Synthesis and biological characterization of novel 2-quinolinecarboxamide ligands of the peripheral benzodiazepine receptors bearing technetium-99m or rhenium

Potential receptor imaging agents based on Tc-99m for the in vivo visualization of the peripheral benzodiazepine receptor (PBR) have been designed on the basis of the information provided by the previously published structure-affinity relationship studies, which suggested the existence of tolerance to voluminous substituents in the receptor area interacting with 3-position of the quinoline nucleus of 2-quinolinecarboxamides 5. In the first step of the investigation, the stereoelectronic features of the above-indicated receptor area were also probed by means of 4-phenyl-3-[(1-piperazinyl)methyl]-2-quinolinecarboxamide derivatives bearing different substituents on the terminal piperazine nitrogen atom (compounds 6a-f). The structure-affinity relationship data confirmed the existence of a tolerance to bulky lipophilic substituents and stimulated the design of bifunctional ligands based on the 4-phenyl-3-[(1-piperazinyl)methyl]-2-quinolinecarboxamide moiety (compounds 6h,j,k,m). The submicromolar PBR affinity of rhenium complexes 6j,m suggests that the presence of their metal-ligand moieties with encaged rhenium is fairly compatible with the interaction with the PBR binding site. Thus, in order to obtain information on the in vivo behavior of these bifunctional ligands, (99m)Tc-labeled compounds 6h,k were synthesized and evaluated in preliminary biodistribution and single photon emission tomography (SPET) studies. The results suggest that both tracers do not present a clear preferential distribution in tissues rich in PBR, probably because of their molecular dimensions, which may hamper both the intracellular diffusion toward PBR and the interaction with the binding site.     

Synthesis and in vivo evaluation of a novel peripheral benzodiazepine receptor PET radioligand

The novel pyrazolopyrimidine ligand, N,N-diethyl-2-[2-(4-methoxyphenyl)-5,7-dimethyl-pyrazolo[1,5-a]pyrimidin-3-yl]-acetamide 1 (DPA-713), has been reported as a potent ligand for the peripheral benzodiazepine receptor (PBR) displaying an affinity of K(i)=4.7 nM. In this study, 1 was successfully synthesised and demethylated to form the phenolic derivative 6 as precursor for labelling with carbon-11 (t(1/2) = 20.4 min). [11C]1 was prepared by O-alkylation of 6 with [11C]methyl iodide. The radiochemical yield of [(11)C]1 was 9% (non-decay corrected) with a specific activity of 36 GBq/micromol at the end of synthesis. The average time of synthesis including formulation was 13.2 min with a radiochemical purity >98%. In vivo assessment of [11C]1 was performed in a healthy Papio hamadryas baboon using positron emission tomography (PET). Following iv administration of [11C]1, significant accumulation was observed in the baboon brain and peripheral organs. In the brain, the radioactivity peaked at 20 min and remained constant for the duration of the imaging experiment. Pre-treatment with the PBR-specific ligand, PK 11195 (5 mg/kg), effectively reduced the binding of [11C]1 at 60 min by 70% in the whole brain, whereas pre-treatment with the central benzodiazepine receptor ligand, flumazenil (1mg/kg), had no inhibitory effect on [11C]1 uptake. These results indicate that accumulation of [11C]1 in the baboon represents selective binding to the PBR. These exceptional in vivo binding properties suggest that [11C]1 may be useful for imaging the PBR in disease states. Furthermore, [11C]1 represents the first ligand of its pharmacological class to be labelled for PET studies and therefore has the potential to generate new information on the pathological role of the PBR in vivo.     

Synthesis and structure-activity relationship studies in peripheral benzodiazepine receptor ligands related to alpidem

The exploration of the structure-affinity relationships concerning a new class of peripheral benzodiazepine receptor (PBR) ligands related to alpidem has been pursued in order to evaluate the consistency of the structure-affinity relationships among different classes (and subclasses) of PBR ligands. The target amide derivatives were prepared following a previously published procedure based on the condensation of pyrrolo[3,4-b]quinoline derivatives 11a,b with glyoxylic acid mono-hydrate and the subsequent amidation of the acids obtained via mixed anhydride. On the other hand, the preparation of compound 9g lacking the pharmacophoric (delta1) carbonyl group involved: (a) the double sequential attack of the dimethylmethyleneammonium salt obtained from bis(dimethylamino)methane and acetyl chloride to pyrrolo[3,4-b]quinoline derivative 11b, (b) the quaternization of the obtained allylamine derivative 13 with methyl iodide, and (c) the palladium-catalyzed allylation of N-methyl-p-anisidine by quaternary allylammonium cation 14. The structure-affinity relationship trends observed in this subclass of tricyclic alpidem-related PBR ligands find correlations in other classes (or subclasses) of PBR ligands. This result supports the initial pharmacophoric hypothesis and suggests a common mode of interaction at the PBR binding site.     

