during the 2 weeks before admission and with urine benzoylecgonine concentration at the first PET scan. Conclusions: These results suggest that chronic cocaine use influences endogenous opioid systems in the human brain and might explain mechanisms of cocaine craving and reinforcement. Key Words: Cocaine, craving, mu-opioid receptor, PET scan, carfentanil, abstinence

C

ocaine addiction is considered a disease related chiefly to brain dopamine dysfunction, because cocaine’s primary psychoactive action is stimulation of the dopaminergic mesolimbic reward circuit by inhibition of the presynaptic dopamine transporter (Dackis and O’Brien 2001); however, other neurotransmitter systems modulate dopamine systems in the brain. In particular, mu-opioid receptors (mOR) are found on mesolimbic dopaminergic neurons (Pollard et al 1977) and are co-localized with dopamine (D2) receptors in the dorsolateral striatum and nucleus accumbens (Ambrose et al 2004). In rat studies, administration of cocaine in a chronic, intermittent (“binge”) pattern for 2 weeks increases brain mOR binding in regions relevant to reward, such as the nucleus accumbens, amygdala, and cingulate cortex (Branch et al 1992; Hammer 1989; Unterwald et al 1994, 2001). One to three days of cocaine treatment is sufficient to increase expression of mOR messenger ribonucleic acid (mRNA) in the nucleus accumbens and amygdala (Azaryan et al 1996a, 1996b; Yuferov et al 1999), although one study found an increase only in the amygdala (Rosin et al 2000). This effect seems to be mediated through dopamine receptors. It is blocked by co-administration of dopamine receptor antagonists (D1, D2, and D3 types) (Azaryan et al 1996a, 1996b). Administration (not necessarily in a binge pattern) of dopamine receptor agonists increases rodent striatal and nucleus

From the Intramural Research Program (DAG, SJB, RN, MC), National Institute on Drug Abuse, National Institutes of Health, Department of Health and Human Services; and Departments of Radiology and Radiological Sciences (YKK, BB, CJE, RFD, JJF), Johns Hopkins University School of Medicine, Baltimore, Maryland. Address reprint requests to David A. Gorelick, M.D., Ph.D., National Institute on Drug Abuse, Intramural Research Program, 5500 Nathan Shock Drive, Baltimore, MD 21224; E-mail: [email protected]. Received November 3, 2004; revised February 11, 2005; accepted February 18, 2005.

0006-3223/05/$30.00 doi:10.1016/j.biopsych.2005.02.026

accumbens mOR levels and decreases striatal enkephalin and pre-enkephalin mRNA levels (Azaryan et al 1996a; Chen et al 1993; George and Kertesz 1987), mimicking the effects of cocaine. Conversely, administration of dopamine (D2 type) receptor antagonists reduces rodent striatal mOR levels and increases striatal enkephalin and pre-enkephalin mRNA levels (Chen et al 1994; George and Kertesz 1987; Steiner and Gerfen 1999). The time course of the mOR change has not been well characterized. One study found decreased mOR binding in rat nucleus accumbens 10 days after the last of two 10-day periods of cocaine treatment (Sharpe et al 2000). There are few human data on the effect of cocaine on mOR. Women who used cocaine during pregnancy were found to have decreased mOR binding in the placenta compared with nondrug-using control subjects, with no change in binding affinity for naloxone (Wang and Schnoll 1987). A postmortem brain study found that cocaine users who died with cocaine and/or its metabolites present at death had decreased mOR binding and enkephalin mRNA levels in the striatum compared with noncocaine-using control subjects (Hurd and Herkenham 1993). We have previously shown that human cocaine users have increased mOR binding in several brain regions and that mOR binding in some regions correlates with self-reported cocaine craving (Zubieta et al 1996). We studied 10 cocaine-dependent men and 7 nondrug-using control subjects, using positron emission tomography (PET) with [11C]carfentanil, a potent, selective mOR agonist. Mu-opioid receptor binding was increased over control levels in the anterior cingulate, frontal and temporal cortex, caudate, and thalamus of the cocaine addicts studied 1– 4 days after their last cocaine use. Binding was positively correlated with the severity of self-reported cocaine craving in the frontal cortex, anterior cingulate, and amygdala. The increased mOR binding persisted after 4 weeks of monitored cocaine abstinence. We now report that brain mOR binding remains increased over the first week of cocaine abstinence, then declines to normal levels or remains relatively constant over the next 3 months, depending on the brain region. Furthermore, the inBIOL PSYCHIATRY 2005;57:1573–1582 © 2005 Society of Biological Psychiatry

1574 BIOL PSYCHIATRY 2005;57:1573–1582 creased binding in some regions is positively correlated with self-reported cocaine craving.

