Profiling learning strategies of medical students: A person-centered approach.

BACKGROUND: Students within a cohort might employ unique subsets of learning strategies (LS) to study. However, little research has aimed to elucidate subgroup-specific LS usage among medical students. Recent methodological developments, particularly person-centred approaches such as latent profile analysis (LPA), offer ways to identify relevant subgroups with dissimilar patterns of LS use. In this paper, we apply LPA to explore subgroups of medical students during preclinical training in anatomy and examine how these patterns are linked with learning outcomes. METHODS: We analysed the LS used by 689 undergraduate, 1st and 2nd-year medical students across 6 German universities who completed the short version of the Learning Strategies of University Students (LIST-K) questionnaire, and answered questions towards external criteria such as learning resources and performance. We used the thirteen different LS facets of the LIST-K (four cognitive, three metacognitive, three management of internal and three management of external resources) as LPA indicators. RESULTS: Based on LPA, students can be grouped into four distinct learning profiles: Active learners (45% of the cohort), collaborative learners (17%), structured learners (29%) and passive learners (9%). Students in each of those latent profiles combine the 13 LS facets in a unique way to study anatomy. The profiles differ in both, the overall level of LS usage, and unique combinations of LS used for learning. Importantly, we find that the facets of LS show heterogeneous and subgroup-specific correlations with relevant outcome criteria, which partly overlap but mostly diverge from effects observed on the population level. CONCLUSIONS: The effects observed by LPA expand results from variable-centered efforts and challenge the notion that LS operate on a linear continuum. These results highlight the heterogeneity between subgroups of learners and help generate a more nuanced interpretation of learning behaviour. Lastly, our analysis offers practical implications for educators seeking to tailor learning experiences to meet individual student needs.

Presynaptic α2δ subunits are key organizers of glutamatergic synapses.

In nerve cells the genes encoding for α2δ subunits of voltage-gated calcium channels have been linked to synaptic functions and neurological disease. Here we show that α2δ subunits are essential for the formation and organization of glutamatergic synapses. Using a cellular α2δ subunit triple-knockout/knockdown model, we demonstrate a failure in presynaptic differentiation evidenced by defective presynaptic calcium channel clustering and calcium influx, smaller presynaptic active zones, and a strongly reduced accumulation of presynaptic vesicle-associated proteins (synapsin and vGLUT). The presynaptic defect is associated with the downscaling of postsynaptic AMPA receptors and the postsynaptic density. The role of α2δ isoforms as synaptic organizers is highly redundant, as each individual α2δ isoform can rescue presynaptic calcium channel trafficking and expression of synaptic proteins. Moreover, α2δ-2 and α2δ-3 with mutated metal ion-dependent adhesion sites can fully rescue presynaptic synapsin expression but only partially calcium channel trafficking, suggesting that the regulatory role of α2δ subunits is independent from its role as a calcium channel subunit. Our findings influence the current view on excitatory synapse formation. First, our study suggests that postsynaptic differentiation is secondary to presynaptic differentiation. Second, the dependence of presynaptic differentiation on α2δ implicates α2δ subunits as potential nucleation points for the organization of synapses. Finally, our results suggest that α2δ subunits act as transsynaptic organizers of glutamatergic synapses, thereby aligning the synaptic active zone with the postsynaptic density.

Interprofessional socialization of first-year medical and midwifery students: effects of an ultra-brief anatomy training.

