Age differences in muscarinic cholinergic receptors assayed with (+)N-[(11)C]methyl-3-piperidyl benzilate in the brains of conscious monkeys.

Age-related changes in muscarinic cholinergic receptors were evaluated with the novel ligand (+)N-[(11)C]methyl-3-piperidyl benzilate ((+)3-MPB) in the living brains of young (5.9 +/- 1.8 years old) and aged (19.0 +/- 3.3 years old) monkeys (Macaca mulatta) in the conscious state using high-resolution positron emission tomography (PET). For quantitative analysis of receptor binding in vivo, metabolite-corrected arterial plasma radioactivity curves were obtained as an input function into the brain, and kinetic analyses using the three-compartment model and graphical Logan plot analysis were applied. Kinetic analyses of [(11)C](+)3-MPB indicated a regionally specific decrease in the receptor binding in vivo determined as binding potential (BP) = k(3)/k(4) in aged animals compared with young animals. Thus, the frontal and temporal cortices as well as the striatum showed age-related reduction of muscarinic cholinergic receptors in vivo, reflecting the reduced receptor density (B(max)) determined by Scatchard plot analysis in vivo. In the hippocampus, although BP of [(11)C](+)3-MPB indicated no significant age-related changes, it showed an inverse correlation with individual cortisol levels in plasma. When the graphical Logan plot analysis was applied, all regions assayed showed significant age-related decrease of [(11)C](+)3-MPB binding. These results demonstrate the usefulness of kinetic three-compartment model analysis of [(11)C](+)3-MPB with metabolite-corrected arterial plasma input as an indicator for the aging process of the cortical muscarinic cholinergic receptors in vivo as measured by PET.     

Evaluation of novel PET ligands (+)N-[11C]methyl-3-piperidyl benzilate ([11C](+)3-MPB) and its stereoisomer [11C](-)3-MPB for muscarinic cholinergic receptors in the conscious monkey brain: a PET study in comparison with

The novel muscarinic cholinergic ligands (+)N-[11C]methyl-3-piperidyl benzilate ([11C](+)3-MPB) and its stereoisomer [11C](-)3-MPB were evaluated in comparison with [11C]4-MPB in the brains of conscious monkeys (Macaca mulatta) using high-resolution positron emission tomography (PET). The regional distribution patterns of [11C](+)3-MPB and [11C]4-MPB at 60-91 min postinjection were almost identical: highest in the striatum and occipital cortex; intermediate in the temporal and frontal cortices, cingulate gyrus, hippocampus, and thalamus; lower in the pons; and lowest in the cerebellum. The uptake of [11C](+)3-MPB in all regions was higher and the dynamic range of regional uptake differences of [11C](+)3-MPB was better than those of [11C]4-MPB. The levels of [11C](-)3-MPB were much lower in all regions of the brain than [11C](+)3-MPB and [11C]4-MPB. Administration of scopolamine, a muscarinic cholinergic antagonist, at a dose of 50 microg/kg reduced the radioactivity of [11C](+)3-MPB and [11C]4-MPB in all regions except the cerebellum. Time-activity curves of [11C](+)3-MPB peaked in all regions, while those of [11C]4-MPB showed gradual increases with time in all regions except the thalamus, pons, and cerebellum. Two graphical analyses (Logan plot and Patlak plot) with plasma radioactivity as an input function into the brain were applied to evaluate receptor binding in vivo. [11C](+)3-MPB showed linear regression curves on Logan plot analysis and nonlinear curves on Patlak plot in all regions, suggesting that [11C](+)3-MPB bound reversibly to the muscarinic receptors. The in vivo binding parameters as well as uptake at 60-91 min postinjection of [11C](+)3-MPB were consistent with muscarinic receptor density in the brain as reported in vitro.     

