Amyloid load and translocator protein 18 kDa in APPswePS1-dE9 mice: a longitudinal study.

We studied concomitantly the level of neuroinflammation and ß-amyloid (Aß) load in the APPswePS1dE9 transgenic mouse model of Alzheimer's disease using positron emission tomography. The translocator protein 18 kDa (TSPO) tracer [(18)F]DPA-714 was used to measure neuroinflammation and [(18)F]AV-45 for Aß load in mice at 6, 9, 12, 15, and 19 months of age. At 19 months, we also analyzed the neuroinflammatory and neuroanatomic status of mice brains. The main affected brain areas were the cortex and hippocampus, with a concomitant progression of neuroinflammation with increased amyloid burden. At 19 months, no increase in TSPO binding was observed in the cerebellum; immunostaining revealed W0-2-positive plaques, indicating that the amyloid deposits seemed not stimulate inflammation. This finding was in agreement with the observed level of microglia and astrocytes staining. Our findings provide a better understanding of the relationships between neuroinflammation and plaque accumulation in the course of the disease in this mouse model. The monitoring of both processes should be of value to validate potential therapeutic approaches.

Translocator protein (18 kDa) mapping with [125I]-CLINDE in the quinolinic acid rat model of excitotoxicity: a longitudinal comparison with microglial activation, astrogliosis, and neuronal death.

Excitotoxicity leads to an inflammatory reaction involving an overexpression of: translocator protein 18 kDa (TSPO) in cerebral microglia and astrocytes. Therefore, we performed ex vivo explorations with [125]-CLINDE, a TSPO-specific radioligand, to follow the time course of TSPO expression, in parallel with lesion progression, over 90 days after induction of cerebral excitotoxicity in rats intrastriatally injected with quinolinic acid. Biodistribution data showed a significant increase in CLINDE uptake on the injured side from 1 days postlesion (dpl); the maximal striatal binding values evidenced a plateau between 7 and 30 dpl. [125I]-CLINDE binding was displaced from the lesion by PK11195, suggesting TSPO specificity. These results were confirmed by ex vivo autoradiography. Combined immunohistochemical studies showed a marked increase in microglial expression in the lesion, peaking at 14 dpl, and astrocytic reactivity enhanced at 7 and 14 dpl, whereas a prominent neuronal cell loss was observed. At 90 dpl, CLINDE binding and immunoreactivity targeting activated microglia, astrogliosis, and neuronal cell density returned to a basal level. These results show that both neuroinflammation and neuronal loss profiles occurred concomitantly and appeared to be transitory processes. These findings provide the possibility of a therapeutic temporal window to compare the differential effects of antiinflammatory treatments in slowing down neurodegeneration in this rodent model, with potential applications to humans.

In vivo PET quantification of the dopamine transporter in rat brain with [¹8F]LBT-999.

INTRODUCTION: We examined whether [(18)F]LBT-999 ((E)-N-(4-fluorobut-2-enyl)2ß-carbomethoxy-3ß-(4'-tolyl)nortropane) is an efficient positron emission tomography (PET) tracer for the quantification of the dopamine transporter (DAT) in the healthy rat brain. METHODS: PET studies were performed using several experimental designs, i.e. test-retest, co-injection with different doses of unlabelled LBT, displacement with GBR12909 and pre-injection of amphetamine. RESULTS: The uptake of [(18)F]LBT-999 confirmed its specific binding to the DAT. The non-displaceable uptake (BP(ND)) in the striatum, between 5.37 and 4.39, was highly reproducible and reliable, and was decreased by 90% by acute injection of GBR12909. In the substantia nigra/ventral tegmental area (SN/VTA), the variability was higher and the reliability was lower. Pre-injection of amphetamine induced decrease of [(18)F]LBT-999 BP(ND) of 50% in the striatum. CONCLUSIONS: [(18)F]LBT-999 allows the quantification of the DAT in living rat brain with high reproducibility, sensitivity and specificity. It could be used to quantify the DAT in rodent models, thereby allowing to study neurodegenerative and neuropsychiatric diseases.

