Neocortical abnormalities of [11C]-flumazenil PET in mesial temporal lobe epilepsy

OBJECTIVE: To characterize abnormalities in neocortical central benzodiazepine receptor (cBZR) binding in patients with mesial temporal lobe epilepsy (mTLE) with unilateral hippocampal sclerosis (HS) using [(11)C]-flumazenil-(FMZ) PET and complementary voxel-based and quantitative volume-of-interest (VOI) methods. METHODS: The authors studied 13 control subjects and 15 patients with refractory mTLE and unilateral HS with [(11)C]-FMZ PET. Data were corrected for partial volume effect in the interactively outlined hippocampus and in 28 cortical VOI using an individualized template. A voxel-based analysis was also performed using statistical parametric mapping (SPM). RESULTS: Fourteen patients with mTLE had reduced [(11)C]-FMZ volume distribution (V(d)) in the hippocampus ipsilateral to the EEG focus, extending into the amygdala in four. Five patients showed additional significant neocortical abnormalities of [(11)C]-FMZ binding: temporal neocortical increases (1), extratemporal decreases (2), extratemporal increases only (1), and temporal and extratemporal neocortical increases (1). Group VOI analysis revealed significant reductions only in the ipsilateral hippocampus. SPM showed decreased [(11)C]-FMZ-V(d) in the ipsilateral hippocampus in 13 of 15 patients, extending into the amygdala in eight. Five patients showed additional neocortical abnormalities: temporal neocortical increases only (3), extratemporal decreases (1), or both temporal neocortical and extratemporal decreases (1). Group analysis showed significant reductions in the ipsilateral hippocampus only. CONCLUSIONS: A combination of VOI- and voxel-based analysis of [(11)C]-FMZ PET detected extrahippocampal changes of cBZR binding in eight of 15 patients with mTLE due to HS. The finding of abnormalities in patients who were thought to have unilateral HS only based on MRI suggests that more widespread abnormalities are present in HS.     

Periventricular white matter flumazenil binding and postoperative outcome in hippocampal sclerosis

PURPOSE: In patients with hippocampal sclerosis (HS), anterior temporal lobe resection offers the possibility of a long-lasting suppression of seizures in two thirds of patients. White matter (WM) [11C]flumazenil volume of distribution (FMZ-Vd) reflects the number of neuronal cell bodies in WM. Our objective was to correlate WM FMZ-Vd in patients with unilateral HS and postsurgical outcome. METHODS: We performed [11C]FMZ-PET in 15 patients with refractory mesial temporal lobe epilepsy (mTLE) and a quantitative MRI diagnosis of unilateral HS subsequently histologically verified in all cases. Median follow-up was 7 years (range, 6-9 years). Metabolite-corrected arterial plasma input functions and spectral analysis were used to generate parametric images of [11C]FMZ-Vd. Statistical parametric mapping (SPM99) with explicit masking was used to investigate the entire brain volume including WM. RESULTS: Eight patients had Engel class IA outcome (completely seizure free since surgery), and seven were not seizure free. Comparison of seizure-free patients with those who continued to have seizures after surgery revealed areas of increased FMZ binding around the posterior horns of the ipsilateral (z=3.7) and contralateral (z=2.7) ventricles in those with suboptimal outcomes. CONCLUSIONS: Preoperative [11C]FMZ-PET can detect periventricular increases of WM FMZ binding, implying heterotopic neurons in WM, in patients with mTLE. The presence of such increases correlates with a poorer outcome.     

In vitro and in vivo characterization of (+)-3-[11C]cyano-dizocilpine

Summary. (+)-3-[¹¹C]Cyano-5-methyl-10,11-dihydro-5H-dibenzo[a,d]-cyclohepten-5,10-imine ([¹¹C]MKC) was successfully synthesized as a potential radiotracer for PET studies on the NMDA receptor channel complex. In vitro binding properties of [¹¹C]MKC were investigated with newly developed techniques for efficient evaluation of ¹¹C-labeled compounds. The association curve of [¹¹C]MKC binding in rat forebrain membranes showed that the specific binding reached an equilibrium within 30 min. Specific binding was saturable with affinity constant K_D = 8.2 ± 0.4 nM and B_max = 1.62 ± 0.04 pmol/mg protein with glutamate and glycine included in the incubation medium. The binding of [¹¹C]MKC was decreased by extensive washing of the membrane preparation. (+)- and (−)-Dizocilpine, 3-cyano-dizocilpine, and ketamine inhibited the specific binding of [¹¹C]MKC with IC₅₀ values of 37.3, 445.0, 65.8 nM and 3.91 μM, respectively. High specific binding in in vitro autoradiography was distributed predominantly in telencephalic regions (the hippocampus, cerebral cortex, and striatum) followed by thalamus. PET studies using rhesus monkeys under anesthesia showed high uptake of [¹¹C]MKC in the temporoparietal and frontal cerebral cortices, striatum, and thalamic regions, although it is problematic to verify the specific binding in vivo by PET. Accepted January 5, 1998; received October 7, 1997     