(R)-N-[11C]methyl-3-pyrrolidyl benzilate,a high-affinity reversible radioligand for PET studies of the muscarinic acetylcholine receptor

We recently reported the synthesis and binding affinity of ligands for the muscarinic acetylcholine receptor (mAChR) based on both the pyrrolidyl and piperidyl benzilate scaffold. One of these, (R)-3-pyrrolidyl benzilate, was successfully radiolabeled with [(11)C]methyl triflate and the resulting compound, (R)-N-[(11)C]methyl-3-pyrrolidyl benzilate (3-[(11)C]NMPYB), was evaluated as a reversible, acetylcholine-sensitive tracer for the mAChR (K(i) of unlabeled 3-NMPYB is 0.72 nM). This compound displayed high, receptor-mediated retention in regions of the mouse and rat brain known to have high concentrations of mAChRs. Moreover, bolus studies in a pigtail monkey showed that this compound had superior clearance from the brain when compared to muscarinic radiotracers previously employed in human PET studies. Infusion studies in the same monkey revealed that it was possible to achieve equilibrium of radiotracer distribution for 3-[(11)C]NMPYB in both the striatum and cortex. Sensitivity to endogenous acetylcholine levels was evaluated by injecting phenserine (5 mg/kg) into rats prior to administration of 3-[(11)C]NMPYB in an equilibrium infusion protocol. This pretreatment produced a modest, statistically significant decrease (9-11%) in the distribution volume ratios for muscarinic receptor rich regions of the rat brain as compared to controls.     

Age-related changes in muscarinic cholinergic receptors in the living brain: a PET study using N-[11C]methyl-4-piperidyl benzilate combined with cerebral blood flow measurement in conscious monkeys

The effects of changes in regional cerebral blood flow (rCBF) with aging on muscarinic cholinergic receptor binding were evaluated with [15O]H(2)O and N-[11C]methyl-4-piperidyl benzilate (4-MPB) in the living brains of young (5.9+/-1.8 years old) and aged (19.0+/-3.3 years old) monkeys (Macaca mulatta) in the conscious state using high-resolution positron emission tomography (PET). For quantitative analysis of receptor binding in vivo with [11C]4-MPB, metabolite-corrected arterial plasma radioactivity curves were obtained as an input function into the brain, and graphical Patlak plot analysis was applied. In addition, two-compartment model analysis using the radioactivity curve in the cerebellum as an input function (reference analysis) was also applied to determine the distribution volume (DV=K(1)/k(2)') for [11C]4-MPB. With metabolite-corrected arterial input, Patlak plot analysis of [11C]4-MPB indicated a regionally specific decrease in muscarinic cholinergic receptor binding in vivo in the frontal and temporal cortices as well as the striatum in aged compared with young animals, showing no correlation with the degree of reduced rCBF. In contrast, on the reference analysis with cerebellar input of [11C]4-MPB, all regions assayed except the pons showed a significant age-related decrease of DV, and the degree of reduction of DV was correlated with that of rCBF. These results demonstrated the usefulness of kinetic analysis of [11C]4-MPB with metabolite-corrected arterial input, not with reference region's input, as an indicator of the aging process of cortical muscarinic cholinergic receptors in vivo measured by PET with less blood flow dependency.     

Different brain kinetics of two sigma 1 receptor ligands, [3H](+)-pentazocine and [11C]SA4503, by P-glycoprotein modulation

We compared the brain kinetics of radiolabeled (+)-pentazocine and SA4503, which have a high and selective affinity for sigma(1) receptors. Brain uptake of [(11)C]SA4503 was high after intravenous injection followed by a gradual decrease in mice, whereas that of [(3)H](+)-pentazocine rapidly decreased. The brain uptake of the two radioligands was dose-dependently reduced, but the reduction of [(3)H](+)-pentazocine was found at higher doses. Percentages of the saturable binding of [(3)H](+)-pentazocine was much lower than that of [(11)C]SA4503. The brain uptake of [(3)H](+)-pentazocine was greatly blocked by SA4503 at a dose of 2 micromol/kg, while that of [(11)C]SA4503 was blocked by (+)-pentazocine at a dose of 20 micromol/kg and over. When mice were treated with cyclosporin A, a P-glycoprotein modulator, the uptake of [(3)H](+)-pentazocine was enhanced, but that of [(11)C]SA4503 was not. Under control and P-glycoprotein-modulated conditions, the brain uptake of both radioligands was reduced by haloperidol, another representative sigma receptor ligand, to a different extent. We concluded that the P-glycoprotein modulation resulted in the different brain kinetics of the two radioligands. The radiolabeled SA4503 is suitable as an in vivo probe, but radiolabeled (+)-pentazocine is not.     