BACKGROUND: Interprofessionalism is considered a key component in modern health profession education. Nevertheless, there remains ongoing debate about when and where to introduce interprofessional trainings in the curriculum. We identified anatomy, a subject commonly shared among health professionals, as a practical choice for initiating early intergroup-contact between first-year medical and midwifery students. Our study examined the effects of a four-hour block course in anatomy on interprofessional socialization and valuing, as well as long-term effects on intergroup contact. METHODS: Based on different concepts and theories of learning, we implemented 12 interprofessional learning stations. Several measures were taken to foster group cohesion: (1) self-directed working in interprofessional tandems on authentic obstetric tasks, (2) competing with other tandems, (3) creating positive interdependencies during task completion, and (4) allowing room for networking. In a pre-post design with a three-month follow-up, we assessed the outcomes of this ultra-brief training with qualitative essays and quantitative scales. RESULTS: After training, both groups improved in interprofessionalism scores with strong effect sizes, mean difference in ISVS-21 = 0.303 [95% CI: 0.120, 0.487], P < .001, η² = 0.171, while the scales measuring uniprofessional identity were unaffected, mean difference in MCPIS = 0.033 [95% CI: -0.236, 0.249], P = .789. A follow-up indicated that these positive short-term effects on the ISVS-21 scale diminished after 12 weeks to baseline levels, yet, positive intergroup contact was still reported. The qualitative findings revealed that, at this initial stage of their professional identity development, both medical and midwifery students considered interprofessionalism, teamwork and social competencies to be of importance for their future careers. CONCLUSION: This study advocates for an early implementation of interprofessional learning objectives in anatomical curricula. Young health profession students are receptive to interprofessional collaboration at this initial stage of their professional identity and derive strong advantages from a concise training approach. Yet, maintaining these gains over time may require ongoing support and reinforcement, such as through longitudinal curricula. We believe that an interprofessional socialization at an early stage can help break down barriers, and help to avoid conflicts that may arise during traditional monoprofessional curricula.

Distinct Alterations in Dendritic Spine Morphology in the Absence of ß-Neurexins.

Dendritic spines are essential for synaptic function because they constitute the postsynaptic compartment of the neurons that receives the most excitatory input. The extracellularly shorter variant of the presynaptic cell adhesion molecules neurexins, ß-neurexin, has been implicated in various aspects of synaptic function, including neurotransmitter release. However, its role in developing or stabilizing dendritic spines as fundamental computational units of excitatory synapses has remained unclear. Here, we show through morphological analysis that the deletion of ß-neurexins in hippocampal neurons in vitro and in hippocampal tissue in vivo affects presynaptic dense-core vesicles, as hypothesized earlier, and, unexpectedly, alters the postsynaptic spine structure. Specifically, we observed that the absence of ß-neurexins led to an increase in filopodial-like protrusions in vitro and more mature mushroom-type spines in the CA1 region of adult knockout mice. In addition, the deletion of ß-neurexins caused alterations in the spine head dimension and an increase in spines with perforations of their postsynaptic density but no changes in the overall number of spines or synapses. Our results indicate that presynaptic ß-neurexins play a role across the synaptic cleft, possibly by aligning with postsynaptic binding partners and glutamate receptors via transsynaptic columns.

PI(4,5)P2 controls slit diaphragm formation and endocytosis in Drosophila nephrocytes.

Drosophila nephrocytes are an emerging model system for mammalian podocytes and proximal tubules as well as for the investigation of kidney diseases. Like podocytes, nephrocytes exhibit characteristics of epithelial cells, but the role of phospholipids in polarization of these cells is yet unclear. In epithelia, phosphatidylinositol(4,5)bisphosphate (PI(4,5)P2) and phosphatidylinositol(3,4,5)-trisphosphate (PI(3,4,5)P3) are asymmetrically distributed in the plasma membrane and determine apical-basal polarity. Here, we demonstrate that both phospholipids are present in the plasma membrane of nephrocytes, but only PI(4,5)P2 accumulates at slit diaphragms. Knockdown of Skittles, a phosphatidylinositol(4)phosphate 5-kinase, which produces PI(4,5)P2, abolished slit diaphragm formation and led to strongly reduced endocytosis. Notably, reduction in PI(3,4,5)P3 by overexpression of PTEN or expression of a dominant-negative phosphatidylinositol-3-kinase did not affect nephrocyte function, whereas enhanced formation of PI(3,4,5)P3 by constitutively active phosphatidylinositol-3-kinase resulted in strong slit diaphragm and endocytosis defects by ectopic activation of the Akt/mTOR pathway. Thus, PI(4,5)P2 but not PI(3,4,5)P3 is essential for slit diaphragm formation and nephrocyte function. However, PI(3,4,5)P3 has to be tightly controlled to ensure nephrocyte development.

Medical imaging training with eye movement modeling examples: A randomized controlled study.

