Structural deviations of the posterior fossa and the cerebellum and their cognitive links in a neurodevelopmental deletion syndrome.

High-impact genetic variants associated with neurodevelopmental disorders provide biologically-defined entry points for mechanistic investigation. The 3q29 deletion (3q29Del) is one such variant, conferring a 40-100-fold increased risk for schizophrenia, as well as high risk for autism and intellectual disability. However, the mechanisms leading to neurodevelopmental disability remain largely unknown. Here, we report the first in vivo quantitative neuroimaging study in individuals with 3q29Del (N = 24) and neurotypical controls (N = 1608) using structural MRI. Given prior radiology reports of posterior fossa abnormalities in 3q29Del, we focused our investigation on the cerebellum and its tissue-types and lobules. Additionally, we compared the prevalence of cystic/cyst-like malformations of the posterior fossa between 3q29Del and controls and examined the association between neuroanatomical findings and quantitative traits to probe gene-brain-behavior relationships. 3q29Del participants had smaller cerebellar cortex volumes than controls, before and after correction for intracranial volume (ICV). An anterior-posterior gradient emerged in finer grained lobule-based and voxel-wise analyses. 3q29Del participants also had larger cerebellar white matter volumes than controls following ICV-correction and displayed elevated rates of posterior fossa arachnoid cysts and mega cisterna magna findings independent of cerebellar volume. Cerebellar white matter and subregional gray matter volumes were associated with visual-perception and visual-motor integration skills as well as IQ, while cystic/cyst-like malformations yielded no behavioral link. In summary, we find that abnormal development of cerebellar structures may represent neuroimaging-based biomarkers of cognitive and sensorimotor function in 3q29Del, adding to the growing evidence identifying cerebellar pathology as an intersection point between syndromic and idiopathic forms of neurodevelopmental disabilities.

Lateralization of major white matter tracts during infancy is time-varying and tract-specific.

Lateralization patterns are a major structural feature of brain white matter and have been investigated as a neural architecture that indicates and supports the specialization of cognitive processing and observed behaviors, e.g. language skills. Many neurodevelopmental disorders have been associated with atypical lateralization, reinforcing the need for careful measurement and study of this structural characteristic. Unfortunately, there is little consensus on the direction and magnitude of lateralization in major white matter tracts during the first months and years of life-the period of most rapid postnatal brain growth and cognitive maturation. In addition, no studies have examined white matter lateralization in a longitudinal pediatric sample-preventing confirmation of if and how white matter lateralization changes over time. Using a densely sampled longitudinal data set from neurotypical infants aged 0-6 months, we aim to (i) chart trajectories of white matter lateralization in 9 major tracts and (ii) link variable findings from cross-sectional studies of white matter lateralization in early infancy. We show that patterns of lateralization are time-varying and tract-specific and that differences in lateralization results during this period may reflect the dynamic nature of lateralization through development, which can be missed in cross-sectional studies.

A comprehensive atlas of white matter tracts in the chimpanzee.

Chimpanzees (Pan troglodytes) are, along with bonobos, humans' closest living relatives. The advent of diffusion MRI tractography in recent years has allowed a resurgence of comparative neuroanatomical studies in humans and other primate species. Here we offer, in comparative perspective, the first chimpanzee white matter atlas, constructed from in vivo chimpanzee diffusion-weighted scans. Comparative white matter atlases provide a useful tool for identifying neuroanatomical differences and similarities between humans and other primate species. Until now, comprehensive fascicular atlases have been created for humans (Homo sapiens), rhesus macaques (Macaca mulatta), and several other nonhuman primate species, but never in a nonhuman ape. Information on chimpanzee neuroanatomy is essential for understanding the anatomical specializations of white matter organization that are unique to the human lineage.

Evolutionary expansion of connectivity between multimodal association areas in the human brain compared with chimpanzees.

