[18F]Nifene test–retest reproducibility in first‐in‐human imaging of α4β2* nicotinic acetylcholine receptors

The aim of this study was to examine the suitability of [¹⁸ F]nifene, a novel α4β2* nicotinic acetylcholine receptor (nAChR) radiotracer, for in vivo brain imaging in a first‐in‐human study. Methods : Eight healthy subjects (4 M,4 F;21–69,44 ± 21 yrs) underwent a [¹⁸F]nifene positron emission tomography scan (200 ± 3.7 MBq), and seven underwent a second scan within 58 ± 31 days. Regional estimates of DVR were measured using the multilinear reference tissue model (MRTM2) with the corpus callosum as reference region. DVR reproducibility was evaluated with test–retest variability (TRV) and intraclass correlation coefficient (ICC). Results : The DVR ranged from 1.3 to 2.5 across brain regions with a TRV of 0–7%, and did not demonstrate a systematic difference between test and retest. The ICCs ranged from 0.2 to 0.9. DVR estimates were stable after 40 min. Conclusion : The binding profile and tracer kinetics of [¹⁸F]nifene make it a promising α4β2* nAChR radiotracer for scientific research in humans, with reliable DVR test–retest reproducibility.     

N-[18F]4'-fluorobenzylpiperidin-4yl-(2-fluorophenyl) acetamide ([18F]FBFPA): a potential fluorine-18 labeled PET radiotracer for imaging sigma-1 receptors in the CNS

A series of brain uptake studies and PET imaging studies were conducted with the sigma(1) selective imaging agent, [(18)F]FBFPA. The results of the study indicate that this radiotracer readily crosses the blood-brain barrier and labels sigma(1) receptors in vivo. In vivo blocking studies with a sigma(1) selective ligand and a nonselective sigma(1)/sigma(2) receptor ligand indicates that [(18)F]FBFPA labels sigma(1) and not sigma(2) receptors in rodent brain. PET imaging studies demonstrated a high uptake in regions of rhesus monkey brain having a high density of sigma(1) receptors. The uptake of [(18)F]FBFPA was displaced by the sigma ligand, haloperidol (1 mg/kg, i.v.). In vivo blocking studies indicate that the progesterone blocked the brain uptake of [(18)F]FBFPA in rat brain. These data indicate that [(18)F]FBFPA is a potential radiotracer for imaging sigma(1) receptors in the CNS in vivo with PET.     

A within‐subject comparison of 6‐[18F]fluoro‐m‐tyrosine and 6‐[18F]fluoro‐L‐dopa in Parkinson's disease

Progression of Parkinson's disease symptoms is imperfectly correlated with positron emission tomography biomarkers for dopamine biosynthetic pathways. The radiopharmaceutical 6‐[¹⁸F]fluoro‐m‐tyrosine is not a substrate for catechol‐O‐methyltransferase and therefore has a more favorable uptake‐to‐background ratio than 6‐[¹⁸F]fluoro‐L ‐dopa. The objective of this study was to evaluate 6‐[¹⁸F]fluoro‐m‐tyrosine relative to 6‐[¹⁸F]fluoro‐L ‐dopa with partial catechol‐O‐methyltransferase inhibition as a biomarker for clinical status in Parkinson's disease. Twelve patients with early‐stage Parkinson's disease, off medication, underwent Unified Parkinson Disease Rating Scale scoring, brain magnetic resonance imaging, and 3‐dimensional dynamic positron emission tomography using equivalent doses of 6‐[¹⁸F]fluoro‐m‐tyrosine and 6‐[¹⁸F]fluoro‐L ‐dopa with tolcapone, a catechol‐O‐methyltransferase inhibitor. Images were realigned within subject, after which the tissue‐derived uptake rate constant was generated for volumes of interest encompassing the caudate nucleus, putamen, and subregions of the putamen. We computed both bivariate (Pearson) and partial (covariate of age) correlations between clinical subscores and tissue‐derived uptake rate constant. Tissue‐derived uptake rate constant values were correlated between the radiopharmaceuticals (r = 0.8). Motor subscores were inversely correlated with the contralateral putamen 6‐[¹⁸F]fluoro‐m‐tyrosine tissue‐derived uptake rate constant (|r| > 0.72, P < .005) but not significantly with the 6‐[¹⁸F]fluoro‐L ‐dopa tissue‐derived uptake rate constant. The uptake rate constants for both radiopharmaceuticals were also inversely correlated with activities of daily living subscores, but the magnitude of correlation coefficients was greater for 6‐[¹⁸F]fluoro‐m‐tyrosine. In this design, 6‐[¹⁸F]fluoro‐m‐tyrosine uptake better reflected clinical status than did 6‐[¹⁸F]fluoro‐L ‐dopa uptake. We attribute this finding to 6‐[¹⁸F]fluoro‐m‐tyrosine's higher affinity for the target, L ‐aromatic amino acid decarboxylase, and the absence of other major determinants of the uptake rate constant. These results also imply that L ‐aromatic amino acid decarboxylase activity is a major determinant of clinical status. © 2011 Movement Disorder Society     