Methods and Materials Subjects Subjects were 17 adult, nontreatment-seeking cocaine users with current cocaine abuse or dependence according to DSM-IV criteria (American Psychiatric Association 1994) who were recruited from the community. The study was approved by the institutional review boards of the National Institute on Drug Abuse (NIDA) intramural research program (IRP) and the Johns Hopkins Bayview Medical Center. All subjects gave written informed consent (when not in acute drug intoxication or withdrawal) and were paid for their study participation. Subjects received a comprehensive medical and psychological evaluation, including medical history and physical examination, clinical laboratory tests, 12-lead electrocardiogram with 3-min rhythm strip, Addiction Severity Index (ASI; McLellan et al 1985), Diagnostic Interview Schedule (Robins et al 1982), and Symptom Check List-90R (SCL-90R; Derogatis 1983). Subject eligibility criteria included current use of cocaine averaging at least 1 g per week over the prior 3 months and at least 10 days of use over the prior 2 weeks; urine drug tests positive for cocaine and negative for other drugs; no clinically significant current abnormalities on the screening evaluation; no history of central nervous system disease, head injury with loss of consciousness greater than 3 min, or adverse event from prior cocaine use; no current psychiatric or substance use disorder except for cocaine and tobacco; not currently taking psychoactive medication; no use of opiates more than three times in the prior 3 months; intelligence quotient of at least 80; and ability to successfully perform study procedures. There were no subject eligibility criteria related to cigarette smoking. Subjects were 11 African American men, 4 African American women, and 2 white men, with a mean (SD) age of 34.0 (3.9) years (range, 24 –39 years). They all smoked cocaine and had used cocaine regularly for 7.2 (4.4) years, on 23.6 (4.5) of the 30 days before screening, and on 11.0 (2.8) of the 14 days before ward admission. They reported spending $569 ($521) on cocaine during those 2 weeks. Their last cocaine use was 16.0 (10.1) hours before admission. Use of other illegal drugs was negligible. No subject reported opiate use during the month before admission; all urine drug tests were negative for opiates. Twelve subjects (71%) were current cigarette smokers, four were never smokers, and one had abstained for 3 years. Analyses involving only the first PET scan included all 17 subjects. Analyses involving other PET scans included only 16 subjects because 1 subject had a technically inadequate second PET scan. Control subjects were 16 nondrug-using, healthy individuals (on the basis of medical history): 5 African American men, 3 African American women, and 8 white men. Their mean (SD) age was 33.6 (5.7) years (range, 24 – 45 years); 27% were current cigarette smokers. Control subjects were selected to be similar to cocaine-using subjects in age and male/female ratio because age and gender are known to influence brain mOR binding (Zubieta et al 1999). Study Design Subjects received a magnetic resonance imaging brain scan and were fitted with a thermoplastic face mask to ensure head immobilization during scanning. They were later admitted to the closed research ward of the NIDA IRP for a 3-month stay. To www.sobp.org/journal

D.A. Gorelick et al prevent unauthorized use of drugs on the ward, no passes or visitors were allowed, all personal belongings and mail were screened by staff, and random urine samples were collected and screened for drugs of abuse. After ward discharge, subjects were followed up periodically for up to 1 year (data to be presented elsewhere). While on the ward, subjects had PET scans done on 3 separate days: 1 day after ward admission (after 2 days for one subject, same day for one subject), approximately 1 week (6 –10 days) after admission (after 18 days for one subject), and approximately 12 weeks (80 – 87 days) after admission (after 13 weeks for one subject). Measures of self-reported cocaine craving (100-mm visual analogue scale), depression (SCL-90 depression subscale), and anxiety (SCL-90 anxiety subscale) were obtained just before each PET scan. Thirteen subjects also had an observed urine sample collected just before the first PET scan. This sample was quantitatively assayed by gas chromatography/mass spectrography for benzoylecgonine, the chief human metabolite of cocaine. Urine concentration of benzoylecgonine reflects the amount of cocaine taken during the preceding 2 to 3 days (Preston et al 1997, 2002). PET Image Acquisition Positron emission tomography scans were acquired in twodimensional mode on a GE 4096 Plus PET scanner (GE Medical Systems, Milwaukee, Wisconsin) with [11C]carfentanil, a muopioid agonist developed for PET imaging (Frost et al 1985). Images were simultaneously collected from 15 contiguous planes with an axial field of view of 10.5 cm. A transmission scan of 10-min duration was obtained with rotating germanium-68 rods before injection of the radiotracer. After intravenous bolus administration of 19.7 ⫾ 1.3 mCi (range, 16.3–21.6 mCi) of [11C]carfentanil (specific activity, 3312 ⫾ 2349 mCi/␮mol; range, 1604 –11,090 mCi/␮mol), 25 sets of images with variable time periods (6 ⫻ 30 sec, 5 ⫻ 60 sec, 5 ⫻ 120 sec, 9 ⫻ 480 sec) were acquired during a 90-min period for each study. After correction for attenuation with the transmission scan, images were reconstructed in a 128 ⫻ 128 ⫻ 15 matrix with pixel size of 2 ⫻ 2 ⫻ 6.5 mm with filtered back projection methods with a ramp filter, and decay corrected. Image Analysis Parameterized images for [11C]-carfentanil were generated with an automated in- house program based on Logan graphical analysis with a reference-tissue input function (occipital lobe), in which the fitted linear slope presents the ratio of the total distribution (DVR) in receptor-rich region to that in reference tissue (Logan et al 1996). The DVR ⫽ DVtissue/DVreference ⫽ (1 ⫹ k3/k4), where k3/k4 represents the binding potential (BP). Parameterized images of BP were computed from BP ⫽ DVR ⫺ 1. All scan data and parametric images were processed with SPM2 (Wellcome Department of Imaging Neuroscience, Institute of Neurology, University College London; http://www.fil.ion. ucl.ac.uk/spm/software/spm2/) and Analyze software packages (version 6.0, Mayo Foundation, Rochester, Minnesota). The 0 –90-min average image of each PET session was created to provide anatomic detail for spatial normalization. This image was then spatially normalized to a [11C]carfentanil template into a standard space (Montreal Neurological Institute) having voxel size equal to 2 ⫻ 2 ⫻ 2 mm. The transformation matrix created was then applied to the corresponding parameterized BP images. Data were then smoothed with a 10-mm full width at half maximum Gaussian smoothing kernel to reduce residual inter-