Evaluation of three quinoline-carboxamide derivatives as potential radioligands for the in vivo pet imaging of neurodegeneration

The peripheral-type benzodiazepine receptors (PBRs) are only minimally expressed in normal brain parenchyma, where they are primarily localized in glial cells. Their basal expression rises in different neurodegenerative disorders, due to the presence of infiltrating inflammatory cells and activated microglia. [11C]PK11195, a selective PBR antagonist, has been used for the in vivo PET monitoring of neurodegeneration in clinical observations. We recently developed and labeled with carbon-11 three new carboxamide derivatives: [11C]VC193M, [11C]VC195 and [11C]VC198M. Aim of this study was to evaluate these ligands for the in vivo measuring of PBRs expression in neurodegenerations and compare their kinetic behavior with that of the reference tracer [11C]PK11195. Radioligands were evaluated in a preclinical model of Huntington's disease consisting in the monolateral striatal injection of quinolinic acid (QA). Activated microglia and astrocytic gliosis was present only within the affected striatum. A concomitant increase in radioactivity accumulation was observed for all the tracers examined (P<0.01). Among the new compounds, [11C]VC195 showed higher levels of lesioned/unlesioned striatum ratios (3.28+/-0.44), in comparison with [11C]VC193M and [11C]VC198M (2.69+/-0.53 and 1.52+/-0.36, respectively), but slightly inferior to that observed for [11C]PK11195 (3.76+/-1.41).In conclusion, the results of the study indicate that [11C]VC195 is a promising candidate for in vivo PET monitoring of neurodegenerative processes but its in vivo behavior overlap that of [11C]PK11195.     

Synthesis, fluorine-18 radiolabeling, and in vitro characterization of 1-iodophenyl-N-methyl-N-fluoroalkyl-3-isoquinoline carboxamide derivatives as potential PET radioligands for imaging peripheral benzodiazepine receptor

The isoquinoline carboxamide derivative 1-(2-chlorophenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinoline carboxamide (PK11195) has been shown to bind strongly and selectively to the peripheral benzodiazepine receptor (PBR) binding sites. A series of PK11195 analogues have been synthesized and biologically characterized. The affinities of the analogues for the PBR were determined using in vitro competitive binding assays with [(3)H]PK11195 in rat kidney mitochondrial membranes. The results showed that the 1-(2-iodophenyl)-N-methyl-N-(3-fluoropropyl)-3-isoquinoline carboxamide (9a) was the most potent compound (K(i)=0.26nM) of this series and is an excellent lead ligand for additional studies for labeling with fluorine-18 to determine whether it possesses the desired in vivo performance in non-human primates by PET imaging. Thus, radiolabeling of 9a with fluorine-18 was developed.     

Effect of noise exposure on rat cardiac peripheral benzodiazepine receptors

Noise is an environmental physical agent, which is regarded as a stressful stimulus: impairment and modifications in biological functions are reported, after loud noise exposure, at several levels in human and animal organs and apparatuses, as well as in the endocrine, cardiovascular and nervous system. In the present study equilibrium binding parameters of peripheral benzodiazepine receptors (PBRs) labelled by the specific radioligand [3H]PK 11195, were evaluated in cardiac tissue of rats submitted to 6 or 12 h noise exposure and of rats treated "in vivo" with PBR ligands such as PK 11195, Ro54864, diazepam and then noise-exposed. Results revealed a statistically significant decrease in the maximum number of binding sites (Bmax) of [3H]PK 11195 in atrial membranes of 6 or 12 h noise exposed rats, compared with sham-exposed animals, without any change in the dissociation constant (Kd). The "in vivo" PBR ligand pre-treatment counteracted the noise-induced modifications of PBR density. As PBRs are mainly located on mitochondria we also investigated whether noise exposure can affect the [3H]PK 11195 binding parameters in isolated cardiac mitochondrial fractions. Results indicated a significant Bmax value decrease in right atrial mitochondrial fractions of rats 6 or 12 h noise-exposed. Furthermore, as PBR has been suggested to be a supramolecular complex that might coincide with the not-yet-established structure of the mitochondrial permeability transition (MPT)-pore, the status of the MPT-pore in isolated heart mitochondria was investigated in noise- and sham-exposed rats. The loss of absorbance associated with the calcium-induced MPT-pore opening was greater in mitochondria isolated from hearts of 6 h noise- than those of sham-exposed rats. In conclusion, these findings represent a further instance for PBR density decrease in response to a stressful stimulus, like noise; in addition they revealed that "in vivo" administration of PBR ligands significantly prevents this decrease. Finally, our data also suggest the involvement of MPT in the response of an organism to noise stress.     