Quantitative imaging of neuroinflammation in human white matter: A positron emission tomography study with translocator protein 18 kDa radioligand, [18F]‐FEPPA

The ability to quantify translocator protein 18 kDa (TSPO) in white matter (WM) is important to understand the role of neuroinflammation in neurological disorders with WM involvement. This article aims to extend the utility of TSPO imaging in WM using a second‐generation radioligand, [¹⁸F]‐FEPPA, and high‐resolution research tomograph (HRRT) positron emission tomography (PET) camera system. Four WM regions of interests (WM‐ROI), relevant to the study of aging and neuroinflammatory diseases, were examined. The corpus callosum, cingulum bundle, superior longitudinal fasciculus, and posterior limb of internal capsule were delineated automatically onto subject's T₁‐weighted magnetic resonance image using a diffusion tensor imaging‐based WM template. The TSPO polymorphism (rs6971) stratified individuals to three genetic groups: high‐affinity binders (HAB), mixed‐affinity binders (MAB), and low‐affinity binders. [¹⁸F]‐FEPPA PET scans were acquired on 32 healthy subjects and analyzed using a full kinetic compartment analysis. The two‐tissue compartment model showed moderate identifiability (coefficient of variation 15–19%) for [¹⁸F]‐FEPPA total volume distribution (V_T) in WM‐ROIs. Noise affects V_T variability, although its effect on bias was small (6%). In a worst‐case scenario, ≤6% of simulated data did not fit reliably. A simulation of increased TSPO density exposed minimal effect on variability and identifiability of [¹⁸F]‐FEPPA V_T in WM‐ROIs. We found no association between age and [¹⁸F]‐FEPPA V_T in WM‐ROIs. The V_T values were 15% higher in HAB than in MAB, although the difference was not statistically significant. This study provides evidence for the utility and limitations of [¹⁸F]‐FEPPA PET to measure TSPO expression in WM. Synapse 68:536–547, 2014 . © 2014 Wiley Periodicals, Inc.     

Attenuation of β‐amyloid‐induced tauopathy via activation of CK2α/SIRT1: Targeting for cilostazol

β‐Amyloid (Aβ) deposits and hyperphosphorylated tau aggregates are the chief hallmarks in the Alzheimer's disease (AD) brains, but the strategies for controlling these pathological events remain elusive. We hypothesized that CK2‐coupled SIRT1 activation stimulated by cilostazol suppresses tau acetylation (Ac‐tau) and tau phosphorylation (P‐tau) by inhibiting activation of P300 and GSK3β. Aβ was endogenously overproduced in N2a cells expressing human APP Swedish mutation (N2aSwe) by exposure to medium containing 1% fetal bovine serum for 24 hr. Increased Aβ accumulation was accompanied by increased Ac‐tau and P‐tau levels. Concomitantly, these cells showed increased P300 and GSK3β P‐Tyr216 expression; their expressions were significantly reduced by treatment with cilostazol (3–30 μM) and resveratrol (20 μM). Moreover, decreased expression of SIRT1 and its activity by Aβ were significantly reversed by cilostazol as by resveratrol. In addition, cilostazol strongly stimulated CK2α phosphorylation and its activity, and then stimulated SIRT1 phosphorylation. These effects were confirmed by using the pharmacological inhibitors KT5720 (1 μM, PKA inhibitor), TBCA (20 μM, inhibitor of CK2), and sirtinol (20 μM, SIRT1 inhibitor) as well as by SIRT1 gene silencing and overexpression techniques. In conclusion, increased cAMP‐dependent protein kinase‐linked CK2/SIRT1 expression by cilostazol can be a therapeutic strategy to suppress the tau‐related neurodegeneration in the AD brain. © 2013 Wiley Periodicals, Inc.     

