The chemogenetic technology designer receptors exclusively activated by designer drugs (DREADDs) afford remotely reversible control of cellular signaling, neuronal activity and behavior. Although the ...combination of muscarinic-based DREADDs with clozapine-N-oxide (CNO) has been widely used, sluggish kinetics, metabolic liabilities and potential off-target effects of CNO represent areas for improvement. Here, we provide a new high-affinity and selective agonist deschloroclozapine (DCZ) for muscarinic-based DREADDs. Positron emission tomography revealed that DCZ selectively bound to and occupied DREADDs in both mice and monkeys. Systemic delivery of low doses of DCZ (1 or 3 μg per kg) enhanced neuronal activity via hM3Dq within minutes in mice and monkeys. Intramuscular injections of DCZ (100 μg per kg) reversibly induced spatial working memory deficits in monkeys expressing hM4Di in the prefrontal cortex. DCZ represents a potent, selective, metabolically stable and fast-acting DREADD agonist with utility in both mice and nonhuman primates for a variety of applications.
A substantial and constitutive expression of translocator protein (TSPO) in cerebral blood vessels hampers the sensitive detection of neuroinflammation characterized by greatly induced TSPO ...expression in activated glia. Here, we conducted in vivo positron emission tomography (PET) and in vitro autoradiographic imaging of normal and TSPO-deficient mouse brains to compare the binding properties of 18F-FEBMP, a relatively novel TSPO radioligand developed for human studies based on its insensitivity to a common polymorphism, with 11C-PK11195, as well as other commonly used TSPO radioligands including 11C-PBR28, 11C-Ac5216 and 18F-FEDAA1106. TSPO in cerebral vessels of normal mice was found to provide a major binding site for 11C-PK11195, 11C-PBR28 and 18F-FEDAA1106, in contrast to no overt specific binding of 18F-FEBMP and 11C-Ac5216 to this vascular component. In addition, 18F-FEBMP yielded PET images of microglial TSPO with a higher contrast than 11C-PK11195 in a tau transgenic mouse modeling Alzheimer’s disease (AD) and allied neurodegenerative tauopathies. Moreover, TSPO expression examined by immunoblotting was significantly increased in AD brains compared with healthy controls, and was well correlated with the autoradiographic binding of 18F-FEBMP but not 11C-PK11195. Our findings support the potential advantage of comparatively glial TSPO-selective radioligands such as 18F-FEBMP for PET imaging of inflammatory glial cells.
Synaptic dysfunction provoking dysregulated cortical neural circuits is currently hypothesized as a key pathophysiological process underlying clinical manifestations in Alzheimer's disease and ...related neurodegenerative tauopathies. Here, we conducted PET along with postmortem assays to investigate time course changes of excitatory and inhibitory synaptic constituents in an rTg4510 mouse model of tauopathy, which develops tau pathologies leading to noticeable brain atrophy at 5-6 months of age. Both male and female mice were analyzed in this study. We observed that radiosignals derived from
Cflumazenil, a tracer for benzodiazepine receptor, in rTg4510 mice were significantly lower than the levels in nontransgenic littermates at 2-3 months of age. In contrast, retentions of (E)-
CABP688, a tracer for mGluR5, were unaltered relative to controls at 2 months of age but then gradually declined with aging in parallel with progressive brain atrophy. Biochemical and immunohistochemical assessment of postmortem brain tissues demonstrated that inhibitory, but not excitatory, synaptic constituents selectively diminished without overt loss of somas of GABAergic interneurons in the neocortex and hippocampus of rTg4510 mice at 2 months of age, which was concurrent with enhanced immunoreactivity of cFos, a well-characterized immediate early gene, suggesting that impaired inhibitory neurotransmission may cause hyperexcitability of cortical circuits. Our findings indicate that tau-induced disruption of the inhibitory synapse may be a critical trigger of progressive neurodegeneration, resulting in massive neuronal loss, and PET assessments of inhibitory versus excitatory synapses potentially offer
indices for hyperexcitability and excitotoxicity early in the etiologic pathway of neurodegenerative tauopathies.
