Simplified Reference Tissue Model Research Articles

The D2-agonist radiotracer [11C]-(+)-PHNO shows a marked increase in sensitivity to amphetamine challenges when compared to [11C]raclopride in the cat brain

Imaging the high affinity state of the D2-receptor using agonist radioligands has aroused considerable interest recently as it could allow for measurements of the functional state of the receptor rather than the total receptor population as measured by antagonist radioligands. Here we report on the evaluation of [11C]-(+)-PHNO as a potential radioligand for in vivo study of the high affinity state D2-receptor using positron emission tomography (PET) in the cat brain. Five cats were anesthetized with isoflurane and scanned using the high resolution PET camera system CPS-HRRT. At baseline conditions, high levels of radioactivity were observed in the striatum whereas low levels of radioactivity were observed in cerebellum. Radioactivity in cerebellum peaked within two minutes post-injection and decreased rapidly thereafter. Radioactivity in striatum peaked at 8–10 minutes post-injection, demonstrating that [11C]-(+)-PHNO binding in this structure is reversible and reaches equilibrium within the time frame of a PET experiment. The striatal binding potential (BP) approximated using the ratio of specific/non-specific (specific = striatum – cerebellum, non-specific = cerebellum) between 31 and 60 minutes post-injection was 3.75 0.51 (n=5). Pretreatment with raclopride (1 mg/kg; i.v.) or haloperidol (0.5 mg/kg; i.v.) reduced [11C]-(+)-PHNO striatal BP by 85% and 94%, respectively, indicating that the majority of radioligand binding in striatum represents specific binding to D2-receptors. Pretreatment with SCH23390 had no effect on [11C]-(+)-PHNO binding. A direct comparison of [11C]-(+)-PHNO and [11C]NPA binding in the same cat showed that [11C]-(+)-PHNO displayed a higher striatal BP than [11C]NPA (3.41 versus 1.59), suggesting that [11C]-(+)-PHNO may be more sensitive than [11C]NPA for in vivo imaging of the high-affinity D2-receptors. Comparison of the dose-effect of amphetamine (0.1, 0.5 and 2 mg/kg i.v.) on both [11C]-(+)-PHNO and [11C]raclopride binding in striatum showed than [11C]-(+)-PHNO was more sensitive to the dopamine releasing effect of the drug. The highest dose of amphetamine induced a 83-88% inhibition of [11C]-(+)-PHNO binding (n=3) and a 60-63% inhibition of [11C]raclopride binding (n=2). Preliminary data showed that the ED50 of amphetamine for inhibiting [11C]-(+)-PHNO and [11C]raclopride striatal bindings were 0.25 mg/kg and 0.63 mg/kg, respectively. Interestingly, the hyperbolic function used to fit the dose-effect data and derive ED50 values best fitted the experimental data when the maximal occupancy value was set at 100% for [11C]-(+)-PHNO and when it was set at only 80% for [11C]raclopride. These results suggest that, in contrast to what is observed with the antagonist radioligand, amphetamine-released dopamine had access to the entire population of D2-receptors recognized by [11C]-(+)-PHNO. Preliminary human PET scans with [11C]-(+)-PHNO are underway. Initial results obtained in 2 healthy volunteers show high striatal uptake with a BP of 2.66 and 2.86, respectively, as approximated using the simplified reference tissue model. These data indicate that [11C]-(+)-PHNO binding in striatum is specific for D2-receptors. The high penetration of [11C]-(+)-PHNO in brain, its high signal to noise ratio, its favourable in vivo kinetics as well as its high sensitivity to amphetamine shows that [11C]-(+)-PHNO has highly suitable characteristics for probing the high-affinity state of D2-receptors with PET.

