Steven Deleye

Towards a reproducible protocol for repetitive and semi-quantitative rat brain imaging with (18) F-FDG: exemplified in a memantine pharmacological challenge.

The standard uptake value (SUV), commonly used to quantify (18)F-FluoroDeoxyGlucose (FDG) uptake in small animal brain PET imaging, is affected by many factors. In this study the influence of fasting times, inter-scan duration and repetitive scanning on the variability of different SUV measures is investigated. Additionally it is demonstrated that these variables could adversely influence the outcome of a pharmacological challenge when not accounted for. Naive Sprague-Dawley rats (n=20) were randomly divided into five different fasting groups (no fasting up to 24h of fasting). SUV brain uptake values were reproducible in naive animals when a fasting period of at least 12h is used and for shorter fasting periods SUV values need to be corrected for the glucose level. Additionally, a separate animal group (n=6) was sufficiently fasted for 16h and in a longitudinal setting being scanned six times in three weeks. Especially with short inter-scan durations, increasing glucose levels were found over time which was attributed to increased stress due to repeated food deprivation, altered food intake or scan manipulations. As a result, even with controlled and sufficient fasting, blood glucose levels should be taken into account for data quantification. Strikingly, even the brain activation effects of an NMDA-antagonist challenge with memantine could not be detected in experiments with a short inter-scan duration if glucose levels were not taken into account. Correcting for glucose levels decreases the inter- and intra-animal variability for rat brain imaging. SUV corrected for glucose levels yields the lowest inter-animal variation. However, if the body weight changes significantly, as in a long experiment, quantification based on the glucose corrected percentage injected dose (and not SUV) is recommendable as this yields the lowest intra-animal variation.

Colonoscopy and µPET/CT are valid techniques to monitor inflammation in the adoptive transfer colitis model in mice.

Background:Preclinical in vivo research on inflammatory bowel diseases requires proper animal models and techniques allowing longitudinal monitoring of colonic inflammation without the need to kill animals. We evaluated colonoscopy and &mgr;-positron emission tomography/computed tomography (&mgr;PET/CT) as monitoring tools in a model for chronic colitis in mice. Methods:Colitis was induced by adoptive transfer of CD4+CD25−CD62L+ T cells in immunocompromised severe combined immunodeficient mice. Three study protocols were designed. In study 1, colonoscopy and µPET/CT were performed once, 4 weeks after transfer. In study 2 and study 3, colitis was sequentially followed up through colonoscopy (study 2) or colonoscopy plus µPET/CT (study 3). Each study included postmortem evaluation of colonic inflammation (macroscopy, microscopy, and myeloperoxidase activity). Results:In study 1, both colonoscopy and µPET/CT detected colitis 4 weeks after transfer. Study 2 showed a gradual increase in colonoscopic score from week 2 (1.4 ± 0.6) to week 8 (6.0 ± 1.1). In study 3, colitis was detected 2 weeks after transfer by µPET/CT (2.0 ± 0.4) but not by colonoscopy, whereas both techniques detected inflammation 4 and 6 weeks after transfer. Colonoscopy correlated with µPET/CT (r = 0.812, 0.884, and 0.781, respectively) and with postmortem analyses in all 3 studies. Conclusions:Adoptive transfer of CD4+CD25−CD62L+ T cells in severe combined immunodeficient mice results in a moderate chronic colitis. Evolution of colitis could be monitored over time by both colonoscopy and µPET/CT. µPET/CT seems to detect inflammation at an earlier time point than colonoscopy. Both techniques represent reliable and safe methods without the need to kill animals.

Neural Substrates of Conversion Deafness in a Cochlear Implant Patient: A Molecular Imaging Study Using H215O-PET