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Table 1. Increased Brain-Normalized Mu-Opioid Receptor Binding in Cocaine-Dependent Subjects over 12 Weeks of Enforced Abstinence: Comparison with 16 Healthy, Non–Drug-Using Control Subjects with Voxel-Based Analysis

Region Anterior Cingulate and Medial Frontal Anterior Frontal Dorsolateral Prefrontal Prefrontal Inferior Frontal Lateral Temporal

Day 1 of Abstinence (n ⫽ 17)

Day 7 of Abstinence (n ⫽ 16)

Day 90 of Abstinence (n ⫽ 16)

Side of Brain

Cluster ka (p)b

Max-z (x, y, z)c

Cluster ka (p)b

Max-z (x, y, z)c

Cluster ka (p)b

Max-z (x, y, z)c

R and L L R L R L R R L

693 (.003) 451 (.011) 229 (.057) 79 (.246) 128 (.144) 80 (.243) 142 (.125) 63 (.299) 94 (.207)

4.15 (4, 50, 16) 3.28 (⫺36, 46, 26) 3.73 (56, 18, 24) 3.28 (⫺56, 16, 18) 4.27 (20, 68, ⫺4) 3.45 (⫺10, 68, ⫺4) 3.38 (44, 46, 4) 3.21 (64, ⫺16, ⫺8) 3.31 (⫺64, ⫺46, ⫺8)

399 (.016) 79 (.246)

3.74 (6, 50, 14) 2.99 (⫺38, 36, 12)

424 (.014) 55 (.332)

8.32 (2, 46, 22) 3.29 (⫺38, 36, 12)

84 (.232)

3.74 (20, 68, ⫺6)

154 (.112)

3.59 (44, 44, 10)

R, right; L, left. Cluster size k represents number of voxels (2 ⫻ 2 ⫻ 2 mm3) at a height threshold of p ⫽ .005, uncorrected. p value for significance at cluster level. c Location of peak expressed as x, y, z coordinates in Montreal Neurological Institute space. a

b

subject anatomic differences, increase the signal-to-noise ratio at the expense of resolution, and increase the validity of statistical inferences by rendering data more normally distributed. The smoothed BP images were proportionally scaled to the same global brain mOR binding value to control for possible individual differences in mOR availability due to factors not assessed in the study. The resulting normalized BP of mOR was used in all statistical analyses. The mean global binding was computed on the subset of data greater than whole mean/8. The analysis threshold was set to ⬎1.3 times the mean global binding for each subject to retain only high specific binding regions in the analyses. The difference in regional mOR binding between the healthy control subjects and the cocaine-using subjects at each of the three time points (day 1, week 1, week 12) was evaluated with the SPM2 PET model “single-subject: condition and covariates,” in which separation conditions were coded for the healthy subjects and for day 1, week 1, and week 12 of monitored cocaine abstinence. Changes in mOR binding over time during cocaine abstinence were tested with a blocked analysis of variance design assuming independent residuals and variance sphericity (PET model of dodgy population main effect), with the three PET studies from each subject treated as three different conditions. Correlational analyses were done to examine whether mOR binding was associated with cocaine craving (visual analogue scale), depression (SCL-90 subscale), or anxiety (SCL-90 subscale) after 1 or 7 days of cocaine abstinence, and to evaluate the relationship between mOR on day 1 and prior cocaine use. Prior cocaine use was measured in terms of lifetime years of cocaine use (ASI), percent days with cocaine use during the 30 days before screening (ASI) or during the 2 weeks before admission, the average amount of cocaine used per day of use during the 2 weeks before admission, and urine benzoylecgonine concentration at the time of the first PET scan. Amounts of cocaine were measured in self-reported dollars-worth of cocaine. The resulting set of values for each statistical comparison constituted a statistical parametric map, with the minimum cluster extent threshold set at 50 contiguous voxels and cluster height set at an ␣ level of p ⬍ .005 to minimize false-positive findings (Friston et al 1995). The mean adjusted value of mOR binding was obtained from all voxels in significant clusters shown by the

correlational analysis in SPM, with the voxel-of-interest function in SPM. P values at the cluster level were adjusted for cluster extent with random field theory (Worsley et al 1996). To further validate the results of the SPM analysis, a separate region of interest analysis was performed with the mean adjusted value of mOR binding for each significant cluster. From each control subject image, which had already been spatially normalized and smoothed with the same methods, the mean value of mOR binding was obtained in the same spatial location as the significant clusters observed in the cocaine-using subjects. The mean values of normalized mOR binding in the cocaine-using and control groups were then compared (independent sample t test, two-tailed ␣ ⫽ .05). The correlation between each craving or mood score and mean mOR binding for each cluster was performed with the regression analysis function in Excel 2002, v. 10.0 (Microsoft, Redmond, Washington).

