Neuronal Growth Cones Research Articles

The TOG5 domain of CKAP5 is required to interact with F-actin and promote microtubule advancement in neurons.

Microtubule (MT) and F-actin cytoskeletal crosstalk and organization are important aspects of axon guidance mechanisms, but how associated proteins facilitate this function remains largely unknown. While the MT-associated protein, CKAP5 (XMAP215/ch-TOG), has been best characterized as a MT polymerase, we have recently highlighted a novel role for CKAP5 in facilitating interactions between MT and F-actin in vitro and in embryonic Xenopus laevis neuronal growth cones. However, the mechanism by which it does so is unclear. Here, using in vitro reconstitution assays coupled with TIRF microscopy, we report that the TOG5 domain of CKAP5 is necessary for its ability to bind to and bundle actin filaments, as well as to crosslink MTs and F-actin in vitro. Additionally, we show that this novel MT/F-actin crosslinking function of CKAP5 is possible even in MT polymerase-incompetent mutants of CKAP5 in vivo. Indeed, CKAP5 requires both MT and F-actin binding, but not MT polymerization, to promote MT-F-actin alignment in growth cones and axon outgrowth. Taken together, our findings provide mechanistic insights into how MT populations penetrate the growth cone periphery through CKAP5-facilitated interaction with F-actin during axon outgrowth and guidance. [Media: see text] [Media: see text] [Media: see text].

TC10 differently controls the dynamics of Exo70 in growth cones of cortical and hippocampal neurons

The exocyst is an octameric protein complex that acts as a tether for GOLGI-derived vesicles at the plasma membrane during exocytosis. It is involved in membrane expansion during axonal outgrowth. Exo70 is a major subunit of the exocyst complex and is controlled by TC10, a Rho family GTPase. How TC10 affects the dynamics of Exo70 at the plasma membrane is not well understood. There is also evidence that TC10 controls Exo70 dynamics differently in nonpolar cells and axons. To address this, we used super-resolution microscopy to study the spatially resolved effects of TC10 on Exo70 dynamics in HeLa cells and the growth cone of cortical and hippocampal neurons. We generated single particle localization and trajectory maps and extracted mean square displacements, diffusion coefficients, and alpha coefficients to characterize Exo70 diffusion. We found that the diffusivity of Exo70 was different in nonpolar cells and in the growth cone of neurons. TC10 stimulated the mobility of Exo70 in HeLa cells, but decreased the diffusion of Exo70 in the growth cone of cortical neurons. In contrast to cortical neurons, TC10 overexpression did not affect the mobility of Exo70 in the axonal growth cone of hippocampal neurons. These data suggest that mainly exocyst tethering in cortical neurons was under the control of TC10.

Cochlear implant/MXene-based electroacoustic stimulation modulates the growth and maturation of spiral ganglion neurons

Cochlear implantation (CI) offers a dependable treatment for sensorineural hearing loss, with precision electroacoustic stimulation parameters showing great potential in improving auditory outcomes in CI patients. Here, we report the attachment of MXene into CI systems which effectively mimic the neural electrode interface due to MXene's excellent electrical conductivity and biocompatibility. Low-frequency short-term biphasic electrical pulses emitted by the MXenes-based CI promoted the outgrowth of spiral ganglion neuron (SGN) neurites and growth cones, substantially boosting the calcium activity in SGNs. This study lays a theoretical foundation for the precision medicine approaches in CI patient care, and informs the selection of materials for cochlear implant electrode materials in the future.

ROCK Inhibitor Enhances Neurite Outgrowth In Vitro and Corneal Sensory Nerve Reinnervation In Vivo.

The intraepithelial corneal nerves are essential to corneal health. Rho kinase or ROCK inhibitors (RIs) have been reported to play a role in neuron survival after injury. Here we assess integrin and extracellular matrix expression in primary mouse neurons and determine whether treating cells with RI impacts neurite outgrowth in vitro and reinnervation after trephine and debridement injury in mice in vivo. Cocultures of human corneal limbal epithelial cells and E11.5 mouse trigeminal neurons and neurons alone were grown on glass coverslips. High-resolution imaging was performed to localize integrins and laminin on neurons and to determine whether RI impacts neurite outgrowth in vitro and in vivo after both 1.5-mm trephine and 1.5-mm debridement injuries. Several integrin α (α3, α6, αv) chains as well as β4 integrin are expressed on neuron axons and growth cones in cocultures. RI treatment of isolated neurons, cocultures, and in conditioned media increases neurite outgrowth. In vivo, RI positively impacts sensory nerve reinnervation after trephine and debridement injury. These studies are the first to demonstrate expression of β4 integrin on trigeminal sensory neurons and preferential adhesion of neurons to the laminin-enriched matrices found in footprints deposited by human corneal limbal epithelial cells. In addition, we also document for the first time the positive impact of RI on neurite outgrowth in vitro and reinnervation in vivo.