Objective Conversion deafness is characterized by sudden hearing loss without any identifiable cause. In the current study, we investigated presumed conversion deafness in a cochlear implant user using H215O–positron emission tomography (PET) scan with speech and noise stimuli in conjunction with audiologic tests such as impedance test and auditory response telemetry. Also, by performing a follow-up PET scan after recovery and comparing prerecovery and postrecovery scans, we attempted to find possible neural substrates of conversion deafness. Patient A 51-year-old man with conversion deafness after 4 years of successful cochlear implant use. Intervention Supportive psychotherapy. Main Outcome Measures Prerecovery and postrecovery H215O-PET scans Results The prerecovery H215O-PET scan revealed auditory cortex activation by sound stimuli, which verified normal stimulation of the central auditory pathway. Notably, compared with the prerecovery state, the postrecovery state showed relative activation in the right auditory cortex both under the speech and noise stimulus conditions. Moreover, the bilateral prefrontal and parietal areas were activated more in the postrecovery state than in the prerecovery state. In other words, relative deactivation of the prefronto-parieto-temporal network, a network responsible for conscious sensory perception, or relative dysfunction of top-down and bottom-up attention shifting mediated by the ventral and the dorsal parietal cortices, may have resulted in conversion deafness in the patient. Conclusion Relative deactivation of the prefronto-parieto-temporal network or dysfunction in the ventral and the dorsal parietal cortices may be related to the development of conversion deafness.

The Effects of Physiological and Methodological Determinants on 18F-FDG Mouse Brain Imaging Exemplified in a Double Transgenic Alzheimer Model

Introduction: In this study, the influence of physiological determinants on 18F-fluoro-d-glucose (18F-FDG) brain uptake was evaluated in a mouse model of Alzheimer disease. Materials and Methods: TASTPM (Tg) and age-matched C57BL/6 J (WT) mice were fasted for 10 hours, while another group was fasted for 20 hours to evaluate the effect of fasting duration. The effect of repeatedly scanning was evaluated by scanning Tg and WT mice at days 1, 4, and 7. Brain 18F-FDG uptake was evaluated in the thalamus being the most indicative region. Finally, the cerebellum was tested as a reference region for the relative standard uptake value (rSUV). Results: When correcting the brain uptake for glucose, the effect of different fasting durations was attenuated and the anticipated hypometabolism in Tg mice was demonstrated. Also, with repeated scanning, the brain uptake values within a group and the hypometabolism of the Tg mice only remained stable over time when glucose correction was applied. Finally, hypometabolism was also observed in the cerebellum, yielding artificially higher rSUV values for Tg mice. Conclusion: Corrections for blood glucose levels have to be applied when semiquantifying 18F-FDG brain uptake in mouse models for AD. Potential reference regions for normalization should be thoroughly investigated to ensure that they are not pathologically affected also by afferent connections.

Continuous flushing of the bladder in rodents reduces artifacts and improves quantification in molecular imaging.

In this study, we evaluated the partial volume effect (PVE) of 2-deoxy-2-[18F]fluoro-D-glucose (18F-FDG) tracer accumulation in the bladder on the positron emission tomographic (PET) image quantification in mice and rats suffering from inflammatory bowel disease. To improve the accuracy, we implemented continuous bladder flushing procedures. Female mice and rats were scanned using microPET/computed tomography (CT) at baseline and after induction of acute colitis by injecting 2,4,6-trinitrobenzene sulfonic acid (TNBS) intrarectally. During the scans, the bladder was continuously flushed in one group, whereas in the other group, no bladder flushing was performed. As a means of in vivo and ex vivo validation of the inflammation, animals also underwent colonoscopy and were sacrificed for gamma counting (subpopulation) and to score the colonic damage both micro- and macroscopically as well as biochemically. At baseline, the microPET signal in the colon of both mice and rats was significantly higher in the nonflushed group compared to the flushed group, caused by the PVE of tracer activity in the bladder. Hence, the colonoscopy and postmortem analyses showed no significant differences at baseline between the flushed and nonflushed animals. TNBS induced significant colonic inflammation, as revealed by colonoscopic and postmortem scores, which was not detected by microPET in the mice without bladder flushing, again because of spillover of bladder activity in the colonic area. MicroPET in bladder-flushed animals did reveal a significant increase in 18F-FDG uptake. Correlations between microPET and colonoscopy, macroscopy, microscopy, and myeloperoxidase yielded higher Spearman rho values in mice with continuously flushed bladders during imaging. Comparable, although somewhat less pronounced, results were shown in the rat. Continuous bladder flushing reduced image artifacts and is mandatory for accurate image quantification in the pelvic region for both mice and rats. We designed and validated experimental protocols to facilitate such.In this study, we evaluated the partial volume effect (PVE) of 2-deoxy-2-[18F]fluoro-D-glucose (18F-FDG) tracer accumulation in the bladder on the positron emission tomographic (PET) image quantification in mice and rats suffering from inflammatory bowel disease. To improve the accuracy, we implemented continuous bladder flushing procedures. Female mice and rats were scanned using microPET/computed tomography (CT) at baseline and after induction of acute colitis by injecting 2,4,6-trinitrobenzene sulfonic acid (TNBS) intrarectally. During the scans, the bladder was continuously flushed in one group, whereas in the other group, no bladder flushing was performed. As a means of in vivo and ex vivo validation of the inflammation, animals also underwent colonoscopy and were sacrificed for gamma counting (subpopulation) and to score the colonic damage both micro- and macroscopically as well as biochemically. At baseline, the microPET signal in the colon of both mice and rats was significantly higher in the nonflushed group compared to the flushed group, caused by the PVE of tracer activity in the bladder. Hence, the colonoscopy and postmortem analyses showed no significant differences at baseline between the flushed and nonflushed animals. TNBS induced significant colonic inflammation, as revealed by colonoscopic and postmortem scores, which was not detected by microPET in the mice without bladder flushing, again because of spillover of bladder activity in the colonic area. MicroPET in bladder-flushed animals did reveal a significant increase in 18F-FDG uptake. Correlations between microPET and colonoscopy, macroscopy, microscopy, and myeloperoxidase yielded higher Spearman rho values in mice with continuously flushed bladders during imaging. Comparable, although somewhat less pronounced, results were shown in the rat. Continuous bladder flushing reduced image artifacts and is mandatory for accurate image quantification in the pelvic region for both mice and rats. We designed and validated experimental protocols to facilitate such.

