Quantification of dopamine transporters in the mouse brain using ultra-high resolution single-photon emission tomography

Functional imaging of small animals, such as mice and rats, using ultra-high resolution positron emission tomography (PET) and single-photon emission tomography (SPET), is becoming a valuable tool for studying animal models of human disease. While several studies have shown the utility of PET imaging in small animals, few have used SPET in real research applications. In this study we aimed to demonstrate the feasibility of using ultra-high resolution SPET in quantitative studies of dopamine transporters (DAT) in the mouse brain. Four healthy ICR male mice were injected with (mean+/-SD) 704+/-154 MBq [(99m)Tc]TRODAT-1, and scanned using an ultra-high resolution SPET system equipped with pinhole collimators (spatial resolution 0.83 mm at 3 cm radius of rotation). Each mouse had two studies, to provide an indication of test-retest reliability. Reference tissue kinetic modeling analysis of the time-activity data in the striatum and cerebellum was used to quantitate the availability of DAT. A simple equilibrium ratio of striatum to cerebellum provided another measure of DAT binding. The SPET imaging results were compared against ex vivo biodistribution data from the striatum and cerebellum. The mean distribution volume ratio (DVR) from the reference tissue kinetic model was 2.17+/-0.34, with a test-retest reliability of 2.63%+/-1.67%. The ratio technique gave similar results (DVR=2.03+/-0.38, test-retest reliability=6.64%+/-3.86%), and the ex vivo analysis gave DVR=2.32+/-0.20. Correlations between the kinetic model and the ratio technique ( R(2)=0.86, P<0.001) and the ex vivo data ( R(2)=0.92, P=0.04) were both excellent. This study demonstrated clearly that ultra-high resolution SPET of small animals is capable of accurate, repeatable, and quantitative measures of DAT binding, and should open up the possibility of further studies of cerebral binding sites in mice using pinhole SPET.     

Simplified reference region model for the kinetic analysis of [99mTc]TRODAT-1 binding to dopamine transporters in nonhuman primates using single-photon emission tomography.

Accurate quantification of neuroreceptors requires full kinetic modeling of the dynamic single-photon emission tomography (SPET) or positron emission tomography (PET) images, with highly invasive arterial blood sampling. This study investigated the application of a reference region kinetic model to the dynamics of [99mTc]TRODAT-1 in nonhuman primates, obviating the need for blood sampling. A series of dynamic SPET scans were performed on two baboons following the injection of approximately 700 MBq of [99mTc]TRODAT-1. Rapid arterial blood samples were taken automatically during scanning. Reconstructed SPET images were coregistered with magnetic resonance imaging (MRI) scans of the baboons, and regions of interest (ROIs) placed on the striatum, cerebellum and cerebral hemispheres. The ROI data were combined with metabolite-corrected blood data, and fitted to a three-compartment kinetic model using nonlinear least squares techniques. The same data were also used in a simplified reference region model, in which the input function was derived from the nondisplaceable tissue compartment. In addition, semiquantitative blinded analysis was performed by three raters to determine the point of transient equilibrium in the specific binding curves. All methods generated values for the ratio of the kinetic rate constants k3/k4, which gives an estimate of the binding potential, BP. These were compared with the full kinetic model. The mean values of k3/k4 for the three different analysis techniques for each baboon were: 1.17 +/- 0.21 and 1.12 +/- 0.13 (full kinetic model), 0.93 +/- 0.13 and 0.90 +/- 0.07 (reference region model), and 0.97 +/- 0.18 and 0.92 +/- 0.08 (equilibrium method). The reference region method gave significantly lower results than the full kinetic model (P = 0.01), but it also produced a much smaller spread and better quality fits to the kinetic data. The reference region model results for k3/k4 correlated very strongly with the full kinetic analysis (r2 = 0.992, P < 0.001), and with the equilibrium model (r2 = 0.88, P = 0.002). The subjectivity inherent in the equilibrium method produces inferior results compared with both kinetic analyses. It is suggested that the self-consistency of the reference region model, which requires no arterial blood sampling, provides a more precise and reliable estimate of the binding of [99mTc]TRODAT-1 to dopamine transporters than full kinetic modeling. The reference region method is also better suited to a routine clinical environment, and would be able to distinguish smaller differences in dopaminergic function between patient groups.     