The development of complex cognitive functions during human evolution coincides with pronounced encephalization and expansion of white matter, the brain's infrastructure for region-to-region communication. We investigated adaptations of the human macroscale brain network by comparing human brain wiring with that of the chimpanzee, one of our closest living primate relatives. White matter connectivity networks were reconstructed using diffusion-weighted MRI in humans (n = 57) and chimpanzees (n = 20) and then analyzed using network neuroscience tools. We demonstrate higher network centrality of connections linking multimodal association areas in humans compared with chimpanzees, together with a more pronounced modular topology of the human connectome. Furthermore, connections observed in humans but not in chimpanzees particularly link multimodal areas of the temporal, lateral parietal, and inferior frontal cortices, including tracts important for language processing. Network analysis demonstrates a particularly high contribution of these connections to global network integration in the human brain. Taken together, our comparative connectome findings suggest an evolutionary shift in the human brain toward investment of neural resources in multimodal connectivity facilitating neural integration, combined with an increase in language-related connectivity supporting functional specialization.

Deep phenotyping in 3q29 deletion syndrome: recommendations for clinical care.

PURPOSE: To understand the consequences of the 3q29 deletion on medical, neurodevelopmental, psychiatric, brain structural, and neurological sequalae by systematic evaluation of affected individuals. To develop evidence-based recommendations using these data for effective clinical care. METHODS: Thirty-two individuals with the 3q29 deletion were evaluated using a defined phenotyping protocol and standardized data collection instruments. RESULTS: Medical manifestations were varied and reported across nearly every organ system. The most severe manifestations were congenital heart defects (25%) and the most common were gastrointestinal symptoms (81%). Physical examination revealed a high proportion of musculoskeletal findings (81%). Neurodevelopmental phenotypes represent a significant burden and include intellectual disability (34%), autism spectrum disorder (38%), executive function deficits (46%), and graphomotor weakness (78%). Psychiatric illness manifests across the lifespan with psychosis prodrome (15%), psychosis (20%), anxiety disorders (40%), and attention deficit-hyperactivity disorder (ADHD) (63%). Neuroimaging revealed structural anomalies of the posterior fossa, but on neurological exam study subjects displayed only mild or moderate motor vulnerabilities. CONCLUSION: By direct evaluation of 3q29 deletion study subjects, we document common features of the syndrome, including a high burden of neurodevelopmental and neuropsychiatric phenotypes. Evidence-based recommendations for evaluation, referral, and management are provided to help guide clinicians in the care of 3q29 deletion patients.

Evolutionary modifications in human brain connectivity associated with schizophrenia.

The genetic basis and human-specific character of schizophrenia has led to the hypothesis that human brain evolution may have played a role in the development of the disorder. We examined schizophrenia-related changes in brain connectivity in the context of evolutionary changes in human brain wiring by comparing in vivo neuroimaging data from humans and chimpanzees, one of our closest living evolutionary relatives and a species with which we share a very recent common ancestor. We contrasted the connectome layout between the chimpanzee and human brain and compared differences with the pattern of schizophrenia-related changes in brain connectivity as observed in patients. We show evidence of evolutionary modifications of human brain connectivity to significantly overlap with the cortical pattern of schizophrenia-related dysconnectivity (P < 0.001, permutation testing). We validated these effects in three additional, independent schizophrenia datasets. We further assessed the specificity of effects by examining brain dysconnectivity patterns in seven other psychiatric and neurological brain disorders (including, among others, major depressive disorder and obsessive-compulsive disorder, arguably characterized by behavioural symptoms that are less specific to humans), which showed no such associations with modifications of human brain connectivity. Comparisons of brain connectivity across humans, chimpanzee and macaques further suggest that features of connectivity that evolved in the human lineage showed the strongest association to the disorder, that is, brain circuits potentially related to human evolutionary specializations. Taken together, our findings suggest that human-specific features of connectome organization may be enriched for changes in brain connectivity related to schizophrenia. Modifications in human brain connectivity in service of higher order brain functions may have potentially also rendered the brain vulnerable to brain dysfunction.

Study protocol for The Emory 3q29 Project: evaluation of neurodevelopmental, psychiatric, and medical symptoms in 3q29 deletion syndrome.

