Online interactive medical neuroimaging exercise to identify human brain structures.

A persisting need remains for developing methods for inspiring and teaching undergraduate medical students to quickly learn to identify the hundreds of human brain structures, tracts and spaces that are clinically relevant (viewed as three-dimensional volumes or two-dimensional neuroimages), and to accomplish this with the option of virtual on-line methods. This notably includes teaching the essentials of recommended diagnostic radiology to allow students to be familiar with patient neuroimages routinely acquired using magnetic resonance imaging (MRI) and computed tomography (CT). The present article includes a brief example video plus details a clinically oriented interactive neuroimaging exercise for first year medical students (MS1s) in small groups, conducted with instructors either in-person or as an entirely online virtual event. This "find-the-brain-structure" (FBS) event included teaching students to identify brain structures and other regions of interest in the central nervous system (and potentially in head and neck gross anatomy), which are traditionally taught using brain anatomy atlases and anatomical specimens. The interactive, small group exercise can be conducted in person or virtually on-line in as little as 30 min depending on the scope of objectives being covered. The learning exercise involves coordinated interaction between MS1s with one or several non-clinical faculty and may include one or several physicians (clinical faculty and/or qualified residents). It further allows for varying degrees of instructor interaction online and is easy to convey to instructors who do not have expertise in neuroimaging. Anonymous pre-event survey (n = 113, 100% response rate) versus post-event surveys (n = 92, 81% response rate) were attained from a cohort of MS1s in a neurobiology course. Results showed multiple statistically significant group-level shifts in response to several of the questions, showing an increase in MS1 confidence with reading MRI images (12% increase shift in mean, p < 0.001), confidence in their approaching physicians for medical training (9%, p < 0.01), and comfort levels in working online with virtual team-based peers and with team-based faculty (6%, p < 0.05). Qualitative student feedback revealed highly positive comments regarding the experience overall, encouraging this virtual medium as a desirable educational approach.

Early construction of the thalamocortical axon pathway requires c-Jun N-terminal kinase signaling within the ventral forebrain.

BACKGROUND: Thalamocortical connectivity is essential for normal brain function. This important pathway is established during development, when thalamic axons extend a long distance through the forebrain before reaching the cerebral cortex. In this study, we identify a novel role for the c-Jun N-terminal kinase (JNK) signaling pathway in guiding thalamocortical axons through intermediate target territories. RESULTS: Complete genetic removal of JNK signaling from the Distal-less 5/6 (Dlx5/6) domain in mice prevents thalamocortical axons from crossing the diencephalon-telencephalon boundary (DTB) and the internal capsule fails to form. Ventral telencephalic cells critical for thalamocortical axon extensions including corridor and guidepost neurons are also disrupted. In addition, corticothalamic, striatonigral, and nigrostriatal axons fail to cross the DTB. Analyses of different JNK mutants demonstrate that thalamocortical axon pathfinding has a non-autonomous requirement for JNK signaling. CONCLUSIONS: We conclude that JNK signaling within the Dlx5/6 territory enables the construction of major axonal pathways in the developing forebrain. Further exploration of this intermediate axon guidance territory is needed to uncover mechanisms of axonal pathfinding during normal brain development and to elucidate how this vital process may be compromised in neurodevelopmental disorders.

JNK Signaling Regulates Cellular Mechanics of Cortical Interneuron Migration.

Aberrant migration of inhibitory interneurons can alter the formation of cortical circuitry and lead to severe neurologic disorders including epilepsy, autism, and schizophrenia. However, mechanisms involved in directing the migration of interneurons remain incompletely understood. Using a mouse model, we performed live-cell confocal microscopy to explore the mechanisms by which the c-Jun NH2-terminal kinase (JNK) pathway coordinates leading process branching and nucleokinesis, two cell biological processes that are essential for the guided migration of cortical interneurons. Pharmacological inhibition of JNK signaling disrupts the kinetics of leading process branching, rate and amplitude of nucleokinesis, and leads to the rearward mislocalization of the centrosome and primary cilium to the trailing process. Genetic loss of Jnk from interneurons also impairs leading process branching and nucleokinesis, suggesting that important mechanics of interneuron migration depend on the intrinsic activity of JNK. These findings highlight key roles for JNK signaling in leading process branching, nucleokinesis, and the trafficking of centrosomes and cilia during interneuron migration, and further implicates JNK signaling as an important mediator of cortical development.