Validation of the semi-quantitative static SUVR method for [18F]-AV45 PET by pharmacokinetic modeling with an arterial input function

Increased brain uptake of 18F-AV45 visualized by PET is a key biomarker for Alzheimer disease (AD). The SUV ratio (SUVR) is widely used for quantification, but is subject to variability based on choice of reference region and changes in cerebral blood flow. Here we validate the SUVR method against the gold standard volume of distribution (VT) to assess cross-sectional differences in plaque load. Methods: Dynamic 60-min 18F-AV45 (291 ± 67 MBq) and 1-min 15O-H2O (370 MBq) scans were obtained in 35 age-matched elderly subjects, including 10 probable AD, 15 amnestic mild cognitive impairment (aMCI), and 10 cognitively healthy controls (HCs). 18F-AV45 VT was determined from 2-tissue-compartment modeling using a metabolite-corrected plasma input function. Static SUVR was calculated at 50–60 min after injection, using either cerebellar gray matter (SUVRCB) or whole subcortical white matter (SUVRWM) as the reference. Additionally, whole cerebellum, pons, centrum semiovale, and a composite region were examined as alternative references. Blood flow was quantified by 15O-H2O SUV. Data are presented as mean ± SEM. Results: There was rapid metabolization of 18F-AV45, with only 35% of unchanged parent remaining at 10 min. Compared with VT, differences in cortical Aβ load between aMCI and AD were overestimated by SUVRWM (+4% ± 2%) and underestimated by SUVRCB (−10% ± 2%). VT correlated better with SUVRWM (Pearson r: from 0.63 for posterior cingulate to 0.89 for precuneus, P < 0.0001) than with SUVRCB (Pearson r: from 0.51 for temporal lobe [P = 0.002] to 0.82 for precuneus [P < 0.0001]) in all tested regions. Correlation results for the alternative references were in between those for CB and WM. 15O-H2O data showed that blood flow was decreased in AD compared with aMCI in cortical regions (−5% ± 1%) and in the reference regions (CB, −9% ± 8%; WM, −8% ± 8%). Conclusion: Increased brain uptake of 18F-AV45 assessed by the simplified static SUVR protocol does not truly reflect Aβ load. However, SUVRWM is better correlated with VT and more closely reflects VT differences between aMCI and AD than SUVRCB.