Validation of reference tissue model of PET ligand [¹¹C]+3-MPB for the muscarinic cholinergic receptor in the living brain of conscious monkey

N-[11C]methyl-3-piperidyl benzilate ([11C]+3-MPB) was developed as a positron emission tomography (PET) ligand for muscarinic cholinergic receptor (mAChR). The aim of the present study was to validate a Logan reference tissue method as an analytical method for in vivo binding of [11C]+3-MPB to mAChR. Seven monkeys (Macaca mulatta) underwent [11C]+3-MPB PET scans with an arterial blood sampling. Logan plot with arterial input function (Logan arterial input method) was performed to determine the binding potential (BP(ND)). The BP(ND) was also determined by Logan plot with the cerebellum as the reference region (Logan reference tissue method). BP(ND) values determined by Logan arterial input method and Logan reference tissue method showed a significant linear relationship. The present study suggests that the cerebellum is a suitable reference region for quantification of mAChR in the living brain with [11C]+3-MPB and PET.     

Age-related changes in muscarinic cholinergic receptors in the living brain: a PET study using N-[C]methyl-4-piperidyl benzilate combined with cerebral blood flow measurement in conscious monkeys

The effects of changes in regional cerebral blood flow (rCBF) with aging on muscarinic cholinergic receptor binding were evaluated with [¹⁵O]H₂O and N-[¹¹C]methyl-4-piperidyl benzilate (4-MPB) in the living brains of young (5.9±1.8 years old) and aged (19.0±3.3 years old) monkeys (Macaca mulatta) in the conscious state using high-resolution positron emission tomography (PET). For quantitative analysis of receptor binding in vivo with [¹¹C]4-MPB, metabolite-corrected arterial plasma radioactivity curves were obtained as an input function into the brain, and graphical Patlak plot analysis was applied. In addition, two-compartment model analysis using the radioactivity curve in the cerebellum as an input function (reference analysis) was also applied to determine the distribution volume (DV=K₁/k₂′) for [¹¹C]4-MPB. With metabolite-corrected arterial input, Patlak plot analysis of [¹¹C]4-MPB indicated a regionally specific decrease in muscarinic cholinergic receptor binding in vivo in the frontal and temporal cortices as well as the striatum in aged compared with young animals, showing no correlation with the degree of reduced rCBF. In contrast, on the reference analysis with cerebellar input of [¹¹C]4-MPB, all regions assayed except the pons showed a significant age-related decrease of DV, and the degree of reduction of DV was correlated with that of rCBF. These results demonstrated the usefulness of kinetic analysis of [¹¹C]4-MPB with metabolite-corrected arterial input, not with reference region’s input, as an indicator of the aging process of cortical muscarinic cholinergic receptors in vivo measured by PET with less blood flow dependency.     

Ketamine alters the availability of striatal dopamine transporter as measured by [(11)C]beta-CFT and [(11)C]beta-CIT-FE in the monkey brain.

The effects of ketamine anesthesia on the binding of [(11)C]-labeled cocaine analogs, [(11)C]beta-CFT (2beta-carbomethoxy-3beta-(4-fluorophenyl)tropane) and [(11)C]beta-CIT-FE (N-(2-fluoroethyl)-2beta-carbomethoxy-3beta-(4-iodophenyl)tropane), to the striatal dopamine transporter (DAT) were evaluated in the monkey brain using positron emission tomography (PET). We sequentially measured the kinetics of these labeled compounds in the brains of five young-adult male rhesus monkeys (Macaca mulatta) in the conscious state, followed by those under ketamine anesthesia with continuous infusion (3 and 10 mg/kg/h). After intravenous injection, [(11)C]beta-CFT and [(11)C]beta-CIT-FE were predominantly accumulated in the striatum in both conscious and ketamine-anesthetized states. In the conscious state, the striatal uptake of [(11)C]beta-CFT and [(11)C]beta-CIT-FE continuously increased with time up to 91 min after injection. Continuous infusion of ketamine-induced higher levels of uptake of [(11)C]beta-CFT and [(11)C]beta-CIT-FE into the brain in a dose-dependent manner as compared with conscious state, and kinetic analysis with metabolite-corrected arterial input function indicated that the binding potentials (BP = k(3)/k(4)) of both compounds were elevated by ketamine. Not only [(11)C]beta-CIT-FE but also [(11)C]beta-CFT reached the equilibrium state of specific binding in the striatum within 40-50 min after injection. The present results demonstrated that ketamine significantly alters the DAT availability as measured with [(11)C]beta-CFT and [(11)C]beta-CIT-FE in the brain.     