A comparison of the high-affinity peripheral benzodiazepine receptor ligands DAA1106 and (R)-PK11195 in rat models of neuroinflammation: implications for PET imaging of microglial activation

Activated microglia are an important feature of many neurological diseases and can be imaged in vivo using 1-(2-chlorophenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinolinecarboxamide (PK11195), a ligand that binds the peripheral benzodiazepine receptor (PBR). N-(2,5-dimethoxybenzyl)-N-(5-fluoro-2-phenoxyphenyl) acetamide (DAA1106) is a new PBR-specific ligand that has been reported to bind to PBR with higher affinity compared with PK11195. We hypothesized that this high-affinity binding of DAA1106 to PBR will enable better delineation of microglia in vivo using positron emission tomography. [(3)H]DAA1106 showed higher binding affinity compared with [(3)H](R)-PK11195 in brain tissue derived from normal rats and the rats injected intrastriatally with 6-hydroxydopamine or lipopolysaccharide at the site of the lesion. Immunohistochemistry combined with autoradiography in brain tissues as well as correlation analyses showed that increased [(3)H]DAA1106 binding corresponded mainly to activated microglia. Finally, ex vivo autoradiography and positron emission tomography imaging in vivo showed greater retention of [(11)C]DAA1106 compared with [(11)C](R)-PK11195 in animals injected with either lipopolysaccaride or 6-hydroxydopamine at the site of lesion. These results indicate that DAA1106 binds with higher affinity to microglia in rat models of neuroinflammation when compared with PK11195, suggesting that [(11)C]DAA1106 may represent a significant improvement over [(11)C](R)-PK11195 for in vivo imaging of activated microglia in human neuroinflammatory disorders.     

Prolactin-stimulated mitogenesis in the Nb2 rat lymphoma cell: lack of protoporphyrin IX effects

Pharmacological characterization of the Nb2 cell peripheral-type benzodiazepine receptor (PBR) was determined using selected 1,4-benzodiazepines, PK 11195, and protoporphyrin IX (PPIX) to compete for specific [3H] Ro5-4864 binding. These data suggest that PPIX possesses an affinity for the Nb2 cell PBR (Ki = 142 nM). We have previously reported that the peripheral benzodiazepine ligands, Ro5-4864 and PK 11195, modulate prolactin-stimulated mitogenesis in the Nb2 cell(1). In contrast, PPIX, a putative endogenous ligand for the PBR had no effect on prolactin-stimulated mitogenesis in the Nb2 cell over the concentration range from 10(-15) M to 10(-6) M. Taken together these data show that PPIX has an affinity for the Nb2 cell PBR but does not modulate prolactin-stimulated mitogenesis at concentrations which should bind to the Nb2 cell PBR.     

2-Arylpyrazolo[1,5-a]pyrimidin-3-yl acetamides. New potent and selective peripheral benzodiazepine receptor ligands

A new class of N,N-diethyl-(2-arylpyrazolo[1,5-a]pyrimidin-3-yl)acetamides (3f-y), as azaisosters of Alpidem, was prepared following a novel synthetic method and their affinities for both the peripheral (PBR) and the central (CBR) benzodiazepine receptors were evaluated. Binding assays were carried out using both [3H]PK 11195 and [3H]Ro 5-4864 as radioligands for PBR, whereas [3H]Ro 15-1788 was used for CBR, in rat kidney and rat cortex, respectively. The tested compounds exhibited a broad range of binding affinities from as low as 0.76 nM to inactivity and most of them proved to be high selective ligands for PBR. The preliminary SAR studies suggested some of the structural features required for high affinity and selectivity; particularly the substituents on the pyrimidine moiety seemed to play an important role in PBR versus CBR selectivity. A subset of the highest affinity compounds was also tested for their ability to stimulate steroid biosynthesis in C6 glioma rat cells and some of these were found to increase pregnenolone formation with potency similar to Ro 5-4864 and PK 11195.     