Mical1 deletion in tyrosinase expressing cells affects mouse running gaits.

Neuronal development is a highly regulated process that is dependent on the correct coordination of cellular responses to extracellular cues. In response to semaphorin axon guidance proteins, the MICAL1 protein is stimulated to produce reactive oxygen species that oxidize actin on specific methionine residues, leading to filamentous actin depolymerization and consequent changes in neuronal growth cone dynamics. Crossing genetically modified mice homozygous for floxed Mical1 (Mical1fl/fl) alleles with transgenic mice expressing Cre recombinase under the control of a tyrosinase gene enhancer/promoter (Tyr::Cre) enabled conditional Mical1 deletion. Immunohistochemical analysis showed Mical1 expression in the cerebellum, which plays a prominent role in the coordination of motor movements, with reduced Mical1 expression in Mical1fl/fl mice co-expressing Tyr::Cre. Analysis of the gaits of mice running on a treadmill showed that both male and female Mical1fl/fl, Tyr::Cre mutant mice had significant alterations to their striding patterns relative to wild-type mice, although the specific aspects of their altered gaits differed between the sexes. Additional motor tests that involved movement on a rotating rod, descending a vertical pole, or crossing a balance beam did not show significant differences between the genotypes, suggesting that the effect of the Mical1fl/fl, Tyr::Cre genetic modifications was only manifested during specific highly coordinated movements that contribute to running. These findings indicate that there is a behavioral consequence in Mical1fl/fl, Tyr::Cre mutant mice that affects motor control as manifested by alterations in their gait.

Ribosomal Protein Dynamics and Its Association with Actin Filaments and Local Translation in Axonal Growth Cones of Dorsal Root Ganglia Neurons.

Local translation in growth cones plays a critical role in responses to extracellular stimuli, such as axon guidance cues. We previously showed that brain-derived neurotrophic factor activates translation and enhances novel protein synthesis through the activation of mammalian target of rapamycin complex 1 signaling in growth cones of dorsal root ganglion neurons. In this study, we focused on 40S ribosomal protein S6 (RPS6), 60S ribosomal protein P0/1/2 (RPP0/1/2), and actin filaments to determine how localization of ribosomal proteins changes with overall protein synthesis induced by neurotrophins. Our quantitative analysis using immunocytochemistry and super-resolution microscopy indicated that RPS6, RPP0/1/2, and actin tend to colocalize in the absence of stimulation, and that these ribosomal proteins tend to dissociate from actin and associate with each other when local protein synthesis is enhanced. We propose that this is because stimulation causes ribosomal subunits to associate with each other to form actively translating ribosomes (polysomes). This study further clarifies the role of cytoskeletal components in local translation in growth cones.

P14 Characterizing the hair follicle–nervous system connection

Abstract Introduction and aims The human hair follicle (HF) is a miniorgan in the skin that possesses a rich microenvironment composed of a complex assembly of sensory nerves that surround it. Recent studies have explored the effects of this perineural niche on the HF, but the influence of HF on the nervous system from a folliculocentric and skin-centric perspective remains underexplored. To explore the potential bidirectionality of the HF–nervous system connection in the context of nerve development, we asked whether the HF influences the differentiation and maturation of sensory neuron subtypes around it through the release of distinct neurotransmitters and neurotrophic factors. Methods To first identify which cell types might be involved in directing sensory neuron development, we reanalysed available skin and HF single-cell RNA sequencing (scRNA-Seq) datasets. Next, to evaluate neurotransmitter release from these cell types, we used fast-scan cyclic voltammetry (FSCV). Lastly, to interrogate the role of the HF on directing neuronal development in vitro, we used induced pluripotent stem cell-derived sensory neurons as a model for developing nerves. Results We found that a higher percentage of KRT14- (epithelial) and PDGFRA-expressing (mesenchymal) cells in the HF coexpressed genes encoding specific neurotrophic factors and enzymatic machinery for neurotransmitter production compared with equivalent cells in the skin, suggesting that the HF possesses a greater capacity to signal to the nervous system compared with interfollicular skin. Using FSCV, we then found that follicular outer root sheath and directionally selective cells are capable of releasing neurotransmitters such as serotonin and histamine. Conclusions We postulate that HF-derived neurotransmitters are important for aspects of sensory neuron development such as axonal growth, as serotonin has previously been shown to be an attractive guidance cue for neuronal growth cones. Overall, our work sheds light on the HF–nervous system connection and how the HF may regulate its microenvironment in the skin.