Quantification of [<Superscript>123</Superscript>I]PE2I binding to dopamine transporters with SPET

The iodinated cocaine derivative [(123)I]PE2I is a new selective ligand for in vivo studies of the dopamine transporter (DAT) with single-photon emission tomography (SPET). The aim of the present study was to describe a method for accurate quantification of binding data following a bolus injection of [(123)I]PE2I. Six healthy subjects (age 51+/-24 years) underwent xenon-133 SPET for quantification of regional CBF and [(123)I]PE2I SPET for quantification of DAT binding. rCBFs were within normal limits in all subjects. Fitting data to a two-tissue compartment model resulted in striatal K(1) values of 0.39+/-0.08 ml ml(-1) min(-1), equal to a first-pass extraction fraction of 0.72+/-0.13. Distribution volumes (DVs) were calculated using compartment analysis, area under the curve analysis and Logan analysis. Logan analysis is preferred since stable DV values were already obtained 120 min after [(123)I]PE2I injection. Mean striatal DV was 37.9+/-9.6 ml ml(-1) and mean occipital cortex DV was 5.5+/-0.7 ml ml(-1). In the absence of local pathology in a reference tissue, Logan analysis without blood sampling is an attractive method for accurate quantification of striatal [(123)I]PE2I binding. The distribution volume ratio (DVR) (6.6+/-1.4) was in good agreement with the DVR calculated with blood (6.7+/-1.4).     

In vivo imaging of serotonin transporters with [99mTc]TRODAT-1 in nonhuman primates

[99mTc]TRODAT-1 was the first 99mTc-labeled imaging agent to show specific binding to dopamine transporters (DAT) in the striatum (STR) of human brain. Additionally, in vitro binding and autoradiographic experiments demonstrated that this tracer also binds to serotonin transporters (SERT) in the midbrain/hypothalamus (MB) area. In this study, [99mTc]TRODAT-1 was investigated as a potentially useful ligand to image SERT in the MB of living brain. A total of eight single-photon emission tomography (SPET) scans were performed in two baboons (Papio anubis) after intravenous (i.v.) injection of 740 MBq (20 mCi) of [99mTc]TRODAT-1 using a triple-head gamma camera equipped with ultra-high-resolution fan-beam collimators (scan time: 0-210 min). In four blocking studies, baboons were pretreated with (+)McN5652 (1 mg/kg, i.v.) or methylphenidate (1 mg/kg, i.v.) to specifically block SERT or DAT, respectively. After co-registration with magnetic resonance images of the same baboon, a region of interest analysis was performed using predefined templates to calculate specific uptake in the midbrain area and the striatum, with the cerebellum as the background region [(MB-CB)/CB, (STR-CB)/CB]. Additionally, two PET scans of the same baboons were performed after i.v. injections of 74-111 MBq (2-3 mCi) of [11C](+)McN5652 to identify the SERT sites. In [99mTc]TRODAT-1/SPET scans, the SERT sites in the MB region were clearly visualized. Semiquantitative analysis revealed a specific uptake in MB ([MB-CB]/CB) of 0.30+/-0.02, which was decreased to 0. 040+/-0.005 after pretreatment with nonradioactive (+)McN5652, a selective SERT ligand. Pretreatment with methylphenidate reduced the specific binding of [99mTc]TRODAT-1 to DAT sites [(STR-CB)/CB] from 2.45+/-0.13 to 0.32+/-0.04 without any effect on its binding to SERT sites [(MB-CB)/CB], which was confirmed by the co-registration of the [11C](+)McN5652/PET scans. This preliminary study suggests that specific binding of [99mTc]TRODAT-1 to SERT sites can be detected by in vivo SPET imaging despite the low target to background ratio. These findings provide impetus for further development of similar compounds with improved binding affinity and selectivity to SERT sites.     

The optimal imaging time for [99Tcm]TRODAT-1/SPET in normal subjects and patients with Parkinson's disease

Imaging of dopamine transporters (DATs) in the brain using [99Tcm]TRODAT-1 showed excellent pharmacokinetics for estimation of transporter concentrations. It has been reported that there may be differences in the binding kinetics of DAT radiotracers to DATs between normal subjects and patients with Parkinson's disease (PD). The aim of this study was to determine an optimal time point for (99Tcm]TRODAT-1 brain single photon emission tomography (SPET) acquisition that provides stable target to non-target ratios reflecting the DAT concentration in the brain. Serial [99Tcm]TRODAT-1 brain SPET images 2, 3 and 4 h after intravenous injection of [99Tcm]TRODAT-1 (925 MBq) were performed in five healthy subjects and nine PD patients. Regions of interests were drawn, and caudate/occipital (C/O) and putamen/occipital (P/O) specific to non-specific [99Tcm]TRODAT-1 binding ratios were calculated. The C/O and P/O ratios in healthy subjects showed consistent increases with time, but in PD patients, the C/O and P/O ratios of [99Tcm]TRODAT-1 reached a stable level at 3 h post-injection. There was a statistically significant difference (P < 0.001) between PD and normal subjects at 4 h post-injection for both the C/O and the P/O ratios. In conclusion, we recommend the acquisition of [99Tcm]TRODAT-1 SPET images at 4 h post-injection, as at this time point the C/O and P/O ratios can be used to discriminate between PD patients and healthy subjects.     