In vivo studies of acetylcholinesterase activity using a labeled substrate,N-[11C]methylpiperdin-4-yl propionate ([11C]PMP)

Two esters, N-[¹¹C]methylpiperidyl acetate ([¹¹C]AMP) and N-[¹¹C]methylpiperidyl propionate ([¹¹C]PMP), were synthesized in no-carrier-added forms and evaluated as in vivo substrates for brain acetylcholinesterase (AChE). After peripheral injection in mice, each ester showed rapid penetration into the brain and a regional retention of radioactivity (striatum > cortex, hippocampus > cerebellum) reflecting known levels of AChE activity in the brain. Regional brain distributions after [¹¹C]PMP administration showed better discrimination between regions of high, intermediate, and low AChE activities. Chromatographic analysis of blood and brain tissue extracts showed rapid and nearly complete hydrolysis of [¹¹C]PMP within 10 min after injection. For both [¹¹C]AMP and [¹¹C]PMP, retention of radioactivity in all regions was reduced by pretreatment with diisopropylfluorophosphate (DFP), a specific irreversible AChE inhibitor. DFP treatment also significantly increased the proportions of unhydrolyzed ester in both blood and brain. Radioactivity localization in brain after peripheral injection was thus dependent on AChE-catalyzed hydrolysis to the hydrophilic product N-[¹¹C]methylpiperidinol. PET imaging of [¹¹C]AMP or [¹¹C]PMP distributions in monkey brain showed clear accumulation of radioactivity in areas of highest AChE activity (striatum, cortex). These esters are thus in vivo substrates for brain AChE, with potential applications as in vivo imaging agents of enzyme action in the human brain. [¹¹C]PMP, the ester with a slower rate of hydrolysis, appears to be the better candidate radiotracer for further development. © 1996 Wiley-Liss, Inc.     

Changes in GABAA receptor properties in amygdala kindled animals: in vivo studies using [11C]flumazenil and positron emission tomography

Purpose: The purpose of the present investigation was to quantify alterations in GABA_A receptor density in vivo in rats subjected to amygdala kindling. Methods: The GABA_A receptor density was quantified by conducting a [¹¹C]flumazenil (FMZ) positron emission tomography (PET) study according to the full saturation method, in which each animal received a single injection of FMZ to fully saturate the GABA_A receptors. Subsequently, the concentration‐time curves of FMZ in blood [using high‐pressure liquid chromatography with UV detector (HPLC‐UV) or high‐performance liquid chromatography coupled to tandem mass spectrometry (LC/MS/MS)] and brain (with PET‐scanning) were analyzed by population modeling using a pharmacokinetic model, containing expressions to describe the time course of FMZ in blood and brain. Results: The GABA_A receptor density (B_max) in kindled rats was decreased by 36% compared with controls. This is consistent with a reduction of 28% in electroencephalography (EEG) effect of midazolam in the same animal model, suggesting that a reduced number of GABA_A receptors underlies the decreased efficacy of midazolam. Furthermore, receptor affinity (K_D) was not changed, but the total volume of distribution in the brain (V_Br), is increased to 178% of control after kindling, which might indicate an alteration in the transport of FMZ across the blood–brain barrier. Conclusions: Both the GABA_A receptor density (B_max), and possibly also the blood–brain barrier transport of FMZ (V_Br) are altered after kindling. Furthermore, this study indicates the feasibility of conducting PET studies for quantifying moderate changes in GABA_A receptor density in a rat model of epilepsy in vivo.     