Potential of [(18)F]beta-CFT-FE (2beta-carbomethoxy-3beta-(4-fluorophenyl)-8-(2-[(18)F]fluoroethyl)nortropane) as a dopamine transporter ligand: A PET study in the conscious monkey brain

A dopamine transporter (DAT) ligand 2beta-carbomethoxy-3beta-(4-fluoro-phenyl)-8-(2-[(18)F]fluoroethyl)nortropane ([(18)F]beta-CFT-FE) was synthesized and evaluated in comparison with [(11)C]beta-CFT in monkey brain using animal positron emission tomography (PET). [(18)F]beta-CFT-FE and [(11)C]beta-CFT were injected intravenously to conscious monkeys for a 91-min PET scan with arterial blood sampling for metabolite analysis. In the conscious state, [(18)F]beta-CFT-FE provided a peak about 20 min after the injection and declined thereafter in the striatum of monkey brain, while [(11)C]beta-CFT continuously increased with time up to 91 min after injection. Metabolite analysis revealed that [(18)F]beta-CFT-FE was more rapidly metabolized in plasma than [(11)C]beta-CFT. The striatal binding of both ligands was dose-dependently displaced by preadministration of a specific DAT inhibitor, GBR12909, at doses of 0.5 and 5 mg/kg; however, the displacement degree of [(11)C]beta-CFT-FE was higher than that of [(18)F]beta-CFT. The effects of the anesthetics, ketamine and isoflurane, on binding were more prominent in [(11)C]beta-CFT than [(18)F]beta-CFT-FE. Specificity and affinity of beta-CFT-FE to DAT were evaluated in an in vitro assay using cloned human DAT, serotonin transporter, and norepinephrine transporter in comparison with other conventional DAT ligands, showing that beta-CFT-FE had lower affinity and higher specificity to DAT than beta-CFT and beta-CIT. These results suggested that [(18)F]beta-CFT-FE could be a potential imaging agent for DAT, providing excellent selectivity and tracer kinetics for quantitative PET imaging.     

乙酰胆碱酯酶活性的PET显像剂-[^11C]MP4A的制备

目的：合成能反映脑乙酰胆碱酯酶活性的PET显像剂[^11C]4-乙酰氧基-N-甲基哌啶（[^11C]MP4A）的前体4-乙酰氧基哌啶（P4A），制备[^11C]MP4A。方法：合成MP4A前体P4A，利用[^11C]碘代甲烷合成模块合成的[^11C]碘代甲烷（[^11C]CH3I）直接与P4A反应制备[^11C]MP4A，或将[^11C]CH3I通过三氟甲基磺酸银柱转化为[^11C]-三氟甲基磺酸甲酯（[^11C]-Triflate-CH3）后再与P4A反应制备[^11C]MP4A。结果：P4A化学纯度达99％以上。采用[^11C]-Triflate-CH3制备的[^11C]MP4A放化纯〉98％，轰击结束后（EOB）放射合成时间为20min，校正衰变后产率为40％-55％。结论：采用[^11C]-Triflate-CH3制备的[^11C]MP4A放化纯度高，产率高。     