Evaluation of prion deposits and microglial activation in scrapie-infected mice using molecular imaging probes.

PURPOSE: A characteristic of prion diseases which affect both animals and humans is the aggregation of PrP amyloid fibrils in the brain, associated with a chronic inflammatory response dominated by microglial activation. In this study, we hypothesised that specific ligands of the 18-kDa translocator protein (TSPO) would be effective in the evaluation of microglial activation related to PrP(sc) deposits in prion disease. PROCEDURES: Chronological studies using in vitro autoradiography were carried out with [(3)H]-PK11195 and [(125)I]-IMPY on frozen cerebral sections from scrapie-infected mice and controls. Accumulation of prion deposits was confirmed by histoblot staining with prion protein-specific monoclonal antibody. Ex vivo autoradiographic studies were carried out with [(125)I]-CLINDE and [(125)I]-IMPY at the terminal stage of infection. RESULTS: Chronological studies using in vitro autoradiography showed that PrP(sc) deposits were co-localised with activated microglia as early as 60 days post-inoculation. Progressive levels of PrP(sc) and TSPO staining were successively observed in the hippocampus, cortex and left thalamus of infected mouse brain sections in the course of the disease and were correlated with the signals obtained by histoblot staining. Significant TSPO labelling was also observed ex vivo in the cortex, hippocampus and thalamus of scrapie-infected mice. In parallel, [(125)I]-IMPY showed labelling in the same cerebral regions but with high background staining. CONCLUSIONS: These findings indicate the ability of [(125)I]-IMPY and [(125)I]-CLINDE to evaluate prion deposits and microglial activation in vitro and ex vivo in scrapie-infected mice at different stages of the disease.

Behavior and serotonergic disorders in rats exposed prenatally to valproate: a model for autism.

In order to explore whether some aspects of the autistic phenotype could be related to impairment of the serotonergic system, we chose an animal model which mimics a potential cause of autism, i.e. rats exposed to valproate (VPA) on the 9th embryonic day (E9). Previous studies have suggested that VPA exposure in rats at E9 caused a dramatic shift in the distribution of serotonergic neurons on postnatal day 50 (PND50). Behavioral studies have also been performed but on rats that were exposed to VPA later (E12.5). Our aim was to test whether VPA exposure at E9 induces comparable behavioral impairments than at E12.5 and causes serotonergic impairments which could be related to behavioral modifications. The results showed significant behavioral impairments such as a lower tendency to initiate social interactions and hyperlocomotor activity in juvenile male rats. The serotonin levels of these animals at PND50 were decreased (-46%) in the hippocampus, a structure involved in social behavior. This study suggests that VPA could have a direct or indirect action on the serotonergic system as early as the progenitor cell stage. Early embryonic exposure to VPA in rats provides a good model for several specific aspects of autism and should help to continue to explore pathophysiological hypotheses.

Docking study, synthesis, and in vitro evaluation of fluoro-MADAM derivatives as SERT ligands for PET imaging.

In order to predict affinity of new diphenylsulfides for the serotonin transporter (SERT), a molecular modeling model was used to compare potential binding affinity of new compounds with known potent ligands. The aim of this study is to identify a suitable PET radioligand for imaging the SERT, new derivatives, and their precursors for a C-11 or F-18 radiolabeling, were synthesized. Two fluorinated derivatives displayed good in vitro affinity for the SERT (K(i)=14.3+/-1 and 10.1+/-2.7 nM) and good selectivity toward the other monoamine transporters as predicted by the docking study.

Evaluation of CLINDE as potent translocator protein (18 kDa) SPECT radiotracer reflecting the degree of neuroinflammation in a rat model of microglial activation.