Attention deficit hyperactivity disorder: binding of [99mTc]TRODAT-1 to the dopamine transporter before and after methylphenidate treatment

Involvement of the dopaminergic system has been suggested in patients suffering from attention deficit hyperactivity disorder (ADHD) since the symptoms can be successfully treated with methylphenidate, a potent blocker of the dopamine transporter (DAT). This study reports the findings on the status of the DAT in adults with ADHD before and after commencement of treatment with methylphenidate, as measured using [99mTc]TRODAT-1. Seventeen patients (seven males, ten females, aged 21-64 years, mean 38 years) were examined before and after the initiation of methylphenidate treatment (3x5 mg/day). All subjects were injected with 800 MBq [99mTc]TRODAT-1 and imaged 3 h p.i. Single-photon emission tomography (SPET) scans were acquired using a triple-headed gamma camera. For semiquantitative evaluation of the DAT, transverse slices corrected for attenuation were used to calculate specific binding in the striatum, with the cerebellum used as background [(STR-BKG)/BKG]. Data were compared with an age-matched control group. It was found that untreated patients presented with a significantly increased specific binding of [99mTc]TRODAT-1 to the DAT as compared with normal controls [(STR-BKG)/BKG: 1.43+/-0.18 vs 1.22+/-0.06, P<0.001]. Under treatment with methylphenidate, specific binding decreased significantly in all patients [(STR-BKG)/BKG: 1.00+/-0.14, P<0.001]. Our findings suggest that the number of DAT binding sites is higher in drug-naive patients suffering from ADHD than in normal controls. The decrease in available DAT binding sites under treatment with methylphenidate correlates well with the improvement in clinical symptoms. The data of this study help to elucidate the complex dysregulation of the dopaminergic neurotransmitter system in patients suffering from ADHD and the effect of treatment with psychoactive drugs.     

Pharmacological effects of dopaminergic drugs on in vivo binding of [99mTc]TRODAT-1 to the central dopamine transporters in rats

R -(exo-exo)]-, [^99mTc]TRODAT-1, to DAT. This paper describes the further characterization of [^99mTc]TRODAT-1 binding sites in rats under conditions which may exist in patients receiving various drug treatments. All experiments were carried out using an i.v. injection of [^99mTc]TRODAT-1 into male Sprague-Dawley rats. Measurements of % dose/gram ratio of (striatum–cerebellum)/cerebellum at 1 h post injection were used as an indicator for specific DAT binding. The biodistribution studies were performed in the presence of drugs which compete for the binding site, such as CFT (WIN 35,428) and methylphenidate, drugs which influence dopamine levels, such as l-DOPA, γ-hydroxybutyrolactone, and α-methyl-p-tyrosine, and d-amphetamine, which both acts as a competitor for DAT binding and increases dopamine levels. Additionally, the influence of dopamine receptor agonists, such as apomorphine and (+)bromocriptine, on biodistribution was tested. Binding of [^99mTc]TRODAT-1 to DAT was found to be inhibited by CFT, methylphenidate, and d-amphetamine in a dose-dependent manner. The specific binding of [^99mTc]TRODAT-1 was not altered by dopamine receptor agonists or by drugs which cause minor changes in dopamine levels. When administered in high doses (634 μmol/kg), l-DOPA also decreased the binding of [^99mTc]TRODAT-1. It is likely that a low dose of l-DOPA (normally needed in the treatment of Parkinson’s disease) will not affect the results on [^99mTc]TRODAT-1 single-photon emission tomographic (SPET) imaging studies. In conclusion, the results clearly demonstrate the specificity of [^99mTc]TRODAT-1 binding to DAT in vivo. Competition for [^99mTc]TRODAT-1 binding was observed only with drug treatment that significantly increases dopamine levels or actively competes for binding at DAT. The results suggest that prior knowledge of whether patients are receiving various drug treatments may assist in the interpretation of DAT status as assessed by SPET imaging studies using [^99mTc]TRODAT-1. Received 7 July and in revised form 27 September 1997     

Quantification of serotonin transporters in nonhuman primates using [(123)I]ADAM and SPECT.

We reported recently a highly selective radioligand, 2-([2-([dimethylamino]methyl)phenyl]thio)-5-[(123)I]iodophenylamine (ADAM), for SPECT imaging of serotonin transporters (SERT). In this article we describe the kinetic modeling of [(123)I]ADAM and its ability to quantitatively and reproducibly measure the concentrations of SERT in the nonhuman primate brain. We also investigate simplified models of tracer behavior that do not require invasive arterial blood sampling. METHODS: Three female baboons each underwent 3 [(123)I]ADAM SPECT studies. The studies consisted of a dynamic sequence of seventy-two 5-min scans after injection of 330 +/- 50 MBq (mean +/- SD) [(123)I]ADAM. Rapid arterial blood samples were obtained and corrected for the presence of labeled metabolites. Dynamic imaging and metabolite-corrected plasma data were analyzed using graphic analysis to give the distribution volumes (DVs) of different brain regions. DV ratios (DVRs) of target to cerebellum were derived and compared against a kinetic reference tissue model and simple target-to-background ratio. RESULTS: Averaged over all 9 scans, the mean DV in the midbrain was 4.86 +/- 1.06 mL/mL and the mean DV in the cerebellum was 2.25 +/- 0.48 mL/mL. The mean test-retest repeatability of the midbrain DV was 14.5%. The reference tissue model gave a mean midbrain DVR of 2.01 +/- 0.17 and correlated strongly with the DVR calculated from the full kinetic model (correlation coefficient [R(2)] = 0.94; P < 0.001), but with much improved repeatability (test-retest, 5.4%; intersubject variability, 5.2%). Similarly, the simple ratio method gave strong correlations with the full kinetic model (R(2) = 0.89; P < 0.001) and a test-retest of 7.6%. CONCLUSION: Accurate, repeatable quantification of SERT in the nonhuman primate brain is possible using kinetic modeling of dynamic [(123)I]ADAM SPECT scans. Simplified models, which do not require arterial blood sampling, gave accurate results that correlated strongly with the full kinetic model. The test-retest reliability of the simplified reference region models was excellent. Quantification of SERT is possible using full kinetic modeling and also with simpler reference region methods.     