BACKGROUND: 3q29 deletion syndrome is caused by a recurrent hemizygous 1.6 Mb deletion on the long arm of chromosome 3. The syndrome is rare (1 in 30,000 individuals) and is associated with mild to moderate intellectual disability, increased risk for autism and anxiety, and a 40-fold increased risk for schizophrenia, along with a host of physical manifestations. However, the disorder is poorly characterized, the range of manifestations is not well described, and the underlying molecular mechanism is not understood. We designed the Emory 3q29 Project to document the range of neurodevelopmental and psychiatric manifestations associated with 3q29 deletion syndrome. We will also create a biobank of samples from our 3q29 deletion carriers for mechanistic studies, which will be a publicly-available resource for qualified investigators. The ultimate goals of our study are three-fold: first, to improve management and treatment of 3q29 deletion syndrome. Second, to uncover the molecular mechanism of the disorder. Third, to enable cross-disorder comparison with other rare genetic syndromes associated with neuropsychiatric phenotypes. METHODS: We will ascertain study subjects, age 6 and older, from our existing registry ( 3q29deletion.org ). Participants and their families will travel to Atlanta, GA for phenotypic assessments, with particular emphasis on evaluation of anxiety, cognitive ability, autism symptomatology, and risk for psychosis via prodromal symptoms and syndromes. Evaluations will be performed using standardized instruments. Structural, diffusion, and resting-state functional MRI data will be collected from eligible study participants. We will also collect blood from the 3q29 deletion carrier and participating family members, to be banked at the NIMH Repository and Genomics Resource (NRGR). DISCUSSION: The study of 3q29 deletion has the potential to transform our understanding of complex disease. Study of individuals with the deletion may provide insights into long term care and management of the disorder. Our project describes the protocol for a prospective study of the behavioral and clinical phenotype associated with 3q29 deletion syndrome. The paradigm described here could easily be adapted to study additional CNV or single gene disorders with high risk for neuropsychiatric phenotypes, and/or transferred to other study sites, providing a means for data harmonization and cross-disorder analysis.

Aberrant interhemispheric homotopic functional and structural connectivity in amyotrophic lateral sclerosis.

OBJECTIVE: Amyotrophic lateral sclerosis (ALS) is an idiopathic and fatal neurodegenerative disease of the human motor system. While microstructural alterations in corpus callosum (CC) have been identified as a consistent feature of ALS, studies directly examining interhemispheric neural connectivity are still lacking. To shed more light on the pathophysiology of ALS, the present study aims to examine alterations of interhemispheric structural and functional connectivity in individuals with ALS. METHODS: Diffusion tensor imaging (DTI) and resting-state functional MRI (rfMRI) data were acquired from 38 individuals with ALS and 35 gender-matched and age-matched control subjects. Indices of interhemispheric functional and structural neural connection were derived with analyses of voxel mirrored homotopic connectivity (VMHC) and probabilistic fibre tracking. RESULTS: The rfMRI has revealed extensive reductions of VMHC associated with ALS in brain regions of the precentral and postcentral gyrus, the paracentral lobule, the superior temporal gyrus, the middle cingulate gyrus, the putamen and the superior parietal lobules. With DTI, the analysis has also revealed reductions of interhemispheric structural connectivity through the CC subregions II, III and V in patients with ALS. Additionally, interhemispheric functional connectivity of the bilateral precentral gyri positively correlated with fractional anisotropy values of the CC subregion III, which structurally connects the bilateral motor cortices. CONCLUSIONS: The present data provided direct evidence confirming and extending the view of impaired interhemispheric neural communications mediated by CC, providing a new perspective for examinations and understanding the pathophysiology of ALS.

Mechanisms of circumferential gyral convolution in primate brains.

Mammalian cerebral cortices are characterized by elaborate convolutions. Radial convolutions exhibit homology across primate species and generally are easily identified in individuals of the same species. In contrast, circumferential convolutions vary across species as well as individuals of the same species. However, systematic study of circumferential convolution patterns is lacking. To address this issue, we utilized structural MRI (sMRI) and diffusion MRI (dMRI) data from primate brains. We quantified cortical thickness and circumferential convolutions on gyral banks in relation to axonal pathways and density along the gray matter/white matter boundaries. Based on these observations, we performed a series of computational simulations. Results demonstrated that the interplay of heterogeneous cortex growth and mechanical forces along axons plays a vital role in the regulation of circumferential convolutions. In contrast, gyral geometry controls the complexity of circumferential convolutions. These findings offer insight into the mystery of circumferential convolutions in primate brains.

Dynamic thalamus parcellation from resting-state fMRI data.