Functional and resting-state characterizations of a periventricular heterotopic nodule associated with epileptogenic activity.

The object of this study was to extensively characterize a region of periventricular nodular heterotopia (PVNH) in an epilepsy patient to reveal its possible neurocognitive functional role(s). The authors used 3-T MRI approaches to exhaustively characterize a single, right hemisphere heterotopion in a high-functioning adult male with medically responsive epilepsy, which had manifested during late adolescence. The heterotopion proved to be spectroscopically consistent with a cortical-like composition and was interconnected with nearby ipsilateral cortical fundi, as revealed by fiber tractography (diffusion-weighted imaging) and resting-state functional connectivity MRI (rsfMRI). Moreover, the region of PVNH demonstrated two novel characterizations for a heterotopion. First, functional MRI (fMRI), as distinct from rsfMRI, showed that the heterotopion was significantly modulated while the patient watched animated video scenes of biological motion (i.e., cartoons). Second, rsfMRI, which demonstrated correlated brain activity during a task-negative state, uniquely showed directionality within an interconnected network, receiving positive path effects from patent cortical and cerebellar foci while outputting only negative path effects to specific brain foci.These findings are addressed in the context of the impact on noninvasive presurgical brain mapping strategies for adult and pediatric patient workups, as well as the impact of this study on an understanding of the functional cortical architecture underlying cognition from a neurodiversity and evolutionary perspective.

JNK signaling is required for proper tangential migration and laminar allocation of cortical interneurons.

The precise migration of cortical interneurons is essential for the formation and function of cortical circuits, and disruptions to this key developmental process are implicated in the etiology of complex neurodevelopmental disorders, including schizophrenia, autism and epilepsy. We have recently identified the Jun N-terminal kinase (JNK) pathway as an important mediator of cortical interneuron migration in mice, regulating the proper timing of interneuron arrival into the cortical rudiment. In the current study, we demonstrate a vital role for JNK signaling at later stages of corticogenesis, when interneurons transition from tangential to radial modes of migration. Pharmacological inhibition of JNK signaling in ex vivo slice cultures caused cortical interneurons to rapidly depart from migratory streams and prematurely enter the cortical plate. Similarly, genetic loss of JNK function led to precocious stream departure ex vivo, and stream disruption, morphological changes and abnormal allocation of cortical interneurons in vivo These data suggest that JNK signaling facilitates the tangential migration and laminar deposition of cortical interneurons, and further implicates the JNK pathway as an important regulator of cortical development.

Mutations in ARL2BP, a protein required for ciliary microtubule structure, cause syndromic male infertility in humans and mice.

Cilia are evolutionarily conserved hair-like structures with a wide spectrum of key biological roles, and their dysfunction has been linked to a growing class of genetic disorders, known collectively as ciliopathies. Many strides have been made towards deciphering the molecular causes for these diseases, which have in turn expanded the understanding of cilia and their functional roles. One recently-identified ciliary gene is ARL2BP, encoding the ADP-Ribosylation Factor Like 2 Binding Protein. In this study, we have identified multiple ciliopathy phenotypes associated with mutations in ARL2BP in human patients and in a mouse knockout model. Our research demonstrates that spermiogenesis is impaired, resulting in abnormally shaped heads, shortened and mis-assembled sperm tails, as well as in loss of axonemal doublets. Additional phenotypes in the mouse included enlarged ventricles of the brain and situs inversus. Mouse embryonic fibroblasts derived from knockout animals revealed delayed depolymerization of primary cilia. Our results suggest that ARL2BP is required for the structural maintenance of cilia as well as of the sperm flagellum, and that its deficiency leads to syndromic ciliopathy.