Presence of an endogenous factor which inhibits binding of α-bungarotoxin 2.2 to its receptor

Cerebral cortical membranes and supernatant from rat were prepared by centrifugation of tissue homogenates at 45,000 g for 10 min. The supernatant fraction thus obtained was found to significantly inhibit α-bungarotoxin binding to the membrane preparation. After a 3 min incubation period, the supernatant inhibited toxin binding by approximately 65%, while the inhibition declined to about 40% after 30 min of incubation, presumably due to the slow reversility of α-bungarotoxin binding. The choice of buffer was found to be an important determinant of the degree of inhibition observed, with 10 mM Tris pH 7.4 providing the most effective condition. This inhibition of toxin binding to cortical membranes by the 45,000 g supernatant was shown not to be due to adsorption of the radiolabeled compound to soluble or residual particulate material in the supernatant fraction. Specificity of the supernatant for the α-bungarotoxin site was demonstrated; a supernatant fr action could be prepared which inhibited α-bungarotoxin binding by 50% but had no effect on [³H]spiroperidol (DA₂ and 5-HT₂), [³H]prazosin, (α₁-adrenergic), [³H]5-hydroxytryptamine (5-HT₁) and [³H]quinuclidinylbenzilate (muscarinic cholinergic) binding. The inhibition of toxin binding also occurred in several other CNS regions including hippocampus, brainstem, spinal cord and cerebellum with an 80 to 90% inhibition of binding occuring in the latter two regions. In addition, the 45,000 g cortical supernatant completely prevented the binding of α-bungarotoxin to extrajunctional neuromuscular receptors and inhibited the binding to junctional receptors by 50%. Supernatants prepared from heart, liver and kidney or bovine serum albumin, at a concentration similar to the supernatant fraction, did not alter radiolabeled toxin binding to cortical membranes, while supernatant prepared from striated muscle tissue was effective. These results suggest there may be an endogenous ligand for the α-bungarotoxin 2.2 binding site in tissues which receive nicotinic cholinergic innervation.     

δ‐Catenin/NPRAP: A new member of the glycogen synthase kinase‐3β signaling complex that promotes β‐catenin turnover in neurons

Through a multiprotein complex, glycogen synthase kinase‐3β (GSK‐3β) phosphorylates and destabilizes β‐catenin, an important signaling event for neuronal growth and proper synaptic function. δ‐Catenin, or NPRAP (CTNND2), is a neural enriched member of the β‐catenin superfamily and is also known to modulate neurite outgrowth and synaptic activity. In this study, we investigated the possibility that δ‐catenin expression is also affected by GSK‐3β signaling and participates in the molecular complex regulating β‐catenin turnover in neurons. Immunofluorescent light microscopy revealed colocalization of δ‐catenin with members of the molecular destruction complex: GSK‐3β, β‐catenin, and adenomatous polyposis coli proteins in rat primary neurons. GSK‐3β formed a complex with δ‐catenin, and its inhibition resulted in increased δ‐catenin and β‐catenin expression levels. LY294002 and amyloid peptide, known activators of GSK‐3β signaling, reduced δ‐catenin expression levels. Furthermore, δ‐catenin immunoreactivity increased and protein turnover decreased when neurons were treated with proteasome inhibitors, suggesting that the stability of δ‐catenin, like that of β‐catenin, is regulated by proteasome‐mediated degradation. Coimmunoprecipitation experiments showed that δ‐catenin overexpression promoted GSK‐3β and β‐catenin interactions. Primary cortical neurons and PC12 cells expressing δ‐catenin treated with proteasome inhibitors showed increased ubiquitinated β‐catenin forms. Consistent with the hypothesis that δ‐catenin promotes the interaction of the destruction complex molecules, cycloheximide treatment of cells overexpressing δ‐catenin showed enhanced β‐catenin turnover. These studies identify δ‐catenin as a new member of the GSK‐3β signaling pathway and further suggest that δ‐catenin is potentially involved in facilitating the interaction, ubiquitination, and subsequent turnover of β‐catenin in neuronal cells. © 2010 Wiley‐Liss, Inc.     