Autoradiographic localization of 5-HT1A receptors in the post-mortem human brain using [H]WAY-100635 and [C]WAY-100635

The distribution of 5-HT_1A receptors was examined in the post-mortem human brain using whole hemisphere autoradiography and the selective 5-HT_1A receptor antagonist [³H]WAY-100635 ([O-methyl-³H]-N-(2-(4-(2-methoxyphenyl)-1-piperazinyl)ethyl)-N-(2-pyridinyl)cyclohexanecarboxamide trihydrochloride). The autoradiograms showed very dense binding to hippocampus, raphe nuclei and neocortex. The labeling in neocortex was slightly lower than in the hippocampus and was mainly at superficial layers, although a faintly labeled band could be seen in deeper neocortical layers. Other regions, such as the amygdala, septum and claustrum, showed low densities of [³H]WAY-100635 binding, reflecting low densities of 5-HT_1A receptors. The labeling was very low in basal ganglia, such as nucleus caudatus and putamen, in cerebellum or in structures of the brain stem except in the raphe nuclei. The labeling of human 5-HT_1A receptors with [³H]WAY-100635 was antagonized by the addition of the 5-HT_1A receptor ligands, 5-HT, buspirone, pindolol or 8-OH-DPAT (10 μM), leaving a very low background of non-specific binding. Saturation analysis of semiquantitative data from several human regions indicated that [³H]WAY-100635 has a K_d of approximately 2.5 nM. The selective labeling of 5-HT_1A receptors with [³H]WAY-100635 clearly show that this compound is useful for further studies of the human 5-HT_1A receptor subtype in vitro. [¹¹C]WAY-100635 is used for the characterization of 5-HT_1A receptors with positron emission tomography (PET). WAY-100635 was also radiolabeled with the short-lived positron-emitting radionuclide carbon-11 (t_1/2=20 min) and used for in vitro autoradiography on human whole hemisphere cryosections. [¹¹C]WAY-100635 gave images qualitatively similar to those of [³H]WAY-100635, although with a lower resolution. Thus, the hippocampal formation was densely labeled, with lower density in the neocortex. Buspirone, pindolol or 8-OH-DPAT (10 μM), blocked all binding of [¹¹C]WAY-100635. The in vitro autoradiography of the distribution of 5-HT_1A receptors obtained with radiolabeled WAY-100635 provide detailed qualitative and quantitative information on the distribution of 5-HT_1A-receptors in the human brain. Moreover, the studies give reference information for the interpretation of previous initial results at much lower resolution in humans with PET and [¹¹C]WAY-100635. These data provide a strong basis for expecting [¹¹C]WAY-100635 to behave as a highly selective radioligand in vivo.     

Central Benzodiazepine/γ-Aminobutyric AcidA Receptors in Idiopathic Generalized Epilepsy: An [11C]Flumazenil Positron Emission Tomography Study

Summary: Purpose: Previous [¹¹C]flumazenil (FMZ) positron emission tomography (PET) investigations in patients with idiopathic generalized epilepsy (IGE) have demonstrated nonsignificant global cortical decreases in central benzodiazepine γ-aminobutyric acid, (GABA_A) receptor (cBZR) binding or focal decreases in the thalamus and increases in the cerebellar nuclei with no changes in cerebral cortex. We previously reported lower [¹¹C]FMZ binding in cerebral cortex of IGE patients treated with valproate (VPA) than in cerebral cortex of controls. We now report high-resolution three-dimensional [¹¹C]FMZ PET studies in a larger number of subjects using an improved method to detect differences in cBZR between IGE patients and controls and a more powerful longitudinal design to determine the functional effect of VPA.
Methods: We compared parametric images of [¹¹C]FMZ volume of distribution (FMZVD) in 10 IGE patients before and after addition of VPA and in 20 normal subjects.
Results: Mean FMZVD was significantly higher in the cerebral cortex (11%, p = 0.009), thalamus (14%, p = 0.018), and cerebellum (15%, p = 0.027) of the 10 IGE patients as compared with that of 20 normal controls. Using statistical parametric mapping, no significant areas of focal abnormality of FMZVD were detected. Addition of VPA was not associated with a significant change in mean FMZVD in any brain area.
Conclusions: Our finding of increased FMZVD in IGE could reflect microdysgenesis or a state of cortical hyperexcitability. Our data suggest that short-term VPA therapy does not affect the number of available cBZR in patients with IGE.     