Simplified reference region model for the kinetic analysis of [99mTc]TRODAT-1 binding to dopamine transporters in nonhuman primates using single-photon emission tomography

Accurate quantification of neuroreceptors requires full kinetic modeling of the dynamic single-photon emission tomography (SPET) or positron emission tomography (PET) images, with highly invasive arterial blood sampling. This study investigated the application of a reference region kinetic model to the dynamics of [^99mTc]TRODAT-1 in nonhuman primates, obviating the need for blood sampling. A series of dynamic SPET scans were performed on two baboons following the injection of approximately 700 MBq of [^99mTc]TRODAT-1. Rapid arterial blood samples were taken automatically during scanning. Reconstructed SPET images were co-registered with magnetic resonance imaging (MRI) scans of the baboons, and regions of interest (ROIs) placed on the striatum, cerebellum and cerebral hemispheres. The ROI data were combined with metabolite-corrected blood data, and fitted to a three-compartment kinetic model using nonlinear least squares techniques. The same data were also used in a simplified reference region model, in which the input function was derived from the nondisplaceable tissue compartment. In addition, semiquantitative blinded analysis was performed by three raters to determine the point of transient equilibrium in the specific binding curves. All methods generated values for the ratio of the kinetic rate constants k ₃ /k ₄, which gives an estimate of the binding potential, BP. These were compared with the full kinetic model. The mean values of k ₃ /k ₄ for the three different analysis techniques for each baboon were: 1.17±0.21 and 1.12±0.13 (full kinetic model), 0.93±0.13 and 0.90±0.07 (reference region model), and 0.97±0.18 and 0.92±0.08 (equilibrium method). The reference region method gave significantly lower results than the full kinetic model (P = 0.01), but it also produced a much smaller spread and better quality fits to the kinetic data. The reference region model results for k ₃ /k ₄ correlated very strongly with the full kinetic analysis (r ² = 0.992, P<0.001), and with the equilibrium model (r ² = 0.88, P = 0.002). The subjectivity inherent in the equilibrium method produces inferior results compared with both kinetic analyses. It is suggested that the self-consistency of the reference region model, which requires no arterial blood sampling, provides a more precise and reliable estimate of the binding of [^99mTc]TRODAT-1 to dopamine transporters than full kinetic modeling. The reference region method is also better suited to a routine clinical environment, and would be able to distinguish smaller differences in dopaminergic function between patient groups. Received 26 October 1998 and in revised form 11 January 1999     

Impaired dopaminergic neurotransmission in patients with traumatic brain injury: a SPECT study using 123I-beta-CIT and 123I-IBZM

Structural imaging suggests that traumatic brain injury (TBI) may be associated with disruption of neuronal networks, including the nigrostriatal dopaminergic pathway. However, to date deficits in pre- and/or postsynaptic dopaminergic neurotransmission have not been demonstrated in TBI using functional imaging. We therefore assessed dopaminergic function in ten TBI patients using [123I]2-beta-carbomethoxy-3-beta-(4-iodophenyl)tropane (beta-CIT) and [123I]iodobenzamide (IBZM) single-photon emission tomography (SPET). Average Glasgow Coma Scale score (+/-SD) at the time of head trauma was 5.8+/-4.2. SPET was performed on average 141 days (SD +/-92) after TBI. The SPET images were compared with structural images using cranial computerised tomography (CCT) and magnetic resonance imaging (MRI). SPET was performed with an ADAC Vertex dual-head camera. The activity ratios of striatal to cerebellar uptake were used as a semiquantitative parameter of striatal dopamine transporter (DAT) and D2 receptor (D2R) binding. Compared with age-matched controls, patients with TBI had significantly lower striatal/cerebellar beta-CIT and IBZM binding ratios (P< or =0.01). Overall, the DAT deficit was more marked than the D2R loss. CCT and MRI studies revealed varying cortical and subcortical lesions, with the frontal lobe being most frequently affected whereas the striatum appeared structurally normal in all but one patient. Our findings suggest that nigrostriatal dysfunction may be detected using SPET following TBI despite relative structural preservation of the striatum. Further investigations of possible clinical correlates and efficacy of dopaminergic therapy in patients with TBI seem justified.     