BACKGROUND: The translocator protein (TSPO; 18 kDa), the new name of the peripheral-type benzodiazepine receptor, is localised in mitochondria of glial cells and expressed in very low concentrations in normal brain. Their expression rises after microglial activation following brain injury. Accordingly, TSPO are potential targets to evaluate neuroinflammatory changes in a variety of CNS disorders. PURPOSE: To date, only a few effective tools are available to explore TSPO by SPECT. We characterised here 6-chloro-2-(4'iodophenyl)-3-(N,N-diethyl)-imidazo[1,2-a]pyridine-3-acetamide or CLINDE in a rat model with different stages of excitotoxic lesion. METHODS: Excitotoxicity was induced in male Wistar rats by unilateral intrastriatal injection of different amounts of quinolinic acid (75, 150 or 300 nmol). Six days later, two groups of rats (n = 5-6/group) were i.v. injected with [(125)I]-CLINDE (0.4 MBq); one group being pre-injected with PK11195 (5 mg/kg). Brains were removed 30 min after tracer injection and the radioactivity of cerebral areas measured. Complementary ex vivo autoradiography, in vitro autoradiography ([(3)H]-PK11195) and immunohistochemical studies (OX-42) were performed on brain sections. RESULTS: In the control group, [(125)I]-CLINDE binding was significantly higher (p < 0.001) in lesioned than that in intact side. This binding disappeared in rats pre-treated with PK11195 (p < 0.001), showing specific binding of CLINDE to TSPO. Ex vivo and in vitro autoradiographic studies and immunohistochemistry were consistent with this, revealing a spatial correspondence between radioactivity signal and activated microglia. Regression analysis yielded a positive relation between the ligand binding and the degree of neuroinflammation. CONCLUSION: These results demonstrate that CLINDE is suitable for TSPO in vivo SPECT imaging to explore their involvement in neurodegenerative disorders associated with microglial activation.

DPA-714, a new translocator protein-specific ligand: synthesis, radiofluorination, and pharmacologic characterization.

UNLABELLED: The translocator protein (18 kDa) (TSPO), formerly known as the peripheral benzodiazepine receptor, is dramatically upregulated under pathologic conditions. Activated microglia are the main cell type expressing the TSPO at sites of central nervous system pathology. Radioligands for the TSPO can therefore measure active disease in the brain. This article details the synthesis, radiofluorination, and pharmacologic evaluation of a new TSPO-specific pyrazolopyrimidine, DPA-714. METHODS: The affinity of DPA-714 for the TSPO was measured in rat kidney membranes with (3)H-PK11195. The in vitro functional activity of DPA-714 was measured in a steroidogenic assay in which the ability of DPA-714 to increase pregnenolone synthesis was measured with rat C6 glioma cells. The radiofluorination of DPA-714 was achieved by nucleophilic (18)F-fluoride displacement of the tosylate precursor. (18)F-DPA-714 was assessed in rats harboring unilateral quinolinic acid (QA) lesions. In addition, pretreatment experiments were performed with PK11195 (5 mg/kg), DPA-714 (1 mg/kg), and DPA-713 (1 mg/kg). The in vivo binding and biodistribution of (18)F-DPA-714 were determined in a baboon with PET. Experiments involving presaturation with PK11195 (1.5 mg/kg) and displacement with DPA-714 (1 mg/kg) were conducted to evaluate the specificity of radioligand binding. RESULTS: In vitro binding studies revealed that DPA-714 displayed a high affinity for the TSPO (dissociation constant, 7.0 nM). DPA-714 stimulated pregnenolone synthesis at levels 80% above the baseline. (18)F-DPA-714 was prepared at a 16% radiochemical yield and a specific activity of 270 GBq/mumol. In rats harboring unilateral QA lesions, an 8-fold-higher level of uptake of (18)F-DPA-714 was observed in the ipsilateral striatum than in the contralateral striatum. Uptake in the ipsilateral striatum was shown to be selective because it was inhibited to the level in the contralateral striatum in the presence of PK11195, nonlabeled DPA-714, or DPA-713. PET studies demonstrated rapid penetration and good retention of (18)F-DPA-714 in the baboon brain. Pretreatment with PK11195 effectively inhibited the uptake of (18)F-DPA-714 in the whole brain, indicating its selective binding to the TSPO. The injection of nonlabeled DPA-714 20 min after the injection of (18)F-DPA-714 resulted in radioligand washout, demonstrating the reversibility of (18)F-DPA-714 binding. CONCLUSION: (18)F-DPA-714 is a specific radioligand for the TSPO, displaying promising in vivo properties and thus warranting further investigation.

