SEARCH · Search NASA
Results for “axon”
Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.
Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.
Changes in microtubule stability and density in myelin-deficient shiverer mouse CNS axons
Altered axon-Schwann cell interactions in PNS myelin-deficient Trembler mice result in changed axonal transport rates, neurofilament and microtubule-associated protein phosphorylation, neurofilament density, and microtubule stability. To determine whether PNS and CNS myelination have equivalent effects on axons, neurofilaments, and microtubules in CNS, myelin-deficient shiverer axons were examined. The genetic defect in shiverer is a deletion in the myelin basic protein (MBP) gene, an essential component of CNS myelin. As a result, shiverer mice have little or no compact CNS myelin. Slow axonal transport rates in shiverer CNS axons were significantly increased, in contrast to the slowing in demyelinated PNS nerves. Even more striking were substantial changes in the composition and properties of microtubules in shiverer CNS axons. The density of axonal microtubules is increased, reflecting increased expression of tubulin in shiverer, and the stability of microtubules is drastically reduced in shiverer axons. Shiverer transgenic mice with two copies of a wild-type myelin basic protein transgene have an intermediate level of compact myelin, making it possible to determine whether the actual level of compact myelin is an important regulator of axonal microtubules. Both increased microtubule density and reduced microtubule stability were still observed in transgenic mouse nerves, indicating that signals beyond synaptogenesis and the mere presence of compact myelin are required for normal regulation of the axonal microtubule cytoskeleton.
Myelinated sensory and alpha motor axon regeneration in peripheral nerve neuromas
Histochemical staining for carbonic anhydrase and cholinesterase (CE) activities was used to analyze sensory and motor axon regeneration, respectively, during neuroma formation in transected and tube-encapsulated peripheral nerves. Median-ulnar and sciatic nerves in the rodent model permitted testing whether a 4 cm greater distance of the motor neuron soma from axotomy site or intrinsic differences between motor and sensory neurons influenced regeneration and neuroma formation 10, 30, and 90 days later. Ventral root radiculotomy confirmed that CE-stained axons were 97% alpha motor axons. Distance significantly delayed axon regeneration. When distance was negligible, sensory axons grew out sooner than motor axons, but motor axons regenerated to a greater quantity. These results indicate regeneration differences between axon subtypes and suggest more extensive branching of motor axons within the neuroma. Thus, both distance from injury site to soma and inherent motor and sensory differences should be considered in peripheral nerve repair strategies.
Partial connectomes of labeled dopaminergic circuits reveal non-synaptic communication and axonal remodeling after exposure to cocaine
Dopaminergic (DA) neurons exert profound influences on behavior including addiction. However, how DA axons communicate with target neurons and how those communications change with drug exposure remains poorly understood. We leverage cell type-specific labeling with large volume serial electron microscopy to detail DA connections in the nucleus accumbens (NAc) of the mouse (Mus musculus) before and after exposure to cocaine. We find that individual DA axons contain different varicosity types based on their vesicle contents. Spatially ordering along individual axons further suggests that varicosity types are non-randomly organized. DA axon varicosities rarely make specific synapses (<2%, 6/410), but instead are more likely to form spinule-like structures (15%, 61/410) with neighboring neurons. Days after a brief exposure to cocaine, DA axons were extensively branched relative to controls, formed blind-ended ‘bulbs’ filled with mitochondria, and were surrounded by elaborated glia. Finally, mitochondrial lengths increased by ~2.2 times relative to control only in DA axons and NAc spiny dendrites after cocaine exposure. We conclude that DA axonal transmission is unlikely to be mediated via classical synapses in the NAc and that the major locus of anatomical plasticity of DA circuits after exposure to cocaine are large-scale axonal re-arrangements with correlated changes in mitochondria.
Identifying motor and sensory myelinated axons in rabbit peripheral nerves by histochemical staining for carbonic anhydrase and cholinesterase activities
Carbonic anhydrase (CA) and cholinesterase (CE) histochemical staining of rabbit spinal nerve roots and dorsal root ganglia demonstrated that among the reactive myeliated axons, with minor exceptions, sensory axons were CA positive and CE negative whereas motor axons were CA negative and CE positive. The high specificity was achieved by adjusting reaction conditions to stain subpopulations of myelinated axons selectively while leaving 50 percent or so unstained. Fixation with glutaraldehyde appeared necessary for achieving selectivity. Following sciatic nerve transection, the reciprocal staining pattern persisted in damaged axons and their regenerating processes which formed neuromas within the proximal nerve stump. Within the neuromas, CA-stained sensory processes were elaborated earlier and in greater numbers than CE-stained regenerating motor processes. The present results indicate that histochemical axon typing can be exploited to reveal heterogeneous responses of motor and sensory axons to injury.