PURPOSE: To determine whether ultrasound training in which the expert's eye movements are superimposed to the underlying ultrasound video (eye movement modeling examples; EMMEs) leads to better learner outcomes than traditional eye movement-free instructions. MATERIALS AND METHODS: 106 undergraduate medical students were randomized in two groups; 51 students in the EMME group watched 5-min ultrasound examination videos combined with the eye movements of an expert performing the task. The identical videos without the eye movements were shown to 55 students in the control group. Performance and behavioral parameters were compared prepost interventional using ANOVAs. Additionally, cognitive load, and prior knowledge in anatomy were surveyed. RESULTS: After training, the EMME group identified more sonoanatomical structures correctly, and completed the tasks faster than the control group. This effect was partly mediated by a reduction of extraneous cognitive load. Participants with greater prior anatomical knowledge benefited the most from the EMME training. CONCLUSION: Displaying experts' eye movements in medical imaging training appears to be an effective way to foster medical interpretation skills of undergraduate medical students. One underlying mechanism might be that practicing with eye movements reduces cognitive load and helps learners activate their prior knowledge.

Neurexin 1 variants as risk factors for suicide death.

Suicide is a significant public health concern with complex etiology. Although the genetic component of suicide is well established, the scope of gene networks and biological mechanisms underlying suicide has yet to be defined. Previously, we reported genome-wide evidence that neurexin 1 (NRXN1), a key synapse organizing molecule, is associated with familial suicide risk. Here we present new evidence for two non-synonymous variants (rs78540316; P469S and rs199784139; H885Y) associated with increased familial risk of suicide death. We tested the impact of these variants on binding interactions with known partners and assessed functionality in a hemi-synapse formation assay. Although the formation of hemi-synapses was not altered with the P469S variant relative to wild-type, both variants increased binding to the postsynaptic binding partner, leucine-rich repeat transmembrane neuronal 2 (LRRTM2) in vitro. Our findings indicate that variants in NRXN1 and related synaptic genes warrant further study as risk factors for suicide death.

Apical-basal polarity regulators are essential for slit diaphragm assembly and endocytosis in Drosophila nephrocytes.

Apical-basal polarity is a key feature of most epithelial cells and it is regulated by highly conserved protein complexes. In mammalian podocytes, which emerge from columnar epithelial cells, this polarity is preserved and the tight junctions are converted to the slit diaphragms, establishing the filtration barrier. In Drosophila, nephrocytes show several structural and functional similarities with mammalian podocytes and proximal tubular cells. However, in contrast to podocytes, little is known about the role of apical-basal polarity regulators in these cells. In this study, we used expansion microscopy and found the apical polarity determinants of the PAR/aPKC and Crb-complexes to be predominantly targeted to the cell cortex in proximity to the nephrocyte diaphragm, whereas basolateral regulators also accumulate intracellularly. Knockdown of PAR-complex proteins results in severe endocytosis and nephrocyte diaphragm defects, which is due to impaired aPKC recruitment to the plasma membrane. Similar, downregulation of most basolateral polarity regulators disrupts Nephrin localization but had surprisingly divergent effects on endocytosis. Our findings suggest that morphology and slit diaphragm assembly/maintenance of nephrocytes is regulated by classical apical-basal polarity regulators, which have distinct functions in endocytosis.

α2δ-4 and Cachd1 Proteins Are Regulators of Presynaptic Functions.

The α2δ auxiliary subunits of voltage-gated calcium channels (VGCC) were traditionally regarded as modulators of biophysical channel properties. In recent years, channel-independent functions of these subunits, such as involvement in synapse formation, have been identified. In the central nervous system, α2δ isoforms 1, 2, and 3 are strongly expressed, regulating glutamatergic synapse formation by a presynaptic mechanism. Although the α2δ-4 isoform is predominantly found in the retina with very little expression in the brain, it was recently linked to brain functions. In contrast, Cachd1, a novel α2δ-like protein, shows strong expression in brain, but its function in neurons is not yet known. Therefore, we aimed to investigate the presynaptic functions of α2δ-4 and Cachd1 by expressing individual proteins in cultured hippocampal neurons. Both α2δ-4 and Cachd1 are expressed in the presynaptic membrane and could rescue a severe synaptic defect present in triple knockout/knockdown neurons that lacked the α2δ-1-3 isoforms (α2δ TKO/KD). This observation suggests that presynaptic localization and the regulation of synapse formation in glutamatergic neurons is a general feature of α2δ proteins. In contrast to this redundant presynaptic function, α2δ-4 and Cachd1 differentially regulate the abundance of presynaptic calcium channels and the amplitude of presynaptic calcium transients. These functional differences may be caused by subtle isoform-specific differences in α1-α2δ protein-protein interactions, as revealed by structural homology modelling. Taken together, our study identifies both α2δ-4 and Cachd1 as presynaptic regulators of synapse formation, differentiation, and calcium channel functions that can at least partially compensate for the loss of α2δ-1-3. Moreover, we show that regulating glutamatergic synapse formation and differentiation is a critical and surprisingly redundant function of α2δ and Cachd1.

