Cheryl Morse

Reversible and regionally selective downregulation of brain cannabinoid CB1 receptors in chronic daily cannabis smokers

Chronic cannabis (marijuana, hashish) smoking can result in dependence. Rodent studies show reversible downregulation of brain cannabinoid CB1 (cannabinoid receptor type 1) receptors after chronic exposure to cannabis. However, whether downregulation occurs in humans who chronically smoke cannabis is unknown. Here we show, using positron emission tomography imaging, reversible and regionally selective downregulation of brain cannabinoid CB1 receptors in human subjects who chronically smoke cannabis. Downregulation correlated with years of cannabis smoking and was selective to cortical brain regions. After ∼4 weeks of continuously monitored abstinence from cannabis on a secure research unit, CB1 receptor density returned to normal levels. This is the first direct demonstration of cortical cannabinoid CB1 receptor downregulation as a neuroadaptation that may promote cannabis dependence in human brain.

In vivo radioligand binding to translocator protein correlates with severity of Alzheimer’s disease

Neuroinflammation is a pathological hallmark of Alzheimers disease, but its role in cognitive impairment and its course of development during the disease are largely unknown. To address these unknowns, we used positron emission tomography with (11)C-PBR28 to measure translocator protein 18 kDa (TSPO), a putative biomarker for inflammation. Patients with Alzheimers disease, patients with mild cognitive impairment and older control subjects were also scanned with (11)C-Pittsburgh Compound B to measure amyloid burden. Twenty-nine amyloid-positive patients (19 Alzheimers, 10 mild cognitive impairment) and 13 amyloid-negative control subjects were studied. The primary goal of this study was to determine whether TSPO binding is elevated in patients with Alzheimers disease, and the secondary goal was to determine whether TSPO binding correlates with neuropsychological measures, grey matter volume, (11)C-Pittsburgh Compound B binding, or age of onset. Patients with Alzheimers disease, but not those with mild cognitive impairment, had greater (11)C-PBR28 binding in cortical brain regions than controls. The largest differences were seen in the parietal and temporal cortices, with no difference in subcortical regions or cerebellum. (11)C-PBR28 binding inversely correlated with performance on Folstein Mini-Mental State Examination, Clinical Dementia Rating Scale Sum of Boxes, Logical Memory Immediate (Wechsler Memory Scale Third Edition), Trail Making part B and Block Design (Wechsler Adult Intelligence Scale Third Edition) tasks, with the largest correlations observed in the inferior parietal lobule. (11)C-PBR28 binding also inversely correlated with grey matter volume. Early-onset (<65 years) patients had greater (11)C-PBR28 binding than late-onset patients, and in parietal cortex and striatum (11)C-PBR28 binding correlated with lower age of onset. Partial volume corrected and uncorrected results were generally in agreement; however, the correlation between (11)C-PBR28 and (11)C-Pittsburgh Compound B binding was seen only after partial volume correction. The results suggest that neuroinflammation, indicated by increased (11)C-PBR28 binding to TSPO, occurs after conversion of mild cognitive impairment to Alzheimers disease and worsens with disease progression. Greater inflammation may contribute to the precipitous disease course typically seen in early-onset patients. (11)C-PBR28 may be useful in longitudinal studies to mark the conversion from mild cognitive impairment or to assess response to experimental treatments of Alzheimers disease.

A Genetic Polymorphism for Translocator Protein 18 Kda Affects both in Vitro and in Vivo Radioligand Binding in Human Brain to this Putative Biomarker of Neuroinflammation

