Culturing of Cerebellar Granule Cells to Study Neuronal Migration: Gradient and Local Perfusion Assays

Guijarro, Patricia; Jiang, Jian; Yuan, Xiao-bing

doi:10.1002/0471142301.ns0326s60

Cited by 2 publications

(2 citation statements)

References 23 publications

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“…To determine the role of IL-1β in leading process pathfinding, we employed a growth cone turning assay. Cerebellar granule cells were used as a model to perform this assay because they constitute a large, homogeneous and easily-cultured population of cells, which maintains migratory behavior, a characteristic that makes them very convenient for in vitro assays [17]. Microscopic gradients of IL-1β were created near the growth cones of isolated neurons by repetitive pulsatile application of picoliters of IL-1β-containing solution with a micropipette.…”

Section: Resultsmentioning

confidence: 99%

Interleukin-1 beta guides the migration of cortical neurons

Zhang

et al. 2014

J Neuroinflammation

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BackgroundProinflammatory cytokine interleukin-1beta (IL-1β) is expressed at high levels in the developing brain and declines to low constitutive levels in the adult. However, the pathophysiological function of IL-1β during brain development remains elusive. In this study, we investigated the role of IL-1β in neuronal migration.MethodsThe Boyden transwell assay was used to examine the effects of IL-1β on the migration of dissociated primary cortical neurons. To determine the role of IL-1β in neuron leading process pathfinding, we employed a growth cone turning assay. In utero electroporation combined with RNAi technology was used to examine the neuronal migration in vivo during brain development in Sprague–Dawley rats.ResultsIL-1β at concentrations ranging from 0.1 to 10 ng/mL in the lower chamber of a transwell induced a significant increase in the number of migrating neurons in a dose-dependent manner. When IL-1β was simultaneously put in both the upper and lower chambers to eliminate the gradient, no significant differences in cell migration were observed. IL-1 receptor antagonist IL-1RA dose-dependently blocked the attractive effect of IL-1β on neuronal migration. Microscopic gradients of IL-1β were created near the growth cones of isolated neurons by repetitive pulsatile application of picoliters of a IL-1β-containing solution with a micropipette. We found that growth cones exhibited a clear bias toward the source of IL-1β at the end of a one hour period in the IL-1β gradient. No significant difference was observed in the rate of neurite extension between IL-1β and controls. We electroporated specific siRNA constructs against IL-1R1 mRNA into cortical progenitors at embryonic day 16 and examined the position and distribution of transfected cells in the somatosensory cortex at postnatal day 5. We found that neurons transfected with IL-1R1-siRNA displayed a severe retardation in radial migration, with about 83% of total cells unable to arrive at the upper cortical layers.ConclusionsOur study suggests an essential contribution of IL-1β to neuronal migration during brain development, which provides a basis to understand the physiological roles of IL-1β in the developing brain and could have significant implications for the prevention of some neurodevelopment disorders due to abnormal neuronal migration.

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Section: Resultsmentioning

confidence: 99%

Interleukin-1 beta guides the migration of cortical neurons

Zhang

et al. 2014

J Neuroinflammation

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“…Using the culturing system and imaging described above we set up a localized microperfusion system to locally deliver NGF to distal axons. The perfusion system was set up as described in Guijarro et al (2012) with modifications. Glass electrodes were pulled to have tips in the 3–5 μm range and filled with NGF containing medium or control medium just prior to use.…”

Section: Methodsmentioning

confidence: 99%

Neurotrophins induce fission of mitochondria along embryonic sensory axons

Armijo-Weingart

Ketschek

Sainath

et al. 2019

eLife

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Neurotrophins are growth factors that have a multitude of roles in the nervous system. We report that neurotrophins induce the fission of mitochondria along embryonic chick sensory axons driven by combined PI3K and Mek-Erk signaling. Following an initial burst of fission, a new steady state of neurotrophin-dependent mitochondria length is established. Mek-Erk controls the activity of the fission mediator Drp1 GTPase, while PI3K may contribute to the actin-dependent aspect of fission. Drp1-mediated fission is required for nerve growth factor (NGF)-induced collateral branching in vitro and expression of dominant negative Drp1 impairs the branching of axons in the developing spinal cord in vivo. Fission is also required for NGF-induced mitochondria-dependent intra-axonal translation of the actin regulatory protein cortactin, a previously determined component of NGF-induced branching. Collectively, these observations unveil a novel biological function of neurotrophins; the regulation of mitochondrial fission and steady state mitochondrial length and density in axons.

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Culturing of Cerebellar Granule Cells to Study Neuronal Migration: Gradient and Local Perfusion Assays

Cited by 2 publications

References 23 publications

Interleukin-1 beta guides the migration of cortical neurons

Interleukin-1 beta guides the migration of cortical neurons

Neurotrophins induce fission of mitochondria along embryonic sensory axons

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