Auxiliary α2δ1 and α2δ3 Subunits of Calcium Channels Drive Excitatory and Inhibitory Neuronal Network Development.

VGCCs are multisubunit complexes that play a crucial role in neuronal signaling. Auxiliary α2δ subunits of VGCCs modulate trafficking and biophysical properties of the pore-forming α1 subunit and trigger excitatory synaptogenesis. Alterations in the expression level of α2δ subunits were implicated in several syndromes and diseases, including chronic neuropathic pain, autism, and epilepsy. However, the contribution of distinct α2δ subunits to excitatory/inhibitory imbalance and aberrant network connectivity characteristic for these pathologic conditions remains unclear. Here, we show that α2δ1 overexpression enhances spontaneous neuronal network activity in developing and mature cultures of hippocampal neurons. In contrast, overexpression, but not downregulation, of α2δ3 enhances neuronal firing in immature cultures, whereas later in development it suppresses neuronal activity. We found that α2δ1 overexpression increases excitatory synaptic density and selectively enhances presynaptic glutamate release, which is impaired on α2δ1 knockdown. Overexpression of α2δ3 increases the excitatory synaptic density as well but also facilitates spontaneous GABA release and triggers an increase in the density of inhibitory synapses, which is accompanied by enhanced axonaloutgrowth in immature interneurons. Together, our findings demonstrate that α2δ1 and α2δ3 subunits play distinct but complementary roles in driving formation of structural and functional network connectivity during early development. An alteration in α2δ surface expression during critical developmental windows can therefore play a causal role and have a profound impact on the excitatory-to-inhibitory balance and network connectivity.SIGNIFICANCE STATEMENT The computational capacity of neuronal networks is determined by their connectivity. Chemical synapses are the main interface for transfer of information between individual neurons. The initial formation of network connectivity requires spontaneous electrical activity and the calcium channel-mediated signaling. We found that, in early development, auxiliary α2δ3 subunits of calcium channels foster presynaptic release of GABA, trigger formation of inhibitory synapses, and promote axonal outgrowth in inhibitory interneurons. In contrast, later in development, α2δ1 subunits promote the glutamatergic neurotransmission and synaptogenesis, as well as strongly enhance neuronal network activity. We propose that formation of connectivity in neuronal networks is associated with a concerted interplay of α2δ1 and α2δ3 subunits of calcium channels.

Missense mutations in CASK, coding for the calcium-/calmodulin-dependent serine protein kinase, interfere with neurexin binding and neurexin-induced oligomerization.

Mutations in the X-linked gene coding for the calcium-/calmodulin-dependent serine protein kinase (CASK) are associated with severe neurological disorders ranging from intellectual disability (in males) to mental retardation and microcephaly with pontine and cerebellar hypoplasia. CASK is involved in transcription control, in the regulation of trafficking of the post-synaptic NMDA and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors, and acts as a presynaptic scaffolding protein. For CASK missense mutations, it is mostly unclear which of CASK's molecular interactions and cellular functions are altered and contribute to patient phenotypes. We identified five CASK missense mutations in male patients affected by neurodevelopmental disorders. These and five previously reported mutations were systematically analysed with respect to interaction with CASK interaction partners by co-expression and co-immunoprecipitation. We show that one mutation in the L27 domain interferes with binding to synapse-associated protein of 97 kDa. Two mutations in the guanylate kinase (GK) domain affect binding of CASK to the nuclear factors CASK-interacting nucleosome assembly protein (CINAP) and T-box, brain, 1 (Tbr1). A total of five mutations in GK as well as PSD-95/discs large/ZO-1 (PDZ) domains affect binding of CASK to the pre-synaptic cell adhesion molecule Neurexin. Upon expression in neurons, we observe that binding to Neurexin is not required for pre-synaptic localization of CASK. We show by bimolecular fluorescence complementation assay that Neurexin induces oligomerization of CASK, and that mutations in GK and PDZ domains interfere with the Neurexin-induced oligomerization of CASK. Our data are supported by molecular modelling, where we observe that the cooperative activity of PDZ, SH3 and GK domains is required for Neurexin binding and oligomerization of CASK.