111In-labelled bleomycin complex for the differentiation of high- and low-grade gliomas

The aim of this study was to evaluate 111In-labelled bleomycin complex (111In-BLMC) SPET in the differentiation of high- and low-grade gliomas. Nineteen glioma patients, 14 with high-grade and five with low-grade tumours, were studied 1, 4 and 24 h after the injection of 111In-BLMC. In the high-grade glioma group, there was significant uptake of 111In-BLMC in 12 patients and no uptake in two patients based on the visual classification of SPET images at 4 and 24 h. In the low-grade glioma group, one patient had low uptake at 4 and 24 h, but the other four patients showed no visible uptake. The mean tumour to extracerebral circulation activity ratio (T/Cr) at 4 h was 0.13 +/- 0.10 (n = 5) in low-grade gliomas and 1.7 +/- 1.0 (n = 14) in high-grade gliomas. At 24 h the T/Cr ratios were 0.56 +/- 0.21 and 3.4 +/- 1.7, respectively. The mean tumour to contralateral normal brain activity ratios (T/Br) were 5.0 +/- 3.9 (4 h) and 3.0 +/- 2.8 (24 h) in low-grade gliomas, and 37.2 +/- 37.3 (4 h) and 8.3 +/- 8.2 (24 h) in high-grade gliomas. These higher T/Br ratios did not, however, result in improved differentiation between the two groups of gliomas; at 4 h the T/Cr and T/Br ratios were of equal value, as two high-grade gliomas would have been misclassified as low-grade, but at 24 h the T/Br ratio resulted in more misclassifications. Our results show that 111In-BLMC can be used in the differentiation of high- and low-grade gliomas and that the selection of the reference area for calculating tumour to non-tumour ratios is important.     

In vivo imaging of serotonin transporters with [99mTc]TRODAT-1 in nonhuman primates

[^99mTc]TRODAT-1 was the first ^99mTc-labeled imaging agent to show specific binding to dopamine transporters (DAT) in the striatum (STR) of human brain. Additionally, in vitro binding and autoradiographic experiments demonstrated that this tracer also binds to serotonin transporters (SERT) in the midbrain/hypothalamus (MB) area. In this study, [^99mTc]TRODAT-1 was investigated as a potentially useful ligand to image SERT in the MB of living brain. A total of eight single-photon emission tomography (SPET) scans were performed in two baboons (Papio anubis) after intravenous (i.v.) injection of 740 MBq (20 mCi) of [^99mTc]TRODAT-1 using a triple-head gamma camera equipped with ultra-high-resolution fan-beam collimators (scan time: 0–210 min). In four blocking studies, baboons were pretreated with (+)McN5652 (1 mg/kg, i.v.) or methylphenidate (1 mg/kg, i.v.) to specifically block SERT or DAT, respectively. After co-registration with magnetic resonance images of the same baboon, a region of interest analysis was performed using predefined templates to calculate specific uptake in the midbrain area and the striatum, with the cerebellum as the background region [(MB–CB)/CB, (STR–CB)/CB]. Additionally, two PET scans of the same baboons were performed after i.v. injections of 74–111 MBq (2–3 mCi) of [¹¹C](+)McN5652 to identify the SERT sites. In [^99mTc]TRODAT-1/SPET scans, the SERT sites in the MB region were clearly visualized. Semiquantitative analysis revealed a specific uptake in MB ([MB–CB]/CB) of 0.30±0.02, which was decreased to 0.040±0.005 after pretreatment with nonradioactive (+)McN5652, a selective SERT ligand. Pretreatment with methylphenidate reduced the specific binding of [^99mTc]TRODAT-1 to DAT sites [(STR-CB)/CB] from 2.45±0.13 to 0.32±0.04 without any effect on its binding to SERT sites [(MB–CB)/CB], which was confirmed by the co-registration of the [¹¹C](+)McN5652/PET scans. This preliminary study suggests that specific binding of [^99mTc]TRODAT-1 to SERT sites can be detected by in vivo SPET imaging despite the low target to background ratio. These findings provide impetus for further development of similar compounds with improved binding affinity and selectivity to SERT sites. Received 15 September and in revised form 15 November 1998     

Dopamine transporter density in the basal ganglia assessed with [<Superscript>123</Superscript>I]IPT SPET in children with attention deficit hyperactivity disorder