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Introduction: In mice, disruption of 5‐HTT function via knockout of the 5‐HTT gene reduces 5‐HT1A receptor density. In humans, the short allele (S) of the serotonin transporter (5‐HTTLPR) gene has been associated with decreased transcriptional activity and reduced 5‐HTT function. Furthermore, a promoter polymorphism in the 5‐HT1A receptor gene (–1018 C>G) inhibits the repression of transcription, which may lead to increased 5‐HT1A autoreceptor expression. We therefore investigated the effects of polymorphisms of the 5‐HTTLPR and 5‐HT1A receptor (–1018 C>G) genes on 5‐HT1A receptor density in humans using PET and [11C] WAY 100635 (a specific ligand for 5‐HT1A receptors). Methods: Thirty‐five healthy volunteers [27 males, mean (SD) age: 46 (13) years], who had previously undergone PET scans with [11C] WAY 100635, underwent blood collection for genotyping (i) the 5‐HT1A receptor gene single nucleotide polymorphism at the site ‐1018 C>G, (ii) the common 44 bp insertion/deletion polymorphism (∼ 1 kb from initiation site) in the upstream regulatory region of 5‐HTTLPR. PET scans were undertaken using the ECAT 953 and 966 scanners. Binding potential (BP) values were obtained using a simplified reference tissue model with a standard PET template. We carried out multivariate repeated measures ANOVA with 21 regions of interest as within‐subject factors and genetic polymorphisms, scanner, and gender as between‐subject factors. Results: Genotype frequencies of all the polymorphisms conformed to the Hardy‐Weinberg equilibrium. The “S” allele of 5‐HTTLPR gene showed significantly lower 11C WAY 100635 BP values when compared to the “L” allele (F = 4.8, df 1.27, P= .037) and this was independent of the scanner and gender effects. The 5‐HT1A receptor gene (–1018 c>g) did not show any significant genetic (allelic or genotypic) effects on 11C WAY 100635 BP values (F = .002, df 1.28, P= .9). Conclusions: This study demonstrates for the first time that a genetic polymorphism of the 5‐HT transporter gene indirectly affects 5‐HT1A receptor density in vivo. This may be mediated by a persistently increased 5‐HT tone associated with the “S” allele of 5‐HTTLPR causing a downregulation of 5‐HT1A receptors. Comment: This PET imaging study, so far available only in abstract form, was done in a group of volunteers and elegantly demonstrates how genetic polymorphism of the 5‐HT transporter gene can modify central 5‐HT1A activity. This is potentially important in migraine and depression, where serotonin levels may be reduced. Could this be another gene locus that may have relevance to migraineurs?—David S. Millson

Reduced midbrain dopamine transporter binding in male adolescents with attention-deficit/hyperactivity disorder: Association between striatal dopamine markers and motor hyperactivity

The hypothesis that altered dopamine transmission underlies hyperactive-inattentive behavior in children with attention-deficit/hyperactivity disorder (ADHD) is based on genetic studies and the efficacy of psychostimulants. Most of previous positron emission tomography (PET) and single photon emission tomography (SPET) studies have shown altered binding of dopamine markers in the basal ganglia. Yet, the functional role of the neurochemical disturbances are poorly understood. The purpose of our study was to examine dopamine transporter (DAT) and dopamine D2 receptor (D2R) binding in adolescents with ADHD and to search for its relationship with cognitive functions as well as locomotor hyperactivity. Twelve adolescents with ADHD and 10 young adults were examined with PET using the selective radioligands [11C]PE2I and [11C]raclopride, indexing DAT and D2R density. The simplified reference tissue model was used to calculate binding potential (BP) values. Attention and motor behavior were investigated with a continuous performance task (CPT) and motion measurements. The BP value for [11C]PE2I and [11C]raclopride in the striatum of children with ADHD did not differ from that of the young adult control subjects. In the midbrain, however, the BP values for DAT were significantly lower (16%; p = .03) in children with ADHD. Dopamine D2 receptor binding in the right caudate nucleus correlated significantly with increased motor activity (r = .70, p = .01). The lower BP values for DAT in the midbrain suggest that dopamine signaling in subjects with ADHD is altered. Altered dopamine signaling might have a causal relationship to motor hyperactivity and might be considered as a potential endophenotype of ADHD.