Traumatic brain injury and epilepsy: Underlying mechanisms leading to seizure.

Post-traumatic epilepsy continues to be a major concern for those experiencing traumatic brain injury. Post-traumatic epilepsy accounts for 10-20% of epilepsy cases in the general population. While seizure prophylaxis can prevent early onset seizures, no available treatments effectively prevent late-onset seizure. Little is known about the progression of neural injury over time and how this injury progression contributes to late onset seizure development. In this comprehensive review, we discuss the epidemiology and risk factors for post-traumatic epilepsy and the current pharmacologic agents used for treatment. We highlight limitations with the current approach and offer suggestions for remedying the knowledge gap. Critical to this pursuit is the design of pre-clinical models to investigate important mechanistic factors responsible for post-traumatic epilepsy development. We discuss what the current models have provided in terms of understanding acute injury and what is needed to advance understanding regarding late onset seizure. New model designs will be used to investigate novel pathways linking acute injury to chronic changes within the brain. Important components of this transition are likely mediated by toll-like receptors, neuroinflammation, and tauopathy. In the final section, we highlight current experimental therapies that may prove promising in preventing and treating post-traumatic epilepsy. By increasing understanding about post-traumatic epilepsy and injury expansion over time, it will be possible to design better treatments with specific molecular targets to prevent late-onset seizure occurrence following traumatic brain injury.

Modeling a model: Mouse genetics, 22q11.2 Deletion Syndrome, and disorders of cortical circuit development.

Understanding the developmental etiology of autistic spectrum disorders, attention deficit/hyperactivity disorder and schizophrenia remains a major challenge for establishing new diagnostic and therapeutic approaches to these common, difficult-to-treat diseases that compromise neural circuits in the cerebral cortex. One aspect of this challenge is the breadth and overlap of ASD, ADHD, and SCZ deficits; another is the complexity of mutations associated with each, and a third is the difficulty of analyzing disrupted development in at-risk or affected human fetuses. The identification of distinct genetic syndromes that include behavioral deficits similar to those in ASD, ADHC and SCZ provides a critical starting point for meeting this challenge. We summarize clinical and behavioral impairments in children and adults with one such genetic syndrome, the 22q11.2 Deletion Syndrome, routinely called 22q11DS, caused by micro-deletions of between 1.5 and 3.0 MB on human chromosome 22. Among many syndromic features, including cardiovascular and craniofacial anomalies, 22q11DS patients have a high incidence of brain structural, functional, and behavioral deficits that reflect cerebral cortical dysfunction and fall within the spectrum that defines ASD, ADHD, and SCZ. We show that developmental pathogenesis underlying this apparent genetic "model" syndrome in patients can be defined and analyzed mechanistically using genomically accurate mouse models of the deletion that causes 22q11DS. We conclude that "modeling a model", in this case 22q11DS as a model for idiopathic ASD, ADHD and SCZ, as well as other behavioral disorders like anxiety frequently seen in 22q11DS patients, in genetically engineered mice provides a foundation for understanding the causes and improving diagnosis and therapy for these disorders of cortical circuit development.

Cortical interneurons require Jnk1 to enter and navigate the developing cerebral cortex.