Partial recovery of dopaminergic pathway after graft of adult mesenchymal stem cells in a rat model of Parkinson's disease.

Cellular therapy with adult stem cells appears as an opportunity for treatment of Parkinson's disease. To validate this approach, we studied the effects of transplantation of rat adult bone-marrow mesenchymal stem cells in a rat model of Parkinson's disease. Animals were unilaterally lesioned in the striatum with 6-hydroxydopamine. Two weeks later, group I did not undergo grafting, group II underwent sham grafting, group III was intra-striatal grafted with cells cultured in an enriched medium and group IV was intra-striatal grafted with cells cultured in a standard medium. Rotational amphetamine-induced behavior was measured weekly until animals were killed 6 weeks later. One week after graft, the number of rotations/min was stably decreased by 50% in groups III and IV as compared with groups I and II. At 8 weeks post-lesion, the density of dopaminergic markers in the nerve terminals and cell bodies, i.e. immunoreactive tyrosine hydroxylase, membrane dopamine transporter and vesicular monoamine transporter-2 was significantly higher in group III as compared with group I. Moreover, using microdialysis studies, we observed that while the rate of pharmacologically induced release of dopamine was significantly reduced in lesioned versus intact striatum in no grafted rats, it was similar in both sides in animals transplanted with mesemchymal stem cells. These data demonstrate that graft of adult mesemchymal stem cells reduces behavioral effects induced by 6-hydroxydopamine lesion and partially restores the dopaminergic markers and vesicular striatal pool of dopamine. This cellular approach might be a restorative therapy in Parkinson's disease.

Improved synthesis and metabolic stability analysis of the dopamine transporter ligand [(18)F]FECT.

INTRODUCTION: [2'-[(18)F]Fluoroethyl (lR-2-exo-3-exe)-8-methyl-3-(4-chlorophenyl)-8-azabicyclo[3.2.1]-octane-2-carboxylate] ([(18)F]FECT) is a positron emission tomography (PET) tracer for imaging the dopamine transporter (DAT) in vivo. We report an improved radiosynthesis procedure and affinity data and have analyzed both brain tissue and plasma samples for the presence of radiometabolites as a function of time post intravenous injection of [(18)F]FECT to rats. METHODS: The radiosynthesis of [(18)F]FECT was carried out using [(18)F]fluoroethyltriflate ([(18)F]FEtOTf) as a labeling agent. The affinity of FECT for DAT was determined in vitro by binding experiments on rat striatal membranes. Three rats were injected with [(18)F]FECT and blood samples were collected at 1 or 3 h post injection (p.i.). Plasma was separated and analyzed using reversed-phase high-performance liquid chromatography (RP-HPLC). Similarly, cerebrum and cerebellum were isolated after sacrifice of the animals at 3 h p.i. of the tracer and homogenized. HPLC analysis was performed on extracts of both samples to examine the presence of metabolites. RESULTS: The radiochemical yield for [(18)F]FECT was 85% relative to the starting activity of [(18)F]FEtOTf. The inhibitory constant (K(i)) of FECT for DAT was found to be 6 nM. The fraction of radioactivity corresponding to intact [(18)F]FECT was 93% in plasma at both 1 and 3 h p.i. and 96% in cerebrum as well as cerebellum samples at 3 h p.i. CONCLUSIONS: FECT has a high affinity for the dopamine transporter. [(18)F]FECT was found to be stable in vivo and the amount of radiolabeled metabolites in plasma and brain at 3 h p.i. is negligible. Hence, [(18)F]FECT can be used for the in vivo quantification of DAT using PET.