The membrane periodic skeleton is an actomyosin network that regulates axonal diameter and conduction
Neurons have a membrane periodic skeleton (MPS) composed of actin rings interconnected by spectrin. Here, combining chemical and genetic gain- and loss-of-function assays, we show that in rat hippocampal neurons the MPS is an actomyosin network that controls axonal expansion and contraction. Using super-resolution microscopy, we analyzed the localization of axonal non-muscle myosin II (NMII). We show that active NMII light chains are colocalized with actin rings and organized in a circular periodic manner throughout the axon shaft. In contrast, NMII heavy chains are mostly positioned along the longitudinal axonal axis, being able to crosslink adjacent rings. NMII filaments can play contractile or scaffolding roles determined by their position relative to actin rings and activation state. We also show that MPS destabilization through NMII inactivation affects axonal electrophysiology, increasing action potential conduction velocity. In summary, our findings open new perspectives on axon diameter regulation, with important implications in neuronal biology.
Formation of compact myelin is required for maturation of the axonal cytoskeleton
Although traditional roles ascribed to myelinating glial cells are structural and supportive, the importance of compact myelin for proper functioning of the nervous system can be inferred from mutations in myelin proteins and neuropathologies associated with loss of myelin. Myelinating Schwann cells are known to affect local properties of peripheral axons (de Waegh et al., 1992), but little is known about effects of oligodendrocytes on CNS axons. The shiverer mutant mouse has a deletion in the myelin basic protein gene that eliminates compact myelin in the CNS. In shiverer mice, both local axonal features like phosphorylation of cytoskeletal proteins and neuronal perikaryon functions like cytoskeletal gene expression are altered. This leads to changes in the organization and composition of the axonal cytoskeleton in shiverer unmyelinated axons relative to age-matched wild-type myelinated fibers, although connectivity and patterns of neuronal activity are comparable. Remarkably, transgenic shiverer mice with thin myelin sheaths display an intermediate phenotype indicating that CNS neurons are sensitive to myelin sheath thickness. These results indicate that formation of a normal compact myelin sheath is required for normal maturation of the neuronal cytoskeleton in large CNS neurons.
A role for cyclin-dependent kinase(s) in the modulation of fast anterograde axonal transport: effects defined by olomoucine and the APC tumor suppressor protein
Proteins that interact with both cytoskeletal and membrane components are candidates to modulate membrane trafficking. The tumor suppressor proteins neurofibromin (NF1) and adenomatous polyposis coli (APC) both bind to microtubules and interact with membrane-associated proteins. The effects of recombinant NF1 and APC fragments on vesicle motility were evaluated by measuring fast axonal transport along microtubules in axoplasm from squid giant axons. APC4 (amino acids 1034-2844) reduced only anterograde movements, whereas APC2 (aa 1034-2130) or APC3 (aa 2130-2844) reduced both anterograde and retrograde transport. NF1 had no effect on organelle movement in either direction. Because APC contains multiple cyclin-dependent kinase (CDK) consensus phosphorylation motifs, the kinase inhibitor olomoucine was examined. At concentrations in which olomoucine is specific for cyclin-dependent kinases (5 microM), it reduced only anterograde transport, whereas anterograde and retrograde movement were both affected at concentrations at which other kinases are inhibited as well (50 microM). Both anterograde and retrograde transport also were inhibited by histone H1 and KSPXK peptides, substrates for proline-directed kinases, including CDKs. Our data suggest that CDK-like axonal kinases modulate fast anterograde transport and that other axonal kinases may be involved in modulating retrograde transport. The specific effect of APC4 on anterograde transport suggests a model in which the binding of APC to microtubules may limit the activity of axonal CDK kinase or kinases in restricted domains, thereby affecting organelle transport.
NELL2-Robo3 complex structure reveals mechanisms of receptor activation for axon guidance
Axon pathfinding is critical for nervous system development, and it is orchestrated by molecular cues that activate receptors on the axonal growth cone. Robo family receptors bind Slit guidance cues to mediate axon repulsion. In mammals, the divergent family member Robo3 does not bind Slits, but instead signals axon repulsion from its own ligand, NELL2. Conversely, canonical Robos do not mediate NELL2 signaling. Here, we present the structures of NELL-Robo3 complexes, identifying a mode of ligand engagement for Robos that is orthogonal to Slit binding. We elucidate the structural basis for differential binding between NELL and Robo family members and show that NELL2 repulsive activity is a function of its Robo3 affinity and is enhanced by ligand trimerization. Our results reveal a mechanism of oligomerization-induced Robo activation for axon guidance and shed light on Robo family member ligand binding specificity, conformational variability, divergent modes of signaling, and evolution.