Attention deficit hyperactivity disorder (ADHD) is a psychiatric disorder in childhood that is known to be associated with dopamine dysregulation. In this study, we investigated dopamine transporter (DAT) density in children with ADHD using iodine-123 labelled N-(3-iodopropen-2-yl)-2beta-carbomethoxy-3beta-(4-chlorophenyl) tropane ([(123)I]IPT) single-photon emission tomography (SPET) and postulated that an alteration in DAT density in the basal ganglia is responsible for dopaminergic dysfunction in children with ADHD. Nine drug-naive children with ADHD and six normal children were included in the study. We performed brain SPET 2 h after the intravenous administration of [(123)I]IPT and carried out both quantitative and qualitative analyses using the obtained SPET data, which were reconstructed for the assessment of the specific/non-specific DAT binding ratio in the basal ganglia. We then investigated the correlation between the severity scores of ADHD symptoms in children with ADHD assessed with ADHD rating scale-IV and the specific/non-specific DAT binding ratio in the basal ganglia. Drug-naive children with ADHD showed a significantly increased specific/non-specific DAT binding ratio in the basal ganglia compared with normal children. However, no significant correlation was found between the severity scores of ADHD symptoms in children with ADHD and the specific/non-specific DAT binding ratio in the basal ganglia. Our findings support the complex dysregulation of the dopaminergic neurotransmitter system in children with ADHD.     

A simplified method to determine [99mTc]TRODAT-1 in human plasma

[99mTc]TRODAT-1 is a useful imaging agent in evaluating changes in presynaptic dopamine transporters (DAT) for Parkinson's disease and other central nervous system (CNS) neurodegenerative diseases, for which a reduction of dopamine neurons is indicated. As part of an effort to establish a quantitative single-photon emission tomography (SPECT) procedure for imaging CNS DAT, measurement of nonmetabolized [99mTc]TRODAT-1 in human plasma was investigated. After an intravenous injection of [99mTc]TRODAT-1, there are three possible radioactive components in human plasma: hydrophilic compounds (pertechnetate, etc.), lipophilic metabolite(s), and unchanged [99mTc]TRODAT-1. Based on the differences in lipophilicity of [99mTc]TRODAT-1 and its lipophilic metabolite [99mTc]BAT, a new quantitative method for measuring [99mTc]TRODAT-1 with a simple solvent extraction method was developed. Various organic solvents or mixtures of solvents were tested, among which cyclohexane gave the best extraction yield for [99mTc]TRODAT-1 (76.06 +/- 3.32%) with a low extraction for [99mTc]BAT (2.43 +/- 0.82%). Extractions of [99mTc]TRODAT-1 and [99mTc]BAT mixtures in different predetermined ratios to simulate the actual human plasma samples with cyclohexane from phosphate buffer (5 mM, pH 8.0) were evaluated. The experimentally obtained ratios were in good agreement with the theoretical ratios. To investigate further the possibility of replacing the previously established high performance liquid chromatography (HPLC) method with the new solvent extraction method for the clinical application, both HPLC and extraction methods were used side by side to determine the unchanged [99mTc]TRODAT-1 in human plasma samples during [99mTc]TRODAT-1/SPECT imaging studies. The results from four human subjects showed that both methods consistently produced similar values for unchanged [99mTc]TRODAT-1 in the plasma samples. This improved solvent extraction method provides an easy and reliable technique to quantify unchanged [99mTc]TRODAT-1 in human plasma, thus making the clinical application of this agent readily available for quantitation of the DAT binding sites in the brain by SPECT imaging.     

Reproducibility of 11C-raclopride binding in the rat brain measured with the microPET R4: effects of scatter correction and tracer specific activity.

A new generation of commercial animal PET cameras may accelerate drug development by streamlining preclinical testing in laboratory animals. However, little information on the feasibility of using these machines for quantitative PET in small animals is available. Here we investigate the reproducibility of microPET imaging of (11)C-raclopride in the rat brain and the effects of tracer-specific activity and photon scatter correction on measures of D2 receptor (D2R) availability. METHODS: Sprague-Dawley rats (422 +/- 29 g; n = 7) were anesthetized with ketamine/xylazine and catheterized for tail vein injection of (11)C-raclopride. Each animal was positioned prone in the microPET, centering the head in the field of view. MicroPET data was collected for 60 min-starting at (11)C-raclopride injection-and binned into 24 time frames (6 x 10 s, 3 x 20 s, 8 x 60 s, 4 x 200 s, 3 x 600 s). In 3 studies, (11)C-raclopride was administered a second time in the same animal, with 2-4 h between injections. In a fourth animal, raclopride (1 mg/kg) was coinjected with (11)C-raclopride for the second injection. Three rats received a single dose of (11)C-raclopride. The range of doses for all studies was 6.11-18.54 MBq (165-501 micro Ci). The specific activity at injection was 4.07-48.1 GBq/ micro mol (0.11-1.3 Ci/ micro mol). Region-of-interest analysis was performed and the distribution volume ratio (DVR) was computed for striatum/cerebellum using sinograms uncorrected and corrected for scatter using a tail-fit method. RESULTS: Test-retest results showed that the (11)C-raclopride microPET DVR was reproducible (change in DVR = -8.3% +/- 4.4%). The average DVR from 6 rats injected with high specific activity (<4 nmol/kg) was 2.43 +/- 0.19 (coefficient of variation = 8%). The DVR for the blocking study was 1.23. The DVR depended on the mass of tracer (11)C-raclopride injected for doses >1.5 nmol/kg. Scatter fractions within the rat head were approximately 25%-45% resulting in an average increase of DVR of 3.5% (range, 0%-10%) after correction. CONCLUSION: This study shows that the (11)C-raclopride microPET-derived DVR is reproducible and suitable for studying D2R availability in the rat brain. MicroPET sensitivity was sufficient to determine reproducible DVRs from (11)C-raclopride injections of 9.25 MBq (approximately 250 micro Ci). However, the effect of tracer mass on the DVR should be considered for studies using more than approximately 1-2 nmol/kg raclopride, and scatter correction has a measurable impact on the results.     

