TLR7 activation in epilepsy of tuberous sclerosis complex.

BACKGROUND: Neuroinflammation and toll-like receptors (TLR) of the innate immune system have been implicated in epilepsy. We previously reported high levels of microRNAs miR-142-3p and miR-223-3p in epileptogenic brain tissue resected for the treatment of intractable epilepsy in children with tuberous sclerosis complex (TSC). As miR-142-3p has recently been reported to be a ligand and activator of TLR7, a detector of exogenous and endogenous single-stranded RNA, we evaluated TLR7 expression and downstream IL23A activation in surgically resected TSC brain tissue. METHODS: Gene expression analysis was performed on cortical tissue obtained from surgery of TSC children with pharmacoresistent epilepsy. Expression of TLRs 2, 4 and 7 was measured using NanoString nCounter assays. Real-time quantitative PCR was used to confirm TLR7 expression and compare TLR7 activation, indicated by IL-23A levels, to levels of miR-142-3p. Protein markers characteristic for TLR7 activation were assessed using data from our existing quantitative proteomics dataset of TSC tissue. Capillary electrophoresis Western blots were used to confirm TLR7 protein expression in a subset of samples. RESULTS: TLR7 transcript expression was present in all TSC specimens. The signaling competent form of TLR7 protein was detected in the membrane fraction of each sample tested. Downstream activation of TLR7 was found in epileptogenic lesions having elevated neuroinflammation indicated by clinical neuroimaging. TLR7 activity was significantly associated with tissue levels of miR-142-3p. CONCLUSION: TLR7 activation by microRNAs may contribute to the neuroinflammatory cascade in epilepsy in TSC. Further characterization of this mechanism may enable the combined of use of neuroimaging and TLR7 inhibitors in a personalized approach towards the treatment of intractable epilepsy.

Trajectory of inflammatory and microglial activation markers in the postnatal rabbit brain following intrauterine endotoxin exposure.

BACKGROUND: Maternal infection is a risk factor for periventricular leukomalacia and cerebral palsy (CP) in neonates. We have previously demonstrated hypomyelination and motor deficits in newborn rabbits, as seen in patients with cerebral palsy, following maternal intrauterine endotoxin administration. This was associated with increased microglial activation, primarily involving the periventricular region (PVR). In this study we hypothesized that maternal intrauterine inflammation leads to a pro-inflammatory environment in the PVR that is associated with microglial activation in the first 2 postnatal weeks. METHODS: Timed pregnant New Zealand white rabbits underwent laparotomy on gestational day 28 (G28). They were randomly divided to receive lipopolysaccharide (LPS; 20µg/kg in 1mL saline) (Endotoxin group) or saline (1mL) (control saline, CS group), administrated along the wall of the uterus. The PVR from the CS and Endotoxin kits were harvested at G29 (1day post-injury), postnatal day1 (PND1, 3day post-injury) and PND5 (7days post-injury) for real-time PCR, ELISA and immunohistochemistry. Kits from CS and Endotoxin groups underwent longitudinal MicroPET imaging, with [11C]PK11195, a tracer for microglial activation. RESULTS: We found that intrauterine endotoxin exposure resulted in pro-inflammatory microglial activation in the PVR of rabbits in the first postnatal week. This was evidenced by increased TSPO (translocator protein) expression co-localized with microglia/macrophages in the PVR, and changes in the microglial morphology (ameboid soma and retracted processes). In addition, CD11b level significantly increased with a concomitant decline in the CD45 level in the PVR at G29 and PND1. There was a significant elevation of pro-inflammatory cytokines and iNOS, and decreased anti-inflammatory markers in the Endotoxin kits at G29, PND1 and PND5. Increased [11C]PK11195 binding to the TSPO measured in vivo by PET imaging in the brain of Endotoxin kits was present up to PND14-17. CONCLUSIONS: Our results indicate that a robust pro-inflammatory microglial phenotype/brain milieu commenced within 24h after LPS exposure and persisted through PND5 and in vivo TSPO binding was found at PND14-17. This suggests that there may be a window of opportunity to treat after birth. Therapies aimed at inducing an anti-inflammatory phenotype in microglia might promote recovery in maternal inflammation induced neonatal brain injury.