3‐[(2,4‐dimethoxy)benzylidene]‐anabaseine dihydrochloride protects against 6‐hydroxydopamine‐induced parkinsonian neurodegeneration through α7 nicotinic acetylcholine receptor stimulation in rats

To explore a novel therapy against Parkinson's disease through enhancement of α7 nicotinic acetylcholine receptor (nAChR), we evaluated the neuroprotective effects of 3‐[(2,4‐dimethoxy)benzylidene]‐anabaseine dihydrochloride (DMXBA; GTS‐21), a functionally selective α7 nAChR agonist, in a rat 6‐hydroxydopamine (6‐OHDA)‐induced hemiparkinsonian model. Microinjection of 6‐OHDA into the nigrostriatal pathway of rats destroys dopaminergic neurons selectively. DMXBA dose dependently inhibited methamphetamine‐stimulated rotational behavior and dopaminergic neuronal loss induced by 6‐OHDA. The protective effects were abolished by methyllycaconitine citrate salt hydrate, an α7 nAChR antagonist. Immunohistochemical study confirmed abundant α7 nAChR expression in the cytoplasm of dopaminergic neurons. These results indicate that DMXBA prevented 6‐OHDA‐induced dopaminergic neuronal loss through stimulating α7 nAChR in dopaminergic neurons. Injection of 6‐OHDA elevated immunoreactivities to glial markers such as ionized calcium binding adaptor molecule 1, CD68, and glial fibrillary acidic protein in the substantia nigra pars compacta of rats. In contrast, these immunoreactivities were markedly inhibited by comicroinjection of DMXBA. Microglia also expressed α7 nAChR in both resting and activated states. Hence, we hypothesize that DMXBA simultaneously affects microglia and dopaminergic neurons and that both actions lead to dopaminergic neuroprotection. The findings that DMXBA attenuates 6‐OHDA‐induced dopaminergic neurodegeneration and glial activation in a rat model of Parkinson's disease raisethe possibility that DMXBA could be a novel therapeutic compound to prevent Parkinson's disease development. © 2012 Wiley Periodicals, Inc.     

α2‐Adrenergic drugs modulate the binding of [18F]fallypride to dopamine D2/3 receptors in striatum of living mouse

Aim. To test for α₂ adrenergic modulation of dopamine D_2/3 receptor availability in striatum of living mice using the high‐affinity ligand [¹⁸F]fallypride and microPET. Methods. Groups of anesthetized mice were pretreated with saline, the α₂‐agonist clonidine (1 mg/kg), and the α₂‐antagonists RX821002 (1 mg/kg) and yohimbine (1 mg/kg). Dynamic microPET recordings lasting 120 min were then initiated upon i.v. tracer injection of [¹⁸F]fallypride. Parametric maps of [¹⁸F]fallypride binding potential (BP_ND) were calculated using the Logan method, with cerebellum serving as the reference region. Results. Mean striatal [¹⁸F]fallypride BP_ND was 10.6 ± 1.7 in the saline control animals, 8.9 ± 1.7 (?16%; P < 0.05) in the RX821002 group, 8.3 ± 2.6 (?22%; P < 0.05) in the yohimbine group and 10.3 ± 2.2 (n.s.) in the clonidine group. Conclusions. These findings are consistent with a tonic inhibition of dopamine release by α₂ adrenergic receptors, such that α₂ blockade increased the competition from endogenous dopamine at D_2/3 receptors, thus reducing the [¹⁸F]fallypride BP_ND by about 20%. Absent effects of clonidine suggest a ceiling effect in the tonic inhibition of dopamine release. This in vivo PET evidence for α₂/dopaminergic interaction may be relevant to putative actions of atypical antipsychotic medications via adrenergic receptors. Synapse, 2010. © 2010 Wiley‐Liss, Inc.     

Methamphetamine‐induced up‐regulation of α2/δ subunit of voltage‐gated calcium channels is regulated by DA receptors

This study was carried out to determine the roles of dopamine D1 and D2 receptors on the up‐regulation of α₂/δ subunit of voltage‐gated Ca²⁺ channels (VGCCs) induced by methamphetamine (METH). In the conditioned place preference paradigm, METH‐induced place preference suppressed with gabapentin, an antagonist for α₂/δ subunit. Under these conditions, the increase in α₂/δ subunit expression was found in the frontal cortex and limbic forebrain. In addition, the METH‐induced place preference was significantly attenuated by dopamine D1 and D2 receptor antagonists, SCH23390 and sulpiride, respectively. The expression of α₂/δ subunit protein and its mRNA was significantly enhanced in the METH‐treated cortical neurons. These increases in protein and mRNA of α₂/δ subunit were completely abolished by SCH23390 and sulpiride with simultaneous exposure to METH. These findings indicate that up‐regulation of α₂/δ subunit is regulated through the activation of dopamine D1 and D2 receptors during METH treatment. Synapse 64:822–828, 2010. © 2010 Wiley‐Liss, Inc.     