Presynaptic α2δ-2 Calcium Channel Subunits Regulate Postsynaptic GABAA Receptor Abundance and Axonal Wiring.

Presynaptic α2δ subunits of voltage-gated calcium channels regulate channel abundance and are involved in glutamatergic synapse formation. However, little is known about the specific functions of the individual α2δ isoforms and their role in GABAergic synapses. Using primary neuronal cultures of embryonic mice of both sexes, we here report that presynaptic overexpression of α2δ-2 in GABAergic synapses strongly increases clustering of postsynaptic GABAARs. Strikingly, presynaptic α2δ-2 exerts the same effect in glutamatergic synapses, leading to a mismatched localization of GABAARs. This mismatching is caused by an aberrant wiring of glutamatergic presynaptic boutons with GABAergic postsynaptic positions. The trans-synaptic effect of α2δ-2 is independent of the prototypical cell-adhesion molecules α-neurexins (α-Nrxns); however, α-Nrxns together with α2δ-2 can modulate postsynaptic GABAAR abundance. Finally, exclusion of the alternatively spliced exon 23 of α2δ-2 is essential for the trans-synaptic mechanism. The novel function of α2δ-2 identified here may explain how abnormal α2δ subunit expression can cause excitatory-inhibitory imbalance often associated with neuropsychiatric disorders.SIGNIFICANCE STATEMENT Voltage-gated calcium channels regulate important neuronal functions such as synaptic transmission. α2δ subunits modulate calcium channels and are emerging as regulators of brain connectivity. However, little is known about how individual α2δ subunits contribute to synapse specificity. Here, we show that presynaptic expression of a single α2δ variant can modulate synaptic connectivity and the localization of inhibitory postsynaptic receptors. Our findings provide basic insights into the development of specific synaptic connections between nerve cells and contribute to our understanding of normal nerve cell functions. Furthermore, the identified mechanism may explain how an altered expression of calcium channel subunits can result in aberrant neuronal wiring often associated with neuropsychiatric disorders such as autism or schizophrenia.

Human CCR5high effector memory cells perform CNS parenchymal immune surveillance via GZMK-mediated transendothelial diapedesis.

Although the CNS is immune privileged, continuous search for pathogens and tumours by immune cells within the CNS is indispensable. Thus, distinct immune-cell populations also cross the blood-brain barrier independently of inflammation/under homeostatic conditions. It was previously shown that effector memory T cells populate healthy CNS parenchyma in humans and, independently, that CCR5-expressing lymphocytes as well as CCR5 ligands are enriched in the CNS of patients with multiple sclerosis. Apart from the recently described CD8+ CNS tissue-resident memory T cells, we identified a population of CD4+CCR5high effector memory cells as brain parenchyma-surveilling cells. These cells used their high levels of VLA-4 to arrest on scattered VCAM1, their open-conformation LFA-1 to crawl preferentially against the flow in search for sites permissive for extravasation, and their stored granzyme K (GZMK) to induce local ICAM1 aggregation and perform trans-, rather than paracellular diapedesis through unstimulated primary brain microvascular endothelial cells. This study included peripheral blood mononuclear cell samples from 175 healthy donors, 29 patients infected with HIV, with neurological symptoms in terms of cognitive impairment, 73 patients with relapsing-remitting multiple sclerosis in remission, either 1-4 weeks before (n = 29), or 18-60 months after the initiation of natalizumab therapy (n = 44), as well as white matter brain tissue of three patients suffering from epilepsy. We here provide ex vivo evidence that CCR5highGZMK+CD4+ effector memory T cells are involved in CNS immune surveillance during homeostasis, but could also play a role in CNS pathology. Among CD4+ T cells, this subset was found to dominate the CNS of patients without neurological inflammation ex vivo. The reduction in peripheral blood of HIV-positive patients with neurological symptoms correlated to their CD4 count as a measure of disease progression. Their peripheral enrichment in multiple sclerosis patients and specific peripheral entrapment through the CNS infiltration inhibiting drug natalizumab additionally suggests a contribution to CNS autoimmune pathology. Our transcriptome analysis revealed a migratory phenotype sharing many features with tissue-resident memory and Th17.1 cells, most notably the transcription factor eomesodermin. Knowledge on this cell subset should enable future studies to find ways to strengthen the host defence against CNS-resident pathogens and brain tumours or to prevent CNS autoimmunity.