A novel statistical methodology for quantifying the spatial arrangements of axons in peripheral nerves
A thorough understanding of the neuroanatomy of peripheral nerves is required for a better insight into their function and the development of neuromodulation tools and strategies. In biophysical modeling, it is commonly assumed that the complex spatial arrangement of myelinated and unmyelinated axons in peripheral nerves is random, however, in reality the axonal organization is inhomogeneous and anisotropic. Present quantitative neuroanatomy methods analyze peripheral nerves in terms of the number of axons and the morphometric characteristics of the axons, such as area and diameter. In this study, we employed spatial statistics and point process models to describe the spatial arrangement of axons and Sinkhorn distances to compute the similarities between these arrangements (in terms of first- and second-order statistics) in various vagus and pelvic nerve cross-sections. We utilized high-resolution transmission electron microscopy (TEM) images that have been segmented using a custom-built high-throughput deep learning system based on a highly modified U-Net architecture. Our findings show a novel and innovative approach to quantifying similarities between spatial point patterns using metrics derived from the solution to the optimal transport problem. We also present a generalizable pipeline for quantitative analysis of peripheral nerve architecture. Our data demonstrate differences between male- and female-originating samples and similarities between the pelvic and abdominal vagus nerves.
Inhibition of PTEN activity promotes IB4‐positive sensory neuronal axon growth
Abstract Traumatic nerve injuries have become a common clinical problem, and axon regeneration is a critical process in the successful functional recovery of the injured nervous system. In this study, we found that peripheral axotomy reduces PTEN expression in adult sensory neurons; however, it did not alter the expression level of PTEN in IB4‐positive sensory neurons. Additionally, our results indicate that the artificial inhibition of PTEN markedly promotes adult sensory axon regeneration, including IB4‐positive neuronal axon growth. Thus, our results provide strong evidence that PTEN is a prominent repressor of adult sensory axon regeneration, especially in IB4‐positive neurons.
Molecular and structural basis of olfactory sensory neuron axon coalescence by Kirrel receptors
Projections from sensory neurons of olfactory systems coalesce into glomeruli in the brain. The Kirrel receptors are believed to homodimerize via their ectodomains and help separate sensory neuron axons into Kirrel2- or Kirrel3-expressing glomeruli. Here, we present the crystal structures of homodimeric Kirrel receptors and show that the closely related Kirrel2 and Kirrel3 have evolved specific sets of polar and hydrophobic interactions, respectively, disallowing heterodimerization while preserving homodimerization, likely resulting in proper segregation and coalescence of Kirrel-expressing axons into glomeruli. We show that the dimerization interface at the N-terminal immunoglobulin (IG) domains is necessary and sufficient to create homodimers and fail to find evidence for a secondary interaction site in Kirrel ectodomains. Furthermore, we show that abolishing dimerization of Kirrel3 in vivo leads to improper formation of glomeruli in the mouse accessory olfactory bulb as observed in Kirrel3 –/– animals. Our results provide evidence for Kirrel3 homodimerization controlling axonal coalescence.
Nanostructure-specific X-ray tomography reveals myelin levels, integrity and axon orientations in mouse and human nervous tissue
Myelin insulates neuronal axons and enables fast signal transmission, constituting a key component of brain development, aging and disease. Yet, myelin-specific imaging of macroscopic samples remains a challenge. Here, we exploit myelin’s nanostructural periodicity, and use small-angle X-ray scattering tensor tomography (SAXS-TT) to simultaneously quantify myelin levels, nanostructural integrity and axon orientations in nervous tissue. Proof-of-principle is demonstrated in whole mouse brain, mouse spinal cord and human white and gray matter samples. Outcomes are validated by 2D/3D histology and compared to MRI measurements sensitive to myelin and axon orientations. Specificity to nanostructure is exemplified by concomitantly imaging different myelin types with distinct periodicities. Finally, we illustrate the method’s sensitivity towards myelin-related diseases by quantifying myelin alterations in dysmyelinated mouse brain. This non-destructive, stain-free molecular imaging approach enables quantitative studies of myelination within and across samples during development, aging, disease and treatment, and is applicable to other ordered biomolecules or nanostructures.