The effect of a 5-HT1A receptor agonist on striatal dopamine release

5-HT1A receptor agonists consistently reduce neuroleptic induced catalepsy in rats. A serotonin-dopamine interaction has been proposed to underlie this effect. Specifically, 5-HT1A receptor agonists may reduce the activity of serotonergic projections that inhibit dopaminergic nigrostriatal neurones, therefore increasing dorsal striatal dopamine levels and partially overcoming the neuroleptic blockade of D2 receptors. We tested the hypothesis that 5-HT1A receptor agonists increase striatal dopamine release in man using PET scanning with the selective D2 receptor radioligand [11C]raclopride, which is sensitive to endogenous dopamine levels. Six healthy volunteers received two PET scans, one after placebo, the other after 1 mg flesinoxan, a selective 5-HT1A receptor agonist. Binding potential values for striatal subdivisions were determined using a simplified reference tissue model. We did not find any difference in striatal [11C]raclopride binding between conditions, even though flesinoxan lead to typical 5-HT1A receptor agonist side effects and produced elevation of growth hormone in five of the six subjects. Our results suggest that the anticataleptic effect of 5-HT1A receptor agonists is not mediated by striatal dopamine release, and indicates a need for further research with other suitable 5-HT1A receptor agonists.

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Activating mutations of the epidermal growth factor receptor (EGFR) occur in multiple tumor types, including non-small cell lung cancer (NSCLC) and malignant glioma, and have become targets for therapeutic intervention. The determination of EGFR mutation status using a noninvasive, molecular imaging approach has the potential for clinical utility. In this study, we investigated [11C]-erlotinib positron emission tomography (PET) imaging as a tool to identify activating mutations of EGFR in both glioma and NSCLC xenografts. Radiotracer specific binding was determined for high and low specific activity (SA) [11C]-erlotinib PET scans in mice bearing synchronous human cancer xenografts with different EGFR expression profiles (PC9, HCC827, U87, U87 ΔEGFR, and SW620). Although xenograft immunohistochemistry demonstrated constitutive EGFR phosphorylation, PET scan analysis using the Simplified Reference Tissue Model showed that only kinase domain mutant NSCLC (HCC827 and PC9) had significantly greater binding potentials in high versus low SA scans. Xenografts with undetectable EGFR expression (SW620), possessing wild-type EGFR (U87), and expressing an activating extracellular domain mutation (U87 ΔEGFR) were indistinguishable under both high and low SA scan conditions. The results suggest that [11C]-erlotinib is a promising radiotracer that could provide a novel clinical methodology for assessing EGFR and erlotinib interactions in patients with tumors that harbor EGFR-activating kinase domain mutations.

SPET imaging of central muscarinic receptors with ( R, R)[ 123I]-I-QNB: methodological considerations

Investigations on the effect of normal healthy ageing on the muscarinic system have shown conflicting results. Also, in vivo determination of muscarinic receptor binding has been hampered by a lack of subtype selective ligands and differences in methods used for quantification of receptor densities. Recent in vitro and in vivo work with the muscarinic antagonist ( R, R)-I-QNB indicates this ligand has selectivity for m 1 and m 4 muscarinic receptor subtypes. Therefore, we used ( R, R)[ 123I]-I-QNB and single photon emission tomography to study brain m 1 and m 4 muscarinic receptors in 25 healthy female subjects (11 younger subjects, age range 26–32 years and 14 older subjects, age range 57–82 years). Our aims were to ascertain the viability of tracer administration and imaging within the same day, and to evaluate whether normalization to whole brain, compared to normalization to cerebellum, could alter the clinical interpretation of results. Images were analyzed using the simplified reference tissue model and by two ratio methods: normalization to whole brain and normalization to cerebellum. Significant correlations were observed between kinetic analysis and normalization to cerebellum, but not to whole brain. Both the kinetic analysis and normalization to cerebellum showed age-related reductions in muscarinic binding in frontal, orbitofrontal, and parietal regions. Normalization to whole brain, however, failed to detect age-related changes in any region. Here we show that, for this radiotracer, normalizing to a region of negligible specific binding (cerebellum) significantly improves sensitivity when compared to global normalization.