The thalamus is a relay center between various subcortical brain areas and the cerebral cortex with delineation of its constituent nuclei being of particular interest in many applications. While previous studies have demonstrated efficacy of connectivity-based thalamus segmentation, they used approaches that do not consider the dynamic nature of thalamo-cortical interactions. In this study, we explicitly exploited the dynamic variation of thalamo-cortical connections to identify different states of functional connectivity and performed state-specific thalamus parcellation. With normalized spectral clustering successively applied in temporal and spatial domains, nine thalamo-cortical connectivity states were identified and the dynamic thalamus parcellation revealed finer thalamic structures with improved atlas correspondence. The present results extend our understanding of thalamo-cortical connectivity and provide a more comprehensive view of the thalamo-cortical interaction.

Comparison of diffusion tractography and tract-tracing measures of connectivity strength in rhesus macaque connectome.

With the mapping of macroscale connectomes by means of in vivo diffusion-weighted MR Imaging (DWI) rapidly gaining in popularity, one of the necessary steps is the examination of metrics of connectivity strength derived from these reconstructions. In the field of human macroconnectomics the number of reconstructed fiber streamlines (NOS) is more and more used as a metric of cortico-cortical interareal connectivity strength, but the link between DWI NOS and in vivo animal tract-tracing measurements of anatomical connectivity strength remains poorly understood. In this technical report, we communicate on a comparison between DWI derived metrics and tract-tracing metrics of projection strength. Tract-tracing information on projection strength of interareal pathways was extracted from two commonly used macaque connectome datasets, including (1) the CoCoMac database of collated tract-tracing experiments of the macaque brain and (2) the high-resolution tract-tracing dataset of Markov and Kennedy and coworkers. NOS and density of reconstructed fiber pathways derived from DWI data acquired across 10 rhesus macaques was found to positively correlate to tract-tracing based measurements of connectivity strength across both the CoCoMac and Markov dataset (both P < 0.001), suggesting DWI NOS to form a valid method of assessment of the projection strength of white matter pathways. Our findings provide confidence of in vivo DWI connectome reconstructions to represent fairly realistic estimates of the wiring strength of white matter projections. Our cross-modal comparison supports the notion of in vivo DWI to be a valid methodology for robust description and interpretation of brain wiring.

Alterations of hippocampal projections in adult macaques with neonatal hippocampal lesions: a Diffusion Tensor Imaging study.

Neuropsychological and brain imaging studies have demonstrated persistent deficits in memory functions and structural changes after neonatal neurotoxic hippocampal lesion in monkeys. However, the relevant microstructural changes in the white matter of affected brain regions following this early insult remain unknown. This study assessed white matter integrity in the main hippocampal projections of adult macaque monkeys with neonatal hippocampal lesions, using diffusion tensor imaging (DTI). Data analysis was performed using tract-based spatial statistics (TBSS) and compared with volume of interest statistics. Alterations of fractional anisotropy (FA) and diffusivity indices were observed in fornix, temporal stem, ventromedial prefrontal cortex and optical radiations. To further validate the lesion effects on the prefrontal cortex, probabilistic diffusion tractography was used to examine the integrity of the fiber connections between hippocampus and ventromedial prefrontal cortex, and alterations were found in these connections. In addition, increased radial diffusivity in the left ventromedial prefrontal cortex correlated negatively with the severity of deficits in working memory in the same monkeys. The findings revealed microstructural changes due to neonatal hippocampal lesion, and confirmed that neonatal neurotoxic hippocampal lesions resulted in significant and enduring functional alterations in the hippocampal projection system.

PCLR: phase-constrained low-rank model for compressive diffusion-weighted MRI.

PURPOSE: This work develops a compressive sensing approach for diffusion-weighted (DW) MRI. THEORY AND METHODS: A phase-constrained low-rank (PCLR) approach was developed using the image coherence across the DW directions for efficient compressive DW MRI, while accounting for drastic phase changes across the DW directions, possibly as a result of eddy current, and rigid and nonrigid motions. In PCLR, a low-resolution phase estimation was used for removing phase inconsistency between DW directions. In our implementation, GRAPPA (generalized autocalibrating partial parallel acquisition) was incorporated for better phase estimation while allowing higher undersampling factor. An efficient and easy-to-implement image reconstruction algorithm, consisting mainly of partial Fourier update and singular value decomposition, was developed for solving PCLR. RESULTS: The error measures based on diffusion-tensor-derived metrics and tractography indicated that PCLR, with its joint reconstruction of all DW images using the image coherence, outperformed the frame-independent reconstruction through zero-padding FFT. Furthermore, using GRAPPA for phase estimation, PCLR readily achieved a four-fold undersampling. CONCLUSION: The PCLR is developed and demonstrated for compressive DW MRI. A four-fold reduction in k-space sampling could be readily achieved without substantial degradation of reconstructed images and diffusion tensor measures, making it possible to significantly reduce the data acquisition in DW MRI and/or improve spatial and angular resolutions.