11C-flumazenil PET in patients with refractory temporal lobe epilepsy and normal MRI

BACKGROUND: Using 11C-flumazenil (FMZ) PET with correction for partial-volume effect, reductions of central benzodiazepine receptor (cBZR) binding can be detected reliably in vivo on remaining neurons in sclerotic hippocampi of patients with mesial temporal lobe epilepsy (TLE). OBJECTIVE: To delineate abnormalities of 11C-FMZ binding in patients with medically refractory TLE and normal quantitative MRI. METHODS: Analysis of parametric images of FMZ volume of distribution (Vd) using two complementary approaches: 1) MRI-based volume of interest (VOI) approach with partial volume effect correction for multiple hippocampal and extrahippocampal VOIs; and 2) statistical parametric mapping (SPM) to localize significant 11C-FMZ binding changes objectively on a voxel-by-voxel basis. RESULTS: Significant abnormalities of absolute FMZ-Vd were found after partial volume effect correction in 5 of 10 patients: unilateral decrease in the amygdala ipsilateral to the EEG focus (1), unilateral hippocampal decreases and bilateral temporal and extratemporal neocortical decreases (2), unilateral increase in the temporal neocortex together with extratemporal neocortical increases (1), and bilateral posterior hippocampal increases together with temporal neocortical increases (1). In the three patients with extratemporal neocortical changes, the concomitant unilateral hippocampal or temporal neocortical changes were contralateral to the presumed epileptic focus. Significant asymmetries of FMZ-Vd between homologous regions were found in six patients. In four of those patients, absolute FMZ-Vd for the homologous regions were within normal limits, with two of the four patients showing relatively higher hippocampal values ipsilateral to the presumed epileptic focus. SPM analysis localized significant abnormalities of FMZ-Vd in similar locations in three of the seven patients in whom VOI analysis detected significant changes. In addition, SPM indicated significant unilateral contralateral hippocampal decreases in an eighth patient. However, both methods failed to localize epileptic foci in two patients identified by depth-EEG recordings. CONCLUSIONS: 11C-FMZ PET showed focal increases as well as decreases of FMZ binding in 80% of patients with refractory TLE and normal high-quality MRI but was not consistently helpful in localizing the epileptic foci.     

In vivo kinetics of [F-18]MEFWAY: a comparison with [C-11]WAY100635 and [F-18]MPPF in the nonhuman primate

[F‐18]Mefway was developed to provide an F‐18 labeled positron emission tomography (PET) neuroligand with high affinity for the serotonin 5‐HT_1A receptor to improve the in vivo assessment of the 5‐HT_1A system. The goal of this work was to compare the in vivo kinetics of [F‐18]mefway, [F‐18]MPPF, and [C‐11]WAY100635 in the rhesus monkey. Methods: Each of four monkeys were given bolus injections of [F‐18]mefway, [C‐11]WAY100635, and [F‐18]MPPF and scans were acquired with a microPET P4 scanner. Arterial blood was sampled to assay parent compound throughout the time course of the PET experiment. Time activity curves were extracted in the high 5‐HT_1A binding areas of the anterior cingulate cortex (ACG), mesial temporal cortex, raphe nuclei, and insula cortex. Time activity curves were also extracted in the cerebellum, which was used as a reference region. The in vivo kinetics of the radiotracers were compared based on the nondisplaceable distribution volume (V_ND) and binding potential (BP_ND). Results: At 30 min, the fraction of radioactivity in the plasma due to parent compound was 19%, 28%, and 29% and cleared from the arterial plasma at rates of 0.0031, 0.0078, and 0.0069 (min^?1) ([F‐18]mefway, [F‐18]MPPF, [C‐11]WAY100635). The BP_ND in the brain regions were mesial temporal cortex: 7.4 ± 0.6, 3.1 ± 0.4, 7.0 ± 1.2, ACG: 7.2 ± 1.2, 2.1 ± 0.2, 7.9 ± 1.2; raphe nuclei: 3.7 ± 0.6, 1.3 ± 0.3, 3.3 ± 0.7; and insula cortex: 4.2 ± 0.6, 1.2 ± 0.1, 4.7 ± 1.0 for [F‐18]mefway, [F‐18]MPPF, and [C‐11]WAY100635 respectively. Conclusions: In the rhesus monkey, [F‐18]mefway has similar in vivo kinetics to [C‐11]WAY100635 and yields greater than 2‐fold higher BP_ND than [F‐18]MPPF. These properties make [F‐18]mefway a promising radiotracer for 5‐HT_1A assay, providing higher counting statistics and a greater dynamic range in BP_ND. Synapse, 2011. © 2010 Wiley‐Liss, Inc.     