Differentiation of extrastriatal dopamine D2 receptor density and affinity in the human brain using PET

Dopaminergic neurotransmission in extrastriatal regions may play a crucial role in the pathophysiology and treatment of neuropsychiatric disorders. The high-affinity radioligands [ 11C]FLB 457, [ 123I]epidepride, and [ 18F]fallypride are now used in clinical studies to measure these low-density receptor populations in vivo. However, a single determination of the regional binding potential (BP) does not differentiate receptor density ( B max) from the apparent affinity ( K D). In this positron emission tomography (PET) study, we measured extrastriatal dopamine D2 receptor density ( B max) and apparent affinity ( K D) in 10 healthy subjects using an in vivo saturation approach. Each subject participated in two to three PET measurements with different specific radioactivity of [ 11C]FLB 457. The commonly used simplified reference tissue model (SRTM) was used in a comparison of BP values with the B max values obtained from the saturation analysis. The calculated regional receptor density values were of the same magnitude (0.33–1.68 nM) and showed the same rank order as reported from postmortem studies, that is, in descending order thalamus, lateral temporal cortex, anterior cinguli, and frontal cortex. The affinity ranged from 0.27 to 0.43 nM, that is, approximately 10–20 times the value found in vitro (20 pM). The area under the cerebellar time activity curve (TAC) was slightly lower (11 ± 8%, mean ± SD, P = 0.004, n = 10) after injection of low as compared with high specific radioactivity, indicating sensitivity to the minute density of dopamine D2 receptors in the this region. The results of the present study support that dopamine D2 receptor density and affinity can be differentiated in low-density regions using a saturation approach. There was a significant ( P < 0.001) correlation between the binding potential calculated with SRTM and the receptor density ( B max), which supports the use of BP in clinical studies where differentiation of B max and K D is not required. In such studies, the mass of FLB 457 has to be less than 0.5 μg injected to avoid a mass effect of the radioligand itself.

Alfentanil increases cortical dopamine D2/D3 receptor binding in healthy subjects

Animal studies have shown that opioids modulate the function of dopaminergic neurons. The effect of alfentanil on cortical and thalamic binding of the D2/D3 receptor ligand [ 11C]FLB 457 was evaluated in eight healthy subjects with positron emission tomography. The simplified reference tissue model was used to calculate tracer binding potential (BP) during a baseline condition and target-controlled infusion of alfentanil, and the results were analyzed using a comparison group not receiving opioid. Behavioral and analgesic effects of alfentanil were also evaluated. In the region-of-interest analysis, alfentanil increased the BP of [ 11C]FLB 457 in the medial frontal cortex ( P=0.028), dorsolateral prefrontal cortex ( P=0.027), superior temporal cortex ( P=0.028), and medial thalamus ( P=0.003). These results were confirmed in a voxel-based analysis, which further revealed an opioid-induced increase in [ 11C]FLB 457 BP in the anterior cingulate cortex ( P<0.001). Alfentanil induced euphoria ( P=0.03) and analgesia ( P=0.006). Cheerfulness ( r=0.982, P<0.001) and euphoria ( r=0.918, P=0.001) were associated with increased BP of [ 11C]FLB 457 in the left posterior cingulate cortex, but the analgesic effect of alfentanil did not correlate with changes in [ 11C]FLB 457 BP. The results of this study demonstrate opioid–dopamine interactions in frontal and temporal cortical regions and the thalamus in healthy subjects. Increased D2/D3 tracer binding during opioid infusion may reflect decreased synaptic dopamine levels. The association of the uplifting effect of alfentanil with increased D2/D3 binding in the posterior cingulate cortex suggests that cortical dopamine may be involved in the behavioral effects of opioids.

Cluster analysis in kinetic modelling of the brain: a noninvasive alternative to arterial sampling

In emission tomography, quantification of brain tracer uptake, metabolism or binding requires knowledge of the cerebral input function. Traditionally, this is achieved with arterial blood sampling. We propose a noninvasive alternative via the use of a blood vessel time–activity curve (TAC) extracted directly from dynamic positron emission tomography (PET) scans by cluster analysis. Five healthy subjects were injected with the 5HT 2A-receptor ligand [ 18F]-altanserin and blood samples were subsequently taken from the radial artery and cubital vein. Eight regions-of-interest (ROI) TACs were extracted from the PET data set. Hierarchical K-means cluster analysis was performed on the PET time series to extract a cerebral vasculature ROI. The number of clusters was varied from K = 1 to 10 for the second of the two-stage method. Determination of the correct number of clusters was performed by the ‘within-variance’ measure and by 3D visual inspection of the homogeneity of the determined clusters. The cluster-determined input curve was then used in Logan plot analysis and compared with the arterial and venous blood samples, and additionally with one of the currently used alternatives to arterial blood sampling, the Simplified Reference Tissue Model (SRTM) and Logan analysis with cerebellar TAC as an input. There was a good agreement ( P < 0.05) between the values of Distribution Volume (DV) obtained from the K-means-clustered input function and those from the arterial blood samples. This work acts as a proof-of-principle that the use of cluster analysis on a PET data set could obviate the requirement for arterial cannulation when determining the input function for kinetic modelling of ligand binding, and that this may be a superior approach as compared to the other noninvasive alternatives.