Coevolution of gyral folding and structural connection patterns in primate brains.

Both cortical folding and structural connection patterns are more elaborated during the evolution of primate neocortex. For instance, cortical gyral shapes and structural connection patterns in humans are more complex and variable than those in chimpanzees and macaques. However, the intrinsic relationship between gyral folding and structural connection and their coevolution patterns across primates remain unclear. Here, our qualitative and quantitative analyses of in vivo diffusion tensor imaging (DTI) and structural magnetic resonance imaging (MRI) data consistently demonstrate that structural fiber connection patterns closely follow gyral folding patterns in the direction "tangent" to the cortical sphere, and this close relationship is well preserved in the neocortices of macaque, chimpanzee, and human brains, despite the progressively increasing complexity and variability of cortical folding and structural connection patterns. The findings suggest a hypothesis that a common axonal fiber pushing mechanism sculpts the curved patterns of gyri in the tangent direction during primate brain evolution. Our DTI/MRI data analysis provides novel insights into the structural architecture of primate brains, a new viewpoint of the relationship between cortical morphology and connection, and a basis for future elucidation of the functional implication of coevolution of cortical folding and structural connection patterns.

Psychosis spectrum symptoms among individuals with schizophrenia-associated copy number variants and evidence of cerebellar correlates of symptom severity.

The 3q29 deletion (3q29Del) is a copy number variant (CNV) with one of the highest effect sizes for psychosis-risk (>40-fold). Systematic research offers avenues for elucidating mechanism; however, compared to CNVs like 22q11.2Del, 3q29Del remains understudied. Emerging findings indicate that posterior fossa abnormalities are common among carriers, but their clinical relevance is unclear. We report the first in-depth evaluation of psychotic symptoms in participants with 3q29Del (N=23), using the Structured Interview for Psychosis-Risk Syndromes, and compare this profile to 22q11.2Del (N=31) and healthy controls (N=279). We also explore correlations between psychotic symptoms and posterior fossa abnormalities. Cumulatively, 48% of the 3q29Del sample exhibited a psychotic disorder or attenuated positive symptoms, with a subset meeting criteria for clinical high-risk. 3q29Del had more severe ratings than controls on all domains and only exhibited less severe ratings than 22q11.2Del in negative symptoms; ratings demonstrated select sex differences but no domain-wise correlations with IQ. An inverse relationship was identified between positive symptoms and cerebellar cortex volume in 3q29Del, documenting the first clinically-relevant neuroanatomical connection in this syndrome. Our findings characterize the profile of psychotic symptoms in the largest 3q29Del sample reported to date, contrast with another high-impact CNV, and highlight cerebellar involvement in psychosis-risk.

Mapping putative hubs in human, chimpanzee and rhesus macaque connectomes via diffusion tractography.

Mapping anatomical brain networks with graph-theoretic analysis of diffusion tractography has recently gained popularity, because of its presumed value in understanding brain function. However, this approach has seldom been used to compare brain connectomes across species, which may provide insights into brain evolution. Here, we employed a data-driven approach to compare interregional brain connections across three primate species: 1) the intensively studied rhesus macaque, 2) our closest living primate relative, the chimpanzee, and 3) humans. Specifically, we first used random parcellations and surface-based probabilistic diffusion tractography to derive the brain networks of the three species under various network densities and resolutions. We then compared the characteristics of the networks using graph-theoretic measures. In rhesus macaques, our tractography-defined hubs showed reasonable overlap with hubs previously identified using anterograde and retrograde tracer data. Across all three species, hubs were largely symmetric in the two hemispheres and were consistently identified in medial parietal, insular, retrosplenial cingulate and ventrolateral prefrontal cortices, suggesting a conserved structural architecture within these regions. However, species differences were observed in the inferior parietal cortex, polar and medial prefrontal cortices. The potential significance of these interspecies differences is discussed.

Axonal fiber terminations concentrate on gyri.

