Dataset Information

Microtubules are not required to generate a nascent axon in embryonic spinal neurons in vivo.

ABSTRACT: Our understanding of the cell behaviours and cytoskeletal requirements of axon formation is largely derived from in vitro models but how these relate to axon formation in vivo is not clear. In vitro, neurons progress through a well-defined multineurite stage to form an axon and both actin and microtubules cooperate to drive the first steps in neurite and axon morphogenesis. However, these steps are not recapitulated in vivo, and it is not clear whether the underlying cell biological mechanisms may differ also. Here, we investigate the mechanisms that regulate axon formation in embryonic zebrafish spinal neurons in vivo. We find microtubule organising centres are located distant from the site of axon initiation, and microtubule plus-ends are not enriched in the axon during axon initiation. Focal F-actin accumulation precedes axon formation, and we find that nocodazole-treated neurons with no detectable microtubules are still able to form nascent axonal protrusions that are approximately 10-μm long, dilated and relatively long-lived. We suggest spinal axon formation in vivo is fundamentally different from axon formation in in vitro models.

SUBMITTER: Moore RE

PROVIDER: S-EPMC9638849 | biostudies-literature | 2022 Nov

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Microtubules are not required to generate a nascent axon in embryonic spinal neurons in vivo.

Moore Rachel E RE Pop Sînziana S Alleyne Caché C Clarke Jonathan D W JDW

EMBO reports 20221004 11

Our understanding of the cell behaviours and cytoskeletal requirements of axon formation is largely derived from in vitro models but how these relate to axon formation in vivo is not clear. In vitro, neurons progress through a well-defined multineurite stage to form an axon and both actin and microtubules cooperate to drive the first steps in neurite and axon morphogenesis. However, these steps are not recapitulated in vivo, and it is not clear whether the underlying cell biological mechanisms m ...[more]

PMID: 36194673

Similar Datasets

Project description:Transected axons fail to regrow across anatomically complete spinal cord injuries (SCI) in adults. Diverse molecules can partially facilitate or attenuate axon growth during development or after injury1-3, but efficient reversal of this regrowth failure remains elusive4. Here we show that three factors that are essential for axon growth during development but are attenuated or lacking in adults-(i) neuron intrinsic growth capacity2,5-9, (ii) growth-supportive substrate10,11 and (iii) chemoattraction12,13-are all individually required and, in combination, are sufficient to stimulate robust axon regrowth across anatomically complete SCI lesions in adult rodents. We reactivated the growth capacity of mature descending propriospinal neurons with osteopontin, insulin-like growth factor 1 and ciliary-derived neurotrophic factor before SCI14,15; induced growth-supportive substrates with fibroblast growth factor 2 and epidermal growth factor; and chemoattracted propriospinal axons with glial-derived neurotrophic factor16,17 delivered via spatially and temporally controlled release from biomaterial depots18,19, placed sequentially after SCI. We show in both mice and rats that providing these three mechanisms in combination, but not individually, stimulated robust propriospinal axon regrowth through astrocyte scar borders and across lesion cores of non-neural tissue that was over 100-fold greater than controls. Stimulated, supported and chemoattracted propriospinal axons regrew a full spinal segment beyond lesion centres, passed well into spared neural tissue, formed terminal-like contacts exhibiting synaptic markers and conveyed a significant return of electrophysiological conduction capacity across lesions. Thus, overcoming the failure of axon regrowth across anatomically complete SCI lesions after maturity required the combined sequential reinstatement of several developmentally essential mechanisms that facilitate axon growth. These findings identify a mechanism-based biological repair strategy for complete SCI lesions that could be suitable to use with rehabilitation models designed to augment the functional recovery of remodelling circuits.

Project description:To assemble the functional circuits of the nervous system, the neuronal axonal growth cones must be precisely guided to their proper targets, which can be achieved through cell-surface guidance receptor activation by ligand binding in the periphery. We investigated the function of paxillin, a focal adhesion protein, as an essential growth cone guidance intermediary in the context of spinal lateral motor column (LMC) motor axon trajectory selection in the limb mesenchyme. Using in situ mRNA detection, we first show paxillin expression in LMC neurons of chick and mouse embryos at the time of spinal motor axon extension into the limb. Paxillin loss-of-function and gain-of-function using in ovo electroporation in chick LMC neurons, of either sex, perturbed LMC axon trajectory selection, demonstrating an essential role of paxillin in motor axon guidance. In addition, a neuron-specific paxillin deletion in mice led to LMC axon trajectory selection errors. We also show that knocking down paxillin attenuates the growth preference of LMC neurites against ephrins in vitro, and erythropoietin-producing human hepatocellular (Eph)-mediated retargeting of LMC axons in vivo, suggesting paxillin involvement in Eph-mediated LMC motor axon guidance. Finally, both paxillin knockdown and ectopic expression of a nonphosphorylable paxillin mutant attenuated the retargeting of LMC axons caused by Src overexpression, implicating paxillin as a Src target in Eph signal relay in this context. In summary, our findings demonstrate that paxillin is required for motor axon guidance and suggest its essential role in the ephrin-Eph signaling pathway resulting in motor axon trajectory selection.SIGNIFICANCE STATEMENT During the development of neural circuits, precise connections need to be established among neurons or between neurons and their muscle targets. A protein family found in neurons, Eph, is essential at different stages of neural circuit formation, including nerve outgrowth and pathfinding, and is proposed to mediate the onset and progression of several neurodegenerative diseases, such as Alzheimer's disease. To investigate how Ephs relay their signals to mediate nerve growth, we investigated the function of a molecule called paxillin and found it important for the development of spinal nerve growth toward their muscle targets, suggesting its role as an effector of Eph signals. Our work could thus provide new information on how neuromuscular connectivity is properly established during embryonic development.

Dataset Information

Microtubules are not required to generate a nascent axon in embryonic spinal neurons in vivo.

Publications

Microtubules are not required to generate a nascent axon in embryonic spinal neurons in vivo.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets