Barry J. Bedell

MicroPET imaging of 5-HT1A receptors in rat brain: a test–retest [18F]MPPF study

PurposeEarlier studies have shown that positron emission tomography (PET) imaging with the radioligand [18F]MPPF allows for measuring the binding potential of serotonin 5-hydroxytryptamine1A (5-HT1A) receptors in different regions of animal and human brain, including that of 5-HT1A autoreceptors in the raphe nuclei. In the present study, we sought to determine if such data could be obtained in rat, with a microPET (R4, Concorde Microsystems).MethodsScans from isoflurane-anaesthetised rats (nu2009=u200918, including six test–retest) were co-registered with magnetic resonance imaging data, and binding potential, blood to plasma ratio and radiotracer efflux were estimated according to a simplified reference tissue model.ResultsValues of binding potential for hippocampus (1.2), entorhinal cortex (1.1), septum (1.1), medial prefrontal cortex (1.0), amygdala (0.8), raphe nuclei (0.6), paraventricular hypothalamic nucleus (0.5) and raphe obscurus (0.5) were comparable to those previously measured with PET in cats, non-human primates or humans. Test–retest variability was in the order of 10% in the larger brain regions (hippocampus, medial prefrontal and entorhinal cortex) and less than 20% in small nuclei such as the septum and the paraventricular hypothalamic, basolateral amygdaloid and raphe nuclei.ConclusionsMicroPET brain imaging of 5-HT1A receptors with [18F]MPPF thus represents a promising avenue for investigating 5-HT1A receptor function in rat.

Metabolic brain activity suggestive of persistent pain in a rat model of neuropathic pain.

Persistent pain is a central characteristic of neuropathic pain conditions in humans. Knowing whether rodent models of neuropathic pain produce persistent pain is therefore crucial to their translational applicability. We investigated the spared nerve injury (SNI) model of neuropathic pain and the formalin pain model in rats using positron emission tomography (PET) with the metabolic tracer [18F]fluorodeoxyglucose (FDG) to determine if there is ongoing brain activity suggestive of persistent pain. For the formalin model, under brief anesthesia we injected one hindpaw with 5% formalin and the FDG tracer into a tail vein. We then allowed the animals to awaken and observed pain behavior for 30min during the FDG uptake period. The rat was then anesthetized and placed in the scanner for static image acquisition, which took place between minutes 45 and 75 post-tracer injection. A single reference rat brain magnetic resonance image (MRI) was used to align the PET images with the Paxinos and Watson rat brain atlas. Increased glucose metabolism was observed in the somatosensory region associated with the injection site (S1 hindlimb contralateral), S1 jaw/upper lip and cingulate cortex. Decreases were observed in the prelimbic cortex and hippocampus. Second, SNI rats were scanned 3weeks post-surgery using the same scanning paradigm, and region-of-interest analyses revealed increased metabolic activity in the contralateral S1 hindlimb. Finally, a second cohort of SNI rats was scanned while anesthetized during the tracer uptake period, and the S1 hindlimb increase was not observed. Increased brain activity in the somatosensory cortex of SNI rats resembled the activity produced with the injection of formalin, suggesting that the SNI model may produce persistent pain. The lack of increased activity in S1 hindlimb with general anesthetic demonstrates that this effect can be blocked, as well as highlights the importance of investigating brain activity in awake and behaving rodents.

High dietary folate in pregnant mice leads to pseudo-MTHFR deficiency and altered methyl metabolism, with embryonic growth delay and short-term memory impairment in offspring.