Results Mu-Opioid Receptor Binding at 1 Day of Cocaine Abstinence Compared with control subjects, mOR binding was increased during initial cocaine abstinence (first PET scan) in several brain regions, including frontal cortex (prefrontal, inferior frontal, and dorsolateral prefrontal regions), lateral temporal cortex, and anterior cingulate cortex (Table 1, Figure 1). This increased binding was positively correlated with self-reported cocaine craving in the left frontal, dorsolateral prefrontal, and anterior cingulate cortex and in the right temporal and parietal cortex (Table 2, Figure 2A and B). Mean (SD) cocaine craving ratings before the first PET scan were 25.6 (29.5) (range, 0 –94). There was no significant relationship between mOR binding and SCL-90 depression (.08 [.15]) or anxiety scores (.04 [.10]). Measures of recent cocaine use showed significant correlations with mOR binding after 1 day of abstinence. Urine benzoylecgonine concentration on the day of the first PET scan was positively correlated with mOR binding in the left dorsolateral prefrontal cortex [maximum z ⫽ 4.13 at x, y, z ⫽ ⫺42, 4, 32, cluster size k ⫽ 63; r2 ⫽.74, F (1,11) ⫽ 30.57, p ⫽ .0001] and negatively correlated with mOR binding in the left insular cortex [maximum z ⫽ 3.20 at x, y, z ⫽ ⫺48, ⫺8, 6, cluster size k ⫽ 53; r2 ⫽ .65, F (1,11) ⫽ 20.75, p ⫽ .008], and bilateral temporal lobes www.sobp.org/journal

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Figure 1. Increased regional brain normalized mu-opioid receptor (mOR, “mu-OR” in figure) binding in cocaine-dependent subjects over 12 weeks of enforced abstinence. Three-dimensional brain renderings of regions (shown in red) with significant increases in mOR binding in cocaine-dependent subjects versus 16 healthy, nondrug-using subjects, after 1 day (n ⫽ 17) (A), 7 days (n ⫽ 16) (B), or 90 days (n ⫽ 16) (C) of enforced cocaine abstinence on a closed research ward. The specific regions indicated are described in the text and in Table 1. (D–F) Means and standard deviations of mOR binding values for regions in the bilateral anterior cingulate cortex (yellow arrow in A), left dorsolateral prefrontal cortex (blue arrow in A), and right lateral temporal cortex (white arrow in A), respectively. Display threshold for mOR binding is p ⬍ .005 (uncorrected); cluster size k ⬎ 50. **p ⬍ .001, *p ⬍ .005 versus control subjects by t test (from region of interest analysis).

[right temporal lobe: maximum z ⫽ 3.72 at x, y, z ⫽ 36, 12, ⫺32, cluster size k ⫽ 278; r2 ⫽ .69, F (1,11) ⫽ 24.9, p ⫽ .0004; left temporal lobe: maximum z ⫽ 3.57 at x, y, z ⫽ ⫺34, 6, ⫺44, cluster size k ⫽ 160; r2 ⫽ .72, F (1,11) ⫽ 27.9, p ⫽ .0002]. Percentage of days with cocaine use during the 2 weeks before ward admission was correlated with higher mOR binding in the right anterior cingulate cortex and medial frontal cortex (Table 3, Figure 3). Amount of cocaine used per day of use during the 2 weeks before ward admission was correlated with higher binding in the right dorsolateral prefrontal cortex (Table 3). One outlier subject seemed to account for some of these correlations (Figure 3D). A repeat analysis omitting this subject confirmed the significant positive correlation in the right anterior cingulate cortex (r2 ⫽ .58, p ⫽ .0005), but the correlation in the right medial frontal cortex was no longer significant. There were no significant correlations between binding and more remote measures of cocaine use, such as percentage of days with cocaine use

in the 30 days before screening, or lifetime years of cocaine use (data not shown). One brain region, amygdala and adjacent temporal cortex, showed a tendency toward decreased mu-opioid receptor binding, but this change did not meet our minimum criteria for number of contiguous voxels (data not shown). mOR Binding Changes at 1 Week and 12 Weeks of Cocaine Abstinence All urine drug tests on the ward were negative for cocaine and other drugs. Mu-opioid receptor binding changes over the 3 months of monitored cocaine abstinence varied by brain region. Binding in the lateral temporal cortex and dorsolateral prefrontal cortex no longer differed from control values after 1 week of abstinence (second PET scan) (Table 1, Figure 1F); binding in anterior frontal, inferior frontal, prefrontal, and anterior cingulate cortex remained increased (Table 1, Figure 1D). Binding in the

Table 2. Correlation of Regional Brain-Normalized Mu-Opioid Receptor Binding with Self-Reported Cocaine Craving (on 100-mm Visual Analogue Scale) During Initial Enforced Abstinence in Cocaine-Dependent Subjects Day 1 of Abstinence (n ⫽ 17)

Brain Region

Side of Brain

AC Frontal

L L

PC Temporal Parietal

L R R

Day 7 of Abstinence (n ⫽ 16)

Correlation Using Mean Value from the Cluster

SPM result MNIa (x, y, z)