The geometry of photopolymerized topography influences neurite pathfinding by directing growth cone morphology and migration

Objective. Cochlear implants provide auditory perception to those with severe to profound sensorineural hearing loss: however, the quality of sound perceived by users does not approximate natural hearing. This limitation is due in part to the large physical gap between the stimulating electrodes and their target neurons. Therefore, directing the controlled outgrowth of processes from spiral ganglion neurons (SGNs) into close proximity to the electrode array could provide significantly increased hearing function. Approach. For this objective to be properly designed and implemented, the ability and limits of SGN neurites to be guided must first be determined. In this work, we engineer precise topographical microfeatures with angle turn challenges of various geometries to study SGN pathfinding and use live imaging to better understand how neurite growth is guided by these cues. Main Results. We find that the geometry of the angled microfeatures determines the ability of neurites to navigate the angled microfeature turns. SGN neurite pathfinding fidelity is increased by 20%–70% through minor increases in microfeature amplitude (depth) and by 25% if the angle of the patterned turn is made obtuse. Further, we see that dorsal root ganglion neuron growth cones change their morphology and migration to become more elongated within microfeatures. Our observations also indicate complexities in studying neurite turning. First, as the growth cone pathfinds in response to the various cues, the associated neurite often reorients across the angle topographical microfeatures. Additionally, neurite branching is observed in response to topographical guidance cues, most frequently when turning decisions are most uncertain. Significance. Overall, the multi-angle channel micropatterned substrate is a versatile and efficient system to assess neurite turning and pathfinding in response to topographical cues. These findings represent fundamental principles of neurite pathfinding that will be essential to consider for the design of 3D systems aiming to guide neurite growth in vivo.

UNC5C: Novel Gene Associated with Psychiatric Disorders Impacts Dysregulation of Axon Guidance Pathways.

The UNC-5 family of netrin receptor genes, predominantly expressed in brain tissues, plays a pivotal role in various neuronal processes. Mutations in genes involved in axon development contribute to a wide spectrum of human diseases, including developmental, neuropsychiatric, and neurodegenerative disorders. The NTN1/DCC signaling pathway, interacting with UNC5C, plays a crucial role in central nervous system axon guidance and has been associated with psychiatric disorders during adolescence in humans. Whole-exome sequencing analysis unveiled two compound heterozygous causative mutations within the UNC5C gene in a patient diagnosed with psychiatric disorders. In silico analysis demonstrated that neither of the observed variants affected the allosteric linkage between UNC5C and NTN1. In fact, these mutations are located within crucial cytoplasmic domains, specifically ZU5 and the region required for the netrin-mediated axon repulsion of neuronal growth cones. These domains play a critical role in forming the supramodular protein structure and directly interact with microtubules, thereby ensuring the functionality of the axon repulsion process. We emphasize that these mutations disrupt the aforementioned processes, thereby associating the UNC5C gene with psychiatric disorders for the first time and expanding the number of genes related to psychiatric disorders. Further research is required to validate the correlation of the UNC5C gene with psychiatric disorders, but we suggest including it in the genetic analysis of patients with psychiatric disorders.

Lithium response in bipolar disorder is associated with focal adhesion and PI3K-Akt networks: a multi-omics replication study

Lithium is the gold standard treatment for bipolar disorder (BD). However, its mechanism of action is incompletely understood, and prediction of treatment outcomes is limited. In our previous multi-omics study of the Pharmacogenomics of Bipolar Disorder (PGBD) sample combining transcriptomic and genomic data, we found that focal adhesion, the extracellular matrix (ECM), and PI3K-Akt signaling networks were associated with response to lithium. In this study, we replicated the results of our previous study using network propagation methods in a genome-wide association study of an independent sample of 2039 patients from the International Consortium on Lithium Genetics (ConLiGen) study. We identified functional enrichment in focal adhesion and PI3K-Akt pathways, but we did not find an association with the ECM pathway. Our results suggest that deficits in the neuronal growth cone and PI3K-Akt signaling, but not in ECM proteins, may influence response to lithium in BD.