α-Neurexins Together with α2δ-1 Auxiliary Subunits Regulate Ca2+ Influx through Cav2.1 Channels.

Action potential-evoked neurotransmitter release is impaired in knock-out neurons lacking synaptic cell-adhesion molecules α-neurexins (αNrxns), the extracellularly longer variants of the three vertebrate Nrxn genes. Ca2+ influx through presynaptic high-voltage gated calcium channels like the ubiquitous P/Q-type (CaV2.1) triggers release of fusion-ready vesicles at many boutons. α2δ Auxiliary subunits regulate trafficking and kinetic properties of CaV2.1 pore-forming subunits but it has remained unclear if this involves αNrxns. Using live cell imaging with Ca2+ indicators, we report here that the total presynaptic Ca2+ influx in primary hippocampal neurons of αNrxn triple knock-out mice of both sexes is reduced and involved lower CaV2.1-mediated transients. This defect is accompanied by lower vesicle release, reduced synaptic abundance of CaV2.1 pore-forming subunits, and elevated surface mobility of α2δ-1 on axons. Overexpression of Nrxn1α in αNrxn triple knock-out neurons is sufficient to restore normal presynaptic Ca2+ influx and synaptic vesicle release. Moreover, coexpression of Nrxn1α together with α2δ-1 subunits facilitates Ca2+ influx further but causes little augmentation together with a different subunit, α2δ-3, suggesting remarkable specificity. Expression of defined recombinant CaV2.1 channels in heterologous cells validates and extends the findings from neurons. Whole-cell patch-clamp recordings show that Nrxn1α in combination with α2δ-1, but not with α2δ-3, facilitates Ca2+ currents of recombinant CaV2.1 without altering channel kinetics. These results suggest that presynaptic Nrxn1α acts as a positive regulator of Ca2+ influx through CaV2.1 channels containing α2δ-1 subunits. We propose that this regulation represents an important way for neurons to adjust synaptic strength.SIGNIFICANCE STATEMENT Synaptic transmission between neurons depends on the fusion of neurotransmitter-filled vesicles with the presynaptic membrane, which subsequently activates postsynaptic receptors. Influx of calcium ions into the presynaptic terminal is the key step to trigger vesicle release and involves different subtypes of voltage-gated calcium channels. We study the regulation of calcium channels by neurexins, a family of synaptic cell-adhesion molecules that are essential for many synapse properties. Using optical measurements of calcium influx in cultured neurons and electrophysiological recordings of calcium currents from recombinant channels, we show that a major neurexin variant facilitates calcium influx through P/Q-type channels by interacting with their α2δ-1 auxiliary subunits. These results propose a novel way how neurons can modulate the strength of distinct synapses.

Protein mutated in paroxysmal dyskinesia interacts with the active zone protein RIM and suppresses synaptic vesicle exocytosis.

Paroxysmal nonkinesigenic dyskinesia (PNKD) is an autosomal dominant episodic movement disorder precipitated by coffee, alcohol, and stress. We previously identified the causative gene but the function of the encoded protein remains unknown. We also generated a PNKD mouse model that revealed dysregulated dopamine signaling in vivo. Here, we show that PNKD interacts with synaptic active zone proteins Rab3-interacting molecule (RIM)1 and RIM2, localizes to synapses, and modulates neurotransmitter release. Overexpressed PNKD protein suppresses release, and mutant PNKD protein is less effective than wild-type at inhibiting exocytosis. In PNKD KO mice, RIM1/2 protein levels are reduced and synaptic strength is impaired. Thus, PNKD is a novel synaptic protein with a regulatory role in neurotransmitter release.

Modulation of synaptic function through the α-neurexin-specific ligand neurexophilin-1.