Max-z

kb

r2

F(1,14)

p

⫺8, ⫺12, 40 ⫺36, 42, 28 ⫺44, 10, 34 ⫺10, ⫺58, 22

3.95 3.3 4.03 4.34

66 44 120 144

.65 .53 .72 .71

25.98 15,56 35.8 34.53

.0001 .001 ⬍.0001 ⬍.0001

44, ⫺52, 32

3.85

68

.67

29

⬍.0001

Correlation Using Mean Value from the Cluster

SPM Result Max-z

k

r2

F(1,14)

p

⫺30, 48, 18

3.5

71

.54

16.55

.001

⫺4, ⫺64, 22 58, ⫺24, ⫺14 48, ⫺50, 28

4.05 3.34 3.38

289 64 74

.62 .51 .54

26.52 14.49 18.37

.0001 .002 .0007

MNI (x, y, z)

SPM, statistical parametric mapping; MNI, Montreal Neurological Institute; AC, anterior cingulate cortex; L, left; R, right; PC, prefrontal cortex. a Location of peak expressed as x, y, z coordinates in MNI space. b Cluster size k represents number of voxels (2 ⫻ 2 ⫻ 2 ⫻ 2 mm3) at a height threshold of p ⫽ .005, uncorrected.

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Figure 2. Association between regional brain normalized mu-opioid receptor (mOR, “mu-OR” in figure) binding and cocaine craving. (A) Left frontal cortex and left anterior and posterior cingulate cortex show significant positive correlation with cocaine craving after 1 day of cocaine abstinence (day 1 positron emission tomography [PET] scan). See Table 2. (B) Association on day 1 between cocaine craving (x-axis ⫽ score on 100-mm visual analogue scale [VAS]) and mOR binding (y-axis) in left dorsolateral prefrontal cortex (closed circles, solid line) and left anterior cingulate cortex (open circles, dotted line). Lines are least squares best fit to data points. (C) Left frontal cortex shows significant positive correlation with cocaine craving after 1 week of cocaine abstinence (day 7 PET scan). See Table 2. (D) Association on day 7 between cocaine craving (x-axis ⫽ score on 100-mm VAS) and mOR binding (y-axis) in left frontal cortex. Display threshold for mOR binding is p ⬍ .005 (uncorrected); cluster size k ⬎ 50.

anterior frontal and anterior cingulate cortex was still increased over control values after 12 weeks (third PET scan) (Table 1, Figure 1D). Within-subject comparisons showed significant decreases in binding in all regions except the anterior cingulate cortex (Table 4, Figure 4). Binding decreased significantly over the first week in the lateral temporal cortex and right anterior frontal cortex. Further decreases in binding occurred over the next 11 weeks in these regions, as well as in the lateral temporal, prefrontal, dorsolateral prefrontal, and inferior frontal regions. After 1 week of abstinence, binding in the left frontal and prefrontal cortex and right temporal and parietal cortex was still significantly associated with self-reported cocaine craving (mean [SD] craving rating ⫽ 20.2 [22.6]) (Table 2, Figure 2C, D) but not with SCL-90 depression (.03 [.08]) or anxiety (.006 [.02]) scores.

Craving scores were minimal after 12 weeks of abstinence, so that meaningful correlational analysis could not be done.

Discussion In this study, we found increased mOR binding in the frontal, lateral temporal, and anterior cingulate cortex of 17 subjects with current cocaine abuse/dependence, consistent with studies showing increased mOR binding in the prefrontal and cingulate cortex of rats treated with cocaine in a binge pattern (Clow et al 1991; Unterwald et al 1994, 2001). The increased binding in several regions was positively correlated with self-reported cocaine craving, suggesting a role for mORs in cocaine craving. There was no significant correlation between mOR binding and mood (depression, anxiety) self-ratings, which might have been

Table 3. Correlation of Regional Brain-Normalized Mu-Opioid Receptor Binding at 1 Day of Cocaine Abstinence with Measures of Cocaine Use in the Prior 2 Weeks in 17 Cocaine-Dependent Subjects Percent Days with Cocaine Use

Brain Region

Side of Brain

AC Frontal DLPC

R R R

Amount of Cocaine Used Per Day of Use

Correlation Using Mean Value from the Cluster

SPM Result MNIa

Max-z

kb

r2

F(1,15)

p

8, 26, 34 16, 50, 20

3.30 3.38

66 52

.52 .51

16.07 15.36

.001 .001

Correlation Using Mean Value from the Cluster

SPM Result MNIa

Max-z

k

r2

F(1,15)

p

50, 12, 24

4.06

119

.61

23.31

.0002

SPM, statistical parametric mapping; MNI, Montreal Neurological Institute; AC, anterior cingulate cortex; L, left; R, right; DLPC, dorsolateral prefrontal cortex. a Location of peak expressed as x, y, z coordinates in MNI space. b Cluster size k represents number of voxels (2 ⫻ 2 ⫻ 2 mm3) at a height threshold of p ⫽ .005, uncorrected.