Abstract Methylenetetrahydrofolate reductase (MTHFR) generates methyltetrahydrofolate for methylation reactions. Severe MTHFR deficiency results in homocystinuria and neurologic impairment. Mild MTHFR deficiency (677C > T polymorphism) increases risk for complex traits, including neuropsychiatric disorders. Although low dietary folate impacts brain development, recent concerns have focused on high folate intake following food fortification and increased vitamin use. Our goal was to determine whether high dietary folate during pregnancy affects brain development in murine offspring. Female mice were placed on control diet (CD) or folic acid‐supplemented diet (FASD) throughout mating, pregnancy and lactation. Three‐week‐old male pups were evaluated for motor and cognitive function. Tissues from E17.5 embryos, pups and dams were collected for choline/methyl metabolite measurements, immunoblotting or gene expression of relevant enzymes. Brains were examined for morphology of hippocampus and cortex. Pups of FASD mothers displayed short‐term memory impairment, decreased hippocampal size and decreased thickness of the dentate gyrus. MTHFR protein levels were reduced in FASD pup livers, with lower concentrations of phosphocholine and glycerophosphocholine in liver and hippocampus, respectively. FASD pup brains showed evidence of altered acetylcholine availability and Dnmt3a mRNA was reduced in cortex and hippocampus. E17.5 embryos and placentas from FASD dams were smaller. MTHFR protein and mRNA were reduced in embryonic liver, with lower concentrations of choline, betaine and phosphocholine. Embryonic brain displayed altered development of cortical layers. In summary, high folate intake during pregnancy leads to pseudo‐MTHFR deficiency, disturbed choline/methyl metabolism, embryonic growth delay and memory impairment in offspring. These findings highlight the unintended negative consequences of supplemental folic acid.

A test-retest study on Parkinson's PPMI dataset yields statistically significant white matter fascicles

In this work, we propose a diffusion MRI protocol for mining Parkinsons disease diffusion MRI datasets and recover robust disease-specific biomarkers. Using advanced high angular resolution diffusion imaging (HARDI) crossing fiber modeling and tractography robust to partial volume effects, we automatically dissected 50 white matter (WM) fascicles. These fascicles connect deep nuclei (thalamus, putamen, pallidum) to different cortical functional areas (associative, motor, sensorimotor, limbic), basal forebrain and substantia nigra. Then, among these 50 candidate WM fascicles, only the ones that passed a test-retest reproducibility procedure qualified for further tractometry analysis. Leveraging the unique 2-timepoints test-retest Parkinsons Progression Markers Initiative (PPMI) dataset of over 600 subjects, we found statistically significant differences in tract profiles along the subcortico-cortical pathways between Parkinsons disease patients and healthy controls. In particular, significant increases in FA, apparent fiber density, tract-density and generalized FA were detected in some locations of the nigro-subthalamo-putaminal-thalamo-cortical pathway. This connection is one of the major motor circuits balancing the coordination of motor output. Detailed and quantifiable knowledge on WM fascicles in these areas is thus essential to improve the quality and outcome of Deep Brain Stimulation, and to target new WM locations for investigation.

Modulation of glucose metabolism and metabolic connectivity by β-amyloid.

Glucose hypometabolism in the pre-clinical stage of Alzheimer’s disease (AD) has been primarily associated with the APOE ɛ4 genotype, rather than fibrillar β-amyloid. In contrast, aberrant patterns of metabolic connectivity are more strongly related to β-amyloid burden than APOE ɛ4 status. A major limitation of previous studies has been the dichotomous classification of subjects as amyloid-positive or amyloid-negative. Dichotomous treatment of a continuous variable, such as β-amyloid, potentially obscures the true relationship with metabolism and reduces the power to detect significant changes in connectivity. In the present work, we assessed alterations of glucose metabolism and metabolic connectivity as continuous function of β-amyloid burden using positron emission tomography scans from the Alzheimer’s Disease Neuroimaging Initiative study. Modeling β-amyloid as a continuous variable resulted in better model fits and improved power compared to the dichotomous model. Using this continuous model, we found that both APOE ɛ4 genotype and β-amyloid burden are strongly associated with glucose hypometabolism at early stages of Alzheimer’s disease. We also determined that the cumulative effects of β-amyloid deposition result in a particular pattern of altered metabolic connectivity, which is characterized by global, synchronized hypometabolism at early stages of the disease process, followed by regionally heterogeneous, progressive hypometabolism.