In vivo SPECT imaging of 5-HT1A receptors with [123I] p-MPPI in nonhuman primates

The in vivo imaging of a novel iodinated phenylpiperazine derivative for 5-HT_1A receptors, [¹²³I]p-MPPI (4-(2′-methoxy-)phenyl-1-[2′-(n-2"-pyridinyl)-p-iodo-benzamido-]ethyl-piperazine), using single photon emission computed tomography (SPECT), was evaluated in nonhuman primates. After an i.v. injection, [¹²³I]p-MPPI penetrated the blood-brain barrier quickly and localized in brain regions where 5-HT_1A receptor density is high (hippocampus, frontal cortex, cingulate gyrus, entorhinal cortex). Maximum ratio of hippocampus to cerebellum was 3 to 1 at 50 min postinjection. The specific binding of the radioligand in the hippocampal region, an area rich in 5-HT_1A receptor density, was blocked by a chasing dose of (±) 8-OH-DPAT (2 mg/kg, i.v.) or non-radioactive p-MPPI (1 mg/kg, i.v.), whereas the regional distribution of [¹²³I]p-MPPI was unaffected by treatment with non 5-HT_1A agents, such as ketanserin. Ex vivo and in vitro autoradiographic studies using monkey brain further confirmed that the specific binding of [¹²³I]p-MPPI is associated with 5-HT_1A receptor sites. However, the initial attempt at [¹²³I]p-MPPI human imaging studies did not display specific localization of 5-HT_1A receptors. This discrepancy observed for [¹²³I]p-MPPI may be due to a dramatic difference in metabolic pathways between humans and monkeys. © 1996 Wiley-Liss, Inc.     

Detecting neuroinflammation in the brain following chronic alcohol exposure in rats: A comparison between in vivo and in vitro TSPO radioligand binding

Excessive alcohol consumption is associated with neuroinflammation, which likely contributes to alcohol‐related pathology. However, positron emission tomography (PET) studies using radioligands for the 18‐kDa translocator protein (TSPO), which is considered a biomarker of neuroinflammation, reported decreased binding in alcohol use disorder (AUD) participants compared to controls. In contrast, autoradiographic findings in alcohol exposed rats reported increases in TSPO radioligand binding. To assess if these discrepancies reflected differences between in vitro and in vivo methodologies, we compared in vitro autoradiography (using [³H]PBR28 and [³H]PK11195) with in vivo PET (using [¹¹C]PBR28) in male, Wistar rats exposed to chronic alcohol‐vapor (dependent n = 10) and in rats exposed to air‐vapor (nondependent n = 10). PET scans were obtained with [¹¹C]PBR28, after which rats were euthanized and the brains were harvested for autoradiography with [³H]PBR28 and [³H]PK11195 (n = 7 dependent and n = 7 nondependent), and binding quantified in hippocampus, thalamus, and parietal cortex. Autoradiography revealed significantly higher binding in alcohol‐dependent rats for both radioligands in thalamus and hippocampus (trend level for [³H]PBR28) compared to nondependent rats, and these group differences were stronger for [³H]PK11195 than [³H]PBR28. In contrast, PET measures obtained in the same rats showed no group difference in [¹¹C]PBR28 binding. Our in vitro data are consistent with neuroinflammation associated with chronic alcohol exposure. Failure to observe similar increases in [¹¹C]PBR28 binding in vivo suggests the possibility that a mechanism mediated by chronic alcohol exposure interferes with [¹¹C]PBR28 binding to TSPO in vivo. These data question the sensitivity of PBR28 PET as a methodology to assess neuroinflammation in AUD.     

Evaluation in monkey of two candidate PET radioligands, [11C]RX‐1 and [18F]RX‐2, for imaging brain 5‐HT4 receptors

The serotonin subtype‐4 (5‐HT₄) receptor, which is known to be involved physiologically in learning and memory, and pathologically in Alzheimer's disease, anxiety, and other neuropsychiatric disorders—has few radioligands readily available for imaging in vivo. We have previously reported two novel 5‐HT₄ receptor radioligands, namely [methoxy‐¹¹C](1‐butylpiperidin‐4‐yl)methyl 4‐amino‐3‐methoxybenzoate; [¹¹C]RX‐1), and the [¹⁸F]3‐fluoromethoxy analog ([¹⁸F]RX‐2), and in this study we evaluated them by PET in rhesus monkey. Brain scans were performed at baseline, receptor preblock or displacement conditions using SB 207710, a 5‐HT₄ receptor antagonist, on the same day for [¹¹C]RX‐1 and on different days for [¹⁸F]RX‐2. Specific‐to‐nondisplaceable ratio (BP_ND) was measured with the simplified reference tissue model from all baseline scans. To determine specific binding, total distribution volume (V_T) was also measured in some monkeys by radiometabolite‐corrected arterial input function after ex vivo inhibition of esterases from baseline and blocked scans. Both radioligands showed moderate to high peak brain uptake of radioactivity (2–6 SUV). Regional BP_ND values were in the rank order of known 5‐HT₄ receptor distribution with a trend for higher BP_ND values from [¹⁸F]RX‐2. One‐tissue compartmental model provided good fits with well identified V_T values for both radioligands. In the highest 5‐HT₄ receptor density region, striatum, 50–60% of total binding was specific. The V_T in receptor‐poor cerebellum reached stable values by about 60 min for both radioligands indicating little influence of radiometabolites on brain signal. In conclusion, both [¹¹C]RX‐1 and [¹⁸F]RX‐2 showed positive attributes for PET imaging of brain 5‐HT₄ receptors, validating the radioligand design strategy. Synapse 68:613–623, 2014 . © 2014 Wiley Periodicals, Inc.     