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Figure 3. Association between regional brain normalized mu-opioid receptor (mOR, “mu-OR” in figure) binding at initial cocaine abstinence (day 1) and percentage of days with cocaine use during the 2 weeks before admission. See Table 4. (A) Right anterior cingulate cortex. (B) Right medial frontal cortex. (C) Association between mOR binding (y-axis) in right anterior cingulate cortex on day 1 and percentage of days with cocaine use (x-axis). (D) Association between mOR binding in right medial frontal cortex (y-axis) on day 1 and percentage of days with cocaine use (x-axis). Display threshold for mOR binding is p ⬍ .005 (uncorrected); cluster size k ⬎ 50.

due to their low values and restricted range. These craving findings confirm our earlier results of mOR binding associated with cocaine craving in the frontal, temporal, and anterior cingulate cortex, and amygdala (Zubieta et al 1996). Our results demonstrate the importance of a dopaminergic– opioid interaction in the human brain and its relevance to cocaine use. The present findings differ in several respects from the previous study (Zubieta et al 1996). We did not find increased mOR binding in the caudate and thalamus and did find a significant correlation between mOR binding and cocaine craving in the parietal cortex. The two samples were generally similar in sociodemographic and drug use characteristics. The differences might have been due to methodologic differences in image analysis and statistical methods. The present study used statistical parametric mapping (SPM2) with a voxel size of 2 ⫻ 2 ⫻ 2 mm and compared cocaine-using and control subjects within the overall SPM analysis. The previous study used region of interest

analysis applied to 8 ⫻ 8-mm square regions and compared the two subject groups by one-way analysis of variance. The present findings differ in some respects from those of rat studies. Almost all rat studies report increased mOR binding in the caudate and amygdala, as well as in the other brain regions mentioned above (Clow and Hammer 1991; Unterwald 2001; Unterwald et al 1992). The present study did not find increased mOR binding in these two brain regions. This might be due, in part, to species differences. A study in guinea pigs found that daily cocaine administration for 7 days produced no change in mOR binding in the caudate and a decrease in the amygdala (Itzhak 1993). A human postmortem study (Hurd and Herkenham 1993) found decreased mOR binding in the caudate/putamen of nine cocaine users compared with six noncocaine-using control subjects. This finding differs from that of our previous human study (Zubieta et al 1996), which found increased binding in the

Table 4. Brain Regions Showing Within-Subject Changes in Normalized Mu-Opioid Receptor Binding over 12 Weeks of Enforced Abstinence in 16 Cocaine-Dependent Subjects

Brain Region

Early Abstinence (Day 1 vs. Day 7)

Side of Brain

Cluster ka (p)b

R

135 (.049)

Anterior and Medial Prefrontal

Dorsolateral Prefrontal and Inferior Frontal Lateral Temporal

Max-z (x, y, z)c 3.56 (24, 24, ⫺16)

Later Abstinence (Day 7 vs. Day 90) Cluster ka (p)b

L R

69 (1.146) 69 (1.146) 84 (.232) 83 (.113) 551 (.000)

L

74 (.133)

R

133 (.051)

3.68 (54, 2, ⫺22)

L

118 (.064) 89 (.102) 371 (.003)

4.11 (⫺50, ⫺46, ⫺18) 3.41 (⫺52, ⫺48, 6) 4.17 (⫺52, ⫺14, ⫺18)

68 (.148) 77 (.126) 123 (.059) 50 (.211)

Max-z (x, y, z)c 4.18 (32, 60, ⫺10) 3.24 (8, 70, 0) 3.74 (20, 68, ⫺6) 2.98 (⫺12, 62, 20) 4.82 (44, 44, 14) 3.48 (⫺36, 32, ⫺16) 4.04 (56, ⫺18, 36) 3.77 (50, ⫺42, 40) 3.24 (52, ⫺52, 12) 3.29 (⫺50, ⫺54, 22)

R, right; L, left Cluster size k represents number of voxels (2 ⫻ 2 ⫻ 2 mm3) at a height threshold of p ⫽ .005, uncorrected. b p value for significance at cluster-level. c Location of peak expressed as x, y, z coordinates in Montreal Neurological Institute space. a

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Total Abstinent Period (Day 1 vs. Day 90) Cluster ka (p)b

Max-z (x, y, z)c

782 (.000)

4.95 (32, 60, ⫺8) 3.50 (8, 70, ⫺2) 3.50 (30, 54, ⫺14) 3.19 (⫺10, 64, ⫺10) 3.90 (44, 48, 8) 3.59 (52, 22, 18) 3.19 (⫺38, 36, 34) 3.05 (⫺44, 52, ⫺2) 4.22 (64, ⫺18, ⫺20) 3.90 (56, ⫺20, 36) 3.54 (52, ⫺52, 14) 4.12 (⫺50, ⫺44, ⫺16) 3.92 (⫺54, ⫺42, 10) 3.87 (⫺50, ⫺24, ⫺28)

121 (.061) 266 (.009) 158 (.035) 54 (.195) 81 (.177) 794 (.000) 99 (.086) 298 (.006) 76 (.128) 286 (.007) 172 (.029)

D.A. Gorelick et al

Figure 4. Within-subject changes in brain normalized mu-opioid receptor (mOR, “mu-OR” in figure) binding in 16 cocaine-dependent subjects over 12 weeks of enforced abstinence. See Table 4. (A) Brain regions (in red) showing significant decreases in mOR binding between 1 day (day 1) and 1 week (day 7) of cocaine abstinence. (B) Brain regions (in red) showing significant decreases in mOR binding between 1 week (day 7) and 12 weeks (day 90) of cocaine abstinence. (C) Brain regions (in red) showing significant decreases in mOR binding between 1 day (day 1) and 12 weeks (day 90) of cocaine abstinence. (D) Changes in mOR binding in right lateral temporal cortex (yellow arrow in C) Each line joins data from an individual subject. (E) Changes in mOR binding in right anterior frontal cortex (blue arrow in C). Each line joins data from an individual subject. Display threshold for mOR binding is p ⬍ .005 (uncorrected); cluster size k ⬎ 50.