PET imaging of ischemic neuronal death in the hippocampus of living monkeys

The aim of the present study was to visualize postischemic hippocampal neuronal death in the living monkey brain, using a high-resolution positron emission tomography (PET) and novel radioligands. In preceding papers, we reported on postischemic hippocampal neuronal death in a model of Japanese monkeys (Macaca fuscata) undergoing a 20-min complete whole-brain ischemia. Using the same model here, we investigated the in vivo bindings of two radiotracers, [11C]Ro15-4513 (a type II benzodiazepine receptor ligand) and [11C](+)3-MPB (a muscarinic cholinergic receptor ligand), in the hippocampus on day 7 after ischemia, as compared to the normal hippocampus. A significant decrease in the in vivo binding of [11C]Ro154513 and [11C(+)3-MPB was observed in the postischemic monkey hippocampus on day 7 after ischemia compared to controls. Light and electron microscopic analyses of postischemic CA1 neurons showed typical features of coagulation necrosis, as associated with a marked reduction of postsynaptic densities and presynaptic vesicles. These results suggest that semiquantification of hippocampal neuronal death is possible in the living primate brain using PET, and that the same procedures can be applied for evaluating neuronal cell loss in patients with ischemic injuries and/or dementia.     

Positron emission tomography imaging of the serotonin transporter in the pig brain using [11C](+)-McN5652 and S-([18F]fluoromethyl)-(+)-McN5652

S-([(18)F]fluoromethyl)-(+)-McN5652 ([(18)F](+)-FMe-McN5652) has recently been synthesized as a new potential radiotracer for positron emission tomography (PET) imaging of the 5-HT transporter. It is an analog of [(11)C](+)McN5652, which has been used in clinical PET studies for 5-HT transporter imaging. This article describes the comparison of these two radiotracers in pigs with respect to their in vivo binding characteristics. PET images revealed that the highest accumulation of both radiotracers was found in the ventral midbrain, thalamus, olfactory lobe, and pons which is consistent with the known density of 5-HT transporters. The specific binding was determined by subtracting the values of the inactive (-) enantiomers or of the occipital cortex from those obtained with [(11)C](+)McN5652 or [(18)F](+)-FMe-McN5652 in the time period between 75 and 115 min after radiotracer injection. The specific binding of the (18)F-labeled derivative was about 40% higher than that of the (11)C-labeled derivative. A strong inhibition of the specific binding was observed for both radiotracers after pretreatment with the selective 5-HT uptake inhibitor citalopram. [(18)F](+)-FMe-McN5652 showed faster kinetics than [(11)C](+)McN5652. It reached the binding equilibrium during a study length of 120 min, which was not the case for [(11)C](+)McN5652. It is concluded that [(18)F](+)-FMe-McN5652 is suitable for 5-HT transporter imaging with PET.     

脑乙酰胆碱酯酶的PET显像剂[^11C]4-乙酰氧基-N-甲基哌啶在大鼠体内分布的研究

目的：研究脑乙酰胆碱酯酶（AChE）活性的正电子断层扫描（PET）诊断显像剂[^11C]4-乙酰氧基-N-甲基哌啶（[^11C]MP4A）在大鼠体内的摄取和分布。方法：正常雌性SD大鼠尾静脉注射[^11C]MP4A，按注射后5、15、25、35、45和60min时间点分组后断头处死动物，迅速取血，分离脏器和脑（大脑皮质、小脑和脑干），用自动γ放射性计数仪计算经衰变校正后不同时间点各脏器放射性摄取率，以％ID／g表示。结果：一次剂量的[^11C]MP4A静脉注射后，血中5min时％ID／g为（13．44±1．88），清除迅速，60min时接近于基线水平；各脏器组织中％ID／g 15-25min达到稳态；大脑皮质5min％ID／g为（2．26±0．29）。脑内各区域的摄取在注射后15～25min达到稳态。皮质的平均％ID/g〉小脑，两者比值范围为1．13～2．08；60min时小脑、脑干和皮质的％ID／g与其最高峰相比分别下降了77％、80％和40％；皮质放射活性清除最慢。结论：[^11C]MP4A血中清除迅速。脑各区域摄取在短时达到稳态，提示[^11C]MP4A有较好的脂溶性，易透过血脑屏障；皮质摄取比小脑高但清除慢，表明[^11C]MP4A能更好反映皮质AChE活性，适合作为脑AChE活性的PET诊断显像剂。     

VMAT2 binding is elevated in dopa-responsive dystonia: visualizing empty vesicles by PET