γ‐Secretase binding sites in aged and Alzheimer's disease human cerebrum: the choroid plexus as a putative origin of CSF Aβ

Deposition of β ‐amyloid (Aβ) peptides, cleavage products of β‐amyloid precursor protein (APP) by β‐secretase‐1 (BACE1) and γ‐secretase, is a neuropathological hallmark of Alzheimer's disease (AD). γ‐Secretase inhibition is a therapeutical anti‐Aβ approach, although changes in the enzyme's activity in AD brain are unclear. Cerebrospinal fluid (CSF) Aβ peptides are thought to derive from brain parenchyma and thus may serve as biomarkers for assessing cerebral amyloidosis and anti‐Aβ efficacy. The present study compared active γ‐secretase binding sites with Aβ deposition in aged and AD human cerebrum, and explored the possibility of Aβ production and secretion by the choroid plexus (CP). The specific binding density of [³H]‐L‐685,458, a radiolabeled high‐affinity γ‐secretase inhibitor, in the temporal neocortex and hippocampal formation was similar for AD and control cases with similar ages and post‐mortem delays. The CP in post‐mortem samples exhibited exceptionally high [³H]‐L‐685,458 binding density, with the estimated maximal binding sites (Bmax) reduced in the AD relative to control groups. Surgically resected human CP exhibited APP, BACE1 and presenilin‐1 immunoreactivity, and β‐site APP cleavage enzymatic activity. In primary culture, human CP cells also expressed these amyloidogenic proteins and released Aβ40 and Aβ42 into the medium. Overall, our results suggest that γ‐secretase activity appears unaltered in the cerebrum in AD and is not correlated with regional amyloid plaque pathology. The CP appears to be a previously unrecognised non‐neuronal contributor to CSF Aβ, probably at reduced levels in AD.     

Human brain serotonin synthesis capacity measured in vivo with α-[C-11]methyl-L-tryptophan

Local cerebral serotonin synthesis capacity was measured with α-[C-11]methyl-L-tryptophan ([C-11]AMT) in normal adult human brain (n = 10; five males, five females; age range, 18–38 years, mean 28.3 years) by using positron emission tomography (PET). [C-11]AMT is an analog of tryptophan, the precursor for serotonin synthesis, and is converted to α-[C-11]methyl-serotonin ([C-11]AM-5HT), which is trapped in serotonergic neurons because [C-11]AM-5HT is not degraded by monoamine oxidase. Kinetic analysis of [C-11] activity in brain after injection of [C-11]AMT confirmed the presence of a compartment with unidirectional uptake that represented approximately 40% of the activity in the brain at 50 min after tracer administration. The undirectional rate constant K, which represents the uptake of [C-11]AMT from the plasma to brain tissue followed by the synthesis and physiologic trapping of [C-11]AM-5HT, was calculated using the Patlak graphic approach on a pixel-by-pixel basis, thus creating parametric images. The rank order of K values for different brain regions corresponded well to the regional concentrations of serotonin in human brain (P < .0001). High serotonin synthesis capacity values were measured in putamen, caudate, thalamus, and hippocampus. Among cortical regions, the highest values were measured in the rectal gyrus of the inferior frontal lobe, followed by transverse temporal gyrus; anterior and posterior cingulate gyrus; middle, superior, and inferior temporal gyri; parietal cortex; occipital cortex, in descending order. Values in women were 10–20% higher (P < .05, MANOVA) throughout the brain than those measured in men. Differences in the serotonin synthesis capacity between men and women measured in this study may reflect gender differences of importance to both normal and pathologic behavior. This study demonstrates the suitability of [C-11]AMT as a tracer for PET scanning of serotonin synthesis capacity in human brain and provides normal adult values for future comparison with patient groups. Synapse 28:33–43, 1998. © 1998 Wiley-Liss, Inc.