Neurotransmission at different synapses is highly variable, and cell-adhesion molecules like α-neurexins (α-Nrxn) and their extracellular binding partners determine synapse function. Although α-Nrxn affect transmission at excitatory and inhibitory synapses, the contribution of neurexophilin-1 (Nxph1), an α-Nrxn ligand with restricted expression in subpopulations of inhibitory neurons, is unclear. To reveal its role, we investigated mice that either lack or overexpress Nxph1. We found that genetic deletion of Nxph1 impaired GABAB receptor (GABA(B)R)-dependent short-term depression of inhibitory synapses in the nucleus reticularis thalami, a region where Nxph1 is normally expressed at high levels. To test the conclusion that Nxph1 supports presynaptic GABA(B)R, we expressed Nxph1 ectopically at excitatory terminals in the neocortex, which normally do not contain this molecule but can be modulated by GABA(B)R. We generated Nxph1-GFP transgenic mice under control of the Thy1.2 promoter and observed a reduced short-term facilitation at these excitatory synapses, representing an inverse phenotype to the knockout. Consistently, the diminished facilitation could be reversed by pharmacologically blocking GABA(B)R with CGP-55845. Moreover, a complete rescue was achieved by additional blocking of postsynaptic GABA(A)R with intracellular picrotoxin or gabazine, suggesting that Nxph1 is able to recruit or stabilize both presynaptic GABA(B)R and postsynaptic GABA(A)R. In support, immunoelectron microscopy validated the localization of ectopic Nxph1 at the synaptic cleft of excitatory synapses in transgenic mice and revealed an enrichment of GABA(A)R and GABA(B)R subunits compared with wild-type animals. Thus, our data propose that Nxph1 plays an instructive role in synaptic short-term plasticity and the configuration with GABA receptors.

Regulated Dynamic Trafficking of Neurexins Inside and Outside of Synaptic Terminals.

Synapses depend on trafficking of key membrane proteins by lateral diffusion from surface populations and by exocytosis from intracellular pools. The cell adhesion molecule neurexin (Nrxn) plays essential roles in synapses, but the dynamics and regulation of its trafficking are unknown. Here, we performed single-particle tracking and live imaging of transfected, epitope-tagged Nrxn variants in cultured rat and mouse wild-type or knock-out neurons. We observed that structurally larger αNrxn molecules are more mobile in the plasma membrane than smaller ßNrxns because αNrxns displayed higher diffusion coefficients in extrasynaptic regions and excitatory or inhibitory terminals. We found that well characterized interactions with extracellular binding partners regulate the surface mobility of Nrxns. Binding to neurexophilin-1 (Nxph1) reduced the surface diffusion of αNrxns when both molecules were coexpressed. Conversely, impeding other interactions by insertion of splice sequence #4 or removal of extracellular Ca(2+) augmented the mobility of αNrxns and ßNrxns. We also determined that fast axonal transport delivers Nrxns to the neuronal surface because Nrxns comigrate as cargo on synaptic vesicle protein transport vesicles (STVs). Unlike surface mobility, intracellular transport of ßNrxn(+) STVs was faster than that of αNrxns, but both depended on the microtubule motor protein KIF1A and neuronal activity regulated the velocity. Large spontaneous fusion of Nrxn(+) STVs occurred simultaneously with synaptophysin on axonal membranes mostly outside of active presynaptic terminals. Surface Nrxns enriched at synaptic terminals where αNrxns and Nxph1/αNrxns recruited GABAAR subunits. Therefore, our results identify regulated dynamic trafficking as an important property of Nrxns that corroborates their function at synapses. SIGNIFICANCE STATEMENT: Synapses mediate most functions in our brains and depend on the precise and timely delivery of key molecules throughout life. Neurexins (Nrxns) are essential synaptic cell adhesion molecules that are involved in synaptic transmission and differentiation of synaptic contacts. In addition, Nrxns have been linked to neuropsychiatric diseases such as autism. Because little is known about the dynamic aspects of trafficking of neurexins to synapses, we investigated this important question using single-molecule tracking and time-lapse imaging. We identify distinct differences between major Nrxn variants both in surface mobility and during intracellular transport. Because their dynamic behavior is highly regulated, for example, by different binding activities, these processes have immediate consequences for the function of Nrxns at synapses.

p140Cap regulates memory and synaptic plasticity through Src-mediated and citron-N-mediated actin reorganization.