Noninvasive Whole-Body Imaging of Phosphatidylethanolamine as a Cell Death Marker Using 99mTc-Duramycin During TNF-Induced SIRS

Systemic inflammatory response syndrome (SIRS) is an inflammatory state affecting the whole body. It is associated with the presence of pro- and antiinflammatory cytokines in serum, including tumor necrosis factor (TNF). TNF has multiple effects and leads to cytokine production, leukocyte infiltration, and blood pressure reduction and coagulation, thereby contributing to tissue damage and organ failure. A sterile mouse model of sepsis, TNF-induced SIRS, was used to visualize the temporal and spatial distribution of damage in susceptible tissues during SIRS. For this, a radiopharmaceutical agent, 99mTc-duramycin, that binds to exposed phosphatidylethanolamine on dying cells was longitudinally visualized using SPECT/CT imaging. Methods: C57BL/6J mice were challenged with intravenous injections of murine TNF or vehicle, and necrostatin-1 was used to interfere with cell death. Two hours after vehicle or TNF treatment, mice received 99mTc-duramycin intravenously (35.44 ± 3.80 MBq). Static whole-body 99mTc-duramycin SPECT/CT imaging was performed 2, 4, and 6 h after tracer injection. Tracer uptake in different organs was quantified by volume-of-interest analysis using PMOD software and expressed as SUVmean. After the last scan, ex vivo biodistribution was performed to validate the SPECT imaging data. Lastly, terminal deoxynucleotidyl-transferase–mediated deoxyuridine triphosphate nick-end labeling (TUNEL) staining was performed to correlate the obtained results to cell death. Results: An increased 99mTc-duramycin uptake was detected in mice injected with TNF, when compared with control mice, in lungs (0.55 ± 0.1 vs. 0.34 ± 0.05), intestine (0.75 ± 0.13 vs. 0.56 ± 0.1), and liver (1.03 ± 0.14 vs. 0.64 ± 0.04) 4 h after TNF and remained significantly elevated until 8 h after TNF. The imaging results were consistent with ex vivo γ-counting results. Significantly increased levels of tissue damage were detected via TUNEL staining in the lungs and intestine of mice injected with TNF. Interestingly, necrostatin-1 pretreatment conferred protection against lethal SIRS and reduced the 99mTc-duramycin uptake in the lungs 8 h after TNF (SUV, 0.32 ± 0.1 vs. 0.51 ± 0.15). Conclusion: This study demonstrated that noninvasive 99mTc-duramycin SPECT imaging can be used to characterize temporal and spatial kinetics of injury and cell death in susceptible tissues during TNF-induced SIRS, making it useful for global, whole-body assessment of tissue damage during diseases associated with inflammation and injury.

[18F]PBR111 PET Imaging in Healthy Controls and Schizophrenia: Test – Retest Reproducibility and Quantification of Neuroinflammation

Activated microglia express the translocator protein (TSPO) on the outer mitochondrial membrane. 18F-PBR111 is a second-generation PET ligand that specifically binds the TSPO, allowing in vivo visualization and quantification of neuroinflammation. The aim of this study was to evaluate whether the test–retest variability of 18F-PBR111 in healthy controls is acceptable to detect a psychosis-associated neuroinflammatory signal in schizophrenia. Methods: Dynamic 90-min 18F-PBR111 scans were obtained in 17 healthy male controls (HCs) and 11 male schizophrenia patients (SPs) during a psychotic episode. Prior genotyping for the rs6917 polymorphism distinguished high-affinity binders (HABs) and mixed-affinity binders (MABs). Total volume of distribution (VT) was determined from 2-tissue-compartment modeling with vascular trapping and a metabolite-corrected plasma input function. A subgroup of HCs (n = 12; 4 HABs and 8 MABs) was scanned twice to assess absolute test–retest variability and intraclass correlation coefficients of the regional VT values. Differences in TSPO binding between HC and SP were assessed using mixed model analysis adjusting for age, genotype, and age*cohort. The effect of using different scan durations (VT-60 min versus VT-90 min) was determined based on Pearson r. Data were mean ± SD. Results: Mean absolute variability in VT ranged from 16% ± 14% (19% ± 20% HAB; 15% ± 11% MAB) in the cortical gray matter to 22% ± 15% (23% ± 15% HAB; 22% ± 16% MAB) in the hippocampus. Intraclass correlation coefficients were consistently between 0.64 and 0.82 for all tested regions. TSPO binding in SP compared with HC depended on age (cohort*age: P < 0.05) and was increased by +14% ± 4% over the regions. There was a significant effect of genotype on TSPO binding, and VT of HABs was 31% ± 8% (HC: 17% ± 5%, SP: 61% ± 14%) higher than MABs. Across all clinical groups, VT-60 min and VT-90 min were strongly correlated (r > 0.7, P < 0.0001). Conclusion: 18F-PBR111 can be used for monitoring of TSPO binding, as shown by medium test–retest variability and reliability of VT in HCs. Microglial activation is present in SPs depending on age and needs to be adjusted for genotype.