Decreased thalamic D2/D3 receptor binding in drug-naive patients with schizophrenia: a PET study with [11C]FLB 457.

The thalamus is a neuroanatomic structure that has reciprocal connections with several brain regions suggested to be involved in the pathophysiology of schizophrenia. Recent studies have reported structural as well as functional abnormalities of the thalamus in schizophrenia. The aim of the present exploratory study was to examine D2/D3 dopamine receptors in the thalamus as well as the anterior cingulate and the frontal and temporal cortices by using the high-affinity radioligand [11C]FLB 457 and positron emission tomography (3D PET) and to explore the data in relation to disease, age and psychopathology. Nine drug-naive patients with schizophrenia and eight control subjects were examined. Regional binding potential (BP) values were calculated using the simplified reference tissue model. The D2/D3 receptor binding was significantly lower in the right medial thalamus in the schizophrenia patients compared to control subjects. In addition, we found a significant negative age effect on the D2/D3 BP in the frontal and temporal cortex for both groups. There was no significant age effect on the D2/D3 BP in the thalamus or in the anterior cingulate. The result provides additional support to the view that the age effect on D2/D3 receptors differ between brain regions. The preliminary finding of low thalamic D2/D3 BP in patients strengthens the hypothesis that the thalamus is a key region in the pathophysiology of schizophrenia.

Linear regression with spatial constraint to generate parametric images of ligand-receptor dynamic PET studies with a simplified reference tissue model

For the quantitative analysis of ligand-receptor dynamic positron emission tomography (PET) studies, it is often desirable to apply reference tissue methods that eliminate the need for arterial blood sampling. A common technique is to apply a simplified reference tissue model (SRTM). Applications of this method are generally based on an analytical solution of the SRTM equation with parameters estimated by nonlinear regression. In this study, we derive, based on the same assumptions used to derive the SRTM, a new set of operational equations of integral form with parameters directly estimated by conventional weighted linear regression (WLR). In addition, a linear regression with spatial constraint (LRSC) algorithm is developed for parametric imaging to reduce the effects of high noise levels in pixel time activity curves that are typical of PET dynamic data. For comparison, conventional weighted nonlinear regression with the Marquardt algorithm (WNLRM) and nonlinear ridge regression with spatial constraint (NLRRSC) were also implemented using the nonlinear analytical solution of the SRTM equation. In contrast to the other three methods, LRSC reduces the percent root mean square error of the estimated parameters, especially at higher noise levels. For estimation of binding potential (BP), WLR and LRSC show similar variance even at high noise levels, but LRSC yields a smaller bias. Results from human studies demonstrate that LRSC produces high-quality parametric images. The variance of R 1 and k 2 images generated by WLR, WNLRM, and NLRRSC can be decreased 30%–60% by using LRSC. The quality of the BP images generated by WLR and LRSC is visually comparable, and the variance of BP images generated by WNLRM can be reduced 10%–40% by WLR or LRSC. The BP estimates obtained using WLR are 3%–5% lower than those estimated by LRSC. We conclude that the new linear equations yield a reliable, computationally efficient, and robust LRSC algorithm to generate parametric images of ligand-receptor dynamic PET studies.

Quantification of brain μ-opioid receptors with [ 11C]carfentanil: reference-tissue methods

[ 11C]Carfentanil (CFN) is a μ-opioid agonist used for in vivo positron emission tomography (PET) studies of μ-opioid receptors. Previously, a tissue-ratio method was validated for the quantification of CFN binding. However, since that initial validation, several other blood independent (reference-tissue) methods have become available. To evaluate these methods, CFN PET studies with arterial blood sampling were acquired in six healthy male control subjects. Specific binding estimates obtained from reference-tissue methods were compared to those obtained with a more rigorous blood input modeling technique. It was determined that both a graphical method, and a simplified reference tissue model, were more accurate than the tissue-ratio method for quantification of CFN binding.