Fully automated synthesis and initial PET evaluation of [11C]PBR28

Fully automated synthesis and initial PET evaluation of a TSPO radioligand, [¹¹C]PBR28 (N-(2-[¹¹C]methoxybenzyl)-N-(4-phenoxypyridin-3-yl)acetamide), are reported. These results facilitate the potential preclinical and clinical PET studies of [¹¹C]PBR28 in animals and humans.     

Synthesis, biodistribution and micro-PET imaging of radiolabeled antimitotic agent T138067 analogues

Radiolabeled antimitotic agents [11C]T138067 and [18F]T138067 have been synthesized for evaluation as new potential positron emission tomography (PET) biomarkers for cancer imaging. In vivo biodistribution and micro-PET imaging of [11C]T138067 were performed in breast cancer animal models MCF-7 transfected with IL-1alpha implanted athymic mice and MDA-MB-435 implanted athymic mice. The results suggest that the uptakes of [11C]T138067 in both MCF-7 transfected with IL-1alpha tumor and MDA-MB-435 tumor are non-specific binding.     

Design, synthesis, and structure-activity relationships of novel tetracyclic compounds as peripheral benzodiazepine receptor ligands

The peripheral benzodiazepine receptor (PBR) is pharmacologically distinct from the central benzodiazepine receptor (CBR) and has been identified in a wide range of peripheral tissues as well as in the central nervous system. Although numerous studies have been performed of it, the physiological roles and functions of the PBR are still unclear. In the present study, in exploring new types of ligands for PBR, we found that a new series of compounds having a tetracyclic ring system, which were designed from FGIN-1-27, exhibited high affinities for PBR. We prepared and evaluated them for PBR affinities. The results of binding tests showed that 12e and 12f were the most potent PBR ligands among them (12e: IC(50)=0.44nM, 12f: IC(50)=0.37nM). In this paper, we present the design, synthesis, and structure-activity relationships (SARs) of novel tetracyclic compounds.     

Design, synthesis and structure-affinity relationships of aryloxyanilide derivatives as novel peripheral benzodiazepine receptor ligands

Since the peripheral benzodiazepine receptor (PBR) has been primarily found as a high-affinity binding site for diazepam in rat kidney, numerous studies of it have been performed. However, the physiological role and functions of PBR have not been fully elucidated. Currently, we presented the pharmacological profile of two high and selective PBR ligands, N-(2,5-dimethoxybenzyl)-N-(4-fluoro-2-phenoxyphenyl)acetamide (7-096, DAA1106) (PBR: IC(50)=0.28 nM) and N-(4-chloro-2-phenoxyphenyl)-N-(2-isopropoxybenzyl)acetamide (7-099, DAA1097) (PBR: IC(50)=0.92 nM). The compounds are aryloxyanilide derivatives, and identified with known PBR ligands such as benzodiazepine (1, Ro5-4864), isoquinoline (2, PK11195), imidazopyridine (3, Alpidem), and indole (5, FGIN-1-27) derivatives. The aryloxyanilide derivatives, which have been derived by opening the diazepine ring of 1, are a novel class as PBR ligands and have exhibited high and selective affinity for peripheral benzodiazepine receptors (PBRs). These novel derivatives would be useful for exploring the functions of PBR. In this paper, the design, synthesis and structure-affinity relationships of aryloxyanilide derivatives are described.     

