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Large-scale microfluidic gradient arrays reveal axon guidance behaviors in hippocampal neurons.

ABSTRACT: High-throughput quantitative approaches to study axon growth behaviors have remained a challenge. We have developed a 1024-chamber microfluidic gradient generator array that enables large-scale investigations of axon guidance and growth dynamics from individual primary mammalian neurons, which are exposed to gradients of diffusible molecules. Our microfluidic method (a) generates statistically rich data sets, (b) produces a stable, reproducible gradient with negligible shear stresses on the culture surface, (c) is amenable to the long-term culture of primary neurons without any unconventional protocol, and (d) eliminates the confounding influence of cell-secreted factors. Using this platform, we demonstrate that hippocampal axon guidance in response to a netrin-1 gradient is concentration-dependent-attractive at higher concentrations and repulsive at lower concentrations. We also show that the turning of the growth cone depends on the angle of incidence of the gradient. Our study highlights the potential of microfluidic devices in producing large amounts of data from morphogen and chemokine gradients that play essential roles not only in axonal navigation but also in stem cell differentiation, cell migration, and immune response.

SUBMITTER: Bhattacharjee N

PROVIDER: S-EPMC6445017 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Large-scale microfluidic gradient arrays reveal axon guidance behaviors in hippocampal neurons.

Bhattacharjee Nirveek N Folch Albert A

Microsystems & nanoengineering 20170508

High-throughput quantitative approaches to study axon growth behaviors have remained a challenge. We have developed a 1024-chamber microfluidic gradient generator array that enables large-scale investigations of axon guidance and growth dynamics from individual primary mammalian neurons, which are exposed to gradients of diffusible molecules. Our microfluidic method (a) generates statistically rich data sets, (b) produces a stable, reproducible gradient with negligible shear stresses on the cult ...[more]

PMID: 31057858

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Project description:During the development of the Drosophila central nervous system the process of midline crossing is orchestrated by a number of guidance receptors and ligands. Many key axon guidance molecules have been identified in both invertebrates and vertebrates, but the transcriptional regulation of growth cone guidance remains largely unknown. One open question is whether transcriptional regulation plays a role in midline crossing, or if local translation can account for the necessary fine tuning of protein levels. To investigate this issue, we conducted a genome wide analysis of transcription in Drosophila embryos using wild type and a number of well-characterized Drosophila guidance mutants and transgenics. We also analyzed a publicly available microarray time course of Drosophila embryonic development with an axon guidance focus. Using hopach, a novel clustering method which is well suited to microarray data analysis, we identified groups of genes with similar expression patterns across guidance mutants and transgenics. We then systematically characterized the resulting clusters with respect to their relevance to axon guidance using two complementary controlled vocabularies: the Gene Ontology (GO) and anatomical annotations of the Atlas of Pattern of Gene Expression (APoGE) in situ hybridization database. The analysis indicates that regulation of gene expression does play a role in the process of axon guidance in Drosophila. We also find a strong link between axon guidance and hemocyte migration, a result that agrees with mounting evidence that axon guidance molecules are co-opted in vertebrate vascularization. Cell cyclin activity in the context of axon guidance is also suggested from our array data. RNA and protein patterns of cell cyclin in axon guidance mutants and transgenics support this possible link. This study provides important insights into the regulation of axon guidance in vivo and suggests that transcription does play a role in control of axon guidance. Keywords: Mutant Analysis

Dataset Information

Large-scale microfluidic gradient arrays reveal axon guidance behaviors in hippocampal neurons.

Publications

Large-scale microfluidic gradient arrays reveal axon guidance behaviors in hippocampal neurons.

Similar Datasets

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