Increased dopamine transporter availability associated with the 9-repeat allele of the SLC6A3 gene.

A polymorphism involving a variable number of tandem repeats (VNTR) has been described in the 3' untranslated region of the gene (SLC6A3) coding for the dopamine transporter (DAT). This polymorphism has 2 common alleles, designated as 10-repeat (*10R) and 9-repeat (*9R), that have been linked with several human clinical phenotypes. Previous investigations of the effects of the SLC6A3 polymorphism on DAT availability in smaller samples of humans have yielded divergent results. METHODS: We assessed genotype at the SLC6A3 promoter VNTR polymorphism in 96 healthy European Americans (age range, 18-88 y) who also underwent SPECT with (123)I-2beta-carbomethoxy-3beta-(4-iodophenyl)tropane (beta-CIT) for measurement of striatal DAT protein availability. A ratio of specific to nondisplaceable brain uptake (i.e., V(3)' = [striatal -occipital]/occipital), a measure proportional to the binding potential, was derived. For this analysis, 9-9 homozygotes and 9-10 heterozygotes were grouped as SLC6A3 *9R carriers and contrasted with SLC6A3 *10R homozygotes. RESULTS: The SLC6A3 *9R carriers had significantly higher striatal DAT availability (V(3)') than did the SLC6A3 *10R homozygotes, controlling for age (F(1,93) = 6.25, P = 0.014, analysis of covariance). The *9R carriers (n = 41, 49.8 +/- 19.5 y) had a mean increase in striatal DAT availability of 8.9% relative to the *10R homozygotes (n = 53, 49.9 +/- 19.2 y). Striatal subregion analysis revealed that the effect of DAT genotype was significant for both the caudate and the putamen. CONCLUSION: These results support the interpretation of higher DAT levels in association with the *9R allele in European Americans and may relate to previously observed associations between DAT genotype and neuropsychiatric diseases.     

Attention deficit hyperactivity disorder: binding of [99mTc]TRODAT-1 to the dopamine transporter before and after methylphenidate treatment

Involvement of the dopaminergic system has been suggested in patients suffering from attention deficit hyperactivity disorder (ADHD) since the symptoms can be successfully treated with methylphenidate, a potent blocker of the dopamine transporter (DAT). This study reports the findings on the status of the DAT in adults with ADHD before and after commencement of treatment with methylphenidate, as measured using [^99mTc]TRODAT-1. Seventeen patients (seven males, ten females, aged 21-64 years, mean 38 years) were examined before and after the initiation of methylphenidate treatment (3ǹ mg/day). All subjects were injected with 800 MBq [^99mTc]TRODAT-1 and imaged 3 h p.i. Single-photon emission tomography (SPET) scans were acquired using a triple-headed gamma camera. For semiquantitative evaluation of the DAT, transverse slices corrected for attenuation were used to calculate specific binding in the striatum, with the cerebellum used as background [(STR-BKG)/BKG]. Data were compared with an age-matched control group. It was found that untreated patients presented with a significantly increased specific binding of [^99mTc]TRODAT-1 to the DAT as compared with normal controls [(STR-BKG)/BKG: 1.43ǂ.18 vs 1.22ǂ.06, P<0.001]. Under treatment with methylphenidate, specific binding decreased significantly in all patients [(STR-BKG)/BKG: 1.00ǂ.14, P<0.001]. Our findings suggest that the number of DAT binding sites is higher in drug-naive patients suffering from ADHD than in normal controls. The decrease in available DAT binding sites under treatment with methylphenidate correlates well with the improvement in clinical symptoms. The data of this study help to elucidate the complex dysregulation of the dopaminergic neurotransmitter system in patients suffering from ADHD and the effect of treatment with psychoactive drugs.     