Mesocorticolimbic Connectivity and Volumetric Alterations in DCC Mutation Carriers

The axon guidance cue receptor DCC (deleted in colorectal cancer) plays a critical role in the organization of mesocorticolimbic pathways in rodents. To investigate whether this occurs in humans, we measured (1) anatomical connectivity between the substantia nigra/ventral tegmental area (SN/VTA) and forebrain targets, (2) striatal and cortical volumes, and (3) putatively associated traits and behaviors. To assess translatability, morphometric data were also collected in Dcc-haploinsufficient mice. The human volunteers were 20 DCC+/− mutation carriers, 16 DCC+/+ relatives, and 20 DCC+/+ unrelated healthy volunteers (UHVs; 28 females). The mice were 11 Dcc+/− and 16 wild-type C57BL/6J animals assessed during adolescence and adulthood. Compared with both control groups, the human DCC+/− carriers exhibited the following: (1) reduced anatomical connectivity from the SN/VTA to the ventral striatum [DCC+/+: p = 0.0005, r(effect size) = 0.60; UHV: p = 0.0029, r = 0.48] and ventral medial prefrontal cortex (DCC+/+: p = 0.0031, r = 0.53; UHV: p = 0.034, r = 0.35); (2) lower novelty-seeking scores (DCC+/+: p = 0.034, d = 0.82; UHV: p = 0.019, d = 0.84); and (3) reduced striatal volume (DCC+/+: p = 0.0009, d = 1.37; UHV: p = 0.0054, d = 0.93). Striatal volumetric reductions were also present in Dcc+/− mice, and these were seen during adolescence (p = 0.0058, d = 1.09) and adulthood (p = 0.003, d = 1.26). Together these findings provide the first evidence in humans that an axon guidance gene is involved in the formation of mesocorticolimbic circuitry and related behavioral traits, providing mechanisms through which DCC mutations might affect susceptibility to diverse neuropsychiatric disorders. SIGNIFICANCE STATEMENT Opportunities to study the effects of axon guidance molecules on human brain development have been rare. Here, the identification of a large four-generational family that carries a mutation to the axon guidance molecule receptor gene, DCC, enabled us to demonstrate effects on mesocorticolimbic anatomical connectivity, striatal volumes, and personality traits. Reductions in striatal volumes were replicated in DCC-haploinsufficient mice. Together, these processes might influence mesocorticolimbic function and susceptibility to diverse neuropsychiatric disorders.

LONGITUDINAL, MULTI-MODALITY, MRI STUDIES OF GAMMA-SECRETASE INHIBITION IN AN APP-SW MOUSE MODEL OF ALZHEIMER'S DISEASE

STUDIESOFGAMMA-SECRETASE INHIBITION IN AN APP-SW MOUSE MODEL OFALZHEIMER’S DISEASE Barry J. Bedell, Simone P. Zehntner, Thomas Rosahl, Kathleen Wood, Kelly R. Bales, Michael A. Marconi, Eric Parker, Felix Carbonell, Alex P. Zijdenbos, Biospective Inc. & McGill University, Montreal, Quebec, Canada; Biospective Inc., Montreal, Quebec, Canada; Merck & Co., Rahway, New Jersey, United States; Pfizer Inc., Cambridge, Massachusetts, United States; 5 Merck Research Laboratories, Kenilworth, New Jersey, United States. Contact e-mail: bbedell@biospective.com

BETA-AMYLOID-RELATED ALTERATIONS OF METABOLIC CONNECTIVITY IN ADNI MCI SUBJECTS: A MULTIVARIATE COVARIANCE ANALYSIS

IC-P-051 LONGITUDINAL, MULTI-MODALITY, MRI STUDIESOFGAMMA-SECRETASE INHIBITION IN AN APP-SW MOUSE MODEL OFALZHEIMER’S DISEASE Barry J. Bedell, Simone P. Zehntner, Thomas Rosahl, Kathleen Wood, Kelly R. Bales, Michael A. Marconi, Eric Parker, Felix Carbonell, Alex P. Zijdenbos, Biospective Inc. & McGill University, Montreal, Quebec, Canada; Biospective Inc., Montreal, Quebec, Canada; Merck & Co., Rahway, New Jersey, United States; Pfizer Inc., Cambridge, Massachusetts, United States; Merck Research Laboratories, Kenilworth, New Jersey, United States. Contact e-mail: bbedell@biospective.com