Second-generation radioligands for translocator protein (TSPO), an inflammation marker, are confounded by the codominant rs6971 polymorphism that affects binding affinity. The resulting three groups are homozygous for high-affinity state (HH), homozygous for low-affinity state (LL), or heterozygous (HL). We tested if in vitro binding to leukocytes distinguished TSPO genotypes and if genotype could affect clinical studies using the TSPO radioligand [11C]PBR28. In vitro binding to leukocytes and [11C]PBR28 brain imaging were performed in 27 human subjects with known TSPO genotype. Specific [3H]PBR28 binding was measured in prefrontal cortex of 45 schizophrenia patients and 47 controls. Leukocyte binding to PBR28 predicted genotype in all subjects. Brain uptake was ~40% higher in HH than HL subjects. Specific [3H]PBR28 binding in LL controls was negligible, while HH controls had ~80% higher binding than HL controls. After excluding LL subjects, specific binding was 16% greater in schizophrenia patients than controls. This difference was insignificant by itself (P = 0.085), but was significant after correcting for TSPO genotype (P = 0.011). Our results show that TSPO genotype influences PBR28 binding in vitro and in vivo. Correcting for this genotype increased statistical power in our postmortem study and is recommended for in vivo positron emission tomography studies.

P-Glycoprotein Function at the Blood–Brain Barrier in Humans Can Be Quantified with the Substrate Radiotracer 11C-N-Desmethyl-Loperamide

Permeability-glycoprotein (P-gp), an efflux transporter in several organs, acts at the blood–brain barrier to protect the brain from exogenous toxins. P-gp almost completely blocks brain entry of the PET radiotracer 11C-N-desmethyl-loperamide (11C-dLop). We examined the ability of 11C-dLop to quantify P-gp function in humans after increasing doses of tariquidar, an inhibitor of P-gp. Methods: Seventeen healthy volunteers had a total of 23 PET scans with 11C-dLop at baseline and after increasing doses of tariquidar (2, 4, and 6 mg/kg intravenously). A subset of subjects received PET with 15O-H2O to measure cerebral blood flow. Brain uptake of 11C-dLop was quantified in 2 ways. Without blood data, uptake was measured as area under the time–activity curve in the brain from 10 to 30 min (AUC10–30). With arterial blood data, brain uptake was quantified with compartmental modeling to estimate the rates of entry into (K1) and efflux from (k2) the brain. Results: Brain uptake of radioactivity was negligible at baseline and increased only slightly (∼30%) after 2 mg of tariquidar per kilogram. In contrast, 4 and 6 mg of tariquidar per kilogram increased brain uptake 2- and 4-fold, respectively. Greater brain uptake reflected greater brain entry (K1), because efflux (k2) and cerebral blood flow did not differ between tariquidar-treated and untreated subjects. In the subjects who received the highest dose of tariquidar (and had the highest brain uptake), regional values of K1 correlated linearly with absolute cerebral blood flow, consistent with high single-pass extraction of 11C-dLop. AUC10–30 correlated linearly with K1. Conclusion: P-gp function at the blood–brain barrier in humans can be quantified using PET and 11C-dLop. A simple measure of brain uptake (AUC10–30) may be used as a surrogate of the fully quantified rate constant for brain entry (K1) and thereby avoid arterial sampling. However, to dissect the function of P-gp itself, both brain uptake and the influx rate constant must be corrected for radiotracer delivery (blood flow).

Cerebellum Can Serve As a Pseudo-Reference Region in Alzheimer Disease to Detect Neuroinflammation Measured with PET Radioligand Binding to Translocator Protein