Compartmental analysis of [11C]flumazenil kinetics for the estimation of ligand transport rate and receptor distribution using positron emission tomography.

The in vivo kinetic behavior of [11C]flumazenil ([11C]FMZ), a non-subtype-specific central benzodiazepine antagonist, is characterized using compartmental analysis with the aim of producing an optimized data acquisition protocol and tracer kinetic model configuration for the assessment of [11C]FMZ binding to benzodiazepine receptors (BZRs) in human brain. The approach presented is simple, requiring only a single radioligand injection. Dynamic positron emission tomography data were acquired on 18 normal volunteers using a 60- to 90-min sequence of scans and were analyzed with model configurations that included a three-compartment, four-parameter model, a three-compartment, three-parameter model, with a fixed value for free plus nonspecific binding; and a two-compartment, two-parameter model. Statistical analysis indicated that a four-parameter model did not yield significantly better fits than a three-parameter model. Goodness of fit was improved for three- versus two-parameter configurations in regions with low receptor density, but not in regions with moderate to high receptor density. Thus, a two-compartment, two-parameter configuration was found to adequately describe the kinetic behavior of [11C]FMZ in human brain, with stable estimates of the model parameters obtainable from as little as 20-30 min of data. Pixel-by-pixel analysis yields functional images of transport rate (K1) and ligand distribution volume (DV"), and thus provides independent estimates of ligand delivery and BZR binding.     

Age-related reduction of [C]MDL100,907 binding to central 5-HT2A receptors:: PET study in the conscious monkey brain

Age-related alterations of serotonin (5-hydroxytryptamine; 5-HT) type 2A receptors (5-HT_2A) in the living brains of young (6.0±1.3 years old) and aged (19.2±3.0 years old) monkeys (Macaca mulatta) were evaluated with [¹¹C]MDL100,907 in the conscious state using high-resolution positron emission tomography (PET). For quantitative analysis of 5-HT_2A binding in vivo, PET scan of [¹¹C]MDL100,907 was performed with arterial blood sampling in each animal, and the metabolic-corrected arterial input function was used for Logan graphical analysis. Higher cerebral binding of [¹¹C]MDL100,907 was observed in the hippocampus, cingulate gyrus, frontal, temporal and occipital cortices, regions known to contain high densities of 5-HT_2A, by in vitro assay. Binding was intermediate in the striatum and thalamus, and lower in the pons and cerebellum in both young and aged monkeys. The age-related decrease in [¹¹C]MDL100,907 binding to 5-HT_2A receptors was prominent in the hippocampus, cingulate gyrus, frontal, temporal and occipital cortices, but not in the striatum, thalamus and pons. These observation demonstrated the usefulness of [¹¹C]MDL100,907 as an labeled compound for assessment of the aging process of the cortical 5-HT_2A measured by PET.     

Autoradiographic characterization of [3H]inositol (1,4,5) trisphosphate and [3H]inositol (1,3,4,5) tetrakisphosphate binding sites in human brain

Summary Autoradiographic techniques were used to investigate the characteristics of tritiated inositol(1,4,5)trisphosphate ([³H]IP₃) and inositol (1,3,4,5)tetrakisphosphate ([³H]IP₄) binding to human brain. In brain sections [³H]IP₃ exhibited a two-site binding with K _d values of 87 nM and 9.3 M respectively for the higher and lower affinity sites. [³H]IP₄ also bound to two sites with K _d values of 43 nM and 1.4 M, respectively. With the conditions fixed in this study, [³H]IP₃ and [³H]IP₄ autoradiography in the cortex, caudate, hippocampus and cerebellum were performed. The most prominent [³H]IP₃ binding among these regions was found in the cerebellum, particularly in the molecular layer. Within the hippocampus, the subiculum and the CA1 region showed much more prominent binding than the other subfields. [³H]IP₄ binding was fairly homogeneous in the regions studied, with the exception of a slightly higher binding in the molecular layer of the cerebellum.     