SPREAD OF BETA-AMYLOID ALONG ANATOMICALLY CONNECTED PATHWAYS IN HAPP TRANSGENIC MICE

Background: The prion-like hypothesis predicts disease spread from propagation of pathogenic misfolded proteins along anatomically-connected pathways. The epidemic spreading model (ESM) is a promising method for predicting this propagation, as was recently demonstrated using data from ADNI. It considers there is an As driving region (epicenter) and that interregional deposition patterns are dependent upon a region’s connectivity to this epicenter. The aim of the present study was to further validate ESM in As overexpressingmice using gold-standard, quantitative immunohistochemistry measures. Methods: 3D quantitative IHC volumes from 12 and 19 month-old transgenic mutant human amyloid precursor protein (hAPP) were spatially normalized to an anatomical template (PERMITS , Biospective Inc.), providing extracted mean values of As deposition in 25 gray matter regions, including 14 cortical regions. C onnectivity between the 25 regions was estimated from publicly-available viral tracer studies (http:// connectivity.brain-map.org/). An anatomical effective distances between each of pair regions i and j (Edist i,j) was then calculated as the shortest path distance between them (i.e. the more connected i and j, the lower the Edist i,j value between them). Using an iterative procedure, we then identified the epicenter region i that maximized the non-linear correlation between regional As deposition values and Edist i,j values (j 1⁄4 1 to 25). Results: The validity of an As epidemic spreading-like behavior hypothesis in mouse models was supported by the identification of the posterior cingulate cortex (PCC) as the epicenter region (same epicenter identified previously for the human subjects). The anatomical effective distance to this epicenter predicted As deposition at 12 and 19 months of age, explaining 60% and 61% of the characteristic As patterns, respectively. Conclusions: The observed correlation between connectivity and As deposition pattern supports the networkdriven hypothesis of misfolded proteins propagation. The epicenter region, PCC, is an central node of the default mode network, which, in humans, has been linked to the progression of Alzheimer’s disease. Further modeling analysis will provide us with a better understanding of disease progression, which will significantly advance translational neurodegenerative research.

RELATIONSHIP BETWEEN CEREBRAL BLOOD FLOW AND CORTICAL LAYER I ASTROCYTES IN A TRANSGENIC MOUSE MODEL OF ALZHEIMER'S DISEASE

(WT) mice (n1⁄48) were studied at 17-19 months-of-age. Whole brain perfusion images (360 mm isotropic spatial resolution) were acquired with a multi-slice, pseudocontinuous ASL pulse sequence on a 7T animal MRI system. Mice were scanned both in the awake state and under sevoflurane anesthesia (a known vasodilator) to assess group differences in resting CBF and vasodilatory response, respectively. The perfusion images were co-registered to anatomical MR images acquired during the same session, and spatially normalized to reference coordinate space to generate regional perfusion measures using a fully-automated image processing pipeline (NIGHTWING TM, Biospective Inc.). Results: The Tg mice exhibited a lower resting CBF (w15%) compared to theWTmice in most brain regions. Under sevoflurane, this difference increased tow30% as the Tgmice exhibited an attenuated vasodilatory response relative to the WT group. The coefficient of variation was higher in the awake mice. Conclusions: This study demonstrated that APPoverexpressing Tg mice have impaired resting cerebrovascular function, which is exacerbated under a challenge condition. The greater effect size of the challenge state suggests that utilization of an appropriate vasodilatory challenge could be a useful paradigm in human AD studies.