A distinct microRNA expression profile is associated with α[11C]-methyl-L-tryptophan (AMT) PET uptake in epileptogenic cortical tubers resected from patients with tuberous sclerosis complex.

Tuberous sclerosis complex (TSC) is characterized by hamartomatous lesions in various organs and arises due to mutations in the TSC1 or TSC2 genes. TSC mutations lead to a range of neurological manifestations including epilepsy, cognitive impairment, autism spectrum disorders (ASD), and brain lesions that include cortical tubers. There is evidence that seizures arise at or near cortical tubers, but it is unknown why some tubers are epileptogenic while others are not. We have previously reported increased tryptophan metabolism measured with α[11C]-methyl-l-tryptophan (AMT) positron emission tomography (PET) in epileptogenic tubers in approximately two-thirds of patients with tuberous sclerosis and intractable epilepsy. However, the underlying mechanisms leading to seizure onset in TSC remain poorly characterized. MicroRNAs are enriched in the brain and play important roles in neurodevelopment and brain function. Recent reports have shown aberrant microRNA expression in epilepsy and TSC. In this study, we performed microRNA expression profiling in brain specimens obtained from TSC patients undergoing epilepsy surgery for intractable epilepsy. Typically, in these resections several non-seizure onset tubers are resected together with the seizure-onset tubers because of their proximity. We directly compared seizure onset tubers, with and without increased tryptophan metabolism measured with PET, and non-onset tubers to assess the role of microRNAs in epileptogenesis associated with these lesions. Whether a particular tuber was epileptogenic or non-epileptogenic was determined with intracranial electrocorticography, and tryptophan metabolism was measured with AMT PET. We identified a set of five microRNAs (miR-142-3p, 142-5p, 223-3p, 200b-3p and 32-5p) that collectively distinguish among the three primary groups of tubers: non-onset/AMT-cold (NC), onset/AMT-cold (OC), and onset/AMT-hot (OH). These microRNAs were significantly upregulated in OH tubers compared to the other two groups, and microRNA expression was most significantly associated with AMT-PET uptake. The microRNAs target a group of genes enriched for synaptic signaling and epilepsy risk, including SLC12A5, SYT1, GRIN2A, GRIN2B, KCNB1, SCN2A, TSC1, and MEF2C. We confirmed the interaction between miR-32-5p and SLC12A5 using a luciferase reporter assay. Our findings provide a new avenue for subsequent mechanistic studies of tuber epileptogenesis in TSC.

Maternal Inflammation Results in Altered Tryptophan Metabolism in Rabbit Placenta and Fetal Brain.

Maternal inflammation has been linked to neurodevelopmental and neuropsychiatric disorders such as cerebral palsy, schizophrenia, and autism. We had previously shown that intrauterine inflammation resulted in a decrease in serotonin, one of the tryptophan metabolites, and a decrease in serotonin fibers in the sensory cortex of newborns in a rabbit model of cerebral palsy. In this study, we hypothesized that maternal inflammation results in alterations in tryptophan pathway enzymes and metabolites in the placenta and fetal brain. We found that intrauterine endotoxin administration at gestational day 28 (G28) resulted in a significant upregulation of indoleamine 2,3-dioxygenase (IDO) in both the placenta and fetal brain at G29 (24 h after treatment). This endotoxin-mediated IDO induction was also associated with intense microglial activation, an increase in interferon gamma expression, and increases in kynurenine and the kynurenine pathway metabolites kynurenine acid and quinolinic acid, as well as a significant decrease in 5-hydroxyindole acetic acid (a precursor of serotonin) levels in the periventricular region of the fetal brain. These results indicate that maternal inflammation shunts tryptophan metabolism away from the serotonin to the kynurenine pathway, which may lead to excitotoxic injury along with impaired development of serotonin-mediated thalamocortical fibers in the newborn brain. These findings provide new targets for prevention and treatment of maternal inflammation-induced fetal and neonatal brain injury leading to neurodevelopmental disorders such as cerebral palsy and autism.

Cortical Tubers: Windows into Dysregulation of Epilepsy Risk and Synaptic Signaling Genes by MicroRNAs.