caudate, and from the present study, which found no change in caudate binding. These differences might be related to differences in duration of cocaine use (not reported for subjects in the postmortem sudy) and/or differences in concomitant substance use. In the postmortem study, one third of the cocaine users had alcohol present at the time of death, as did half of the control subjects. Alcohol is known to influence brain mORs in animals (Oswald and Wand 2004; Rosin et al 2003; Turchan et al 1999) and humans (Bencherif et al 2004). Subjects in the prior (Zubieta et al 1996) and present studies were free of all other substances at the time of PET scanning. This study extends the duration of our earlier findings. The increased mOR binding persisted for 1 week in the inferior frontal and prefrontal cortex and for 12 weeks in the left anterior frontal cortex and bilateral anterior cingulate cortex. Because increased mOR binding is associated with cocaine craving, mORs might mediate the high rates of relapse to cocaine use during early abstinence. Assessment of mOR function could help predict individual differences in treatment outcome and optimize patient–treatment matching. Modulation of mOR function might

BIOL PSYCHIATRY 2005;57:1573–1582 1579 offer a useful treatment approach. A controlled clinical trial found that high-dose (16 mg daily) buprenorphine, a partial mOR agonist, significantly reduced cocaine use, as well as opiate use, in outpatients dependent on both drugs (Montoya et al 2004). This dose of buprenorphine would have occupied up to 95% of brain mOR (Zubieta et al 2000). Mu-opioid receptor occupancy by buprenorphine might ameliorate the effects of decreased endogenous opioid levels due to chronic cocaine use. Two brain regions showing increased mOR binding in this study (anterior cingulate and frontal cortex) have shown increased metabolic or functional activity or increased regional cerebral blood flow (rCBF) in other brain imaging studies of cocaine users (Daglish and Nutt 2003; Goldstein and Volkow 2002; Kilts et al 2004). Both the increased rCBF and increased metabolic activity were associated with cocaine craving. These findings suggest an important role for mOR in these brain regions, which are thought to mediate the salience of a reinforcer (such as cocaine) and the ability to inhibit prepotent responses (such as drug-seeking and -taking behavior) (Goldstein and Volkow 2002). We are aware of only one other research group that has studied brain imaging in cocaine users after long-term monitored abstinence. Volkow et al used PET scanning to study a sample of seven cocaine-abusing men who completed a 3- to 4-month inpatient rehabilitation program. Compared with healthy, nondrug-using control subjects, the cocaine abusers had decreased glucose metabolism in the left dorsolateral and dorsomedial prefrontal cortex (Volkow et al 1992) and decreased N-methylspiperidol binding (suggestive of decreased dopamine D2 receptor availability) in the orbitofrontal cortex and cingulate cortex (Volkow et al 1993). In the present study, we did not observe persisting increases in mOR binding in these regions, except for cingulate cortex. Taken together, however, these findings do confirm that chronic cocaine use can lead to long-lasting neuropharmacologic changes in the brain. These brain changes might underlie the persisting risk of relapse experienced by abstinent cocaine abusers. In particular, the increased mOR binding shown in this study and the decreased dopamine D2 receptor availability shown in another study (Volkow et al 1993) in the cingulate cortex might be important in the pathogenesis of cocaine addiction, given the role of this brain region in cocaine craving (see above) and in animal models of relapse (Kalivas and McFarland 2003). Our data provide evidence as to the degree of cocaine exposure and temporal proximity to PET scanning needed to produce measurable changes in mOR binding. Mu-opioid receptor binding in some brain regions showed significant positive correlations with measures of cocaine use (percent days with cocaine use, amount of cocaine used per day of use) over the 2 weeks before PET scanning (Table 3, Figure 3), but not with more remote or longer-term measures of cocaine use (days of use in the month before screening, years of lifetime use). This pattern suggests a close temporal relationship between changing usage of cocaine and regulation of mOR binding. In rodent studies, 1–3 days of thrice-daily cocaine administration is sufficient to significantly increase brain levels of mOR mRNA (Yuferov et al 1999, 2003), although not to increase mORs themselves (Branch et al 1992; Hammer 1989; Rosin et al 2003; Unterwald et al 1994, 2001). The shortest-term measure of prior cocaine use, urine benzoylecgonine concentration, showed both positive and negative correlations with regional brain mOR binding. Positive correlations might be due to increased numbers of mOR, www.sobp.org/journal