WITHDRAWN: Prenatal exposure to MDMA induces sex-dependent and region-specific modifications in brain dopamine systems at adult age in the rat.

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(E)-[125I]-5-AOIBV: a SPECT radioligand for the vesicular acetylcholine transporter.

The premise that, over the course of Alzheimer's disease (AD), changes in the levels of the vesicular acetylcholine transporter (VAChT) occur in parallel with changes to other cholinergic marker proteins provides the basis for the applicability of benzovesamicol derivatives as radioligands for AD studies by single photon emission computed tomography or positron emission tomography. We report the synthesis of enantiopure benzovesamicol derivatives: (R,R) or (S,S)-(E)-2-hydroxy-5-(3-iodoprop-2-en-1-oxy)-3-(4-phenylpiperidino)tetralin [(R,R)-AOIBV: Kd=0.45 nM or (S,S)-5-AOIBV: Kd=4.3 nM] and their corresponding tributyltin precursors for radioiodination. (R,R or S,S)-5-AOIBV was labeled with iodine-125 from their corresponding n-tributyltin precursors. Both compounds were obtained with radiochemical and optical purity greater than 97% and in radiochemical yields ranging 34-36%. To determine if these compounds could provide an advantage when compared to [125I]-iodo benzovesamicol (IBVM), IBVM was also labeled and used as the reference compound in all ex vivo experiments. Ex vivo biodistribution experiments in rats revealed that [125I]-(R,R)-5-AOIBV displayed the most suitable pharmacological profile as the radioactivity distribution corresponded well with the known VAChT brain density. Moreover, pre-injection of vesamicol prevented the uptake of [125I]-(R,R)-5-AOIBV in striatum, cortex and hippocampus, demonstrating selectivity for the VAChT. However, even if time activity curves of [125I]-(R,R)-5-AOIBV confirmed that this compound could be used to visualize the VAChT in vivo, at each point of the kinetic study, [125I]-(R,R)-5-AOIBV showed a lower specific binding compared to [125I]-IBVM. These results made [125I]-( R,R)-5-AOIBV inferior to [125I]-IBVM for the VAChT exploration in vivo.

Ex vivo and in vivo evaluation of (2R,3R)-5-[(18)F]-fluoroethoxy- and fluoropropoxy-benzovesamicol, as PET radioligands for the vesicular acetylcholine transporter.

Molecular imaging of the vesicular acetylcholine transporter (VAChT) using positron emission tomography (PET) may provide insights into early diagnosis and better understanding of Alzheimer's disease. We further characterized the VAChT ligand (2R,3R)-5-FEOBV (1) and developed new fluoropropoxy analogues. Ex vivo studies of the new nonradiolabeled analogues (2R,3R)-5-FPOBV (2) (k(D) = 0.7 nM) and (2S,3S)-5-FPOBV (3) (k(D) = 8.8 nM) were performed in rat brain and showed an enantioselective inhibition of (-)-5-[(125)I]-IBVM uptake in striatum, cortex, and hippocampus (e.g., 74% for 2 and only 54% for 3 in the cortex). Radiochemical procedures were developed to produce [(18)F]1 and [(18)F]2 as potential imaging agent for the VAChT. The radiochemistry was carried out in a one step procedure, with radiolabeling yields of 17 and 2.6% (range: 1-5.4), respectively, nondecay corrected with good specific activity: 124-338 GBq/micromol. The radiochemical purity was greater than 98%. The biological (ex vivo and in vivo) properties of these radioligands were evaluated in rats and showed a low (less then 0.1% of the injected dose) and homogeneous brain uptake. The in vivo PET study of [(18)F]2 performed in baboon also revealed rapid defluorination as the main problem. Therefore [(18)F]1 and [(18)F]2 appear to be unsuitable for in vivo imaging of the VAChT using PET.