In vivo imaging of peripheral benzodiazepine receptors in mouse lungs: a biomarker of inflammation

The ability to visualize the immune response with radioligands targeted to immune cells will enhance our understanding of cellular responses in inflammatory diseases. Peripheral benzodiazepine receptors (PBR) are present in monocytes and neutrophils as well as in lung tissue. We used lipopolysaccharide (LPS) as a model of inflammation to assess whether the PBR could be used as a noninvasive marker of inflammation in the lungs. Planar imaging of mice administrated 10 or 30 mg/kg LPS showed increased [(123)I]-(R)-PK11195 radioactivity in the thorax 2 days after LPS treatment relative to control. Following imaging, lungs from control and LPS-treated mice were harvested for ex vivo gamma counting and showed significantly increased radioactivity above control levels. The specificity of the PBR response was determined using a blocking dose of nonradioactive PK11195 given 30 min prior to radiotracer injection. Static planar images of the thorax of nonradioactive PK11195 pretreated animals showed a significantly lower level of radiotracer accumulation in control and in LPS-treated animals (p < .05). These data show that LPS induces specific increases in PBR ligand binding in the lungs. We also used in vivo small-animal PET studies to demonstrate increased [(11)C]-(R)-PK11195 accumulation in the lungs of LPS-treated mice. This study suggests that measuring PBR expression using in vivo imaging techniques may be a useful biomarker to image lung inflammation.     

Cloning and expression of a pharmacologically unique bovine peripheral-type benzodiazepine receptor isoquinoline binding protein.

High affinity binding of isoquinolines, such as PK 11195, is a conserved feature of peripheral-type benzodiazepine receptors (PBR) across species. However, species differences in PBR ligand binding have been described based on the affinity for N1-alkyl-1,4-benzodiazepines, such as Ro5-4864. Ro5-4864 binds with high affinity to the rat receptor but has low affinity for the bovine PBR. Photolabeling with an isoquinoline ligand, [3H]PK 14105, identifies a 17-kDa protein, the PBR isoquinoline binding protein (PBR/IBP), in both species. To further elucidate the role of the PBR/IBP in determining PBR benzodiazepine and isoquinoline binding characteristics, the bovine PBR/IBP was cloned and expressed. Using a cDNA encoding a rat PBR/IBP to screen a fetal bovine adrenal cDNA library, a bovine cDNA encoding a polypeptide of 169 residues was cloned. The bovine and rat PBR/IBPs had similar hydropathy profiles exhibiting five potential transmembrane domains. Transfecting the cloned bovine PBR/IBP cDNA into COS-7 cells resulted in an 11-fold increase in the density of high affinity [3H]PK 11195 binding sites which had only low affinity for Ro5-4864. Expression of the bovine PBR/IBP yields a receptor which is pharmacologically distinct from both endogenous COS-7 PBR and the rat PBR based on the affinity for several N1-alkyl-1,4-benzodiazepine ligands. These results suggest the PBR/IBP is the minimal functional component required for PBR ligand binding characteristics and the different protein sequences account for the species differences in PBR benzodiazepine ligand binding.     

Synthesis and evaluation of C-11, F-18 and I-125 small molecule radioligands for detecting oxytocin receptors

Compounds 1-4 were synthesized and investigated for selectivity and potency for the oxytocin receptor (OTR) to determine their viability as radioactive ligands. Binding assays determined 1-4 to have high binding affinity for both the human and rodent OTR and also have high selectivity for the human OTR over human vasopressin V1a receptors (V1aR). Inadequate selectivity for OTR over V1aR was found for rodent receptors in all four compounds. The radioactive (C-11, F-18, and I-125) derivatives of 1-4 were synthesized and investigated for use as autoradiography and positron emission tomography (PET) ligands. Receptor autoradiography performed with [(125)I]1 and [(125)I]2 on rodent brain slices provided the first small molecule radioligand images of the OTR and V1aR. Biodistribution studies determined [(125)I]1 and [(125)I]2 were adequate for in vivo peripheral investigations, but not for central investigations due to low uptake within the brain. A biodistribution study with [(18)F]3 suggested brain uptake occurred slowly over time. PET imaging studies with [(18)F]3 and [(11)C]4 using a rat model provided insufficient uptake in the brain over a 90 and 45 min scan times respectively to merit further investigations in non-human primates.     