In this study, we examined the in vivo status of excitatory and inhibitory synapses in the brain of the rTg4510 tauopathy mouse model by PET imaging with (E)-
CABP688 and
Cflumazenil, respectively. We identified inhibitory synapse as being significantly dysregulated before brain atrophy at 2 months of age, while excitatory synapse stayed relatively intact at this stage. In line with this observation, postmortem assessment of brain tissues demonstrated selective attenuation of inhibitory synaptic constituents accompanied by the upregulation of cFos before the formation of tau pathology in the forebrain at young ages. Our findings indicate that selective degeneration of inhibitory synapse with hyperexcitability in the cortical circuit constitutes the critical early pathophysiology of tauopathy.
Colony-stimulating factor 1 receptor (CSF1R) is a specific biomarker for microglia. In this study, we developed a novel PET radioligand for CSF1R, 11C-GW2580, and compared it to a reported CSF1R ...tracer, 11C-CPPC, in mouse models of acute and chronic neuroinflammation and a rhesus monkey. Dynamic 11C-GW2580- and 11C-CPPC-PET images were quantified by reference tissue-based models and standardized uptake value ratio. Both tracers exhibited increased uptake in the lesioned striata of lipopolysaccharide-injected mice and in the forebrains of AppNL-G-F/NL-G-F-knock-in mice, spatially in agreement with an increased 18-kDa translocator protein radioligand retention. Moreover, 11C-GW2580 captured changes in CSF1R availability more sensitively than 11C-CPPC, with a larger dynamic range and a smaller inter-individual variability, in these model animals. PET imaging of CSF1R in a rhesus monkey displayed moderate-to-high tracer retention in the brain at baseline. Homologous blocker (i. e. unlabeled tracer) treatment reduced the uptake of 11C-GW2580 by ∼30% in all examined brain regions except for centrum semi-ovale white matter, but did not affect the retention of 11C-CPPC. In summary, our results demonstrated that 11C-GW2580-PET captured inflammatory microgliosis in the mouse brain with higher sensitivity than a reported radioligand, and displayed saturable binding in the monkey brain, potentially providing an imaging-based quantitative biomarker for reactive microgliosis.
Positron emission tomography (PET) allows biomolecular tracking but PET monitoring of brain networks has been hampered by a lack of suitable reporters. Here, we take advantage of bacterial ...dihydrofolate reductase, ecDHFR, and its unique antagonist, TMP, to facilitate in vivo imaging in the brain. Peripheral administration of radiofluorinated and fluorescent TMP analogs enabled PET and intravital microscopy, respectively, of neuronal ecDHFR expression in mice. This technique can be used to the visualize neuronal circuit activity elicited by chemogenetic manipulation in the mouse hippocampus. Notably, ecDHFR‐PET allows mapping of neuronal projections in non‐human primate brains, demonstrating the applicability of ecDHFR‐based tracking technologies for network monitoring. Finally, we demonstrate the utility of TMP analogs for PET studies of turnover and self‐assembly of proteins tagged with ecDHFR mutants. These results establish opportunities for a broad spectrum of previously unattainable PET analyses of mammalian brain circuits at the molecular level.
SYNOPSIS
ecDHFR‐based reporter system can be utilized for bimodal fluorescence and Positron emission tomography (PET) imaging of expression and dynamics of its fused protein of interest in living animal brains, offering broad‐spectrum analyses of a mammalian deep brain circuit at molecular levels.
We established a genetically encoded ecDHFR‐based reporter system applicable for bimodal optical and PET imaging in living animal brains.
The reporter gene expression driven by an activity‐dependent promoter illuminates neuronal ensemble activities elicited by chemogenetic manipulation in the mouse hippocampal circuit.
ecDHFR/TMP systems enable visualization of neuronal tracts in deep brain regions of non‐human primates.
The utility of TMP analogs for PET monitoring of aggregation and turnover of proteins tagged with mutant forms of ecDHFR.
Application of bacterial dihydrofolate reductase ecDHFR and its unique antagonist TMP achieves a broad spectrum of previously unattainable in vivo PET analyses of mammalian brain circuits at the molecular level.