Convoluted cortical folding and neuronal wiring are 2 prominent attributes of the mammalian brain. However, the macroscale intrinsic relationship between these 2 general cross-species attributes, as well as the underlying principles that sculpt the architecture of the cerebral cortex, remains unclear. Here, we show that the axonal fibers connected to gyri are significantly denser than those connected to sulci. In human, chimpanzee, and macaque brains, a dominant fraction of axonal fibers were found to be connected to the gyri. This finding has been replicated in a range of mammalian brains via diffusion tensor imaging and high-angular resolution diffusion imaging. These results may have shed some lights on fundamental mechanisms for development and organization of the cerebral cortex, suggesting that axonal pushing is a mechanism of cortical folding.

Functional maturation in visual pathways predicts attention to the eyes in infant rhesus macaques: Effects of social status.

Differences in looking at the eyes of others are one of the earliest behavioral markers for social difficulties in neurodevelopmental disabilities, including autism. However, it is unknown how early visuo-social experiences relate to the maturation of infant brain networks that process visual social stimuli. We investigated functional connectivity (FC) within the ventral visual object pathway as a contributing neural system. Densely sampled, longitudinal eye-tracking and resting state fMRI (rs-fMRI) data were collected from infant rhesus macaques, an important model of human social development, from birth through 6 months of age. Mean trajectories were fit for both datasets and individual trajectories from subjects with both eye-tracking and rs-fMRI data were used to test for brain-behavior relationships. Exploratory findings showed infants with greater increases in FC between left V1 to V3 visual areas have an earlier increase in eye-looking before 2 months. This relationship was moderated by social status such that infants with low social status had a stronger association between left V1 to V3 connectivity and eye-looking than high status infants. Results indicated that maturation of the visual object pathway may provide an important neural substrate supporting adaptive transitions in social visual attention during infancy.

Quantitative assessment of a framework for creating anatomical brain networks via global tractography.

Interregional connections of the brain measured with diffusion tractography can be used to infer valuable information regarding both brain structure and function. However, different tractography algorithms can generate networks that exhibit different characteristics, resulting in poor reproducibility across studies. Therefore, it is important to benchmark different tractography algorithms to quantitatively assess their performance. Here we systematically evaluated a newly introduced tracking algorithm, global tractography, to derive anatomical brain networks in a fiber phantom, 2 post-mortem macaque brains, and 20 living humans, and compared the results with an established local tracking algorithm. Our results demonstrated that global tractography accurately characterized the phantom network in terms of graph-theoretic measures, and significantly outperformed the local tracking approach. Results in brain tissues (post-mortem macaques and in vivo humans), however, showed that although the performance of global tractography demonstrated a trend of improvement, the results were not vastly different than that of local tractography, possibly resulting from the increased fiber complexity of real tissues. When using macaque tracer-derived connections as the ground truth, we found that both global and local algorithms generated non-random patterns of false negative and false positive connections that were probably related to specific fiber systems and largely independent of the tractography algorithm or tissue type (post-mortem vs. in vivo) used in the current study. Moreover, a close examination of the transcallosal motor connections, reconstructed via either global or local tractography, demonstrated that the lateral transcallosal fibers in humans and macaques did not exhibit the denser homotopic connections found in primate tracer studies, indicating the need for more robust brain mapping techniques based on diffusion MRI data.

The effects of connection reconstruction method on the interregional connectivity of brain networks via diffusion tractography.

Estimating the interregional structural connections of the brain via diffusion tractography is a complex procedure and the parameters chosen can affect the outcome of the connectivity matrix. Here, we investigated the influence of different connection reconstruction methods on brain connectivity networks. Specifically, we applied three connection reconstruction methods to the same set of diffusion MRI data, initiating tracking from deep white matter (method #1, M1), from the gray matter/white matter interface (M2), and from the gray/white matter interface with thresholded tract volume rather than the connection probability as the connectivity index (M3). Small-world properties, hub identification, and hemispheric asymmetry in connectivity patterns were then calculated and compared across methods. Despite moderate to high correlations in the graph-theoretic measures across different methods, significant differences were observed in small-world indices, identified hubs, and hemispheric asymmetries, highlighting the importance of reconstruction method on network parameters. Consistent with the prior reports, the left precuneus was identified as a hub region in all three methods, suggesting it has a prominent role in brain networks.