A modified motor-clutch model reveals that neuronal growth cones respond faster to soft substrates.

Neuronal growth cones sense a variety of cues including chemical and mechanical ones to establish functional connections during nervous system development. Substrate-cytoskeletal coupling is an established model for adhesion-mediated growth cone advance; however, the detailed molecular and biophysical mechanisms underlying the mechanosensing and mechanotransduction process remain unclear. Here, we adapted a motor-clutch model to better understand the changes in clutch and cytoskeletal dynamics, traction forces, and substrate deformation when a growth cone interacts with adhesive substrates of different stiffnesses. Model parameters were optimized using experimental data from Aplysia growth cones probed with force-calibrated glass microneedles. We included a reinforcement mechanism at both motor and clutch level. Furthermore, we added a threshold for retrograde F-actin flow that indicates when the growth cone is strongly coupled to the substrate. Our modeling results are in strong agreement with experimental data with respect to the substrate deformation and the latency time after which substrate-cytoskeletal coupling is strong enough for the growth cone to advance. Our simulations show that it takes the shortest time to achieve strong coupling when substrate stiffness was low at 4 pN/nm. Taken together, these results suggest that Aplysia growth cones respond faster and more efficiently to soft than stiff substrates.

Interrogating the Molecular Clutch in Neuronal Growth Cones: Measuring Traction Forces, F-actin Retrograde Flow, and Point Contact Demographics.

Growth cone-dependent outgrowth of neuronal processes is essential for the development, plasticity, and regenerative capacity of the nervous system. This process involves the attachment of the growth cone to the substrate and the cyclical engagement/disengagement of the molecular clutch at the sites of adhesive contact. In this chapter, we describe protocols for traction force microscopy, measurement of F-actin retrograde flow velocities, and the assessment of adhesive point contacts by immunofluorescence. These complementary techniques collectively facilitate investigations into the regulation of the molecular clutch in neuronal growth cones.

Measuring Retrograde Actin Flow in Neuronal Growth Cones.

Actin flow refers to the motion of the F-actin cytoskeleton and has been observed in many different cell types, especially in motile cells including neuronal growth cones. The direction of the actin flow is generally retrograde from the periphery toward the center of the cell. Actin flow can be harnessed for forward movement of the cell through substrate-cytoskeletal coupling; thus, a key function of actin flow is in cell locomotion. In this chapter, we illustrate three different methods of quantifying retrograde F-actin flow in growth cones derived from cultured Aplysia bag cell neurons. These methods include tracking the movement of surface marker beads as well as kymograph analysis of time-lapse sequences acquired by differential interference contrast (DIC) imaging or fluorescent speckle microscopy (FSM). Due to their large size, Aplysia neuronal growth cones are uniquely suited for these methods; however, they can also be applied to any other growth cones with clear F-actin-rich peripheral domains.

The role of microtubule-associated protein tau in netrin-1 attractive signaling.

Direct binding of netrin receptors with dynamic microtubules (MTs) in the neuronal growth cone plays an important role in netrin-mediated axon guidance. However, how netrin-1 (NTN1) regulates MT dynamics in axon turning remains a major unanswered question. Here, we show that the coupling of netrin-1 receptor DCC with tau (MAPT)-regulated MTs is involved in netrin-1-promoted axon attraction. Tau directly interacts with DCC and partially overlaps with DCC in the growth cone of primary neurons. Netrin-1 induces this interaction and the colocalization of DCC and tau in the growth cone. The netrin-1-induced interaction of tau with DCC relies on MT dynamics and TUBB3, a highly dynamic β-tubulin isotype in developing neurons. Netrin-1 increased cosedimentation of DCC with tau and TUBB3 in MTs, and knockdown of either tau or TUBB3 mutually blocked this effect. Downregulation of endogenous tau levels by tau shRNAs inhibited netrin-1-induced axon outgrowth, branching and commissural axon attraction in vitro, and led to defects in spinal commissural axon projection in vivo. These findings suggest that tau is a key MT-associated protein coupling DCC with MT dynamics in netrin-1-promoted axon attraction.