Tuberous sclerosis complex (TSC) is a multisystem genetic disorder caused by mutations in the TSC1 and TSC2 genes. Over 80% of TSC patients are affected by epilepsy, but the molecular events contributing to seizures in TSC are not well understood. Recent reports have demonstrated that the brain is enriched with microRNA activity, and they are critical in neural development and function. However, little is known about the role of microRNAs in TSC. Here, we report the characterization of aberrant microRNA activity in cortical tubers resected from 5 TSC patients surgically treated for medically intractable epilepsy. By comparing epileptogenic tubers with adjacent nontuber tissue, we identified a set of 4 coordinately overexpressed microRNAs (miRs 23a, 34a, 34b*, 532-5p). We used quantitative liquid chromatography-tandem mass spectrometry (LC-MS/MS) proteomic profiling to investigate the combined effect of the 4 microRNAs on target proteins. The proportion of repressed proteins among the predicted targets was significantly greater than in the overall proteome and was highly enriched for proteins involved in synaptic signal transmission. Among the combinatorial targets were TSC1, coding for the protein hamartin, and several epilepsy risk genes. We found decreased levels of hamartin in epileptogenic tubers and confirmed targeting of the TSC1 3' UTR by miRs-23a and 34a.

Efficacy of Low-Dose Buspirone for Restricted and Repetitive Behavior in Young Children with Autism Spectrum Disorder: A Randomized Trial.

OBJECTIVES: To determine safety and efficacy of the 5HT1A serotonin partial agonist buspirone on core autism and associated features in children with autism spectrum disorder (ASD). STUDY DESIGN: Children 2-6 years of age with ASD (N = 166) were randomized to receive placebo or 2.5 or 5.0 mg of buspirone twice daily. The primary objective was to evaluate the effects of 24 weeks of buspirone on the Autism Diagnostic Observation Schedule (ADOS) Composite Total Score. Secondary objectives included evaluating the effects of buspirone on social competence, repetitive behaviors, language, sensory dysfunction, and anxiety and to assess side effects. Positron emission tomography measures of tryptophan metabolism and blood serotonin concentrations were assessed as predictors of buspirone efficacy. RESULTS: There was no difference in the ADOS Composite Total Score between baseline and 24 weeks among the 3 treatment groups (P = .400); however, the ADOS Restricted and Repetitive Behavior score showed a time-by-treatment effect (P = .006); the 2.5-mg buspirone group showed significant improvement (P = .003), whereas placebo and 5.0-mg buspirone groups showed no change. Children in the 2.5-mg buspirone group were more likely to improve if they had fewer foci of increased brain tryptophan metabolism on positron emission tomography (P = .018) or if they showed normal levels of blood serotonin (P = .044). Adverse events did not differ significantly among treatment groups. CONCLUSIONS: Treatment with 2.5 mg of buspirone in young children with ASD might be a useful adjunct therapy to target restrictive and repetitive behaviors in conjunction with behavioral interventions. TRIAL REGISTRATION: ClinicalTrials.gov: NCT00873509.

Independent component analysis tractography combined with a ball-stick model to isolate intravoxel crossing fibers of the corticospinal tracts in clinical diffusion MRI.

The independent component analysis (ICA) tractography method has improved the ability to isolate intravoxel crossing fibers; however, the accuracy of ICA is limited in cases with voxels in local clusters lacking sufficient numbers of fibers with the same orientations. To overcome this limitation, the ICA was combined with a ball-stick model (BSM) ["ICA+BSM"]. An ICA approach is applied to identify crossing fiber components in voxels of small cluster, which are maximally independent in orientation. The eigenvectors of these components are numerically optimized via the subsequent BSM procedure. Simulation studies for two or three crossing fibers demonstrate that ICA+BSM overcomes the limitation of the original ICA method by refining regional ICA solutions in diffusion measurement of a single voxel. It shows 2°-5° of angular errors to isolate two or three fibers, providing a better recovery of simulated fibers compared with ICA alone. Human studies show that ICA+BSM achieves high anatomical correspondence of corticospinal tracts compared with postmortem corticospinal histology, yielding 92.2% true positive detection including both lateral and medial projections, compared with 84.1% for ICA alone. This study demonstrates that the intravoxel crossing fiber problem in clinical diffusion MRI may be sorted out more efficiently by combining ICA with BSM.

Concurrent quantification of tryptophan and its major metabolites.