Dopa-responsive dystonia (DRD) is a lifelong disorder in which dopamine deficiency is not associated with neuronal loss and therefore it is an ideal human model for investigating the compensatory changes that occur in response to this biochemical abnormality. Using positron emission tomography (PET), we examined the (+/-)-alpha-[(11)C]dihydrotetrabenazine ([(11)C]DTBZ) binding potential of untreated DRD patients and normal controls. Two other PET markers of presynaptic nigrostriatal function, d-threo-[(11)C]methylphenidate ([(11)C]MP) and 6-[(18)F]fluoro-L-dopa ([(18)F]-dopa), and [(11)C]raclopride were also used in the study. We found increased [(11)C]DTBZ binding potential in the striatum of DRD patients. By contrast, no significant changes were detected in either [(11)C]MP binding potential or [(18)F]-dopa uptake rate constant. In addition, we found evidence for increased dopamine turnover in one DRD patient by examining changes in [(11)C]raclopride binding potential in relation to levodopa treatment. We propose that the increase in [(11)C]DTBZ binding likely reflects the dramatic decrease in the intravesicular concentration of dopamine that occurs in DRD; upregulation of vesicular monoamine transporter type 2 (VMAT2) expression may also contribute. Our findings suggest that the striatal expression of VMAT2 (as estimated by [(11)C]DTBZ binding) is not coregulated with dopamine synthesis. This is in keeping with a role for VMAT2 in other cellular processes (i.e., sequestration and release from the cell of potential toxic products), in addition to its importance for the quantal release of monoamines.     

Occupancy of dopamine D2/3 receptors in rat brain by endogenous dopamine measured with the agonist positron emission tomography radioligand [11C]MNPA

Estimates of dopamine D(2/3) receptor occupancy by endogenous dopamine using positron emission tomography (PET) in animals have varied almost threefold. This variability may have been caused by incomplete depletion of dopamine or by the use of antagonist radioligands, which appear less sensitive than agonist radioligands to changes in endogenous dopamine. PET scans were performed in rats with the agonist PET radioligand [(11)C]MNPA ([O-methyl-(11)C]2-methoxy-N-propylnorapomorphine). [(11)C]MNPA was injected as a bolus plus constant infusion to achieve steady-state concentration in the body and equilibrium receptor binding in the brain. Radioligand binding was compared at baseline and after treatment with reserpine plus alpha-methyl-para-tyrosine, which cause approximately 95% depletion of endogenous dopamine. Depletion of dopamine increased radioligand binding in striatum but had little effect in cerebellum. Striatal [(11)C]MNPA binding potential was 0.93 +/- 0.12 at baseline and increased to 1.99 +/- 0.25 after dopamine depletion. Occupancy of D(2/3) receptors by endogenous dopamine at baseline was calculated to be approximately 53%. Striatal binding was displaceable with raclopride, but not with BP 897 (a selective D(3) compound), thus confirming the D(2) receptor specificity of [(11)C]MNPA binding. Radioactivity extracted from rat brain contained only 8-10% radiometabolites and was insignificantly altered by administration of reserpine plus alpha-methyl-para-tyrosine. Hence, dopamine depletion did not increase the PET measurements via an effect on radiotracer metabolism. Our in vivo estimate of dopamine's occupancy of D(2/3) receptors at baseline is higher than that previously reported using antagonist radioligands and PET, but is similar to that reported using agonist radioligands and ex vivo measurements.     

In vivo vulnerability to competition by endogenous dopamine: comparison of the D2 receptor agonist radiotracer (-)-N-[11C]propyl-norapomorphine ([11C]NPA) with the D2 receptor antagonist radiotracer [11C]-raclopride