A major challenge in the neuroscience field is the identification of molecules and pathways that control synaptic plasticity and memory. Dendritic spines play a pivotal role in these processes, as the major sites of excitatory synapses in neuronal communication. Previous studies have shown that the scaffold protein p140Cap localizes into dendritic spines and that its knockdown negatively modulates spine shape in culture. However, so far, there is no information on its in vivo relevance. By using a knock-out mouse model, we here demonstrate that p140Cap is a key element for both learning and synaptic plasticity. Indeed, p140Cap(-/-) mice are impaired in object recognition test, as well as in LTP and in LTD measurements. The in vivo effects of p140Cap loss are presumably attenuated by noncell-autonomous events, since primary neurons obtained from p140Cap(-/-) mice show a strong reduction in number of mushroom spines and abnormal organization of synapse-associated F-actin. These phenotypes are most likely caused by a local reduction of the inhibitory control of RhoA and of cortactin toward the actin-depolymerizing factor cofilin. These events can be controlled by p140Cap through its capability to directly inhibit the activation of Src kinase and by its binding to the scaffold protein Citron-N. Altogether, our results provide new insight into how protein associated with dynamic microtubules may regulate spine actin organization through interaction with postsynaptic density components.

Dystroglycan binding to α-neurexin competes with neurexophilin-1 and neuroligin in the brain.

α-Neurexins (α-Nrxn) are mostly presynaptic cell surface molecules essential for neurotransmission that are linked to neuro-developmental disorders as autism or schizophrenia. Several interaction partners of α-Nrxn are identified that depend on alternative splicing, including neuroligins (Nlgn) and dystroglycan (αDAG). The trans-synaptic complex with Nlgn1 was extensively characterized and shown to partially mediate α-Nrxn function. However, the interactions of α-Nrxn with αDAG, neurexophilins (Nxph1) and Nlgn2, ligands that occur specifically at inhibitory synapses, are incompletely understood. Using site-directed mutagenesis, we demonstrate the exact binding epitopes of αDAG and Nxph1 on Nrxn1α and show that their binding is mutually exclusive. Identification of an unusual cysteine bridge pattern and complex type glycans in Nxph1 ensure binding to the second laminin/neurexin/sex hormone binding (LNS2) domain of Nrxn1α, but this association does not interfere with Nlgn binding at LNS6. αDAG, in contrast, interacts with both LNS2 and LNS6 domains without inserts in splice sites SS#2 or SS#4 mostly via LARGE (like-acetylglucosaminyltransferase)-dependent glycans attached to the mucin region. Unexpectedly, binding of αDAG at LNS2 prevents interaction of Nlgn at LNS6 with or without splice insert in SS#4, presumably by sterically hindering each other in the u-form conformation of α-Nrxn. Thus, expression of αDAG and Nxph1 together with alternative splicing in Nrxn1α may prevent or facilitate formation of distinct trans-synaptic Nrxn·Nlgn complexes, revealing an unanticipated way to contribute to the identity of synaptic subpopulations.

Transfer of learning in histology: Insights from a longitudinal study.

All anatomical educators hope that students apply past training to both similar and new tasks. This two-group longitudinal study investigated the development of such transfer of learning in a histology course. After 0, 10, and 20 sessions of the 10-week-long course, medical students completed theoretical tasks, examined histological slides trained in the course (retention task), and unfamiliar histological slides (transfer task). The results showed that students in the histology group gradually outperformed the control group in all tasks, especially in the second half of the course, η2 = 0.268 (p < 0.001). The best predictor of final transfer performance was students' retention performance after 10 sessions, ß = 0.32 (p = 0.028), and theoretical knowledge after 20 sessions, ß = 0.46 (p = 0.003). Results of eye tracking methodology further revealed that the histology group engaged in greater "visual activity" when solving transfer tasks, as indicated by an increase in the total fixation count, η2 = 0.103 (p = 0.014). This longitudinal study provides evidence that medical students can use what they learn in histology courses to solve unfamiliar problems but cautions that positive transfer effects develop relatively late in the course. Thus, course time and the complex relationship between theory, retention, and transfer holds critical implications for anatomical curricula seeking to foster the transfer of learning.