Pharmacological characterization of (E)-N-(4-fluorobut-2-enyl)-2beta-carbomethoxy-3beta-(4'-tolyl)nortropane (LBT-999) as a highly promising fluorinated ligand for the dopamine transporter.

In the aim to develop an efficient fluorinated probe for positron emission tomography (PET) exploration of the dopamine transporter (DAT), we studied several in vitro and in vivo characteristics of the phenyltropane derivative (E)-N-(4-fluorobut-2-enyl)-2beta-carbomethoxy-3beta-(4'-tolyl)nortropane (LBT-999). In vitro on rat striatal membrane, [(3)H]LBT-999 bound to a single site with a K(d) of 9 nM, B(max) of 17 pmol/mg protein, and a very high selectivity for the DAT [IC(50) for 1-{2-[bis-(4-fluorophenyl)-methoxy]ethyl}-4-(3-phenylpropyl)piperazine (GBR 12909) and (E)-N-(3-iodoprop-2-enyl)-2beta-carbomethoxy-3beta-(4'-methylphenyl)nortropane (PE2I): 2.4 and 18 nM, respectively; IC(50) for paroxetine, citalopram, N,N-dimethyl-2-(2-amino-4-methylphenyl thio)benzylamine, nisoxetine, and desipramine >1 muM]. In vitro on post-mortem human brain sections, LBT-999 bound with high intensity to the caudate-putamen, weakly to the thalamus, and not in the neocortex and cerebellum. This binding was totally abolished in the presence of PE2I. Ex vivo cerebral biodistribution of [(11)C]LBT-999 in rats showed striatum/cerebellum radioactivity ratios of 18 and 25 at 30 and 60 min postinjection, respectively. This accumulation was strongly prevented by preinjection of GBR 12909, whereas paroxetine and nisoxetine had no effect. An in vivo kinetic PET study in three baboons showed a fast and very high uptake in the striatum, with a plateau at 30 min postinjection and a maximal putamen/cerebellum ratio of 30. Taken together, these findings demonstrate that LBT-999 is a highly promising agent for in vivo exploration of the DAT. This probe is currently labeled with (18)F for further characterizations.

ADAM is an effective tool for in vivo study of serotonergic function: validation in rat models.

ADAM, 2-((2-((dimethylamino)methyl)phenyl)thio)-5-iodophenylamine, is a recently described SPECT tracer for exploration of the serotonin transporter. We evaluated its potential to detect abnormalities in serotonergic function in the rat using 1) a model of serotonergic neuron lesion induced with 5,7-dihydroxytryptamine (5,7-DHT), and 2) experimental induction of acute decrease in endogenous brain serotonin levels. Cerebral biodistribution studies of [125I]ADAM were performed in normal conditions, in 5,7-DHT-lesioned rats, and after acute serotonin depletion obtained with p-chlorophenylalanine (pCPA). Around 50% reduction in accumulation of ADAM was observed in the hypothalamus and hippocampus 3 weeks after lesion of serotonergic neurons, whereas a more modest decrease of 15-30% occurred in the thalamus, frontal cortex, and striatum. This demonstrated the ability of the tracer to detect serotonergic neuron loss in vivo. After inducing acute 5-HT depletion with pCPA, we observed an increase in in vivo [125I]ADAM binding in all brain areas studied. The higher in vivo binding of [125I]ADAM in pCPA-treated rats than in controls was mainly due to an increase in specific binding to the SERT, as demonstrated by greatly reduced binding in the presence of a saturating dose of paroxetine. This may indicate in vivo competition between ADAM and 5-HT for binding to the SERT. The present findings thus demonstrate that ADAM is a specific SERT radioligand which can be used for in vivo study of central serotonin systems, and supports its use as a tracer for SPECT studies in human disorders involving dysfunction of serotonergic neurotransmission.

Pharmacological characterization of N,N-dimethyl-2-(2-amino-4-methylphenyl thio)benzylamine as a ligand of the serotonin transporter with high affinity and selectivity.