1580 BIOL PSYCHIATRY 2005;57:1573–1582 as suggested above. Negative correlations might reflect occupancy of mOR by endogenous ligands released by recent cocaine use. Cocaine administration in rats increases brain ␤-endorphin (mOR ligand) levels for at least several hours (Olive et al 2001; Roth-Deri et al 2003). In our subjects, greater recent cocaine use in the 2 to 3 days before the first PET scan, reflected in higher urine benzoylecgonine concentrations, might have resulted in greater mOR occupancy by endogenous ligands, reflected in less binding of the PET radiotracer. This could account for the observed negative correlation in some brain regions. The mechanism of the brain mOR binding changes cannot be determined from our data. The PET scan analysis does not distinguish between changes in receptor number/density (Bmax) and in receptor affinity (Kd). In rat studies, cocaine increases mOR numbers rather than mOR affinity (Unterwald et al 1992). The technique used also cannot distinguish between changes in the mOR and changes in its occupancy by endogenous ligands. Most rat studies find that treatment with cocaine increases mOR mRNA levels in relevant brain regions (Azaryan et al 1996a, 1998; Yuferov et al 1999), suggesting that chronic cocaine use would increase mOR number. Some (but not all) animal studies suggest that shorter-term cocaine exposure increases brain mRNA levels of precursors of enkephalin (an endogenous mOR ligand), whereas longer exposure results in reduced levels (Daunais et al 1997; Laforge et al 2003; Przewlocka and Lason 1995). Monkeys self-administering cocaine for 2 years have significantly decreased preproenkephalin mRNA levels in caudate (Daunais et al 1997). Our subjects had been using cocaine regularly for 2–16 years (mean, 7.2 years), so it is plausible that they had reduced brain levels of enkephalin compared with control subjects. Such reduced levels could result in compensatory upregulation of mORs and would have left more receptors available for occupancy by the radiotracer [11C]carfentanil during PET scanning. Both processes would produce a finding of increased receptor binding. It is possible that the mOR binding changes observed in this study were confounded by the effect of changes in physical exercise, social stress, or other lifestyle factors associated with being on the research ward, rather than in the community. We are not aware of any human data showing that such lifestyle variables can affect brain mOR binding. In rodent studies, social stress increases mOR mRNA levels in the ventral tegmental area (Nikulina et al 1999), whereas physical exercise increases some regional brain levels of enkephalin and enkephalin mRNA (Persson et al 2004; Werme et al 2000). Assuming that these effects occur in humans, the decreased physical exercise likely associated with being on a closed ward might decrease brain enkephalin levels, resulting in increased receptor availability for carfentanil and increased mOR binding on the PET scans, the opposite of what was actually observed. The effect of social stress is less straightforward, depending on whether subjects on the research ward are considered to experience more (from living with strangers in close quarters in a new environment) or less (from being in a safe environment) social stress than when in the community. Both factors are probably at play, making it unlikely that there was a robust effect of social stress in this study. Our findings are only partially consistent with a recent report of significant correlations between alcohol craving and regional brain mOR binding during early alcohol abstinence (Bencherif et al 2004). That study found that lower (than in healthy control www.sobp.org/journal

D.A. Gorelick et al subjects) mOR binding in the right dorsolateral prefrontal cortex, right anterior frontal cortex, and right parietal cortex was associated with higher alcohol craving and higher depression scores. The failure of the present study to find any significant association between depression scores and mOR binding or craving is probably due to the very low levels and restricted range of depression scores in the cocaine-using subjects. In contrast, the alcohol-dependent subjects in the prior study had Beck Depression Inventory scores within the range associated with clinically significant depression. The difference in the direction of the association between mOR binding and craving between the two studies might be due to differences in the neuropharmacologic actions of cocaine and alcohol. Cocaine influences mORs and their endogenous opioid ligands through its (indirect) actions on dopamine receptors (Azaryan et al 1996a, 1996b; Chen et al 1993; George and Kertesz 1987), whereas alcohol influences them through actions on ␥-aminobutyric acid and N-methyl-D-aspartate receptors (Oswald and Wand 2004). Some rodent studies directly comparing the effects of cocaine and alcohol administration on brain mOR binding have found different effects (e.g., a single dose of alcohol increases mOR binding in the nucleus accumbens, basolateral amygdala, and other brain areas, whereas a single dose of cocaine produces no changes [Rosin et al 2003]). Chronic alcohol administration (in drinking water for 1 month) reduces mOR binding in the nucleus accumbens, a decrease that persists for at least 96 hours into abstinence, whereas daily cocaine administration for 5 days produces no changes except for a decrease at 24 hours into abstinence (Turchan et al 1999). In summary, our findings show that cocaine use is associated with elevations in regional brain mOR binding and that these elevations are related to craving for cocaine. Mu-opioid receptors decreased to normal levels over 12 weeks of enforced abstinence in frontal and temporal cortex but remained elevated in cingulate cortex. This mixed pattern of change during abstinence suggests that there might be both state and trait relationships with regional brain elevations in mOR binding. Future research should examine the relationship between persistently elevated mOR binding in the cingulate cortex and long-term clinical phenomena, such as craving and relapse. These findings provide a better understanding of the neurobiological basis of cocaine addiction and provide leads to improved treatment and prediction of treatment outcome.

This research was supported by National Institute on Drug Abuse intramural funds and National Institutes of Health grant RO1 DA-09479 (JJF). A preliminary version of this work was presented at the Collegium Internationale Neuropsychopharmacologicum XXIII congress, Montreal, Quebec, Canada, June 23-27, 2002. We thank Dr. Ismail Alhamrawy for help with data management and analysis. BB is currently at the Department of Radiology University of Pittsburgh School of Medicine. SJB is currently at the Department of Psychiatry, University of Maryland School of Medicine. Ambrose LM, Unterwald EM, Van Bockstaele EJ (2004): Ultrastructural evidence for co-localization of dopamine D2 and mu-opioid receptors in the rat dorsolateral striatum. Anat Rec 279:583–591. American Psychiatric Association (1994): Diagnostic and Statistical Manual of Mental Disorders, 4th ed. Washington, DC: American Psychiatric Association.

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