Alzheimer disease (AD) is associated with an increase in the brain of the 18-kDa translocator protein (TSPO), which is overexpressed in activated microglia and reactive astrocytes. Measuring the density of TSPO with PET typically requires absolute quantitation with arterial blood sampling, because a reference region devoid of TSPO does not exist in the brain. We sought to determine whether a simple ratio method could substitute for absolute quantitation of binding with 11C-PBR28, a second-generation radioligand for TSPO. Methods: 11C-PBR28 PET imaging was performed in 21 healthy controls, 11 individuals with mild cognitive impairment, and 25 AD patients. Group differences in 11C-PBR28 binding were compared using 2 methods. The first was the gold standard method of calculating total distribution volume (VT), using the 2-tissue-compartment model with the arterial input function, corrected for plasma-free fraction of radiotracer (fP). The second method used a ratio of brain uptake in target regions to that in cerebellum—that is, standardized uptake value ratio (SUVR). Results: Using absolute quantitation, we confirmed that TSPO binding (VT/fP) was greater in AD patients than in healthy controls in expected temporoparietal regions and was not significantly different among the 3 groups in the cerebellum. When the cerebellum was used as a pseudo-reference region, the SUVR method detected greater binding in AD patients than controls in the same regions as absolute quantification and in 1 additional region, suggesting SUVR may have greater sensitivity. Coefficients of variation of SUVR measurements were about two-thirds lower than those of absolute quantification, and the resulting statistical significance was much higher for SUVR when comparing AD and healthy controls (e.g., P < 0.0005 for SUVR vs. P = 0.023 for VT/fP in combined middle and inferior temporal cortex). Conclusion: To measure TSPO density in AD patients and control subjects, a simple ratio method SUVR can substitute for, and may even be more sensitive than, absolute quantitation. The SUVR method is expected to improve subject tolerability by allowing shorter scanning time and not requiring arterial catheterization. In addition, this ratio method allows smaller sample sizes for comparable statistical significance because of the relatively low variability of the ratio values.

Reduced cannabinoid CB 1 receptor binding in alcohol dependence measured with positron emission tomography

Brain cannabinoid CB1 receptors contribute to alcohol-related behaviors in experimental animals, but their potential role in humans with alcohol dependence is poorly understood. We measured CB1 receptors in alcohol dependent patients in early and protracted abstinence, and in comparison with control subjects without alcohol use disorders, using positron emission tomography and [18F]FMPEP-d2, a radioligand for CB1 receptors. We scanned 18 male in-patients with alcohol dependence twice, within 3–7 days of admission from ongoing drinking, and after 2–4 weeks of supervised abstinence. Imaging data were compared with those from 19 age-matched healthy male control subjects. Data were also analyzed for potential influence of a common functional variation (rs2023239) in the CB1 receptor gene (CNR1) that may moderate CB1 receptor density. On the first scan, CB1 receptor binding was 20–30% lower in patients with alcohol dependence than in control subjects in all brain regions and was negatively correlated with years of alcohol abuse. After 2–4 weeks of abstinence, CB1 receptor binding remained similarly reduced in these patients. Irrespective of the diagnostic status, C allele carriers at rs2023239 had higher CB1 receptor binding compared with non-carriers. Alcohol dependence is associated with a widespread reduction of cannabinoid CB1 receptor binding in the human brain and this reduction persists at least 2–4 weeks into abstinence. The correlation of reduced binding with years of alcohol abuse suggests an involvement of CB1 receptors in alcohol dependence in humans.

Increased In Vivo Expression of an Inflammatory Marker in Temporal Lobe Epilepsy

Animal studies and clinical observations suggest that epilepsy is associated with inflammation. Translocator protein (TSPO) (18 kDa), a marker of inflammation, is increased in vitro in surgical samples from patients with temporal lobe epilepsy. TSPO can be measured in the living human brain with PET and the novel radioligand 11C-PBR28. In this study, we sought to determine whether in vivo expression of TSPO is increased ipsilateral to the seizure focus in patients with temporal lobe epilepsy. Methods: Sixteen patients with unilateral temporal lobe epilepsy and 30 healthy subjects were studied with 11C-PBR28 PET and MRI. Uptake of radioactivity after injection of 11C-PBR28 was measured from regions of interest drawn bilaterally onto MR images. Brain uptake from ipsilateral and contralateral hemispheres was compared using a paired-samples t test. Results: We found that brain uptake was higher ipsilateral to the seizure focus in the hippocampus, parahippocampal gyrus, amygdala, fusiform gyrus, and choroid plexus but not in other brain regions. This asymmetry was more pronounced in patients with hippocampal sclerosis than in those without. Conclusion: We found increased uptake of radioactivity after injection of 11C-PBR28 ipsilateral to the seizure focus in patients with temporal lobe epilepsy, suggesting increased expression of TSPO. Studies in larger samples are required to confirm this finding and determine the clinical utility of imaging TSPO in temporal lobe epilepsy.