Trabid patient mutations impede the axonal trafficking of adenomatous polyposis coli to disrupt neurite growth

ZRANB1 (human Trabid) missense mutations have been identified in children diagnosed with a range of congenital disorders including reduced brain size, but how Trabid regulates neurodevelopment is not understood. We have characterized these patient mutations in cells and mice to identify a key role for Trabid in the regulation of neurite growth. One of the patient mutations flanked the catalytic cysteine of Trabid and its deubiquitylating (DUB) activity was abrogated. The second variant retained DUB activity, but failed to bind STRIPAK, a large multiprotein assembly implicated in cytoskeleton organization and neural development. Zranb1 knock-in mice harboring either of these patient mutations exhibited reduced neuronal and glial cell densities in the brain and a motor deficit consistent with fewer dopaminergic neurons and projections. Mechanistically, both DUB-impaired and STRIPAK-binding-deficient Trabid variants impeded the trafficking of adenomatous polyposis coli (APC) to microtubule plus-ends. Consequently, the formation of neuronal growth cones and the trajectory of neurite outgrowth from mutant midbrain progenitors were severely compromised. We propose that STRIPAK recruits Trabid to deubiquitylate APC, and that in cells with mutant Trabid, APC becomes hyperubiquitylated and mislocalized causing impaired organization of the cytoskeleton that underlie the neuronal and developmental phenotypes.

Trabid patient mutations impede the axonal trafficking of adenomatous polyposis coli to disrupt neurite growth.

ZRANB1 (human Trabid) missense mutations have been identified in children diagnosed with a range of congenital disorders including reduced brain size, but how Trabid regulates neurodevelopment is not understood. We have characterized these patient mutations in cells and mice to identify a key role for Trabid in the regulation of neurite growth. One of the patient mutations flanked the catalytic cysteine of Trabid and its deubiquitylating (DUB) activity was abrogated. The second variant retained DUB activity, but failed to bind STRIPAK, a large multiprotein assembly implicated in cytoskeleton organization and neural development. Zranb1 knock-in mice harboring either of these patient mutations exhibited reduced neuronal and glial cell densities in the brain and a motor deficit consistent with fewer dopaminergic neurons and projections. Mechanistically, both DUB-impaired and STRIPAK-binding-deficient Trabid variants impeded the trafficking of adenomatous polyposis coli (APC) to microtubule plus-ends. Consequently, the formation of neuronal growth cones and the trajectory of neurite outgrowth from mutant midbrain progenitors were severely compromised. We propose that STRIPAK recruits Trabid to deubiquitylate APC, and that in cells with mutant Trabid, APC becomes hyperubiquitylated and mislocalized causing impaired organization of the cytoskeleton that underlie the neuronal and developmental phenotypes.

CKAP5 enables formation of persistent actin bundles templated by dynamically instable microtubules

Cytoskeletal rearrangements and crosstalk between microtubules and actin filaments are vital for living organisms. Recently, an abundantly present microtubule polymerase, CKAP5 (XMAP215 homolog), has been reported to play a role in mediating crosstalk between microtubules and actin filaments in the neuronal growth cones. However, the molecular mechanism of this process is unknown. Here, we demonstrate, in a reconstituted system, that CKAP5 enables the formation of persistent actin bundles templated by dynamically instable microtubules. We explain the templating by the difference in CKAP5 binding to microtubules and actin filaments. Binding to the microtubule lattice with higher affinity, CKAP5 enables the formation of actin bundles exclusively on the microtubule lattice, at CKAP5 concentrations insufficient to support any actin bundling in the absence of microtubules. Strikingly, when the microtubules depolymerize, actin bundles prevail at the positions predetermined by the microtubules. We propose that the local abundance of available CKAP5-binding sites in actin bundles allows the retention of CKAP5, resulting in persisting actin bundles. In line with our observations, we found that reducing CKAP5 levels invivo results in a decrease in actin-microtubule co-localization in growth cones and specifically decreases actin intensity at microtubule plus ends. This readily suggests a mechanism explaining how exploratory microtubules set the positions of actin bundles, for example, in cytoskeleton-rich neuronal growth cones.

The Plasma Membrane Polarity Is Higher in the Neuronal Growth Cone than in the Cell Body of Hippocampal and Cerebellar Granule Neurons.