(-)-N-Propyl-norapomorphine (NPA) is a full dopamine (DA) D2 receptor agonist and [11C]NPA is a suitable radiotracer to image D2 receptors configured in a state of high affinity for agonists with positron emission tomography (PET). In this study the vulnerability of the in vivo binding of [11C]NPA to acute fluctuation in synaptic DA was assessed with PET in baboons and compared to that of the reference D2 receptor antagonist radiotracer [11C]raclopride. Three male baboons were studied with [11C]raclopride and [11C]NPA under baseline conditions and following administration of the potent DA releaser amphetamine (0.3, 0.5, and 1.0 mg kg(-1) i.v.). Kinetic modeling with an arterial input function was used to derive the striatal specific-to-nonspecific equilibrium partition coefficient (V3"). [11C]Raclopride V3" was reduced by 24 +/- 10%, 32 +/- 6%, and 44 +/- 9% following amphetamine doses of 0.3, 0.5, and 1.0 mg kg(-1), respectively. [11C]NPA V3" was reduced by 32 +/- 2%, 45 +/- 3%, and 53 +/- 9% following amphetamine doses of 0.3, 0.5, and 1.0 mg kg(-1), respectively. Thus, endogenous DA was more effective at competing with [11C]NPA binding compared to [11C]raclopride binding, a finding consistent with the pharmacology of these tracers (agonist vs. antagonist). These results also suggest that 71% of D2 receptors are configured in a state of high affinity for agonists in vivo. In conclusion, [11C]NPA might provide a superior radiotracer to probe presynaptic DA function with PET in health and disease.     

Behaviour of [11C]R(-)- and [11C]S(+)-rolipram in vitro and in vivo, and their use as PET radiotracers for the quantificative assay of PDE4

Cyclic AMP (cAMP) is a continually produced nucleotide which is inactivated by hydrolysis to 5'AMP via phosphodiesterase 4 (PDE4) enzymes. Rolipram is a selective PDE4 inhibitor which exists in two enantiomeric forms, R(-) and S(+). Both of these enantiomers have previously been labelled with carbon-11 and used as positron emission tomography (PET) ligands for measuring PDE4 expression and function, and indirectly to explore the function of the cAMP second messenger, in vivo, using PET. The aim of these studies was to relate the in vitro affinities of the two rolipram enantiomers using standard pharmacological assays with the in vivo behaviour of the two enantiomers using PET. In vitro competition assays were performed using rat cortical membranes and [(3)H]R(-)- and [(3)H]S(+)-rolipram with increasing concentrations of either unlabelled R(-)- or S(+)-rolipram. In vivo, a series of PET studies were performed in the porcine brain using [(11)C]R(-)-rolipram with co-administration of increasing doses of either unlabelled R(-)- or S(+)-rolipram. Additional in vivo PET studies were performed using [(11)C]S(+)-rolipram with saturating doses of rolipram. In all studies, R(-)-rolipram exhibited a higher affinity for the PDE4 enzyme than S(+)-rolipram. The calculated affinity ratios were 7.97 from the in vitro studies; 12.5 from the in vivo studies using [(11)C]R(-)-rolipram; and 14.7 from the in vivo studies using [(11)C]S(+)-rolipram. To conclude, the in vitro affinities of R(-)- and S(+)-rolipram predict their apparent in vivo behaviour in the porcine brain, as measured by PET.     

Facilitation of dopaminergic neural transmission does not affect [(11)C]SCH23390 binding to the striatal D(1) dopamine receptors, but the facilitation enhances phosphodiesterase type-IV activity through D(1) receptors: PET studies in the conscious monkey brain.

The present study evaluated the effects of methamphetamine and scopolamine on the striatal dopamine D(1) receptor binding, measured by [(11)C]SCH23390, and D(1) receptor-coupled cAMP messenger system, determined as phosphodiesterase type-IV (PDE-IV) activity, were evaluated in the brains of conscious monkeys using positron emission tomography (PET) with microdialysis. When methamphetamine (0.1, 0.3, and 1 mg/kg) or scopolamine (0.01, 0.03, and 0.1 mg/kg) was systemically administered, [(11)C]SCH23390 binding to D(1) receptors was not affected. With administration of methamphetamine, the striatal PDE-IV activity, as measured with R-[(11)C]rolipram (active form) and S-[(11)C]rolipram (inactive form), was dose-dependently facilitated with enhanced dopamine level in the striatal ECF. Administration of scopolamine also induced facilitated PDE-IV activity without any apparent changes in the ECF dopamine. These facilitations of PDE-IV activity were abolished by preadministration of SCH23390, but not by raclopride. These results demonstrate that, as evaluated by PDE-IV activity, the inhibition of muscarinic cholinergic receptors actually facilitated dopamine neuronal signal transduction through D(1) receptors, as observed previously on D(2) receptors with no apparent increase in the striatal ECF dopamine level, but the enhanced dopamine transmission could not detected by [(11)C]SCH23390.     