Brain uptake of α-[14C]methyl-para-tyrosine in the rat

The blood-brain permeabilities of L-[³H]tyrosine and the tyrosine hydroxylase (TH) inhibitor β-[¹⁴C]methyl-para-tyrosine ([¹⁴C]AMPT) were determined in rat striatum, a brain region rich in TH activity, and in other brain regions containing relatively little TH activity. In striatum, the unidirectional clearance rate (K₁) for L-[³H]tyrosine (6.2 ml hg- ^?1 min^?1) was significantly greater than the rates for L-[¹⁴C]AMPT (2.8 ml hg^?1 min^?1) and D-[¹⁴C]AMPT (0.8 ml hg^?1 min^?1). The apparent volume of distribution (V_f) for L-[¹⁴C]AMPT in striatum (72.5 ± 4.0 ml hg-¹) did not differ from the V_f in other brain regions. The homogeneous distribution of L-[¹⁴C]AMPT in rat brain indicates that labeled AMPT is unsuitable for the study of TH in vivo by quantitative autoradiography. © 1994 Wiley-Liss, Inc.     

Comparative evaluation of two glycine transporter 1 radiotracers [11C]GSK931145 and [18F]MK‐6577 in baboons

Glycine transporter type‐1 (GlyT1) has been proposed as a target for drug development for schizophrenia. PET imaging with a GlyT1 specific radiotracer will allow for the measurement of target occupancy of GlyT1 inhibitors, and for in vivo investigation of GlyT1 alterations in schizophrenia. We conducted a comparative evaluation of two GlyT1 radiotracers, [¹¹C]GSK931145, and [¹⁸F]MK‐6577, in baboons. Two baboons were imaged with [¹¹C]GSK931145 and [¹⁸F]MK‐6577. Blocking studies with GSK931145 (0.3 or 0.2 mg/kg) were conducted to determine the level of tracer specific binding. [¹¹C]GSK931145 and [¹⁸F]MK‐6577 were synthesized in good yield and high specific activity. Moderately fast metabolism was observed for both tracers, with ～30% of parent at 30 min post‐injection. In the brain, both radiotracers showed good uptake and distribution profiles consistent with regional GlyT1 densities. [¹⁸F]MK‐6577 displayed higher uptake and faster kinetics than [¹¹C]GSK931145. Time activity curves were well described by the two‐tissue compartment model. Regional volume of distribution (V_T) values were higher for [¹⁸F]MK‐6577 than [¹¹C]GSK931145. Pretreatment with GSK931145 reduced tracer uptake to a homogeneous level throughout the brain, indicating in vivo binding specificity and lack of a reference region for both radiotracers. Linear regression analysis of V_T estimates between tracers indicated higher specific binding for [¹⁸F]MK‐6577 than [¹¹C]GSK931145, consistent with higher regional binding potential (BP_ND) values of [¹⁸F]MK‐6577 calculated using V_T from the baseline scans and non‐displaceable distribution volume (V_ND) derived from blocking studies. [¹⁸F]MK‐6577 appears to be a superior radiotracer with higher brain uptake, faster kinetics, and higher specific binding signals than [¹¹C]GSK931145. Synapse 70:112–120, 2016. © 2016 Wiley Periodicals, Inc.     

Pharmacokinetics and in vivo specificity of [LLC]dl-threo-methylphenidate for the presynaptic dopaminergic neuron