Imaging and Quantitation of Cannabinoid CB1 Receptors in Human and Monkey Brains Using 18F-Labeled Inverse Agonist Radioligands

We recently demonstrated that 11C-MePPEP, a PET ligand for CB1 receptors, has such high uptake in the human brain that it can be imaged for 210 min and that receptor density can be quantified as distribution volume (VT) using the gold standard of compartmental modeling. However, 11C-MePPEP had relatively poor retest and intersubject variabilities, which were likely caused by errors in the measurements of radioligand in plasma at low concentrations by 120 min. We sought to find an analog of 11C-MePPEP that would provide more accurate plasma measurements. We evaluated several promising analogs in the monkey brain and chose the 18F-di-deutero fluoromethoxy analog (18F-FMPEP-d2) to evaluate further in the human brain. Methods: 11C-FMePPEP, 18F-FEPEP, 18F-FMPEP, and 18F-FMPEP-d2 were studied in 5 monkeys with 10 PET scans. We calculated VT using compartmental modeling with serial measurements of unchanged parent radioligand in arterial plasma and radioactivity in the brain. Nonspecific binding was determined by administering a receptor-saturating dose of rimonabant, an inverse agonist at the CB1 receptor. Nine healthy human subjects participated in 17 PET scans using 18F-FMPEP-d2, with 8 subjects having 2 PET scans to assess retest variability. To identify sources of error, we compared intersubject and retest variability of brain uptake, arterial plasma measurements, and VT. Results: 18F-FMPEP-d2 had high uptake in the monkey brain, with greater than 80% specific binding, and yielded less radioactivity uptake in bone than did 18F-FMPEP. High brain uptake with 18F-FMPEP-d2 was also observed in humans, in whom VT was well identified within approximately 60 min. Retest variability of plasma measurements was good (16%); consequently, VT had a good retest variability (14%), intersubject variability (26%), and intraclass correlation coefficient (0.89). VT increased after 120 min, suggesting an accumulation of radiometabolites in the brain. Radioactivity accumulated in the skull throughout the entire scan but was thought to be an insignificant source of data contamination. Conclusion: Studies in monkeys facilitated our development and selection of 18F-FMPEP-d2, compared with 18F-FMPEP, as a radioligand demonstrating high brain uptake, high percentage of specific binding, and reduced uptake in bone. Retest analysis in human subjects showed that 18F-FMPEP-d2 has greater precision and accuracy than 11C-MePPEP, allowing smaller sample sizes to detect a significant difference between groups.

Neuroinflammation in Temporal Lobe Epilepsy Measured Using Positron Emission Tomographic Imaging of Translocator Protein