Effects of aging on serotonin transporter availability and its response to fluvoxamine in the living brain: PET study with [(11)C](+)McN5652 and [(11)C](-)McN5652 in conscious monkeys

Age-related changes in the serotonin transporter (SERT) in the living brains of conscious young (5.9 +/- 1.8 years old) and aged (19.0 +/- 3.3 years old) monkeys (Macaca mulatta) were evaluated in combination with [(11)C](+)McN5652 and its inactive enantiomer [(11)C](-)McN5652 by high-resolution positron emission tomography (PET). For the quantitative analysis of SERT binding in vivo, two serial PET scans with [(11)C](+)McN5652 and [(11)C](-)McN5652 were performed in the same animals in a day and the differences in radioactivities of [(11)C](+)McN5652 vs. [(11)C](-)McN5652 measured from 41-91 min postinjection were calculated as an estimate of specific ligand binding. Higher specific binding of SERT was observed in the thalamus and striatum, regions known to contain high densities of SERT by in vitro assay, with intermediate levels in the pons, hippocampus, cingulate gyrus, and cortical regions and lower levels in the cerebellum in both young and aged monkeys. Almost all regions assayed except the cerebellum showed significant age-related decreases in the specific binding of SERT, which showed reverse correlation with cortisol level in plasma. When the SERT blocker fluvoxamine (1 mg/kg) was administered intravenously 30 min after tracer injection, specific binding of SERT was displaced in both age groups. However, the degree of displacement was more marked in young than in aged monkeys. Cortisol level in plasma was significantly higher in aged than in young animals. These observations demonstrate the usefulness of the combined use of [(11)C](+)McN5652 and [(11)C](-)McN5652 as an indicator for the age-related changes in cortical SERT measured noninvasively by PET. In addition, these observations suggest that the age-related impairment of SERT sensitivity for fluvoxamine might be related to the reduced efficacy of antidepressant therapy in elderly patients with depression.     

Visualization of alpha5 subunit of GABAA/benzodiazepine receptor by 11C Ro15-4513 using positron emission tomography

Although [(11)C]Ro15-4513 and [(11)C]flumazenil both bind to the central benzodiazepine (BZ) receptors, the distributions of the two ligands are not identical in vivo. Moreover, the in vivo pharmacological properties of [(11)C]Ro15-4513 have not been thoroughly examined. In the present study, we examined the pharmacological profile of [(11)C]Ro15-4513 binding in the monkey brain using positron emission tomography (PET). [(11)C]Ro15-4513 showed relatively high accumulation in the anterior cingulate cortex, hippocampus, and insular cortex, with the lowest uptake being observed in the pons. Accumulation in the cerebral cortex was significantly diminished by the BZ antagonist flumazenil (0.1 mg/kg, i.v.), but not that in the pons. Using the pons as a reference region, the specific binding of [(11)C]Ro15-4513 in most of the cerebral cortex including the limbic regions clearly revealed two different affinity sites. On the other hand, specific binding in the occipital cortex and cerebellum showed only a low affinity site. Zolpidem with affinity for alpha1, alpha2, and alpha3 subunits of GABA(A)/BZ receptor fully inhibited [(11)C]Ro15-4513 binding in the occipital cortex and cerebellum, while only about 23% of the binding was blocked in the anterior cingulate cortex. Diazepam with affinity for alpha1, alpha2, alpha3, and alpha5 subunits inhibited the binding in all brain regions. Since Ro15-4513 has relatively high affinity for the alpha5 subunit in vitro, these in vivo bindings of [(11)C]Ro15-4513 can be interpreted as the relatively high accumulation in the fronto-temporal limbic regions representing binding to the GABA(A)/BZ receptor alpha5 subunit.