Serotonin transporter has a key-role in regulation of serotoninergic function, and is involved in numerous neurodegenerative and psychiatric disorders. To obtain an efficient radioactive ligand allowing the study of this transporter in vitro and in vivo, we synthesized a new diphenyl sulfide derivative, N,N-dimethyl-2-(2-amino-4-methylphenylthio)benzylamine or MADAM. We present here extensive pharmacological characterization of this compound. [3H]MADAM bound to serotonin transporters with a very high affinity in vitro on rat cortical membranes, at least 2 times better than the most commonly used radioactive probes (Kd, 60 pM; Bmax, 543 fmol/mg of protein). Competition studies showed few inhibitory effect of nisoxetine (Ki = 270 nM), no inhibitory effect of desipramine or 1-[2-(diphenylmethoxy) ethyl]-4-(3-phenylpropyl)piperazine (GBR 12935) (Ki >1000 nM), and strong effect of paroxetine (Ki = 0.32 nM) and citalopram (Ki = 1.57 nM). Therefore, MADAM has around 1000-fold better selectivity for the serotonin transporter than for other transporters. Autoradiographic studies both on rat and postmortem human brain slices demonstrated that the distribution of [3H]MADAM parallels the localization of serotonin transporters and is prevented by known inhibitors of them. The high affinity and selectivity of [3H]MADAM for the serotonin transporter show that it is very valuable for studies using in vitro approaches. The high selectivity and low nonspecific binding of [3H]MADAM on the postmortem human brain, together with preliminary in vivo results with [11C]MADAM, is a new argument for future use of this ligand in in vivo studies of the distribution, pharmacology, and pathophysiology of the serotonin transporter in the human brain with positron emission tomography.

Synthesis, radiolabeling and preliminary biological evaluation of radiolabeled 5-methyl-6-nitroquipazine, a potential radioligand for the serotonin transporter.

5-Methyl-6-nitroquipazine, a novel analogue of the potent and selective serotonin transporter inhibitor 6-nitroquipazine was synthesized and radiolabeled with tritium and the positron emitter carbon-11. [3H]5-methyl-6-nitroquipazine was found to have a K(d)=51+/-7 pM. The high affinity and the facile labeling of [11C]5-methyl-6-nitroquipazine makes it a promising radioligand for visualization of the serotonin transporter with positron emission tomography.

Relevance of different striatal markers in assessment of the MPP+-induced dopaminergic nigrostriatal injury in rat.

Many striatal dopaminergic markers are available for estimating the degree of the nigrostriatal lesion by MPTP/MPP+, but the changes of these markers are not perfectly matched. In this study we investigated different striatal markers and determined which ones closely reflected the nigrostriatal alteration. The in vivo binding of (E)-N-(3-iodoprop-2-enyl)-2-beta-carbomethoxy-3beta-(4'-methylphenyl)nortropane (PE2I), a selective and potent inhibitor of the neuronal dopamine transporter (DAT) was considered as the reference index of injury of striatal dopaminergic nerve-endings. Rats received a 10-microg MPP+ injection in the right substantia nigra and were killed at 7 days after lesion. The results were as follows: (i) a decrease (66%) of the biodistribution of [125I]PE2I; (ii) a great reduction of the DAT expression measured by the binding of [125I]PE2I in striatal membranes (Bmax decreased by 54%) and in cerebral slices (88%); (iii) an 80% inhibition of the vesicular monoamine transporter expression revealed by the binding of [3H]dihydrotetrabenazine in cerebral slices; (iv) a robust decrease in the quantity of DA and its metabolites (about 50-60%); (v) a slight modification of the DAT activity with a decreased number of functional sites (Vmax decreased by 12%, p < 0.05) without change of the affinity in striatal synaptosomes. Among these markers the binding of [125I]PE2I in membrane homogenates and the content of DA, and its metabolites, in striatum could be the most relevant in vitro indexes of the degenerative state of the nigrostriatal pathway after MPP+ lesion.