Synthesis, radiosynthesis and in vivo preliminary evaluation of [11C]LBT-999, a selective radioligand for the visualisation of the dopamine transporter with PET

LBT-999 (8-((E)-4-fluoro-but-2-enyl)-3beta-p-tolyl-8-aza-bicyclo[3.2.1]octane-2beta-carboxylic acid methyl ester), a cocaine derivative belonging to a new generation of highly selective dopamine transporter (DAT) ligands, and its corresponding carboxylic acid derivative, the latter used as precursor for labelling both with tritium and the positron-emitter carbon-11 (half-life: 20.38 min), were synthesized from (R)-cocaine. [(3)H]LBT-999 (>99% radiochemically pure, specific radioactivity of 3.1 TBq/mmol) was prepared from [(3)H]methyl iodide, allowing its in vitro pharmacological evaluation (K(D): 9 nM for DAT and IC(50) > 1000 nM for SERT and NET). Routine production batches of 4.5-9.0 GBq of iv injectable solutions of [(11)C]LBT-999 (with specific radioactivities ranging from 30 to 45 GBq/mumol) were prepared in 25-30 min (HPLC purification and formulation included) using the efficient methylation reagent [(11)C]methyl triflate. The preliminary in vivo pharmacological evaluation of [(11)C]LBT-999, using both biodistributions in rats and brain imaging in monkeys with positron emission tomography (PET), clearly illustrates that this ligand is an excellent candidate for quantification with PET of DAT in humans.     

Synthesis and receptor binding studies of halogenated N,N-dialkylel-(2-phenyl-1H-indol-3-yl)glyoxylamides to visualize peripheral benzodiazepine receptors with SPECT or PET

A library of halogenated 2-arylindolyl-3-oxocarboxamides was prepared to develop radioligands to visualize cerebral PBR by SPECT and PET imaging. In vitro evaluation showed that most of the synthesized compounds were selective,high-affinity PBR ligands with adequate lipophilicity (log D7.4 in the range of 1.6-2.4). The iodinated derivative 11 (Ki = 2.6 nM) and the fluorinated analog 26 (Ki = 6.2 nM) displayed higher affinity than reference compounds.     

[18F]FEAC and [18F]FEDAC: Two novel positron emission tomography ligands for peripheral-type benzodiazepine receptor in the brain

[¹⁸F]FEAC ([¹⁸F]4a) and [¹⁸F]FEDAC ([¹⁸F]4b) were developed as two novel positron emission tomography (PET) ligands for peripheral-type benzodiazepine receptor (PBR). [¹⁸F]4a and [¹⁸F]4b were synthesized by fluoroethylation of precursors 8a and 8b with [¹⁸F]FCH₂CH₂Br ([¹⁸F]9), respectively. Small-animal PET scan for a neuroinflammatory rat model showed that the two radioligands had high uptakes of radioactivity in the kainic acid-infused striatum, a brain region where PBR density was increased.     

An alteration in the lateral geniculate nucleus of experimental glaucoma monkeys: in vivo positron emission tomography imaging of glial activation

We examined lateral geniculate nucleus (LGN) degeneration as an indicator for possible diagnosis of glaucoma in experimental glaucoma monkeys using positron emission tomography (PET). Chronic intraocular pressure (IOP) elevation was induced by laser trabeculoplasty in the left eyes of 5 cynomolgus monkeys. Glial cell activation was detected by PET imaging with [(11)C]PK11195, a PET ligand for peripheral-type benzodiazepine receptor (PBR), before and at 4 weeks after laser treatment (moderate glaucoma stage). At mild, moderate, and advanced experimental glaucoma stages (classified by histological changes based on the extent of axonal loss), brains were stained with cresyl violet, or antibodies against PBR, Iba-1 (a microglial marker), and GFAP (an activated astrocyte marker). In laser-treated eyes, IOP was persistently elevated throughout all observation periods. PET imaging showed increased [(11)C]PK11195 binding potential in the bilateral LGN at 4 weeks after laser treatment; the increase in the ipsilateral LGN was statistically significant (P<0.05, n = 4). Immunostaining showed bilateral activations of microglia and astrocytes in LGN layers receiving input from the laser-treated eye. PBR-positive cells were observed in LGN layers receiving input from laser-treated eye at all experimental glaucoma stages including the mild glaucoma stage and their localization coincided with Iba-1 positive microglia and GFAP-positive astrocytes. These data suggest that glial activation occurs in the LGN at a mild glaucoma stage, and that the LGN degeneration could be detected by a PET imaging with [(11)C]PK11195 during the moderate experimental glaucoma stage after unilateral ocular hypertension. Therefore, activated glial markers such as PBR in the LGN may be useful in noninvasive molecular imaging for diagnosis of glaucoma.