Noise reduction in the simplified reference tissue model for neuroreceptor functional imaging.

The Simplified Reference Tissue Model (SRTM) produces functional images of receptor binding parameters using an input function derived from a reference region and assuming a model with one tissue compartment. Three parameters are estimated: binding potential (BP), relative delivery (R1), and the reference region clearance constant k'2. Since k'2 should not vary across brain pixels, the authors developed a two-step method (SRTM2) using a global value of k'2. Whole-brain simulations were performed using human input functions and rate constants for [18F]FCWAY, [11C]flumazenil, and [11C]raclopride, and parameter SD and bias were determined for SRTM and SRTM2. The global mean of k'2 was slightly biased (2% to 6%), but the median was unbiased (<1%) and was used as the global value. Binding potential noise reductions with SRTM2 were 4% to 14%, 20% to 53%, and 10% to 30% for [18F]FCWAY, [11C]flumazenil, and [11C]raclopride, respectively, with larger reductions for shorter scans. R1 noise reduction was larger than that of BP. Simulations were also performed to assess bias when the reference and/or tissue regions followed a two-tissue compartment model. Owing to the constrained k'2, SRTM2 showed somewhat larger biases due to violations of the one-compartment model assumption. These studies demonstrate that SRTM2 should be a useful method to improve the quality of neuroreceptor functional images.

Noise Reduction in the Simplified Reference Tissue Model for Neuroreceptor Functional Imaging

Noise Reduction in the Simplified Reference Tissue Model for Neuroreceptor Functional Imaging

A Database of [11C]WAY-100635 Binding to 5-HT1A Receptors in Normal Male Volunteers: Normative Data and Relationship to Methodological, Demographic, Physiological, and Behavioral Variables

PET studies of [11C]WAY-100635 binding are proving to be a useful tool to evaluate 5-HT1A receptor function in vivo in humans. We describe the pattern of [11C]WAY-100635 binding in 61 healthy male brains and examine its variability. For all PET scans, binding potential (BP) values for [11C]WAY-100635 in different regions were calculated using a simplified reference tissue model, with the cerebellum as reference region. Specifically we describe (1) region of interest and SPM databases of PET [11C]WAY-100635 binding, including test–retest variability; (2) the sensitivity of [11C]WAY-100635 binding to manipulations of endogenous 5-HT; and (3) correlations between [11C]WAY-100635 binding and radiochemical, demographic, physiological, and behavioral variables. The regional distribution of [11C]WAY-100635 binding in healthy human brain was similar to that reported in vitro. The test–retest variability was ∼12% (range 9–16%) and was similar for all methods of regional sampling. The binding of [11C]WAY-100635 was insensitive to changes in brain 5-HT induced by tryptophan infusion and depletion. Although BP values varied greatly across subjects (range 2.9–6.8), there were no significant correlations of regional and global BP with common radiochemical, demographic, physiological, and personality variables. Specifically, in contrast with two recent small studies, we found no decline of [11C]WAY-100635 binding with age in our large cohort over the age range of 24 to 53 years. Assessment of 5-HT1A receptors in vivo using PET and [11C]WAY-100635 gives reliable measures of 5-HT1A binding. The large between-subject variability observed could not be explained by common methodological, physiological, or behavioral factors and hence the biological basis of this variability remains to be clarified.

Potentials and Pitfalls Using High Affinity Radioligands in PET and SPET Determinations on Regional Drug Induced D2 Receptor Occupancy—A Simulation Study Based on Experimental Data