An imbalance in tryptophan (TRP) metabolites is associated with several neurological and inflammatory disorders. Therefore, analytical methods allowing for simultaneous quantification of TRP and its major metabolites would be highly desirable, and may be valuable as potential biomarkers. We have developed a HPLC method for concurrent quantitative determination of tryptophan, serotonin, 5-hydroxyindoleacetic acid, kynurenine, and kynurenic acid in tissue and fluids. The method utilizes the intrinsic spectroscopic properties of TRP and its metabolites that enable UV absorbance and fluorescence detection by HPLC, without additional labeling. The origin of the peaks related to analytes of interest was confirmed by UV-Vis spectral patterns using a PDA detector and mass spectrometry. The developed methods were validated in rabbit fetal brain and amniotic fluid at gestational day 29. Results are in excellent agreement with those reported in the literature for the same regions. This method allows for rapid quantification of tryptophan and four of its major metabolites concurrently. A change in the relative ratios of these metabolites can provide important insights in predicting the presence and progression of neuroinflammation in disorders such as cerebral palsy, autism, multiple sclerosis, Alzheimer disease, and schizophrenia.

Automatic detection of primary motor areas using diffusion MRI tractography: comparison with functional MRI and electrical stimulation mapping.

PURPOSE: As an alternative tool to identify cortical motor areas for planning surgical resection in children with focal epilepsy, the present study proposed a maximum a posteriori probability (MAP) classification of corticospinal tract (CST) visualized by diffusion MR tractography. METHODS: Diffusion-weighted imaging (DWI) was performed in 17 normally developing children and 20 children with focal epilepsy. An independent component analysis tractography combined with ball-stick model was performed to identify unique CST pathways originating from mouth/lip, finger, and leg areas determined by functional magnetic resonance imaging (fMRI) in healthy children and electrical stimulation mapping (ESM) in children with epilepsy. Group analyses were performed to construct stereotaxic probability maps of primary motor pathways connecting precentral gyrus and posterior limb of internal capsule, and then utilized to design a novel MAP classifier that can sort individual CST fibers associated with three classes of interest: mouth/lip, fingers, and leg. A systematic leave-one-out approach was applied to train an optimal classifier. A match was considered to occur if classified fibers contacted or surrounded true areas localized by fMRI and ESM. KEY FINDINGS: It was found that the DWI-MAP provided high accuracy for the CST fibers terminating in proximity to the localization of fMRI/ESM: 78%/77% for mouth/lip, 77%/76% for fingers, 78%/86% for leg (contact), and 93%/89% for mouth/lip, 91%/89% for fingers, and 92%/88% for leg (surrounded within 2 cm). SIGNIFICANCE: This study provides preliminary evidence that in the absence of fMRI and ESM data, the DWI-MAP approach can effectively retrieve the locations of cortical motor areas and underlying CST courses for planning epilepsy surgery.

Successful surgical treatment of an inflammatory lesion associated with new-onset refractory status epilepticus.

New-onset refractory status epilepticus (NORSE) has high morbidity and mortality. The authors describe the successful surgical treatment of a 56-year-old man presenting with NORSE. Magnetic resonance imaging showed a left temporal lobe lesion suspicious for a low-grade tumor, while PET imaging with the alpha[(11)C]methyl-L-tryptophan (AMT) radiotracer showed increased cortical uptake extending beyond this lesion and partly overlapping with epileptogenic cortex mapped by chronic intracranial electroencephalographic monitoring. Resection of the epileptic focus resulted in long-term seizure freedom, and the nonresected portion of the PET-documented abnormality normalized. Histopathology showed reactive gliosis and inflammatory markers in the AMT-PET-positive cortex. Molecular imaging of neuroinflammation can be instrumental in the management of NORSE by guiding placement of intracranial electrodes or assessing the extent and severity of inflammation for antiinflammatory interventions.

Altered white matter structure of the dentatorubrothalamic pathway in children with autistic spectrum disorders.