The polarity of the biological membrane, or lipid order, regulates many cellular events. It is generally believed that the plasma membrane polarity is regulated according to cell type and function, sometimes even within a cell. Neurons have a variety of functionally specialized subregions, each of which bears distinct proteins and lipids, and the membrane polarity of the subregions may differ accordingly. However, no direct experimental evidence of it has been presented to date. In the present study, we used a cell-impermeable solvatochromic membrane probe NR12A to investigate the local polarity of the plasma membrane of neurons. Both in hippocampal and cerebellar granule neurons, growth cones have higher membrane polarity than the cell body. In addition, the overall variation in the polarity value of each pixel was greater in the growth cone than in cell bodies, suggesting that the lateral diffusion and/or dynamics of the growth cone membrane are greater than other parts of the neuron. These tendencies were much less notably observed in the lamellipodia of a non-neuronal cell. Our results suggest that the membrane polarity of neuronal growth cones is unique and this characteristic may be important for its structure and function.

Abstract IA004: Identification of potential new therapeutic targets in kidney cancer

Abstract VHL is the most important tumor suppressor in clear cell renal cell carcinoma (ccRCC). In the majority of ccRCC, as a result of VHL loss, HIF2a stabilization is sufficient and necessary for promoting ccRCC tumor growth. Despite the development of HIF2a specific inhibitors, the intrinsic and extrinsic resistance do exist, highlighting the importance of identifying additional therapeutic vulnerabilities in ccRCC. We have recently performed some genome-wide screening and identified new signaling pathways regulated by VHL, including ZHX2 and SFMBT1 that play important roles in ccRCC. In addition, we also identified some synthetic lethality binding partners for VHL loss in kidney cancer, including TBK1. However, it is challenging to target TBK1 in clinical practice due to lack of selectivity for TBK1 inhibitors and its role involved in innate immune signaling. To overcome this difficulty, one approach would be to identify a critical upstream regulator of TBK1 that can be targeted therapeutically, especially in ccRCC displaying hyper-activated TBK1 activity due to VHL loss. We have performed a kinome-wide screening and identified Doublecortin like kinase 2 (DCLK2), an enigmatic kinase involved in neuronal survival and growth cone regeneration upon injury, as a novel TBK1 regulator in ccRCC. Our functional assays demonstrate that DCLK2 is necessary and sufficient to promote ccRCC tumorigenesis through activating TBK1. In addition, we also characterized a DCLK2 short isoform (DCLK2203) that play a predominant role in ccRCC tumorigenesis. Our work suggests that DCLK2 is a novel TBK1 activator and potential therapeutic target in ccRCC. Citation Format: Qing Zhang. Identification of potential new therapeutic targets in kidney cancer [abstract]. In: Proceedings of the AACR Special Conference: Advances in Kidney Cancer Research; 2023 Jun 24-27; Austin, Texas. Philadelphia (PA): AACR; Cancer Res 2023;83(16 Suppl):Abstract nr IA004.

CRMP2 conditional knockout changes axonal function and ultrastructure of axons in mice corpus callosum

Collapsin response mediator protein 2 (CRMP2) is a member of a protein family, which is highly involved in neurodevelopment, but most of its members become heavily downregulated in adulthood. CRMP2 is an important factor in neuronal polarization, axonal formation and growth cone collapse. The protein remains expressed in adulthood, but is more region specific. CRMP2 is present in adult corpus callosum (CC) and in plastic areas like prefrontal cortex and hippocampus. CRMP2 has been implicated as one of the risk-genes for Schizophrenia (SZ). Here, a CRMP2 conditional knockout (CRMP2-cKO) mouse was used as a model of SZ to investigate how it could affect the white matter and therefore brain connectivity.Multielectrode electrophysiology (MEA) was used to study the function of corpus callosum showing an increase in conduction velocity (CV) measured as Compound Action Potentials (CAPs) in acute brain slices. Light- and electron-microscopy, specifically Serial Block-face Scanning Electron Microscopy (SBF-SEM), methods were used to study the structure of CC in CRMP2-cKO mice. A decrease in CC volume of CRMP2-cKO mice as compared to controls was observed. No differences were found in numbers nor in the size of CC oligodendrocytes (OLs). Similarly, no differences were found in myelin thickness or in node of Ranvier (NR) structure. In contrast, abnormally smaller axons were measured in the CRMP2-cKO mice.Using these state-of-the-art methods it was possible to shed light on specific parts of the dysconnectivity aspect of deletion of CRMP2 related to SZ and add details to previous findings helping further understanding the disease. This paper substantiates the white matter changes in the absence of CRMP2 and ties it to the role it plays in this complex disorder.