dl-threo-Methylphenidate (Ritalin) was labeled with carbon-11 (t_1/2:20.4 minutes) in order to measure its pharmacokinetics, to evaluate it as a radiotracer for the presynaptic dopaminergic neuron, and to examine its sensitivity to the loss of dopaminergic neurons. Positron emission tomographic (PET) studies were carried out in the baboon to determine specificity for the presynaptic dopaminergic neuron and in humans to assess sensitivity to neuronal loss. Studies with [^llC]dl-threo-methylphenidate ([^llC]MP) in baboon demonstrated high regional uptake in the striatum. Peak uptake (0.04%/cc) occurred at 5-15 minutes post-injection. The half-time for clearance from peak uptake for [¹¹C]MP was 60 minutes and the ratio between the radioactivity in the striatum and that in the cerebellum (ST/CB) ranged from 2.2 to 2.6 at 40 minutes. Repeated measures in the same baboon showed ≤8% variability in the ST/CB ratio. Pretreatment with unlabeled methylphenidate (0.5 mg/kg) or GBR12909 (1.5 mg/kg) 30 minutes prior to [¹¹C]MP injection markedly reduced the striatal but not the cerebellar uptake of [¹¹C]MP, demonstrating the saturable and specific binding of [¹¹C]MP to a site on the dopamine transporter in the brain. In both cases, the ratio of striatum to cerebellum (ST/CB) after pretreatment was reduced by about 43% The ratios of distribution volumes at the steady-state for the striatum to cerebellum (ST/CB) for these two separate studies in the same baboon were reduced by 37 and 38%, respectively. In contrast, pretreatment with tomoxetine (1.5 mg/kg) or citalopram (2.0 mg/kg), inhibitors of the norepinephrine and serotonin transporter, respectively, did not produce a significant change in the kinetics of [¹¹C]MP and the ST/CB after pretreatment was similar to that for control. In one patient with Parkinson's disease, the striatum to cerebellum ratio of [¹¹C]MP was markedly lower than that of an elderly normal subject. These results demonstrate the saturable [¹¹C]MP binding to the dopamine transporter in the baboon brain and that [¹¹C]MP is sensitive to dopamine neuron degeneration in Parkinson's disease. © 1994 Wiley-Liss, Inc

1 This article is a US Government work and, as such, is in the public domain in the United States of America

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Lamina-specific changes in hippocampal GABAA/cBZR and mossy fibre sprouting during and following amygdala kindling in the rat

Alterations in hippocampal GABA_A/central benzodiazepine receptor (GABA_A/cBZR) expression and mossy fibre sprouting (MFS) may have aetiological significance in temporal lobe epileptogenesis. Their relationship with each other is also unknown. We utilised [³H]-flumazenil autoradiography to quantify changes in GABA_A/cBZR density and affinity in all hippocampal laminae, and Timm's staining for MFS at different stages of epileptogenesis in the amygdala kindling rat model (after 24 stimulations, 48 stimulations and two weeks post-kindling). During kindling, receptor density was significantly elevated within the dentate stratum moleculare and granulosum, but decreased within the stratum radiatum of CA3 and CA2. Two weeks post-kindling, receptor density remained upregulated in the dentate stratum moleculare and was also upregulated in CA3 stratum oriens and CA1 stratum moleculare. MFS was significantly increased in the dentate stratum moleculare at two weeks post-kindling, with a strong inverse correlation between MFS and GABA_A/cBZR density in this region. No changes in GABA_A/cBZR binding affinity were detected for any hippocampal subregion at any time point. Our results demonstrate that changes in hippocampal GABA_A/cBZR expression are lamina- and time-specific. Within the dentate gyrus, receptor density is upregulated throughout epileptogenesis, whilst within the hippocampus proper, receptor density is downregulated early in epileptogenesis but upregulated at the chronic phase. A novel association between MFS and GABA_A/cBZR density has been demonstrated by this study, which could represent an important compensatory or pathological mechanism associated with epileptogenesis.     

[11C]deschloroclozapine is an improved PET radioligand for quantifying a human muscarinic DREADD expressed in monkey brain

Previous work found that [¹¹C]deschloroclozapine ([¹¹C]DCZ) is superior to [¹¹C]clozapine ([¹¹C]CLZ) for imaging Designer Receptors Exclusively Activated by Designer Drugs (DREADDs). This study used PET to quantitatively and separately measure the signal from transfected receptors, endogenous receptors/targets, and non-displaceable binding in other brain regions to better understand this superiority. A genetically-modified muscarinic type-4 human receptor (hM₄Di) was injected into the right amygdala of a male rhesus macaque. [¹¹C]DCZ and [¹¹C]CLZ PET scans were conducted 2–24 months later. Uptake was quantified relative to the concentration of parent radioligand in arterial plasma at baseline (n = 3 scans/radioligand) and after receptor blockade (n = 3 scans/radioligand). Both radioligands had greater uptake in the transfected region and displaceable uptake in other brain regions. Displaceable uptake was not uniformly distributed, perhaps representing off-target binding to endogenous receptor(s). After correction, [¹¹C]DCZ signal was 19% of that for [¹¹C]CLZ, and background uptake was 10% of that for [¹¹C]CLZ. Despite stronger [¹¹C]CLZ binding, the signal-to-background ratio for [¹¹C]DCZ was almost two-fold greater than for [¹¹C]CLZ. Both radioligands had comparable DREADD selectivity. All reference tissue models underestimated signal-to-background ratio in the transfected region by 40%–50% for both radioligands. Thus, the greater signal-to-background ratio of [¹¹C]DCZ was due to its lower background uptake.