Neuropathological studies have demonstrated decreased Purkinje cells in cerebellar cortex and changes in the dentate nucleus of the cerebellum, the projection target for the Purkinje cells, in autistic spectrum disorders (ASD). The dentatorubrothalamic tract is formed by efferents from the dentate nucleus projecting toward the red nucleus with axon collaterals to this nucleus and continuing to innervate the ventral lateral and ventral anterior nuclei of the thalamus. In the current study, we assessed whether the dentatorubrothalamic tract is altered in ASD using Q-ball imaging (QBI). The QBI tractography was performed in 13 children with high functioning ASD (HFA), 11 children with low functioning ASD (LFA), and 14 typically developing children (TD). Regions of interest in dentate nucleus and red nucleus in both hemispheres were objectively placed to sort bilateral dorsal-rostral (DR), dorsal-caudal (DC), ventral-rostral (VR), and ventral-caudal (VC) portions of the dentatorubrothalamic pathway. Group differences in fractional anisotropy (FA), axial diffusivity, radial diffusivity, and fiber volume of individual pathways were analyzed. Significantly reduced FA was found in children with LFA and HFA, compared to the TD group in tracts originating in all four subdivisions of the right dentate nucleus. Tract-based morphometry (TBM) analysis demonstrated significant reductions of FA in caudal midbrain (p<0.0001), dorsal-caudal dentate (p=0.0013), and ventral-caudal dentate (p=0.0061) on the right in the LFA group. The FA values in TBM segments of right VR and VC pathways were significantly correlated with communication skills in the combined HFA/LFA group, while there was a significant correlation found between TBM segments of right DR pathway and daily living skills (r=0.76; p=0.004). Decreased white matter integrity in dorsal portions of the dentatorubrothalamic tract may be related to motor features in ASD, while changes in the ventral portions are related more to communication behavior.

Increased tryptophan transport in epileptogenic dysembryoplastic neuroepithelial tumors.

Dysembryoplastic neuroepithelial tumors (DNTs) are typically hypometabolic but can show increased amino acid uptake on positron emission tomography (PET). To better understand mechanisms of amino acid accumulation in epileptogenic DNTs, we combined quantitative α-[(11)C]methyl-L: -tryptophan (AMT) PET with tumor immunohistochemistry. Standardized uptake values (SUVs) of AMT and glucose were measured in 11 children with temporal lobe DNT. Additional quantification for AMT transport and metabolism was performed in 9 DNTs. Tumor specimens were immunostained for the L: -type amino acid transporter 1 (LAT1) and indoleamine 2,3-dioxygenase (IDO), a key enzyme of the immunomodulatory kynurenine pathway. All 11 tumors showed glucose hypometabolism, while mean AMT SUVs were higher than normal cortex in eight DNTs. Further quantification showed increased AMT transport in seven and high AMT metabolic rates in three DNTs. Two patients showing extratumoral cortical increases of AMT SUV had persistent seizures despite complete tumor resection. Resected DNTs showed moderate to strong LAT1 and mild to moderate IDO immunoreactivity, with the strongest expression in tumor vessels. These results indicate that accumulation of tryptophan in DNTs is driven by high amino acid transport, mediated by LAT1, which can provide the substrate for tumoral tryptophan metabolism through the kynurenine pathway, that can produce epileptogenic metabolites. Increased AMT uptake can extend to extratumoral cortex, and presence of such cortical regions may increase the likelihood of recurrent seizures following surgical excision of DNTs.

Congruence of happy and sad emotion in music and faces modifies cortical audiovisual activation.

BACKGROUND: The powerful emotion inducing properties of music are well-known, yet music may convey differing emotional responses depending on environmental factors. We hypothesized that neural mechanisms involved in listening to music may differ when presented together with visual stimuli that conveyed the same emotion as the music when compared to visual stimuli with incongruent emotional content. METHODS: We designed this study to determine the effect of auditory (happy and sad instrumental music) and visual stimuli (happy and sad faces) congruent or incongruent for emotional content on audiovisual processing using fMRI blood oxygenation level-dependent (BOLD) signal contrast. The experiment was conducted in the context of a conventional block-design experiment. A block consisted of three emotional ON periods, music alone (happy or sad music), face alone (happy or sad faces), and music combined with faces where the music excerpt was played while presenting either congruent emotional faces or incongruent emotional faces. RESULTS: We found activity in the superior temporal gyrus (STG) and fusiform gyrus (FG) to be differentially modulated by music and faces depending on the congruence of emotional content. There was a greater BOLD response in STG when the emotion signaled by the music and faces was congruent. Furthermore, the magnitude of these changes differed for happy congruence and sad congruence, i.e., the activation of STG when happy music was presented with happy faces was greater than the activation seen when sad music was presented with sad faces. In contrast, incongruent stimuli diminished the BOLD response in STG and elicited greater signal change in bilateral FG. Behavioral testing supplemented these findings by showing that subject ratings of emotion in faces were influenced by emotion in music. When presented with happy music, happy faces were rated as more happy (p=0.051) and sad faces were rated as less sad (p=0.030). When presented with sad music, happy faces were rated as less happy (p=0.008) and sad faces were rated as sadder (p=0.002). INTERPRETATION: Happy-sad congruence across modalities may enhance activity in auditory regions while incongruence appears to impact the perception of visual affect, leading to increased activation in face processing regions such as the FG. We suggest that greater understanding of the neural bases of happy-sad congruence across modalities can shed light on basic mechanisms of affective perception and experience and may lead to novel insights in the study of emotion regulation and therapeutic use of music.