The D2 dopamine receptor density ranges from 0.2 to 40 nM among human brain regions. For high density regions radioligands like [11C]raclopride provide accurate and reliable estimates of the receptor density. In research on neuropsychiatric disorders there is, however, a growing need for quantitative approaches that accurately measure D2 dopamine receptor occupancy induced by drugs or endogenous dopamine in regions with low receptor density. The new high affinity radioligands [11C]FLB 457 and [123I]epidepride have been shown to provide a signal for extrasriatal D2 dopamine receptor populations in the human brain in vivo. Initial observations indicate, however, that the time required to reach equilibrium is dependent on receptor density. Ratio analyses may thus not be readily used for comparisons among different brain regions. The aim of the present simulation study was to examine commonly used approaches for calculation of drug induced D2 dopamine receptor occupancy among regions with widely different receptor density. The input functions and the rate constants of [11C]FLB 457 and the reference ligand [11C]raclopride were first used in a simulation estimating the effect of receptor density on equilibrium time. In a second step we examined how errors produced by inaccurate determination of the binding potential parameter propagate to calculations of drug induced receptor occupancy. The simulations showed a marked effect of receptor density on equilibrium time for [11C]FLB 457, but not for [11C]raclopride. For [11C]FLB 457, a receptor density above about 7 nM caused the time of equilibrium to fall beyond time of data acquisition (1 h). The use of preequilibrium data caused the peak equilibrium and the end time ratio approaches but not the simplified reference tissue model (SRTM) approach to underestimate the binding potential and thus also the drug occupancy calculated for high-density regions. The study supports the use of ratio and SRTM analyses in extrastriatal low-density receptor regions for which the high affinity ligand [11C]FLB 457 was developed. However, in high-density regions such as the human striatum simple ratio approaches cannot be validly applied, whereas the SRTM approach has higher potential to provide valid estimates. Interestingly, the results suggest that published data on a proposed extrastriatal selectivity for the antipsychotic drugs clozapine and olanzapine may be due to erroneous estimations of the binding potential when using ratio approaches.

Quantifying drug-related 5-HT1A receptor occupancy with.

There is an increased interest in measuring the interaction of new or established drugs with their targets, in order to gain a better understanding of their mechanisms of action. PET can provide this information if an appropriate radioligand is available. [18F]MPPF (4-(2'-methoxyphenyl)-1-[2'-(N-2"-pyridinyl)-p-[18F]fluorobenzamido]ethylpiperazine) is a selective radioligand for serotonin 5-HT1A receptors. We have established that the binding potential (BP=Bmax/KD) of [18F]MPPF for cerebral 5-HT1A receptors can be assessed in human brain without arterial sampling. The aim of this study was to assess if 5-HT1A receptor occupancy can be measured through calculation of a drug-related decrease in BP with [18F]MPPF and PET. Six volunteers were scanned twice using a Siemens Exact HR+ camera following injection of 70+/-18 MBq [18F]MPPF (baseline and medicated conditions). Before the second scan, volunteers orally received either 3x10 mg pindolol at T=-15.5 h, T=-6.5 h, and T=-1.5 h (n=3) or 10 mg buspirone in a single dose at T=-1.5 h (n=3). Binding potentials were calculated using the simplified reference tissue model with the cerebellum as reference. Administration of 30 mg pindolol led to a significant reduction in [18F]MPPF binding potential of 42+/-17%. In contrast, no significant reduction of [18F]MPPF binding potential was observed following administration of buspirone (5+/-17%). These results show that [18F]MPPF can be used for measurement of drug-related 5-HT1A receptor occupancy and may be of particular interest in determining the 5-HT1A receptor interaction of new or established drugs in phase 1 and early phase 2 drug trials. Apparently, the 5-HT1A partial agonist buspirone is already clinically effective at low levels of 5-HT1A receptor occupancy.

Quantitative measurement of cerebral acetylcholinesterase using.

[11C]physostigmine, an acetylcholinesterase inhibitor, has been shown to be a promising positron emission tomography ligand to quantify the cerebral concentration of the enzyme in animals and humans in vivo. Here, a quantitative and noninvasive method to measure the regional acetylcholinesterase concentration in the brain is presented. The method is based on the observation that the ratio between regions rich in acetylcholinesterase and white matter, a region almost entirely deprived of this enzyme, was found to become approximately constant after 20 to 30 minutes, suggesting that at late time points the uptake mainly contains information about the distribution volume. Taking the white matter as the reference region, a simplified reference tissue model, with effectively one reversible tissue compartment and three parameters, was found to give a good description of the data in baboons. One of these parameters, the ratio between the total distribution volumes in the target and reference regions, showed a satisfactory correlation with the acetylcholinesterase concentration measured postmortem in two baboon brains. Eight healthy male subjects were also analyzed and the regional enzyme concentrations obtained again showed a good correlation with the known acetylcholinesterase concentrations measured in postmortem studies of human brain.