Purpose
Phosphodiesterase (PDE) 7 is a potential therapeutic target for neurological and inflammatory diseases, although in vivo visualization of PDE7 has not been successful. In this study, we aimed ...to develop
11
CMTP38 as a novel positron emission tomography (PET) ligand for PDE7.
Methods
11
CMTP38 was radiosynthesized by
11
C-cyanation of a bromo precursor with
11
CHCN. PET scans of rat and rhesus monkey brains and in vitro autoradiography of brain sections derived from these species were conducted with
11
CMTP38. In monkeys, dynamic PET data were analyzed with an arterial input function to calculate the total distribution volume (
V
T
). The non-displaceable binding potential (
BP
ND
) in the striatum was also determined by a reference tissue model with cerebellar reference. Finally, striatal occupancy of PDE7 by an inhibitor was calculated in monkeys according to changes in
BP
ND
.
Results
11
CMTP38 was synthesized with radiochemical purity ≥99.4% and molar activity of 38.6 ± 12.6 GBq/μmol. Autoradiography revealed high radioactivity in the striatum and its reduction by non-radiolabeled ligands, in contrast with unaltered autoradiographic signals in other regions. In vivo PET after radioligand injection to rats and monkeys demonstrated that radioactivity was rapidly distributed to the brain and intensely accumulated in the striatum relative to the cerebellum. Correspondingly, estimated
V
T
values in the monkey striatum and cerebellum were 3.59 and 2.69 mL/cm
3
, respectively. The cerebellar
V
T
value was unchanged by pretreatment with unlabeled MTP38. Striatal
BP
ND
was reduced in a dose-dependent manner after pretreatment with MTP-X, a PDE7 inhibitor. Relationships between PDE7 occupancy by MTP-X and plasma MTP-X concentration could be described by Hill’s sigmoidal function.
Conclusion
We have provided the first successful preclinical demonstration of in vivo PDE7 imaging with a specific PET radioligand.
11
CMTP38 is a feasible radioligand for evaluating PDE7 in the brain and is currently being applied to a first-in-human PET study.
The tendency to avoid punishment, called behavioral inhibition system, is an essential aspect of motivational behavior. Behavioral inhibition system is related to negative affect, such as anxiety, ...depression and pain, but its neural basis has not yet been clarified. To clarify the association between individual variations in behavioral inhibition system and brain 5-HT
2A
receptor availability and specify which brain networks were involved in healthy male subjects, using
18
Faltanserin positron emission tomography and resting-state functional magnetic resonance imaging. Behavioral inhibition system score negatively correlated with 5-HT
2A
receptor availability in anterior cingulate cortex. A statistical model indicated that the behavioral inhibition system score was associated with 5-HT
2A
receptor availability, which was mediated by the functional connectivity between anterior cingulate cortex and left middle frontal gyrus, both of which involved in the cognitive control of negative information processing. Individuals with high behavioral inhibition system displays low 5-HT
2A
receptor availability in anterior cingulate cortex and this cognitive control network links with prefrontal-cingulate integrity. These findings have implications for underlying the serotonergic basis of physiologies in aversion.
Although aberrations in the number and function of glutamate AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid) receptors are thought to underlie neuropsychiatric disorders, no methods are ...currently available for visualizing AMPA receptors in the living human brain. Here we developed a positron emission tomography (PET) tracer for AMPA receptors. A derivative of 4-2-(phenylsulfonylamino)ethylthio-2,6-difluoro-phenoxyacetamide radiolabeled with
C (
CK-2) showed specific binding to AMPA receptors. Our clinical trial with healthy human participants confirmed reversible binding of
CK-2 in the brain according to Logan graphical analysis (UMIN000020975; study design: non-randomized, single arm; primary outcome: dynamics and distribution volumes of
CK-2 in the brain; secondary outcome: adverse events of
CK-2 during the 4-10 d following dosing; this trial met prespecified endpoints). In an exploratory clinical study including patients with epilepsy, we detected increased
CK-2 uptake in the epileptogenic focus of patients with mesial temporal lobe epilepsy, which was closely correlated with the local AMPA receptor protein distribution in surgical specimens from the same individuals (UMIN000025090; study design: non-randomized, single arm; primary outcome: correlation between
CK-2 uptake measured with PET before surgery and AMPA receptor protein density examined by biochemical study after surgery; secondary outcome: adverse events during the 7 d following PET scan; this trial met prespecified endpoints). Thus,
CK-2 is a potent PET tracer for AMPA receptors, potentially providing a tool to examine the involvement of AMPA receptors in neuropsychiatric disorders.