Magnitude of [(11)C]PK11195 binding is related to severity of motor deficits in a rabbit model of cerebral palsy induced by intrauterine endotoxin exposure.

Intrauterine inflammation is known to be a risk factor for the development of periventricular leukomalacia (PVL) and cerebral palsy. In recent years, activated microglial cells have been implicated in the pathogenesis of PVL and in the development of white matter injury. Clinical studies have shown the increased presence of activated microglial cells diffusely throughout the white matter in brains of patients with PVL. In vitro studies have reported that activated microglial cells induce oligodendrocyte damage and white matter injury by release of inflammatory cytokines, reactive nitrogen and oxygen species and the production of excitotoxic metabolites. PK11195 [1-(2-chlorophenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinoline carboxamide] is a ligand that is selective for the 18-kDa translocator protein expressed on the outer mitochondrial membrane of activated microglia and macrophages. When labeled with carbon-11, [(11)C]PK11195 can effectively be used as a ligand in positron emission tomography (PET) studies for the detection of activated microglial cells in various neuroinflammatory and neurodegenerative conditions. In this study, we hypothesized that the magnitude of [(11)C]-(R)-PK11195 uptake in the newborn rabbit brain, as measured using a small-animal PET scanner, would match the severity of motor deficits resulting from intrauterine inflammation-induced perinatal brain injury. Pregnant New Zealand white rabbits were intrauterinely injected with endotoxin or saline at 28 days of gestation. Kits were born spontaneously at 31 days and underwent neurobehavioral testing and PET imaging following intravenous injection of the tracer [(11)C]-(R)-PK11195 on the day of birth. The neurobehavioral scores were compared with the change in [(11)C]PK11195 uptake over the time of scanning, for each of the kits. Upon analysis using receiver operating characteristic curves, an optimal combined sensitivity and specificity for detecting abnormal neurobehavioral scores suggestive of cerebral palsy in the neonatal rabbit was noted for a positive change in [(11)C]PK11195 uptake in the brain over time on PET imaging (sensitivity of 100% and area under the curve of >0.82 for all parameters tested). The strongest agreements were noted between a positive uptake slope - indicating increased [(11)C]PK11195 uptake over time - and worsening scores for measures of locomotion (indicated by hindlimb movement, forelimb movement, circular motion and straight- line motion; Cohen's κ >0.75 for each) and feeding (indicated by ability to suck and swallow and turn the head during feeding; Cohen's κ >0.85 for each). This was also associated with increased numbers of activated microglia (mean ratio ± SD of activated to total microglia: 0.96 ± 0.16 in the endotoxin group vs. 0.13 ± 0.08 in controls; p < 0.001) in the internal capsule and corona radiata. Our findings indicate that the magnitude of [(11)C]PK11195 binding measured in vivo by PET imaging matches the severity of motor deficits in the neonatal rabbit. Molecular imaging of ongoing neuroinflammation in the neonatal period may be helpful as a screening biomarker for detecting patients at risk of developing cerebral palsy due to a perinatal insult.

Differential kinetics of α-[¹¹C]methyl-L-tryptophan on PET in low-grade brain tumors.