Derivation of [ 11C]WAY-100635 binding parameters with reference tissue models: effect of violations of model assumptions

In several positron-emission tomography studies of human subjects, analyses of data from the 5-hydroxytryptamine 1A (5-HT 1A) receptor radioligand, [ 11C]WAY-100635 {[ carbonyl- 11C] N-(2-(4-methoxy-phenyl)-1-piperazin-1-yl)ethyl)- N-(2-pyridyl)cyclohexanecarboxamide} have shown a discrepancy between the outcome measure k 3/k 4 (binding potential normalized to cerebellum) as estimated by the simplified reference region method and results obtained by conventional kinetic modeling with an arterial input function. The reference region method has yielded results that are lower than the conventional approach, with the relative underestimation appearing to be an increasing function of k 3/k 4. We performed simulations on idealized data to identify the source of the discrepancy. Both the simplified reference tissue model (SRTM) and the original full reference tissue model (FRTM) were tested to determine (a) if the error in estimated k 3/k 4 is dependent on the blood flow in the region of interest relative to the blood flow in the region of reference (R 1) and on the receptor density in the region of interest (true k 3/k 4), and (b) which violation of the reference model assumptions were responsible for this effect. FRTM returned parameter estimates that were independent and accurate if the reference region was constructed precisely as a one-tissue compartment model. SRTM overestimated k 3/k 4 when the reference region was constructed as a one-tissue compartment model and underestimated k 3/k 4 when the reference region was constructed as a two-tissue compartment model (which is the case for [ 11C]WAY-100635). In both cases, the magnitude of the error in k 3/k 4 returned by SRTM was dependent on true R 1 and true k 3/k 4. In conclusion, the SRTM is associated with a bias in the derivation of k 3/k 4 that is not a simple scaling factor. This magnitude of these errors should be carefully evaluated for each new radioligand.

Use of PET and the radioligand [ carbonyl- 11C]WAY-100635 in psychotropic drug development

Positron-emission tomography (PET) provides potential in neuropsychiatric drug development by expanding knowledge of drug action in the living human brain and reducing time consumption and costs. The 5-hydroxytryptamine 1A (5-HT 1A) receptor is of central interest as a target for the treatment of anxiety, depression, and schizophrenia. Research on the clinical significance of the 5-HT 1A receptor now benefits from the highly selective radioligand [ carbonyl- 11C]WAY-100635 (WAY) for quantitative determination of 5-HT 1A receptors in the primate and human brain in vivo using PET. In this paper, three studies are reviewed to demonstrate the suitability of WAY as radioligand for quantification of central 5-HT 1A receptors in brain and as an applicable tool for drug development. In the first study a monkey model was used to characterize WAY binding. It was confirmed that the reference ligand 8-OH-DPAT and psychoactive drugs such as buspirone and pindolol occupies 5-HT 1A receptors in the primate brain. Pindolol is an β-adrenoreceptor antagonist with a high affinity to 5-HT 1A receptors. This drug has been suggested in combination with selective serotonin reuptake inhibitors for the treatment of depression and was given to healthy males in the second study. Pindolol induced a marked inhibition of central 5-HT 1A receptors as calculated by the ratio-analysis method and simplified reference tissue model, 2 h after administration of 10 mg as a single oral dose. This observation suggests that pindolol may have a role for the suggested potentiation of selective serotonin reuptake inhibitor treatment of depression. The third study was on robalzotan (NAD-299), a recently developed 5-HT 1A receptor antagonist and putative drug with implications for the treatment of depression. In the cynomolgus monkey brain, robalzotan in the dose range 2–100 μg/kg IV occupied 5-HT 1A receptors in a dose-dependent and saturable manner with a maximal calculated occupancy of 70–80%. The relationship between robalzotan plasma concentration and 5-HT 1A receptor occupancy could be described by a hyperbolic function that was used to guide the selection of appropriate doses in man. In a subsequent PET study of robalzotan binding to 5-HT 1A receptors in the living human brain, similar results have been replicated recently. These studies reviewed here illustrate and corroborate that quantitative neuroimaging of receptor binding has potential for the evaluation and dose finding of new central nervous system drugs.