Monoacylglycerol lipase (MAGL) is a cytosolic serine hydrolase involved in endocannabinoid and inflammatory signaling. Positron-emission tomography (PET) imaging of MAGL serves to validate target ...engagement of therapeutic MAGL inhibitors as well as to investigate MAGL levels under normal and disease conditions. However, PET radioligands with reversible binding kinetics for MAGL, which allow quantitative assessment of MAGL, are hitherto unavailable. In this study, we designed and synthesized fluoro-containing PET probes starting from a recently identified piperazinyl pyrrolidine-2-one derivative with reversible binding to MAGL. By tailoring the lipophilicity of the molecule to optimize nonspecific binding and blood–brain barrier permeability, we successfully identified two compounds that show high uptake to regions enriched with MAGL. PET imaging of wild-type and MAGL-deficient mice as well as a macaque monkey indicated that 18F5 ((4R)-1-{3-2-(18F)fluoro-4-methylpyridin-3-ylphenyl}-4-4-(1,3-thiazol-2-ylcarbonyl)piperazin-1-ylpyrrolidin-2-one, 18FT-401) specifically binds to MAGL with adequate reversibility, yielding a high contrast for MAGL within an appropriate imaging time.
•18F-PM-PBB3 PET captures diverse tau lesions in FTLD-tau as well as in AD.•We developed a new workflow to extract optimal reference regions using histograms.•Reference regions extracted from gray ...matter increased the sensitivity of FTLD-tau.•The methodology enabled accurate quantification of tau lesions in diverse illnesses.
Positron emission tomography (PET) with 18F-PM-PBB3 (18F-APN-1607, 18F-Florzolotau) enables high-contrast detection of tau depositions in various neurodegenerative dementias, including Alzheimer's disease (AD) and frontotemporal lobar degeneration (FTLD). A simplified method for quantifying radioligand binding in target regions is to employ the cerebellum as a reference (CB-ref) on the assumption that the cerebellum has minimal tau pathologies. This procedure is typically valid in AD, while FTLD disorders exemplified by progressive supranuclear palsy (PSP) are characterized by occasional tau accumulations in the cerebellum, hampering the application of CB-ref. The present study aimed to establish an optimal method for defining reference tissues on 18F-PM-PBB3-PET images of AD and non-AD tauopathy brains. We developed a new algorithm to extract reference voxels with a low likelihood of containing tau deposits from gray matter (GM-ref) or white matter (WM-ref) by a bimodal fit to an individual, voxel-wise histogram of the radioligand retentions and applied it to 18F-PM-PBB3-PET data obtained from age-matched 40 healthy controls (HCs) and 23 CE, 40 PSP, and five other tau-positive FTLD patients. PET images acquired at 90–110 min after injection were averaged and co-registered to corresponding magnetic resonance imaging space. Subsequently, we generated standardized uptake value ratio (SUVR) images estimated by CB-ref, GM-ref and WM-ref, respectively, and then compared the diagnostic performances. GM-ref and WM-ref covered a broad area in HCs and were free of voxels located in regions known to bear high tau burdens in AD and PSP patients. However, radioligand retentions in WM-ref exhibited age-related declines. GM-ref was unaffected by aging and provided SUVR images with higher contrast than CB-ref in FTLD patients with suspected and confirmed corticobasal degeneration. The methodology for determining reference tissues as optimized here improves the accuracy of 18F-PM-PBB3-PET measurements of tau burdens in a wide range of neurodegenerative illnesses.
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