Proper assembly of cortical circuitry relies on the correct migration of cortical interneurons from their place of birth in the ganglionic eminences to their place of terminal differentiation in the cerebral cortex. Although molecular mechanisms mediating cortical interneuron migration have been well studied, intracellular signals directing their migration are largely unknown. Here we illustrate a novel and essential role for c-Jun N-terminal kinase (JNK) signaling in guiding the pioneering population of cortical interneurons into the mouse cerebral cortex. Migrating cortical interneurons express Jnk proteins at the entrance to the cortical rudiment and have enriched expression of Jnk1 relative to noninterneuronal cortical cells. Pharmacological blockade of JNK signaling in ex vivo slice cultures resulted in dose-dependent and highly specific disruption of interneuron migration into the nascent cortex. Time-lapse imaging revealed that JNK-inhibited cortical interneurons advanced slowly and assumed aberrant migratory trajectories while traversing the cortical entry zone. In vivo analyses of JNK-deficient embryos supported our ex vivo pharmacological data. Deficits in interneuron migration were observed in Jnk1 but not Jnk2 single nulls, and those migratory deficits were further exacerbated when homozygous loss of Jnk1 was combined with heterozygous reduction of Jnk2. Finally, genetic ablation of Jnk1 and Jnk2 from cortical interneurons significantly perturbed migration in vivo, but not in vitro, suggesting JNK activity functions to direct their guidance rather than enhance their motility. These data suggest JNK signaling, predominantly mediated by interneuron expressed Jnk1, is required for guiding migration of cortical interneurons into and within the developing cerebral cortex.

Cxcr4 regulation of interneuron migration is disrupted in 22q11.2 deletion syndrome.

Interneurons are thought to be a primary pathogenic target for several behavioral disorders that arise during development, including schizophrenia and autism. It is not known, however, whether genetic lesions associated with these diseases disrupt established molecular mechanisms of interneuron development. We found that diminished 22q11.2 gene dosage-the primary genetic lesion in 22q11.2 deletion syndrome (22q11.2 DS)-specifically compromises the distribution of early-generated parvalbumin-expressing interneurons in the Large Deletion (LgDel) 22q11.2DS mouse model. This change reflects cell-autonomous disruption of interneuron migration caused by altered expression of the cytokine C-X-C chemokine receptor type 4 (Cxcr4), an established regulator of this process. Cxcr4 is specifically reduced in LgDel migrating interneurons, and genetic analysis confirms that diminished Cxcr4 alters interneuron migration in LgDel mice. Thus, diminished 22q11.2 gene dosage disrupts cortical circuit development by modifying a critical molecular signaling pathway via Cxcr4 that regulates cortical interneuron migration and placement.

Proliferative and transcriptional identity of distinct classes of neural precursors in the mammalian olfactory epithelium.

Neural precursors in the developing olfactory epithelium (OE) give rise to three major neuronal classes - olfactory receptor (ORNs), vomeronasal (VRNs) and gonadotropin releasing hormone (GnRH) neurons. Nevertheless, the molecular and proliferative identities of these precursors are largely unknown. We characterized two precursor classes in the olfactory epithelium (OE) shortly after it becomes a distinct tissue at midgestation in the mouse: slowly dividing self-renewing precursors that express Meis1/2 at high levels, and rapidly dividing neurogenic precursors that express high levels of Sox2 and Ascl1. Precursors expressing high levels of Meis genes primarily reside in the lateral OE, whereas precursors expressing high levels of Sox2 and Ascl1 primarily reside in the medial OE. Fgf8 maintains these expression signatures and proliferative identities. Using electroporation in the wild-type embryonic OE in vitro as well as Fgf8, Sox2 and Ascl1 mutant mice in vivo, we found that Sox2 dose and Meis1 - independent of Pbx co-factors - regulate Ascl1 expression and the transition from lateral to medial precursor state. Thus, we have identified proliferative characteristics and a dose-dependent transcriptional network that define distinct OE precursors: medial precursors that are most probably transit amplifying neurogenic progenitors for ORNs, VRNs and GnRH neurons, and lateral precursors that include multi-potent self-renewing OE neural stem cells.

Diminished dosage of 22q11 genes disrupts neurogenesis and cortical development in a mouse model of 22q11 deletion/DiGeorge syndrome.