Evaluation of μPET outcome measures to detect disease modification induced by BACE inhibition in a transgenic mouse model of Alzheimer’s disease.

In this study, we investigated the effects of chronic administration of an inhibitor of the β-site amyloid precursor protein-cleaving enzyme 1 (BACE1) on Alzheimer-related pathology by multitracer PET imaging in transgenic APPPS1-21 (TG) mice. Methods: Wild-type (WT) and TG mice received vehicle or BACE inhibitor (60 mg/kg) starting at 7 wk of age. Outcome measures of brain metabolism, neuroinflammation, and amyloid-β pathology were obtained through small-animal PET imaging with 18F-FDG, 18F-peripheral benzodiazepine receptor (18F-PBR), and 18F-florbetapir (18F-AV45), respectively. Baseline scans were acquired at 6–7 wk of age and follow-up scans at 4, 7, and 12 mo. 18F-AV45 uptake was measured at 8 and 13 mo of age. After the final scans, histologic measures of amyloid-β (4G8), microglia (ionized calcium binding adaptor molecule 1), astrocytes (glial fibrillary acidic protein), and neuronal nuclei were performed. Results: TG mice demonstrated significant age-associated increases in 18F-AV45 uptake. An effect of treatment was observed in the cortex (P = 0.0014), hippocampus (P = 0.0005), and thalamus (P < 0.0001). Histology confirmed reduction of amyloid-β pathology in TG-BACE mice. Regardless of treatment, TG mice demonstrated significantly lower 18F-FDG uptake than WT mice in the thalamus (P = 0.0004) and hippocampus (P = 0.0332). Neuronal nucleus staining was lower in both TG groups in the thalamus and cortex. 18F-PBR111 detected a significant age-related increase in TG mice (P < 0.0001) but did not detect the treatment-induced reduction in activated microglia as demonstrated by histology. Conclusion: Although 18F-FDG, 18F-PBR111, and 18F-AV45 all detected pathologic alterations between TG and WT mice, only 18F-AV45 could detect an effect of BACE inhibitor treatment. However, changes in WT binding of 18F-AV45 undermine the specificity of this effect.

A three-dimensional digital neurological atlas of the mustached bat (Pteronotus parnellii)

&NA; Substantial knowledge of auditory processing within mammalian nervous systems emerged from neurophysiological studies of the mustached bat (Pteronotus parnellii). This highly social and vocal species retrieves precise information about the velocity and range of its targets through echolocation. Such high acoustic processing demands were likely the evolutionary pressures driving the over‐development at peripheral (cochlea), metencephalic (cochlear nucleus), mesencephalic (inferior colliculus), diencephalic (medial geniculate body of the thalamus), and telencephalic (auditory cortex) auditory processing levels in this species. Auditory researchers stand to benefit from a three dimensional brain atlas of this species, due to its considerable contribution to auditory neuroscience. Our MRI‐based atlas was generated from 2 sets of image data of an ex‐vivo male mustached bats brain: a detailed 3D‐T2‐weighted‐RARE scan [(59 × 63 x 85) &mgr;m3] and track density images based on super resolution diffusion tensor images [(78) &mgr;m3] reconstructed from a set of low resolution diffusion weighted images using Super‐Resolution‐Reconstruction (SRR). By surface‐rendering these delineations and extrapolating from cortical landmarks and data from previous studies, we generated overlays that estimate the locations of classic functional subregions within mustached bat auditory cortex. This atlas is freely available from our website and can simplify future electrophysiological, microinjection, and neuroimaging studies in this and related species.