IMPORTANCE Neuroinflammation may play a role in epilepsy. Translocator protein 18 kDa (TSPO), a biomarker of neuroinflammation, is overexpressed on activated microglia and reactive astrocytes. A preliminary positron emission tomographic (PET) imaging study using carbon 11 ([11C])-labeled PBR28 in patients with temporal lobe epilepsy (TLE) found increased TSPO ipsilateral to seizure foci. Full quantitation of TSPO in vivo is needed to detect widespread inflammation in the epileptic brain. OBJECTIVES To determine whether patients with TLE have widespread TSPO overexpression using [11C]PBR28 PET imaging, and to replicate relative ipsilateral TSPO increases in patients with TLE using [11C]PBR28 and another TSPO radioligand, [11C]DPA-713. DESIGN, SETTING, AND PARTICIPANTS In a cohort study from March 2009 through September 2013 at the Clinical Epilepsy Section of the National Institute of Neurological Disorders and Stroke, participants underwent brain PET and a subset had concurrent arterial sampling. Twenty-three patients with TLE and 11 age-matched controls were scanned with [11C]PBR28, and 8 patients and 7 controls were scanned with [11C]DPA-713. Patients with TLE had unilateral temporal seizure foci based on ictal electroencephalography and structural magnetic resonance imaging. Participants with homozygous low-affinity TSPO binding were excluded. MAIN OUTCOMES AND MEASURES The [11C]PBR28 distribution volume (VT) corrected for free fraction (fP) was measured in patients with TLE and controls using FreeSurfer software and T1-weighted magnetic resonance imaging for anatomical localization of bilateral temporal and extratemporal regions. Side-to-side asymmetry in patients with TLE was calculated as the ratio of ipsilateral to contralateral [11C]PBR28 and [11C]DPA-713 standardized uptake values from temporal regions. RESULTS The [11C]PBR28 VT to fp ratio was higher in patients with TLE than in controls for all ipsilateral temporal regions (27%-42%; P < .05) and in contralateral hippocampus, amygdala, and temporal pole (approximately 30%-32%; P < .05). Individually, 12 patients, 10 with mesial temporal sclerosis, had asymmetrically increased hippocampal [11C]PBR28 uptake exceeding the 95% confidence interval of the controls. Binding of [11C]PBR28 was increased significantly in thalamus. Relative [11C]PBR28 and [11C]DPA-713 uptakes were higher ipsilateral than contralateral to seizure foci in patients with TLE ([11C]PBR28: 2%-6%; [11C]DPA-713: 4%-9%). Asymmetry of [11C]DPA-713 was greater than that of [11C]PBR28 (F = 29.4; P = .001). CONCLUSIONS AND RELEVANCE Binding of TSPO is increased both ipsilateral and contralateral to seizure foci in patients with TLE, suggesting ongoing inflammation. Anti-inflammatory therapy may play a role in treating drug-resistant epilepsy.

Brain and Whole-Body Imaging of Nociceptin/Orphanin FQ Peptide Receptor in Humans Using the PET Ligand 11C-NOP-1A

Nociceptin/orphanin FQ peptide (NOP) receptor is a new class of opioid receptor that may play a pathophysiologic role in anxiety and drug abuse and is a potential therapeutic target in these disorders. We previously developed a high-affinity PET ligand, 11C-NOP-1A, which yielded promising results in monkey brain. Here, we assessed the ability of 11C-NOP-1A to quantify NOP receptors in human brain and estimated its radiation safety profile. Methods: After intravenous injection of 11C-NOP-1A, 7 healthy subjects underwent brain PET for 2 h and serial sampling of radial arterial blood to measure parent radioligand concentrations. Distribution volume (VT; a measure of receptor density) was determined by compartmental (1- and 2-tissue) and noncompartmental (Logan analysis and Ichises bilinear analysis [MA1]) methods. A separate group of 9 healthy subjects underwent whole-body PET to estimate whole-body radiation exposure (effective dose). Results: After 11C-NOP-1A injection, the peak concentration of radioactivity in brain was high (∼5–7 standardized uptake values), occurred early (∼10 min), and then washed out quickly. The unconstrained 2-tissue-compartment model gave excellent VT identifiability (∼1.1% SE) and fitted the data better than a 1-tissue-compartment model. Regional VT values (mL·cm−3) ranged from 10.1 in temporal cortex to 5.6 in cerebellum. VT was well identified in the initial 70 min of imaging and remained stable for the remaining 50 min, suggesting that brain radioactivity was most likely parent radioligand, as supported by the fact that all plasma radiometabolites of 11C-NOP-1A were less lipophilic than the parent radioligand. Voxel-based MA1 VT values correlated well with results from the 2-tissue-compartment model, showing that parametric methods can be used to compare populations. Whole-body scans showed radioactivity in brain and in peripheral organs expressing NOP receptors, such as heart, pancreas, and spleen. 11C-NOP-1A was significantly metabolized and excreted via the hepatobiliary route. Gallbladder had the highest radiation exposure (21 μSv/MBq), and the effective dose was 4.3 μSv/MBq. Conclusion: 11C-NOP-1A is a promising radioligand that reliably quantifies NOP receptors in human brain. The effective dose in humans is low and similar to that of other 11C-labeled radioligands, allowing multiple scans in 1 subject.