Effect of temporal sampling on evaluation of left ventricular ejection fraction by means of thallium-201 gated SPET: comparison of 16- and 8-interval gating, with reference to equilibrium radionuclide angiography

Gated myocardial single-photon emission tomography (SPET) allows the evaluation of left ventricular ejection fraction (LVEF), but temporal undersampling may lead to systolic truncation and ejection fraction underestimation. The aim of this study was to evaluate the impact of temporal sampling on thallium gated SPET LVEF measurements. Fifty-five consecutive patients (46 men, mean age 62+/-12 years) with a history of myocardial infarction (anterior 31, inferior 24) were studied. All patients underwent equilibrium radionuclide angiography (ERNA) and gated SPET 4 h after a rest injection of 185 MBq (5 mCi) of thallium-201 using either 8-interval (group 1, n=25) or 16-interval gating (group 2, n=30). In group 2, gated SPET acquisitions were automatically resampled to an 8-interval data set. Projection data were reconstructed using filtered back-projection (Butterworth filter, order 5, cut-off 0.20). LVEF was then calculated using commercially available software (QGS). A higher correlation between gated SPET and ERNA was obtained with 16-interval gating (r=0.94) compared with the resampled data set (r=0.84) and 8-interval gating (r=0.71). Bland-Altman plots showed a dramatic improvement in the agreement between gated SPET and ERNA with 16-interval gating (mean difference: -0.10%+/-5%). Using multiple ANOVA, temporal sampling was the only parameter to influence the difference between the two methods. When using 8-interval gating, gated SPET LVEF was overestimated in women and underestimated in men (ERNA minus gated SPET = -4.0%+/-9.6% in women and 3.6%+/-7.6% in men, P=0.01). In conclusion, 16-interval thallium gated SPET offered the best correlation and agreement with ERNA, and should be preferred to 8-interval gated acquisition for LVEF measurement.     

Imaging early-stage corticobasal degeneration with [99mTc]TRODAT-1 SPET

AIM: The aim of this study was to investigate the nigrostriatal dopaminergic function in patients with corticobasal degeneration (CBD) using [2-[[2-[[[3-(4-chlorophenyl)-8-methyl-8-azabicyclo[3.2.1]oct-2-yl]methyl](2-mercaptoethyl)amino]ethyl]amino]ethanethiolato(3-)-N2,N2',S2,S2']oxo-[1R-(exo-exo)]-[99mTc]technetium ([99mTc]TRODAT-1) brain single-photon emission tomography (SPET). METHODS: Five patients with probable CBD, 10 age- and duration-matched patients with idiopathic Parkinson's disease (IPD) and 10 age-matched healthy volunteers completed the SPET study. The images were obtained 4 h after intravenous injection of 925 MBq of [99mTc]TRODAT-1. Using a magnetic resonance imaging atlas of the striatum, the ratios of specific striatal binding to non-specific occipital binding were calculated. RESULTS: Clinical analysis showed that the CBD patients obtained significantly higher scores on the Unified Parkinson's Disease Rating Scale and a significantly worse score for activities of daily living. In the CBD and IPD groups, striatum-occipital/occipital, caudate nucleus-occipital/occipital and putamen-occipital/occipital ratios decreased significantly relative to those of healthy subjects. No statistical difference could be found between the CBD and IPD groups for these ratios, although relatively even, decreased uptakes in the caudate nucleus and putamen were found in the CBD group. On further analysis of the index of binding reduction, the differences between the caudate nucleus and putamen were significantly lower in the CBD group than in the IPD group. The striatal uptake of [99mTc]TRODAT-1 showed a distinct asymmetry in both the CBD and IPD patients. CONCLUSION: From this study, it can be concluded that early-stage CBD patients have a worse performance and more difficulties with daily activities than IPD patients. CBD patients demonstrated essentially similar patterns of [99mTc]TRODAT-1 binding as those with IPD. However, there was relatively more homogeneous involvement of the caudate nucleus and putamen in the CBD patients. This provides information about the differences between these patients in the early stages.     

Dopamine transporter density of basal ganglia assessed with [<Superscript>123</Superscript>I]IPT SPET in obsessive-compulsive disorder

It has been suggested that dopamine, as well as serotonin, is associated with the pathophysiology of obsessive-compulsive disorder (OCD). Thus, many studies have been performed on brain regions associated with dopamine in patients with OCD. In the present study, we investigated the DAT density of the basal ganglia using iodine-123 labelled N-(3-iodopropen-2-yl)-2-carbomethoxy-3-(4-chlorophenyl) tropane ([¹²³I]IPT) single-photon emission tomography (SPET) and evaluated the activity of the presynaptic dopamine function in patients with OCD. Fifteen patients with OCD and 19 normal control adults were included in the study. We performed brain SPET 2 h after the intravenous administration of [¹²³I]IPT and carried out both quantitative and qualitative analyses using the obtained SPET data, which were reconstructed for the assessment of the specific/non-specific dopamine transporter (DAT) binding ratio in the basal ganglia. We then investigated the correlation between the severity scores of OCD symptoms assessed with the Yale-Brown Obsessive-Compulsive Scale (Y-BOCS) and the specific/non-specific DAT binding ratio of the basal ganglia. Compared with normal control adults, patients with OCD showed a significantly increased specific/non-specific DAT binding ratio in the right basal ganglia and a tendency towards an increased specific/non-specific DAT binding ratio in the left basal ganglia. No significant correlation was found between the total scores on the Y-BOCS and the specific/non-specific DAT binding ratio of the basal ganglia. These findings suggest that the dopaminergic neurotransmitter system of the basal ganglia in patients with OCD could be involved in the pathophysiology of OCD.