The 22q11 deletion (or DiGeorge) syndrome (22q11DS), the result of a 1.5- to 3-megabase hemizygous deletion on human chromosome 22, results in dramatically increased susceptibility for "diseases of cortical connectivity" thought to arise during development, including schizophrenia and autism. We show that diminished dosage of the genes deleted in the 1.5-megabase 22q11 minimal critical deleted region in a mouse model of 22q11DS specifically compromises neurogenesis and subsequent differentiation in the cerebral cortex. Proliferation of basal, but not apical, progenitors is disrupted, and subsequently, the frequency of layer 2/3, but not layer 5/6, projection neurons is altered. This change is paralleled by aberrant distribution of parvalbumin-labeled interneurons in upper and lower cortical layers. Deletion of Tbx1 or Prodh (22q11 genes independently associated with 22q11DS phenotypes) does not similarly disrupt basal progenitors. However, expression analysis implicates additional 22q11 genes that are selectively expressed in cortical precursors. Thus, diminished 22q11 gene dosage disrupts cortical neurogenesis and interneuron migration. Such developmental disruption may alter cortical circuitry and establish vulnerability for developmental disorders, including schizophrenia and autism.

Molecular specification and patterning of progenitor cells in the lateral and medial ganglionic eminences.

We characterized intrinsic and extrinsic specification of progenitors in the lateral and medial ganglionic eminences (LGE and MGE). We identified seven genes whose expression is enriched or restricted in either the LGE [biregional cell adhesion molecule-related/downregulated by oncogenes binding protein (Boc), Frizzled homolog 8 (Fzd8), Ankrd43 (ankyrin repeat domain-containing protein 43), and Ikzf1 (Ikaros family zinc finger 1)] or MGE [Map3k12 binding inhibitory protein 1 (Mbip); zinc-finger, SWIM domain containing 5 (Zswim5); and Adamts5 [a disintegrin-like and metallopeptidase (reprolysin type) with thrombospondin type 1 motif, 5]]. Boc, Fzd8, Mbip, and Zswim5 are apparently expressed in LGE or MGE progenitors, whereas the remaining three are seen in the postmitotic mantle zone. Relative expression levels are altered and regional distinctions are lost for each gene in LGE or MGE cells propagated as neurospheres, indicating that these newly identified molecular characteristics of LGE or MGE progenitors depend on forebrain signals not available in the neurosphere assay. Analyses of Pax6(Sey/Sey), Shh(-/-), and Gli3(XtJ/XtJ) mutants suggests that LGE and MGE progenitor identity does not rely exclusively on previously established forebrain-intrinsic patterning mechanisms. Among a limited number of additional potential patterning mechanisms, we found that extrinsic signals from the frontonasal mesenchyme are essential for Shh- and Fgf8-dependent regulation of LGE and MGE genes. Thus, extrinsic and intrinsic forebrain patterning mechanisms cooperate to establish LGE and MGE progenitor identity, and presumably their capacities to generate distinct classes of neuronal progeny.

Position and time specify the migration of a pioneering population of olfactory bulb interneurons.

We defined the cellular mechanisms for genesis, migration, and differentiation of the initial population of olfactory bulb (OB) interneurons. This cohort of early generated cells, many of which become postmitotic on embryonic day (E) 14.5, differentiates into a wide range of mature OB interneurons by postnatal day (P) 21, and a substantial number remains in the OB at P60. Their precursors autonomously acquire a distinct identity defined by their position in the lateral ganglionic eminence (LGE). The progeny migrate selectively to the OB rudiment in a pathway that presages the rostral migratory stream. After arriving in the OB rudiment, these early generated cells acquire cellular and molecular hallmarks of OB interneurons. Other precursors--including those from the medial ganglionic eminence (MGE) and OB--fail to generate neuroblasts with similar migratory capacity when transplanted to the LGE. The positional identity and migratory specificity of the LGE precursors is rigidly established between E12.5 and E14.5. Thus, the pioneering population of OB interneurons is generated from spatially and temporally determined LGE precursors whose progeny uniquely recognize a distinct migratory trajectory.