Increased tryptophan metabolism via the kynurenine pathway is a major mechanism of tumor immuno-resistance. α-[(11)C]Methyl-L: -tryptophan (AMT) is a positron emission tomography (PET) tracer for tryptophan catabolism, and increased AMT uptake has been demonstrated in brain tumors. In this study we evaluated the use of AMT PET for detection of low-grade gliomas and glioneuronal tumors, and determined if kinetic parameters of AMT uptake can differentiate among tumor types. AMT PET images were obtained in 23 patients with newly diagnosed low-grade brain tumors (WHO grade II gliomas and WHO grade I dysembryoplastic neuroepithelial tumors [DNETs]). Kinetic variables, including the unidirectional uptake rate (K-complex) and volume of distribution (VD; which characterizes tracer transport), were measured using a graphical approach from tumor dynamic PET and blood-input data, and metabolic rates ([Formula: see text]) were also calculated. These values as well as tumor/cortex ratios were compared across tumor types. AMT PET showed increased tumor/cortex K-complex (n = 16) and/or VD ratios (n = 15) in 21/23 patients (91%), including 11/13 tumors with no gadolinium enhancement on MRI. No increases in AMT were seen in an oligodendroglioma and a DNET. Astrocytomas and oligoastrocytomas showed higher [Formula: see text] tumor/cortex ratios (1.66 ± 0.46) than oligodendrogliomas (0.96 ± 0.21; P = 0.001) and DNETs (0.75 ± 0.39; P < 0.001). These results demonstrate that AMT PET identifies most low-grade gliomas and DNETs by high uptake, even if these tumors are not contrast-enhancing on MRI. Kinetic analysis of AMT uptake shows significantly higher tumor/cortex tryptophan metabolic ratios in astrocytomas and oligoastrocytomas in comparison with oligodendrogliomas and DNETs.

Alterations in frontal lobe tracts and corpus callosum in young children with autism spectrum disorder.

Major frontal lobe tracts and corpus callosum (CC) were investigated in 32 children with autism spectrum disorder (ASD, mean age: 5 years), 12 nonautistic developmentally impaired children (DI, mean age: 4.6 years), and 16 typically developing children (TD, mean age: 5.5 years) using diffusion tensor imaging tractography and tract-based spatial statistics. Various diffusion and geometric properties were calculated for uncinate fasciculus (UF), inferior fronto-occipital fasciculus (IFO), arcuate fasciculus (AF), cingulum (Cg), CC, and corticospinal tract. Fractional anisotropy was lower in the right UF, right Cg and CC in ASD and DI children; in right AF in ASD children; and in bilateral IFO in DI children, compared with TD children. Apparent diffusion coefficient was increased in right AF in both ASD and DI children. The ASD group showed shorter length of left UF and increased length, volume, and density of right UF; increased length and density of CC; and higher density of left Cg, compared with the TD group. Compared with DI group, ASD group had increased length, volume, and density of right UF; higher volume of left UF; and increased length of right AF and CC. Volume of bilateral UF and right AF and fiber density of left UF were positively associated with autistic features.

Association Between Gray Matter Volume Variations and Energy Utilization in the Brain: Implications for Developmental Stuttering.

Purpose The biological mechanisms underlying developmental stuttering remain unclear. In a previous investigation, we showed that there is significant spatial correspondence between regional gray matter structural anomalies and the expression of genes linked to energy metabolism. In the current study, we sought to further examine the relationship between structural anomalies in the brain in children with persistent stuttering and brain regional energy metabolism. Method High-resolution structural MRI scans were acquired from 26 persistent stuttering and 44 typically developing children. Voxel-based morphometry was used to quantify the between-group gray matter volume (GMV) differences across the whole brain. Group differences in GMV were then compared with published values for the pattern of glucose metabolism measured via F18 fluorodeoxyglucose uptake in the brains of 29 healthy volunteers using positron emission tomography. Results A significant positive correlation between GMV differences and F18 fluorodeoxyglucose uptake was found in the left hemisphere (ρ = .36, p < .01), where speech-motor and language processing are typically localized. No such correlation was observed in the right hemisphere (ρ = .05, p = .70). Conclusions Corroborating our previous gene expression studies, the results of the current study suggest a potential connection between energy metabolism and stuttering. Brain regions with high energy utilization may be particularly vulnerable to anatomical changes associated with stuttering. Such changes may be further exacerbated when there are sharp increases in brain energy utilization, which coincides with the developmental period of rapid speech/language acquisition and the onset of stuttering during childhood. Supplemental Material https://doi.org/10.23641/asha.14110454.