A study of axonal degeneration in the optic nerves of aging mice
The optic nerves of C57BL/6J mice ranging from 3 to 30 months were examined by electron microscopy. At all ages investigated, optic nerve axons contained enlarged mitochondria with abnormal cristae. With increasing age, a large number of necrotic axons were observed and were in the process of being phagocytized. The abnormal mitochondria may represent preliminary changes that eventually lead to necrosis of the axon.
Fray, a Drosophila serine/threonine kinase homologous to mammalian PASK, is required for axonal ensheathment
Fray is a serine/threonine kinase expressed by the peripheral glia of Drosophila, whose function is required for normal axonal ensheathment. Null fray mutants die early in larval development and have nerves with severe swelling and axonal defasciculation. The phenotype is associated with a failure of the ensheathing glia to correctly wrap peripheral axons. When the fray cDNA is expressed in the ensheathing glia of fray mutants, normal nerve morphology is restored. Fray belongs to a novel family of Ser/Thr kinases, the PF kinases, whose closest relatives are the PAK kinases. Rescue of the Drosophila mutant phenotype with PASK, the rat homolog of Fray, demonstrates a functional homology among these proteins and suggests that the Fray signaling pathway is widely conserved.
Vinculin is required for neuronal mechanosensing but not for axon outgrowth
Integrin receptors are transmembrane proteins that bind to the extracellular matrix (ECM). In most animal cell types integrins cluster together with adaptor proteins at focal adhesions that sense and respond to external mechanical signals. In the central nervous system (CNS), ECM proteins are sparsely distributed, the tissue is comparatively soft and neurons do not form focal adhesions. Thus, how neurons sense tissue stiffness is currently poorly understood. Here, we found that integrins and the integrin-associated proteins talin and focal adhesion kinase (FAK) are required for the outgrowth of neuronal processes. Vinculin, however, whilst not required for neurite outgrowth was a key regulator of integrin-mediated mechanosensing of neurons. During growth, growth cones of axons of CNS derived cells exerted dynamic stresses of around 10–12 Pa on their environment, and axons grew significantly longer on soft (0.4 kPa) compared to stiff (8 kPa) substrates. Depletion of vinculin blocked this ability of growth cones to distinguish between soft and stiff substrates. These data suggest that vinculin in neurons acts as a key mechanosensor, involved in the regulation of growth cone motility.
Axon-like active signal transmission
Any electrical signal propagating in a metallic conductor loses amplitude due to the natural resistance of the metal. Compensating for such losses presently requires repeatedly breaking the conductor and interposing amplifiers that consume and regenerate the signal. This century-old primitive severely constrains the design and performance of modern interconnect-dense chips. Here we present a fundamentally different primitive based on semi-stable edge of chaos (EOC), a long-theorized but experimentally elusive regime that underlies active (self-amplifying) transmission in biological axons. By electrically accessing the spin crossover in LaCoO 3 , we isolate semi-stable EOC, characterized by small-signal negative resistance and amplification of perturbations. In a metallic line atop a medium biased at EOC, a signal input at one end exits the other end amplified, without passing through a separate amplifying component. While superficially resembling superconductivity, active transmission offers controllably amplified time-varying small-signal propagation at normal temperature and pressure, but requires an electrically energized EOC medium. Operando thermal mapping reveals the mechanism of amplification—bias energy of the EOC medium, instead of fully dissipating as heat, is partly used to amplify signals in the metallic line, thereby enabling spatially continuous active transmission, which could transform the design and performance of complex electronic chips.
Release of kinesin from vesicles by hsc70 and regulation of fast axonal transport
The nature of kinesin interactions with membrane-bound organelles and mechanisms for regulation of kinesin-based motility have both been surprisingly difficult to define. Most kinesin is recovered in supernatants with standard protocols for purification of motor proteins, but kinesin recovered on membrane-bound organelles is tightly bound. Partitioning of kinesin between vesicle and cytosolic fractions is highly sensitive to buffer composition. Addition of either N-ethylmaleimide or EDTA to homogenization buffers significantly increased the fraction of kinesin bound to organelles. Given that an antibody against kinesin light chain tandem repeats also releases kinesin from vesicles, these observations indicated that specific cytoplasmic factors may regulate kinesin release from membranes. Kinesin light tandem repeats contain DnaJ-like motifs, so the effects of hsp70 chaperones were evaluated. Hsc70 released kinesin from vesicles in an MgATP-dependent and N-ethylmaleimide-sensitive manner. Recombinant kinesin light chains inhibited kinesin release by hsc70 and stimulated the hsc70 ATPase. Hsc70 actions may provide a mechanism to regulate kinesin function by releasing kinesin from cargo in specific subcellular domains, thereby effecting delivery of axonally transported materials.