Bidirectional influences between neurons and glial cells in the developing olfactory system.

Olfactory systems serve as excellent model systems for the study of numerous widespread aspects of neural development and also for the elucidation of features peculiar to the formation of neural circuits specialized to process odor inputs. Accumulated research reveals a fine balance between developmental autonomy of olfactory structures and intercellular interactions essential for their normal development. Recent findings have uncovered evidence for more autonomy than previously realized, but simultaneously have begun to reveal the complex cellular and molecular underpinnings of key interactions among neurons and glial cells at several important steps in olfactory development. Striking similarities in the functional organization of olfactory systems across vertebrate and invertebrate species allow the advantages of different species to be used to address common issues. Our own work in the moth Manduca sexta has demonstrated reciprocal neuron-glia interactions that have key importance in two aspects of development, the sorting of olfactory receptor axons into fascicles targeted for specific glomeruli and the creation of glomeruli. Studies in vertebrate species suggest that similar neuron-glia interactions may underlie olfactory development, although here the roles have not been tested so directly. Similar cellular interactions also are likely to play roles in development of some other systems in which axons of intermixed neurons must sort according to target specificity and systems in which reiterated modules of synaptic neuropil develop.

In vitro analyses of interactions between olfactory receptor growth cones and glial cells that mediate axon sorting and glomerulus formation.

During development, the axons of olfactory receptor neurons project to the CNS and converge on glomerular targets. For vertebrate and invertebrate olfactory systems, neuron-glia interactions have been hypothesized to regulate the sorting and targeting of olfactory receptor axons and the development of glomeruli. In the moth Manduca sexta, glial reduction experiments have directly implicated two types of central olfactory glia, the sorting zone- and neuropil-associated glia, in key events in olfactory development, including axon sorting and glomerulus stabilization. By using cocultures containing central olfactory glial cells and explants of olfactory receptor epithelium, we show that olfactory receptor growth cones elaborate extensively and cease advancement following contact with sorting zone- and neuropil-associated glial cells. These effects on growth cone behavior were specific to central olfactory glia; peripheral glial cells of the olfactory nerve failed to elicit similar responses in olfactory receptor growth cones. We propose that sorting zone- and neuropil-associated glial cells similarly modify axon behavior in vitro by altering the adhesive properties and cytoskeleton of olfactory receptor growth cones and that these in vitro changes may underlie functionally relevant changes in growth cone behavior in vivo.

Reciprocal interactions between olfactory receptor axons and olfactory nerve glia cultured from the developing moth Manduca sexta.

In olfactory systems, neuron-glia interactions have been implicated in the growth and guidance of olfactory receptor axons. In the moth Manduca sexta, developing olfactory receptor axons encounter several types of glia as they grow into the brain. Antennal nerve glia are born in the periphery and enwrap bundles of olfactory receptor axons in the antennal nerve. Although their peripheral origin and relationship with axon bundles suggest that they share features with mammalian olfactory ensheathing cells, the developmental roles of antennal nerve glia remain elusive. When cocultured with antennal nerve glial cells, olfactory receptor growth cones readily advance along glial processes without displaying prolonged changes in morphology. In turn, olfactory receptor axons induce antennal nerve glial cells to form multicellular arrays through proliferation and process extension. In contrast to antennal nerve glia, centrally derived glial cells from the axon sorting zone and antennal lobe never form arrays in vitro, and growth-cone glial-cell encounters with these cells halt axon elongation and cause permanent elaborations in growth cone morphology. We propose that antennal nerve glia play roles similar to olfactory ensheathing cells in supporting axon elongation, yet differ in their capacity to influence axon guidance, sorting, and targeting, roles that could be played by central olfactory glia in Manduca.