Exosomes in Epilepsy of Tuberous Sclerosis Complex: Carriers of Pro-Inflammatory MicroRNAs.

Exosomes are a class of small, secreted extracellular vesicles (EV) that have recently gained considerable attention for their role in normal cellular function, disease processes and potential as biomarkers. Exosomes serve as intercellular messengers and carry molecular cargo that can alter gene expression and the phenotype of recipient cells. Here, we investigated alterations of microRNA cargo in exosomes secreted by epileptogenic tissue in tuberous sclerosis complex (TSC), a multi-system genetic disorder that includes brain lesions known as tubers. Approximately 90% of TSC patients suffer from seizures that originate from tubers, and ~60% are resistant to antiseizure drugs. It is unknown why some tubers cause seizures while others do not, and the molecular basis of drug-resistant epilepsy is not well understood. It is believed that neuroinflammation is involved, and characterization of this mechanism may be key to disrupting the "vicious cycle" between seizures, neuroinflammation, and increased seizure susceptibility. We isolated exosomes from epileptogenic and non-epileptogenic TSC tubers, and we identified differences in their microRNA cargo using small RNA-seq. We identified 12 microRNAs (including miR-142-3p, miR-223-3p and miR-21-5p) that are significantly increased in epileptogenic tubers and contain nucleic acid motifs that activate toll-like receptors (TLR7/8), initiating a neuroinflammatory cascade. Exosomes from epileptogenic tissue caused induction of key pathways in cultured cells, including innate immune signaling (TLR), inflammatory response and key signaling nodes SQSTM1 (p62) and CDKN1A (p21). Genes induced in vitro were also significantly upregulated in epileptogenic tissue. These results provide new evidence on the role of exosomes and non-coding RNA cargo in the neuroinflammatory cascade of epilepsy and may help advance the development of novel biomarkers and therapeutic approaches for the treatment of drug-resistant epilepsy.

PET imaging of medulloblastoma with an 18F-labeled tryptophan analogue in a transgenic mouse model.

In vivo positron emission tomography (PET) imaging is a key modality to evaluate disease status of brain tumors. In recent years, tremendous efforts have been made in developing PET imaging methods for pediatric brain tumors. Carbon-11 labelled tryptophan derivatives are feasible as PET imaging probes in brain tumor patients with activation of the kynurenine pathway, but the short half-life of carbon-11 limits its application. Using a transgenic mouse model for the sonic hedgehog (Shh) subgroup of medulloblastoma, here we evaluated the potential of the newly developed 1-(2-[18F]fluoroethyl)-L-tryptophan (1-L-[18F]FETrp) as a PET imaging probe for this common malignant pediatric brain tumor. 1-L-[18F]FETrp was synthesized on a PETCHEM automatic synthesizer with good chemical and radiochemical purities and enantiomeric excess values. Imaging was performed in tumor-bearing Smo/Smo medulloblastoma mice with constitutive actvation of the Smoothened (Smo) receptor using a PerkinElmer G4 PET-X-Ray scanner. Medulloblastoma showed significant and specific accumulation of 1-L-[18F]FETrp. 1-L-[18F]FETrp also showed significantly higher tumor uptake than its D-enantiomer, 1-D-[18F]FETrp. The uptake of 1-L-[18F]FETrp in the normal brain tissue was low, suggesting that 1-L-[18F]FETrp may prove a valuable PET imaging probe for the Shh subgroup of medulloblastoma and possibly other pediatric and adult brain tumors.

Automated production of 1-(2-[18F]fluoroethyl)-l-tryptophan for imaging of tryptophan metabolism.

Automated production of an fluorine-18 labeled tryptophan analogue, 1-(2-[18F]fluoroethyl)-l-tryptophan (1-L-[18F]FETrp) in a current Good Manufacturing Practice facility was achieved. 1-L-[18F]FETrp was produced by a one-pot, two-step strategy with an overall synthesis time of approximately 100 min, a radiochemical yield of 20 ± 5% (decay corrected), radiochemical purity and enantiomeric excess over 90%, and a molar activity of 103 ± 15 GBq/µmol at the end of synthesis (EOS). The dose mass of 1-L-FETrp in four consecutive batches was less than 5 µg. The radiopharmaceutical product met all quality control criteria for clinical use.